Gram positive

Mol Microbiol, 2001 Aug, 41(3), 665 - 73
Identification of a cell wall channel of Streptomyces griseus: the channel contains a binding site for streptomycin; Kim BH et al.; Cells of the Gram-positive actinomycete Streptomyces griseus were disrupted and the cell envelope was subjected to sucrose step-gradient centrifugation . The different fractions were analysed for NADH-oxidase activity and the formation of ion-permeable channels in lipid bilayers . Highest channel-forming activity and highest NADH-oxidase activity were found in different fractions . The cell wall fraction contained an ion-permeable channel with a single-channel conductance of 850 pS in 1 M KCl . The channel-forming protein, with an apparent molecular mass of 28 kDa, was purified to homogeneity using fast protein liquid chromatography after the extraction of whole cells with detergent . Single-channel experiments suggest that the cell wall channel is wide and water-filled . Titration experiments with streptomycin produced by S . griseus suggested that the cell wall channel binds this antibiotic with a half saturation constant of about 6 mM in 1 M KCl . The binding of streptomycin was found to be ionic strength dependent and the half saturation constant decreased to 60 microM at 0.1 M KCl . The results indicate that the 28 kDa protein represents the hydrophilic pathway through the cell wall of the Gram-positive bacterium S . griseus.

Shock, 2001 Sep, 16(3), 178 - 82
Peptidoglycan primes for LPS-induced release of proinflammatory cytokines in whole human blood; Wang JE et al.; The pathophysiological mechanisms involved in mixed bacterial infections caused by gram-positive and gram-negative bacteria are largely unknown . The present study examines the potential interaction between lipopolysaccharide (LPS) and peptidoglycan (PepG) in the induction of the sepsis-associated cytokines tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), and IL-10 in whole human blood . Plasma values of these cytokines were measured by enzyme immunoassays and a TNF bioassay . Co-administration of PepG (10 microg/mL) or muramyl dipeptide (MDP, 1 microg/mL) with LPS (10 ng/mL) caused significantly elevated values of TNF-alpha and IL-6 in the blood that could not be obtained by the sum of the values obtained by each stimulant alone, or by 3-fold higher doses of either bacterial component alone . This phenomenon was observed 1 h after stimulation, throughout the experimental period (24 h), and with different doses of LPS and PepG . In contrast, the release of IL-10 was not influenced by the co-administration of PepG or MDP with LPS . The TNF-alpha release induced by co-administration of LPS and PepG was abrogated after pretreatment with a monoclonal antibody against CD14 (18D11) . Addition of PepG or MDP to whole blood caused a 2-fold increase in the surface expression of CD14 on monocytes, as measured by flow cytometry . In contrast, LPS caused decreased expression of this receptor . Our data suggest that PepG and MDP primes human whole blood leukocytes for LPS-induced release of proinflammatory cytokines . We speculate that synergy between PepG and LPS may contribute to the pathogenesis in sepsis caused by mixed bacterial infections.

Clin Diagn Lab Immunol, 2001 Sep, 8(5), 972 - 9
Lipoteichoic acid inhibits interleukin-2 (IL-2) function by direct binding to IL-2; Plitnick LM et al.; Lipoteichoic acid (LTA) is associated with the cell envelope of most gram-positive bacteria . Although previously thought to act mainly as a virulence factor by virtue of its adhesive nature, evidence is now provided that LTA can also suppress the function of interleukin-2 (IL-2), an autocrine growth factor for T cells . LTA from four separate bacterial strains lowered the levels of detectable IL-2 during a peripheral blood mononuclear cell response to the antigen tetanus toxoid (TT) . T-cell proliferation in response to TT was similarly inhibited by LTA . In contrast, levels of detectable gamma interferon increased . In addition, LTA inhibited IL-2 detection by enzyme-linked immunosorbent assay (ELISA) and blocked the proliferative response of an IL-2-dependent T-cell line to soluble IL-2 . Further studies using ELISA demonstrated that LTA blocks IL-2 detection and function by binding directly to IL-2 . Flow cytometric analysis revealed that IL-2 binding to T cells is inhibited in the presence of purified LTA but not LTA plus anti-LTA monoclonal antibody . In summary, these studies demonstrate a novel effect of LTA on the immune response through direct binding to IL-2 and inhibition of IL-2 function . Importantly, gram-positive organisms from which LTA is obtained not only play an important role in the pathology of diseases such as bacterial endocarditis, septic shock, acute respiratory distress syndrome, and multiple organ failure but also comprise a significant portion of commensal populations within the human host . Inhibition of IL-2 function by LTA may represent yet another mechanism by which gram-positive bacteria dampen the host immune response and facilitate survival . Thus, LTA provides a potential target for therapeutic intervention when gram-positive organisms are involved.

J Endotoxin Res, 2000, 6(5), 401 - 5
Role of MD-2 in TLR2- and TLR4-mediated recognition of Gram-negative and Gram-positive bacteria and activation of chemokine genes; Dziarski R et al.; MD-2 is associated with TLR4 on the cell surface and enables TLR4 to respond to LPS . TLR2 without MD-2 does not respond to pure protein-free endotoxic LPS, ReLPS, and lipid A . MD-2 enables TLR2 to respond to non-activating LPS, ReLPS, and lipid A, and enhances TLR2-mediated responses to Gram-negative and Gram-positive bacteria, protein-containing LPS, peptidoglycan, and lipoteichoic acid . MD-2 enables TLR4 to respond to a wide variety of endotoxic LPS partial structures, Gram-negative bacteria, and Gram-positive lipoteichoic acid, but not to Gram-positive bacteria, peptidoglycan, and lipopeptide . MD-2 physically associates with both TLR4 and TLR2, but the association with TLR2 is weaker than with TLR4 . Also, MD-2 and TLR2 and TLR4 enhance each other's expression . The highest induced genes in human monocytes stimulated with Gram-positive and Gram-negative bacterial cell wall components are chemokine genes, and IL-8 is the highest induced chemokine . Both Gram-positive and Gram-negative bacteria activate TLR2-->MyD88-->IRAK-->TRAF-->NIK-->IKK-->NF-->kappaB signal transduction pathway that induces transcription of the IL-8 gene . Therefore, TLR2 is a functional receptor for both Gram-positive and Gram-negative bacteria and it induces activation of IL-8.

J Endotoxin Res, 2000, 6(5), 383 - 7
The role of Toll-like receptors and MyD88 in innate immune responses; Akira S et al.; Toll-like receptors (TLRs) are phylogenetically conserved receptors that recognize pathogen associated molecular patterns (PAMPS) . We previously generated mice lacking TLR2 and TLR4 and showed the differential role of TLR2 and TLR4 in microbial recognition . TLR4 functions as the transmembrane component of the lipopolysaccharide (LPS) receptor, while TLR2 recognizes peptidoglycan from Gram-positive bacteria and lipoprotein . We also generated mice lacking MyD88, an adaptor involved in IL-1R/TLR signalings . The responses to a variety of bacterial components were completely abrogated in MyD88-deficient cells . However, unlike the signaling mediated by other bacterial components such as lipoprotein and bacterial DNA, activation of NF-kappaB and MAP kinases was induced in response to LPS even in the absence of MyD88, which indicates the existence of a MyD88-independent pathway . We have recently found that the MyD88-independent pathway is involved in LPS-induced maturation of dendritic cells (DCs).

Pediatr Res, 2001 Sep, 50(3), 315 - 21
Toll-like receptors as sensors of pathogens; Hallman M et al.; Initial recognition of microbes, as they enter the body, is based on germ line-encoded pattern recognition receptors that selectively bind to essential components of pathogens . This allows the body to respond immediately to the microbial invasion before the development of active immunity . The signal-transducing receptors that trigger the acute inflammatory cascade have been elusive until very recently . On the basis of their genetic similarity to the Toll signaling pathway in Drosophila, mammalian Toll-like receptors (TLRs) have been identified . By now, nine transmembrane proteins in the TLR family have been described . Mammalian TLR4 is the signal-transducing receptor activated by the bacterial lipopolysaccharide . The activation of TLR4 leads to DNA binding of the transcription factor NF-kappaB, resulting in activation of the inflammatory cascade . Activation of other TLRs is likely to have similar consequences . TLR2 mediates the host response to Gram-positive bacteria and yeast . TLR1 and TLR6 may participate in the activation of macrophages by Gram-positive bacteria, whereas TLR9 appears to respond to a specific sequence of bacterial DNA . The TLRs that control the onset of an acute inflammatory response are critical antecedents for the development of adaptive acquired immunity . Genetic and developmental variation in the expression of microbial pattern recognition receptors may affect the individual's predisposition to infections in childhood and may contribute to susceptibility to severe neonatal inflammatory diseases, allergies, and autoimmune diseases.

Hypertension, 2001 Aug, 38(2), 177 - 82
Downregulation of angiotensin II type 1 receptors during sepsis; Bucher M et al.; Our study aimed to characterize the mechanisms underlying the attenuated cardiovascular responsiveness toward the renin-angiotensin system during sepsis . For this purpose, we determined the effects of experimental Gram-negative and Gram-positive sepsis in rats . We found that sepsis led to a ubiquitous upregulation of NO synthase isoform II expression and to pronounced hypotension . Despite increased plasma renin activity and plasma angiotensin (Ang) II levels, plasma aldosterone concentrations were normal, and the blood pressure response to exogenous Ang II was markedly diminished in septic rats . Mimicking the fall of blood pressure during sepsis by short-term infusion of the NO donor sodium nitroprusside in normal rats did not alter their blood pressure response to exogenous Ang II . Therefore, we considered the possibility of an altered expression of Ang II receptors during sepsis . It turned out that Ang II type 1 receptor expression was markedly downregulated in all organs of septic rats . Further in vitro studies with rat renal mesangial cells showed that NO and a combination of proinflammatory cytokines (interleukin-1beta, tumor necrosis factor-alpha, and interferon-gamma) downregulated Ang II type 1 receptor expression in a synergistic fashion . In summary, our data suggest that sepsis causes a systemic downregulation of Ang II type 1 receptors that is likely mediated by proinflammatory cytokines and NO . We suggest that this downregulation of Ang II type 1 receptors is the main reason for the attenuated responsiveness of blood pressure and of aldosterone formation to Ang II and, therefore, contributes to the characteristic septic shock.

Curr Opin Ophthalmol, 2001 Aug, 12(4), 282 - 7
Keratoprosthesis: an update; Khan B et al.; Porous polytetrafluoroethylene and polyurethane skirt materials, as well as copolymers of poly (2-hydroxyethyl methacrylate), have shown promise in approaching the goal of a "biointegratable" keratoprosthesis . A novel fixation method that uses scleral haptics also has been introduced to increase stability . An all silicone device that can be sewn into position has been used successfully, temporarily, during vitreoretinal procedures . The prognosis of keratoprosthesis (KPro) procedures depends on the preoperative diagnosis: graft failure-noncicatrizing disease>ocular cicatricial pemphigoid>chemical burns>Stevens-Johnson syndrome . The likelihood of endophthalmitis after KPro surgery follows this scheme . Causative organisms tend to be gram-positive . Modified vitreoretinal techniques also have been developed, allowing successful treatment of posterior segment complications . The science of keratoprosthesis is advancing more rapidly than in previous years . However, use of KPro to address complicated corneal blindness worldwide remains limited . The authors conducted an English language literature review from January 1, 2000 to April 1, 2001 and describe advances of note in the field of keratoprosthesis design, materials, and medical and surgical management.

FEMS Microbiol Lett, 2001 Aug 7, 202(1), 1 - 7
Triclosan: a widely used biocide and its link to antibiotics; Schweizer HP; Triclosan is the active ingredient in a multitude of health care and consumer products with germicidal properties, which have flooded the market in recent years in response to the public's fear of communicable bacteria . Although originally thought to kill bacteria by attacking multiple cellular targets, triclosan was recently shown to target a specific bacterial fatty acid biosynthetic enzyme, enoyl-{acyl-carrier protein} reductase, in Gram-negative and Gram-positive bacteria, as well as in the Mycobacteria . Triclosan resistance mechanisms include target mutations, increased target expression, active efflux from the cell, and enzymatic inactivation/degradation . These are the same types of mechanisms involved in antibiotic resistance and some of them account for the observed cross-resistance with antibiotics in laboratory isolates . Therefore, there is a link between triclosan and antibiotics, and the widespread use of triclosan-containing antiseptics and disinfectants may indeed aid in development of microbial resistance, in particular cross-resistance to antibiotics.

Acta Odontol Scand, 2001 Jun, 59(3), 124 - 30
Toll-like receptors and their signaling mechanism in innate immunity; Kaisho T et al.; In Drosophila the Toll family, a group of transmembrane proteins, plays crucial roles in the host defense against invading pathogens . Mammalian species also conserve this system as the Toll-like receptor (TLR) family, which includes more than 10 members that have been identified so far . Both the Toll and TLR families recognize various kinds of microorganisms through pathogen-associated molecular patterns . Mammalian TLRs are expressed on macrophages and dendritic cells and mediate the signal for cytokine release or upregulation of costimulatory molecules . These activities cooperatively generate host defense mechanisms . Recently, gene targeting experiments, including ours, have contributed much to clarifying not only the function but also the signaling mechanism of TLRs . TLR2 is essential for recognizing lipopeptides and lipoproteins from several microorganisms and also peptidoglycans derived from gram-positive bacteria . TLR4 recognizes lipopolysaccharides and lipoteichoic acids from gram-negative and- positive bacteria, respectively . Furthermore, TLR9 is critical for recognizing bacterial DNAs . Thus, TLRs distinguish various immunostimulatory molecular patterns . Although TLR9 can produce similar biological responses, studies with mutant mice lacking a TLR-associating protein, MyD88, showed that TLR signaling is differentially regulated among TLR family members . Here, we describe recent progress in elucidating the function and signaling mechanisms of the TLR family.

Folia Microbiol (Praha), 2001, 46(1), 71 - 2
Restriction and modification systems of ruminal bacteria; Pristas P et al.; A high frequency of type II restriction endonuclease activities was detected in Selenomonas ruminantium but not in other rumen bacteria tested . Eight different restriction endonucleases were characterized in 17 strains coming from genetically homogeneous local population . Chromosomal DNA isolated from S . ruminantium strains was found to be refractory to cleavage by various restriction enzymes, implying the presence of methylase activities additional to those required for protection against the cellular endonucleases . The presence of Dam methylation was detected in S . ruminantium strains as well as in several other species belonging to the Sporomusa subbranch of low G + C Gram-positive bacteria (Megasphaera elsdenii, Mitsuokella multiacidus).

Folia Microbiol (Praha), 2001, 46(1), 45 - 8
Phenotypic and genetic data supporting reclassification of Butyrivibrio fibrisolvens isolates; Kopecny J et al.; Among 55 Butyrivibrio fibrisolvens strains five ribotypes of B . fibrisolvens were described on the basis of RFLP profiles of 16S rDNA regions obtained with restriction endonuclease HaeIII . In the phylogenetic tree, these ribotypes were located in the XIVa cluster of Gram-positive bacteria . Phenotypic differences of selected ribotype groups became the basis for further reclassification of B . fibrisolvens.

Transfus Clin Biol, 2001 Jun, 8(3), 278 - 81
Mechanisms of severe transfusion reactions; Kopko PM et al.; Serious adverse effects of transfusion may be immunologically or non-immunologically mediated . Currently, bacterial contamination of blood products, particularly platelets, is one of the most significant causes of transfusion-related morbidity and mortality . Septic transfusion reactions can present with clinical symptoms similar to immune-mediated hemolytic transfusion reactions and transfusion-related acute lung injury . Extremely high fever and/or gastrointestinal symptoms, in a transfusion recipient, may be indicative of sepsis . The diagnosis is based upon culturing the same organism from both the patient and the transfused blood component . Numerous organisms have been implicated as the cause of septic transfusion reactions . Due to different storage conditions, gram negative organisms are more often isolated from red blood cell components; gram positive organisms are more often isolated from platelets . Prevention of septic transfusion reactions is primarily dependent on an adequate donor history and meticulous preparation of the donor phlebotomy site . Visual inspection of blood components prior to transfusion is also vital to preventing these reactions . Several methods of detection of bacterial contamination and inactivation of pathogens are currently under active investigation.

Drug Resist Updat, 2000 Jun, 3(3), 178 - 189
beta -Lactamases: which ones are clinically important?
Rice LB, Bonomo RA.
The introduction of a large array of beta-lactam antibiotics has spawned the emergence of an even larger variety of beta-lactamases designed to confer resistance to these agents . beta-lactamases are produced by both gram-positive and gram-negative bacteria, but their clinical importance is far greater among the gram-negatives . The virtual explosion in our knowledge about the variety of these enzymes can often create confusion and frustration among those not well versed in the field . In this paper, we attempt to focus the discussion of beta-lactamases on those enzymes that are of the greatest clinical importance, the Ambler Class A and C enzymes . We also discuss the growing importance of the Ambler Class B metallo beta-lactamases, which hydrolyze carbapenems and are increasing in prevalence in areas of significant carbapenem usage .

Nippon Yakurigaku Zasshi, 2001 Jul, 118(1), 5 - 14
{Induction of histidine decarboxylase in inflammation and immune responses}; Endo Y; Histamine is a classical, but still interesting inflammatory mediator . Many people have long believed that histamine is derived from mast cells or basophils alone . However, the histamine-forming enzyme, histidine decarboxylase (HDC), is induced in a variety of tissues in response (i) to gram-positive and gram-negative bacterial components (lipopolysaccharides, peptidoglycan, and enterotoxin A) and (ii) to various cytokines (IL-1, IL-3, IL-12, IL-18, TNF, G-CSF, and GM-CSF) . HDC is induced even in mast-cell-deficient mice . The histamine newly formed via the induction of HDC is released immediately and may be involved in a variety of immune responses . Reviewing our work and that of Schayer and Kahlson, the pioneers in this field, lead us to the conclusion that nowadays we need to understand that histamine can be produced via the induction of HDC by a mechanism coupled with the cytokine network . We call this histamine "neohistamine", to distinguish it from the classical histamine derived from mast cells or basophils . Neohistamine is involved in physiological reactions, inflammation, immune responses and a variety of diseases such as periodontitis, muscle fatigue (or temporomandibular disorders), stress- or drug-induced gastric ulcers, rheumatoid arthritis, complications in diabetes, hepatitis, allograft rejection, allergic reactions, tumor growth, and inflammatory side effects of aminobisphosphonates.

Clin Exp Rheumatol, 2001 Jul-Aug, 19(4), 459 - 62
Widespread nocardiosis in two patients with Behçet's disease; Korkmaz C et al.; Nocardia spp., a group of gram-positive variably acid-fast aerobic bacteria, are opportunistic pathogens in immunocompromised hosts . We here-within describe 2 cases of widespread nocardiosis in patients with Behcet's disease . In addition to endogen endophthalmitis in case 1, both cases developed lung, brain and skin involvement . Despite brain involvement, the prognosis was good, although vision was completely lost in case 1, which was directly attributable to a delay in both diagnosis and treatment.

Int J Syst Evol Microbiol, 2001 Jul, 51(Pt 4), 1401 - 3
Facklamia miroungae sp . nov., from a juvenile southern elephant seal (Mirounga leonina); Hoyles L et al.; An unusual gram-positive, catalase-negative, facultatively anaerobic, coccus-shaped organism that originated from a juvenile elephant seal was characterized by phenotypic and molecular taxonomic methods . Comparative 16S rRNA gene sequencing showed that the unknown coccus represents a new subline within the genus Facklamia . The unknown strain was readily distinguishable from all currently recognized species of the genus Facklamia (Facklamia hominis, Facklamia languida, Facklamia ignava, Facklamia sourekii and Facklamia tabacinasalis) by biochemical tests and electrophoretic analysis of whole-cell proteins . Based on phylogenetic and phenotypic evidence, it is proposed that the unknown bacterium be classified as Facklamia miroungae sp . nov . The type strain of F . miroungae is CCUG 42728T (= CIP 106764T) . F . miroungae is the first member of the genus Facklamia to be isolated from an animal other than man.

Int J Syst Evol Microbiol, 2001 Jul, 51(Pt 4), 1245 - 56
Alkaliphilus transvaalensis gen . nov., sp . nov., an extremely alkaliphilic bacterium isolated from a deep South African gold mine; Takai K et al.; A novel extreme alkaliphile was isolated from a mine water containment dam at 3.2 km below land surface in an ultra-deep gold mine near Carletonville, South Africa . The cells of this bacterium were straight to slightly curved rods, motile by flagella and formed endospores . Growth was observed over the temperature range 20-50 degrees C (optimum 40 degrees C; 45 min doubling time) and pH range 8.5-12.5 (optimum pH 10.0) . The novel isolate, one of the most alkaliphilic micro-organisms yet described, was a strictly anaerobic chemo-organotroph capable of utilizing proteinaceous substrates such as yeast extract, peptone, tryptone and casein . Elemental sulfur, thiosulfate or fumarate, when included as accessory electron acceptors, improved growth . The G+C content of genomic DNA was 36.4 mol % . Phylogenetic analysis based on the 16S rDNA sequence indicated that the isolate is a member of cluster XI within the low G+C gram-positive bacteria, but only distantly related to previously described members . On the basis of physiological and molecular properties, the isolate represents a novel species, for which the name Alkaliphilus transvaalensis gen . nov., sp . nov . is proposed (type strain SAGM1T = JCM 10712T = ATCC 700919T) . The mechanism of generation of the highly alkaline microbial habitat and the possible source of the alkaliphile are discussed.

Clin Infect Dis, 2001 Sep 1, 33 Suppl 2, S84 - 93
Clinical presentations of soft-tissue infections and surgical site infections; Nichols RL et al.; Skin and soft-tissue infections that usually follow minor traumatic events or surgical procedures are caused by a wide spectrum of bacteria . Less frequently, the infections occur spontaneously, which often is clinically confusing and leads to delays in diagnosis . Most of the infections are self-limited and easily treated with local measures and/or antibiotics . Others are life-threatening, requiring prompt diagnosis and aggressive surgical debridement in addition to the wise choice of antibiotic agents to limit tissue loss and preserve life . Many survivors experience critical tissue losses that may require changes in lifestyle as well as major reconstructive cosmetic surgery . Involvement of antibiotic-resistant gram-positive microorganisms in these infections only increases the difficulty of their treatment and may have a significant influence on the ultimate outcome.

J Ethnopharmacol, 2001 Sep, 77(1), 5 - 9
An ethnobotanical study of plants used for the treatment of sexually transmitted diseases (njovhera) in Guruve District, Zimbabwe; Kambizi L et al.; The use of medicinal plants in Guruve District, Zimbabwe, contributes significantly to primary health care of the people of the area . This paper presents findings of a survey of plants used for the treatment of sexually transmitted diseases (STDs) in the district . Ethnobotanical information obtained from traditional herbalists and other knowledgeable rural dwellers, has revealed 15 plant species belonging to 10 families as medicinal plants used for the treatment of these infections in the area . Six of these are the commonest and most frequently prescribed by the healers . Roots are the most frequently used parts of the plants constituting 53% of preparations while oral administration of extracts is the main method of prescription . Based on the information gathered from the traditional healers, Acacia nilotica (L.) Willd . ex Delile, Cassia abbreviata Oliv . Dichrostachys cinerea Wight and Arn, Solanum incanum L., Vernonia amygdalina Del . and Zanha africana (Radlk) Excell . are the most frequently used plants for the treatment of STDs . The methanol extracts of Cassia abbreviata, Zanha africana and Acacia nilotica showed significant inhibition against Gram-positive and Gram-negative bacteria, while acetone extracts of these plants inhibited most of the species . Generally the water extracts show less activity than acetone and methanol extracts.

Farmaco, 2001 May-Jul, 56(5-7), 447 - 9
Effect of the exposure to gentamicin and diltiazem on the permeability of model membranes; Trombetta D et al.; Preliminary observations showed that the calcium-antagonist diltiazem enhances the 'in vitro' bactericidal action of the aminoglycoside gentamicin, especially against Gram-positive bacteria . To verify if a non-specific interaction of these two drugs with biomembranes may play a role in their synergic effect on bacterial cells, we have studied the effect of exposure to gentamicin, in the absence or presence of diltiazem, on the release of carboxyfluorescein (CF) trapped in phosphatidylcholine (PC) unilamellar vesicles (LUVs) used as model membranes . A significant leakage of trapped CF from PC LUVs was registered when liposomes were treated with gentamicin and diltiazem together, employed at doses (50 and 100 microg/ml, respectively) unable to affect CF release if applied alone; the combined effect of gentamicin and diltiazem was synergic and not cumulative . The present findings demonstrate that the simultaneous exposure to gentamicin and diltiazem may induce significant alterations in the permeability of phospholipid membranes and, so, very likely, in functional properties of bacterial membranes, targets of their action.

J Antibiot (Tokyo), 2001 May, 54(5), 441 - 7
TPU-0031-A and B, new antibiotics of the novobiocin group produced by Streptomyces sp . TP-A0556; Sasaki T et al.; Two novel antibiotics, TPU-0031-A and B, were isolated from the culture broth of an actinomycete strain . The producing strain, TP-A0556, was identified as Streptomyces sp . based on the taxonomic study . The new antibiotics were obtained by solvent extraction and chromatographic purification . Spectroscopic analyses showed that TPU-0031-A and B were 7'-demethylnovobiocin and 5"-demethylnovobiocin, respectively . These compounds showed antibiotic activity against Gram-positive and -negative bacteria.

FEMS Microbiol Lett, 2001 Jul 24, 201(2), 151 - 5
Methyl chloride utilising bacteria are ubiquitous in the natural environment; McAnulla C et al.; Enrichment and isolation of methyl chloride utilising bacteria from a variety of pristine terrestrial, freshwater, estuarine and marine environments resulted in the detection of six new methyl chloride utilising Hyphomicrobium strains, strain CMC related to Aminobacter spp . and to two previously isolated methyl halide utilising bacteria CC495 and IMB-1, and a Gram-positive isolate SAC-4 phylogenetically related to Nocardioides spp . All the pristine environments sampled for enrichment resulted in the successful isolation of methyl chloride utilising organisms.

J Vet Intern Med, 2001 Jul-Aug, 15(4), 394 - 400
Ability of hematologic and serum biochemical variables to differentiate gram-negative and gram-positive mastitis in dairy cows; Smith GW et al.; Medical records of 142 dairy cows with clinical mastitis were examined to determine whether hematologic or serum biochemical results could be used to distinguish between mastitis episodes caused by gram-negative bacteria (n = 78) from those caused by gram-positive bacteria (n = 64) . Signalment, historic information, hematologic and serum biochemical results, milk culture results, and outcome (discharged from hospital or died) were obtained from the medical records . Cows with gram-negative mastitis had significantly (P < .01) lower blood leukocyte, segmented neutrophil, monocyte, and lymphocyte counts and had higher blood hemoglobin concentrations and hematocrits than did cows with gram-positive mastitis . Serum urea nitrogen was the only serum biochemical result associated with pathogen type, and it was higher in cows with gram-negative mastitis than in those with gram-positive mastitis . Mortality rate (25% overall) did not differ between groups . Logistic regression indicated that routine hematologic analysis (segmented neutrophil count, monocyte count, and hemoglobin concentration) was an accurate predictor of gram-negative mastitis, with a sensitivity of .93, a specificity of .89, and an overall accuracy of 91% . The values for sensitivity and specificity were higher than those previously reported for clinical tests differentiating mastitis episodes caused by gram-negative bacteria from those caused by gram-positive bacteria . Our results indicate that routine hematologic analysis is useful for predicting pathogen type in dairy cows with clinical mastitis, thereby facilitating treatment decisions.

J Biol Chem, 2001 Sep 14, 276(37), 34686 - 94 Epub 2001 Jul 18.
Peptidoglycan recognition proteins: a novel family of four human innate immunity pattern recognition molecules; Liu C et al.; The innate immune system recognizes microorganisms through a series of pattern recognition receptors that are highly conserved in evolution . Insects have a family of 12 peptidoglycan recognition proteins (PGRPs) that recognize peptidoglycan, a ubiquitous component of bacterial cell walls . We report cloning of three novel human PGRPs (PGRP-L, PGRP-Ialpha, and PGRP-Ibeta) that together with the previously cloned PGRP-S, define a new family of human pattern recognition molecules . PGRP-L, PGRP-Ialpha, and PGRP-Ibeta have 576, 341, and 373 amino acids coded by five, seven, and eight exons on chromosomes 19 and 1, and they all have two predicted transmembrane domains . All mammalian and insect PGRPs have at least three highly conserved C-terminal PGRP domains located either in the extracellular or in the cytoplasmic (or in both) portions of the molecules . PGRP-L is expressed in liver, PGRP-Ialpha and PGRP-Ibeta in esophagus (and to a lesser extent in tonsils and thymus), and PGRP-S in bone marrow (and to a lesser extent in neutrophils and fetal liver) . All four human PGRPs bind peptidoglycan and Gram-positive bacteria . Thus, these PGRPs may play a role in recognition of bacteria in these organs.

J Fr Ophtalmol, 2001 Jun, 24(6), 597 - 602
{Infectious keratitis after penetrating keratoplasty}; Tixier J et al.; PURPOSE: To investigate the prevalence of microbial keratitis, predisposing risk factors, the spectrum of pathogens and the prognosis for graft survival and visual outcome in patients who developed microbial keratitis following penetrating keratoplasty (PK) . MATERIAL: and methods: We reviewed 16 cases (15 patients) of microbial keratitis after PK . In all cases, corneal scrapings were obtained and microbiologically analyzed . Efficacy of treatment was evaluated by anatomical (clarity of graft) and visual recovery . RESULTS: Principal indications for PK were pseudophakic bullous keratopathy (50%) and microbial keratitis in the previous graft (25%) . Sixty-three per cent of infections occurred within 1 year of PK . Principal predisposing risk factors were suture-related problems (44%) and microbial keratitis in the previous graft (25%) . All of the scrapings were positive according to the microbiological evaluation with gram-positive cocci (64%), gram-positive rods (12%), fungi (18%), and Acanthamoeba (6%) . We found 1 case of polymicrobial infection . Best visual and anatomical results were observed in nonadvanced cases and/or these treated early . After medical and surgical treatments, 8 patients (50%) had a clear graft and 10 patients (63%) had visual acuity less than 20/200 . CONCLUSION: Postoperative control of risk factors and early recognition of infectious complications may decrease the incidence of severe microbial keratitis after PK.

Brain, 2001 Aug, 124(Pt 8), 1544 - 54
Bacterial peptidoglycan and immune reactivity in the central nervous system in multiple sclerosis; Schrijver IA et al.; Multiple sclerosis is believed to result from a CD4+ T-cell response against myelin antigens . Peptidoglycan, a major component of the Gram-positive bacterial cell wall, is a functional lipopolysaccharide analogue with potent proinflammatory properties and is conceivably a mediator of sterile inflammation . Here we demonstrate that peptidoglycan is present within antigen-presenting cells in the brain of multiple sclerosis patients . These cells have macrophage and dendritic cell characteristics, and are immunocompetent as evidenced by co-expression of inflammatory cytokines and co-stimulatory molecules . In addition, intrathecal plasma cells specific for peptidoglycan are present in multiple sclerosis brain tissue, and antibodies binding peptidoglycan are present in CSF during active disease . Peptidoglycan may thus contribute to T- and B-cell activity during brain inflammation without a requirement for local bacterial replication.

FEMS Microbiol Ecol, 2001 Jul, 36(2-3), 211 - 222
Isolation and characterisation of new Gram-negative and Gram-positive atrazine degrading bacteria from different French soils; Rousseaux S et al.; The capacity of 12 soils to degrade atrazine was studied in laboratory incubations using radiolabelled atrazine . Eight soils showed enhanced degradation of this compound . Twenty-five bacterial strains able to degrade atrazine were isolated by an enrichment method from 10 of these soils . These soils were chosen for their wide range of physico-chemical characteristics . Their history of treatment with atrazine was also variable . The genetic diversity of atrazine degraders was determined by amplified ribosomal restriction analysis (ARDRA) of the 16S rDNA gene with three restriction endonucleases . The 25 bacterial strains were grouped into five ARDRA types . By sequencing and aligning the 16S rDNA genes, the isolates were shown to belong to the Gram-negative species Chelatobacter heintzii, Aminobacter aminovorans, Stenotrophomonas maltophilia and to the Gram-positive genus Arthrobacter crystallopoietes . These species were not described previously as being capable of atrazine degradation . Most Gram-negative bacteria could mineralise (14)C ring labelled atrazine and carried the atzA, atzB, atzC and trzD genes . Gram-positive strains could convert atrazine to cyanuric acid and carried only the atzB and atzC genes . In this study, we describe the atrazine degradation capacities and corresponding genes in bacterial species that were not known as atrazine degraders . We report for the first time the occurrence of the trzD gene in these atrazine-mineralising bacteria and we demonstrate the potential use of colony hybridisation to isolate bacteria involved in atrazine degradation.

J Membr Biol, 2001 Jul 15, 182(2), 147 - 57
Study of the properties of a channel-forming protein of the cell wall of the gram-positive bacterium Mycobacterium phlei; Riess FG et al.; The gram-positive bacterium Mycobacterium phlei was treated with detergents . Reconstitution experiments using lipid bilayers suggested that the detergent extracts contain a channel forming protein . The protein was purified to homogeneity by preparative SDS-PAGE and identified as a protein with an apparent molecular mass of about 135 kDa . The channel-forming unit dissociated into subunits with a molecular mass of about 22 kDa when it was boiled in 80% dimethylsulfoxid (DMSO) . The channel has on average a single channel conductance of 4.5 nS in 1 m KCl and is highly voltage-dependent in an asymmetric fashion when the protein is added to only one side of the membrane . Zero-current membrane potential measurements with different salts implied that the channel is highly cation-selective because of negative point charges in or near the channel mouth . Analysis of the single-channel conductance as a function of the hydrated cation radii using the Renkin correction factor and the effect of the negative point charges on the single-channel conductance suggest that the diameter of the cell wall channel is about 1.8 to 2.0 nm . The channel properties were compared with those of other members of the mycolata and suggest that these channels share common features . Southern blots demonstrated that the chromosome of M . phlei and other mycolata tested contain homologous sequences to mspA (gene of the cell wall porin of Mycobacterium smegmatis).

Vet Surg, 2001 Jul-Aug, 30(4), 366 - 73
Comparison of two methods for presurgical disinfection of the equine hoof; Hennig GE et al.; OBJECTIVES: To determine for equine hooves the normal resident aerobic bacterial population and the efficacy of 2 methods of disinfection . Study Design-Measurement of total bacterial, gram-positive bacterial, and gram-negative bacterial surface populations from the frog, sole, and hoof wall after each step of 2 different preoperative surgical disinfection techniques . ANIMALS: Six adult horses . METHODS: Hoof wall, sole, and frog samples were collected for quantitative bacteriology before, during, and after 2 multistep antiseptic preparation techniques: Method A-6-minute scrub with povidone-iodine soap, followed by 24-hour submersion in povidone-iodine solution-soaked cotton; and Method B-initial removal of superficial layer of hoof capsule before completing Method A disinfection procedures . RESULTS: Removal of the superficial hoof layer, application of the povidone iodine scrub, and completion of the povidone-iodine soak all significantly (P < .0008) decreased total bacterial numbers . Method B had significantly lower bacterial counts than method A at each consecutive step . Final total bacterial counts remained greater than 10(5) bacteria per gram of tissue regardless of preparation method . CONCLUSIONS: The hoof surface hosts a broad spectrum of aerobic gram-positive and -negative bacteria, many of which are potential pathogens . Bacterial numbers can be significantly reduced by removal of the superficial hoof surface, by application of a povidone-iodine scrub, and by use of a 24-hour povidone-iodine soak . However, bacterial populations >10(5) g per tissue persist after these disinfection procedures . CLINICAL RELEVANCE: Regardless of the preparation methods used in this study, bacterial populations capable of inducing wound infection remain on the hoof capsule .

Biotech Histochem, 2001 Jan, 76(1), 15 - 22
Gram staining and lectin binding properties of Myxosporea and Sporozoea; Schachner et al.; The staining method developed by Christian Gram was introduced as a simple and highly selective tool for demonstrating myxosporean and coccidian sporogonic stages . When using standard blood staining procedures for those enigmatic parasites it is sometimes difficult to distinguish them from fish host tissue . They clearly exhibit a partial gram-positive reaction in histological sections, but staining is variable in air dried fish organ imprints . To visualize the gram-negative background of different host tissue components in histological sections, the conventional safranin counterstain of the gram protocol may be modified as follows: after application of 2% crystal violet (basic violet 3) and Lugol's solution, sections are stained with 0.1% nuclear fast red-5% aluminum sulfate and 0.35% aniline blue (acid blue 22) dissolved in saturated aqueous picric acid . Replacement of the gram-specific dye crystal violet with 2% malachite green gave similar results in organ imprints containing myxospores or coccidia, but only in sections containing myxosporea . Staining for 1 min with an aqueous solution of 0.5% malachite green and followed 1 min washing was sufficient for rapidly demonstrating the parasite spores in organ imprints of both myxosores and oocysts . With regard to the role of acid mucopolysaccharides and other carbohydrates in the gram reaction of spores, alcian blue 8GX staining was compared to the binding of FITC-labeled WGA, GS I and GS II . Each lectin was applied at 20 microl/ml PBS, HEPES for 1 hr . Whereas WGA yielded a nonspecific pattern like the alcian blue staining, GS II resulted in a pattern similar to the gram staining results . This binding was weak in untreated specimens, but was significantly enhanced when digested first within trypsin overnight in a humid chamber at 37 degrees C . The binding of GS II to both myxosporidian and coccidian spores suggests that they are both composed of polymers containing N-acetyl-D-glucosamine residues . Furthermore, the results suggest that this hexosamine plays a key role in the gram reaction.

Int J Med Microbiol, 2001 May, 291(2), 107 - 17
Regulation of transcription in Helicobacter pylori: simple systems or complex circuits?
Scarlato V, Delany I, Spohn G, Beier D.
A common strategy used by both Gram-negative and Gram-positive bacterial pathogens is based on the synchronisation of virulence gene expression using a variety of regulatory systems and networks to overcome host defence . During the last decade an exponentially growing number of studies on Helicobacter pylori, a human pathogen associated with diverse stomach diseases, have mainly focussed on the elucidation of mechanisms and functions of virulence factors . A subset of these studies were focussed on the molecular mechanisms regulating gene transcription in H . pylori with the aim of understanding the profound physiological changes that this pathogen, as well as other bacteria, undergoes during infection . Despite the limited number of putative regulatory proteins, as deduced from genome sequence analyses, evidence is accumulating for the existence of new and complex circuits regulating gene transcription and virulence of this bacterium . Here we will focus on the molecular mechanisms used by H . pylori to control gene transcription.

Structure (Camb), 2001 Jun, 9(6), 527 - 37
A structural genomics approach to the study of quorum sensing: crystal structures of three LuxS orthologs; Lewis HA et al.; BACKGROUND: Quorum sensing is the mechanism by which bacteria control gene expression in response to cell density . Two major quorum-sensing systems have been identified, system 1 and system 2, each with a characteristic signaling molecule (autoinducer-1, or AI-1, in the case of system 1, and AI-2 in system 2) . The luxS gene is required for the AI-2 system of quorum sensing . LuxS and AI-2 have been described in both Gram-negative and Gram-positive bacterial species and have been shown to be involved in the expression of virulence genes in several pathogens . RESULTS: The structure of the LuxS protein from three different bacterial species with resolutions ranging from 1.8 A to 2.4 A has been solved using an X-ray crystallographic structural genomics approach . The structure of LuxS reported here is seen to have a new alpha-beta fold . In all structures, an equivalent homodimer is observed . A metal ion identified as zinc was seen bound to a Cys-His-His triad . Methionine was found bound to the protein near the metal and at the dimer interface . CONCLUSIONS: These structures provide support for a hypothesis that explains the in vivo action of LuxS . Specifically, acting as a homodimer, the protein binds a methionine analog, S-ribosylhomocysteine (SRH) . The zinc atom is in position to cleave the ribose ring in a step along the synthesis pathway of AI-2.

Biochemistry, 2001 Jul 10, 40(27), 7964 - 72
Biosynthesis of D-alanyl-lipoteichoic acid: the tertiary structure of apo-D-alanyl carrier protein; Volkman BF et al.; The D-alanylation of lipoteichoic acid (LTA) allows the Gram-positive organism to modulate its surface charge, regulate ligand binding, and control the electromechanical properties of the cell wall . The incorporation of D-alanine into LTA requires the D-alanine:D-alanyl carrier protein ligase (AMP-forming) (Dcl) and the carrier protein (Dcp) . The high-resolution solution structure of the 81-residue (8.9 kDa) Dcp has been determined by multidimensional heteronuclear NMR . An ensemble of 30 structures was calculated using the torsion angle dynamics approach of DYANA . These calculations utilized 3288 NOEs containing 1582 unique nontrivial NOE distance constraints . Superposition of residues 4-81 on the mean structure yields average atomic rmsd values of 0.43 +/- 0.08 and 0.86 +/- 0.09 A for backbone and non-hydrogen atoms, respectively . The solution structure is composed of three alpha-helices in a bundle with additional short 3(10)- and alpha-helices in intervening loops . Comparisons of the three-dimensional structure with the acyl carrier proteins involved in fatty acid, polyketide, and nonribosomal peptide syntheses support the conclusion that Dcp is a homologue in this family . While there is conservation of the three-helix bundle fold, Dcp has a higher enthalpy of unfolding and no apparent divalent metal binding site(s), features that distinguish it from the fatty acid synthase acyl carrier protein of Escherichia coli . This three-dimensional structure also provides insights into the D-alanine ligation site recognized by Dcl, as well as the site which may bind the poly(glycerophosphate) acceptor moiety of membrane-associated LTA.

Turk J Pediatr, 2001 Apr-Jun, 43(2), 105 - 9
Monotherapy with meropenem versus combination therapy with piperacillin plus amikacin as empiric therapy for neutropenic fever in children with lymphoma and solid tumors; Duzova A et al.; The purpose of this study was to compare meropenem monotherapy with combination therapy for empirical treatment of neutropenic fever in children with lymphoma and solid tumors . Ninety episodes of neutropenic fever in children (0.7-16.0; mean age 7.7 years) with solid tumors in a single center were randomized to receive either meropenem (50 mg/kg/dose-maximum 1 g, every 8 hours) or piperacillin (200 mg/kg/dose, every 6 hours) plus amikacin (15 mg/kg daily) . Failure was defined as treatment modification . Non-Hodgkin's lymphoma (NHL) accounted for 62.2 percent of all episodes, and solid tumors (37.8%) for the rest . Blood cultures were positive in 23 percent of all episodes . Sixty-seven percent of all isolated microorganisms stained Gram-positive . Overall success was 70.0 percent (63/90) . The success with meropenem was comparable to that seen with piperacillin plus amikacin: 76.6 versus 64.6 percent (p = 0.25) . The failure rate was 33 percent with Gram-positive culture and 78 percent with Gram-negative or mixed cultures . The solid tumor group had significantly less bacteremia (4/34 versus 17/56; p < 0.05) and treatment failure (3/34 versus 24/56; p < 0.001) than the NHL group . No serious drug-related adverse event was noticed . Meropenem monotherapy was as effective as piperacillin plus amikacin combination in the empirical treatment of neutropenic fever in children with lymphoma and solid tumors.

Am J Kidney Dis, 2001 Jul, 38(1), 127 - 31
Cefepime versus vancomycin plus netilmicin therapy for continuous ambulatory peritoneal dialysis-associated peritonitis; Wong KM et al.; Cefepime is a cephalosporin with a broad spectrum of activity against most gram-positive and gram-negative pathogens . In this study, we attempted to compare the safety and efficacy of cefepime monotherapy against the potentially more toxic combination of vancomycin and netilmicin in the treatment of continuous ambulatory peritoneal dialysis (CAPD)-associated bacterial peritonitis . Eighty-one consecutive CAPD patients who presented with peritonitis from January 1, 1998, to June 30, 2000, were recruited for study . Patients were randomized to be administered either intraperitoneal (IP) cefepime, 1 g once daily (group A), or intravenous vancomycin and netilmicin at conventional doses (group B) for 10 days . Bacterial growth was obtained in 52 episodes (66%), and pathogens identified included gram-positive organisms (30 episodes; 38%), gram-negative organisms (14 episodes; 18%), mixed organisms (2 episodes; 2.5%), and fungus (6 episodes; 8%) . Eight patients were excluded after randomization for various reasons (6 patients, fungal peritonitis; 2 patients, wrong diagnoses) . Because of the relatively low peritonitis rate after the use of a disconnect system, the sample size of this study was relatively small, giving a power of 0.45 . There were no significant differences in primary response rates and cure rates (no relapse >28 days after completion of antibiotic therapy) between both groups of patients (group A versus group B, 82% {32 of 39 patients} versus 85% {29 of 34 patients} and 72% {28 of 39 patients} versus 76% {26 of 34 patients}, respectively; P = not significant) . No significant side effect was encountered in either group . Total peritonitis-related hospitalizations were 84 patient-days (1, 7, 8, 11, 20, and 37 patient-days) and 115 patient-days (3, 6, 9, 14, 21, 21, and 41 patient-days), whereas total costs per patient cure were estimated to be US $1,039 and US $1,371 in groups A and B, respectively . We conclude that once-daily 1-g IP cefepime monotherapy is a simple, safe, and cost-effective alternative to vancomycin and netilmicin therapy in the treatment of CAPD-associated bacterial peritonitis.

Orthopedics, 2001 Jun, 24(6), 587 - 9
Culture results and amputation rates in high-pressure paint gun injuries of the hand; Mirzayan R et al.; High-pressure paint gun injuries have been well described in the literature, and the use of antibiotics is recommended as part of their management . However, there is no scientific evidence to support the use of antibiotics . In addition, the type of paint injected (water- versus oil-based) has never been investigated to determine the extent of morbidity resulting from these injuries . This study examines the organisms cultured in wounds resulting from these injuries and whether the type of paint injected had an influence on amputation rates . Charts of 35 patients with high-pressure paint gun injuries to their hands were reviewed . The amputation rate was 50% with oil-based paints and 0% with water-based paints . Forty-seven percent of wound cultures were positive, with gram-negative bacteria found in 58% of isolates . Our findings support the use of antibiotics, which should cover both gram-positive and gram-negative organisms.

J Mol Biol, 2001 Jul 6, 310(2), 291 - 7
Modular structure, local flexibility and cold-activity of a novel chitobiase from a psychrophilic Antarctic bacterium; Lonhienne T et al.; The gene archb encoding for the cell-bound chitobiase from the Antarctic Gram-positive bacterium Arthrobacter sp . TAD20 was cloned and expressed in Escherichia coli in a soluble form . The mature chitobiase ArChb possesses four functionally independent domains: a catalytic domain stabilized by Ca(2+), a galactose-binding domain and an immunoglobulin-like domain followed by a cell-wall anchorage signal, typical of cell-surface proteins from Gram-positive bacteria . Binding of saccharides was analyzed by differential scanning calorimetry, allowing to distinguish unequivocally the catalytic domain from the galactose-binding domain and to study binding specificities . The results suggest that ArChb could play a role in bacterium attachment to natural hosts . Kinetic parameters of ArChb demonstrate perfect adaptation to catalysis at low temperatures, as shown by a low activation energy associated with unusually low K(m) and high k(cat) values . Thermodependence of these parameters indicates that discrete amino acid substitutions in the catalytic center have optimized the thermodynamic properties of weak interactions involved in substrate binding at low temperatures . Microcalorimetry also reveals that heat-lability, a general trait of psychrophilic enzymes, only affects the active site domain of ArChb .

Appl Environ Microbiol, 2001 Jul, 67(7), 2922 - 6
Photosynthetic and phylogenetic primers for detection of anoxygenic phototrophs in natural environments; Achenbach LA et al.; Primer sets were designed to target specific 16S ribosomal DNA (rDNA) sequences of photosynthetic bacteria, including the green sulfur bacteria, the green nonsulfur bacteria, and the members of the Heliobacteriaceae (a gram-positive phylum) . Due to the phylogenetic diversity of purple sulfur and purple nonsulfur phototrophs, the 16S rDNA gene was not an appropriate target for phylogenetic rDNA primers . Thus, a primer set was designed that targets the pufM gene, encoding the M subunit of the photosynthetic reaction center, which is universally distributed among purple phototrophic bacteria . The pufM primer set amplified DNAs not only from purple sulfur and purple nonsulfur phototrophs but also from Chloroflexus species, which also produce a reaction center like that of the purple bacteria . Although the purple bacterial reaction center structurally resembles green plant photosystem II, the pufM primers did not amplify cyanobacterial DNA, further indicating their specificity for purple anoxyphototrophs . This combination of phylogenetic- and photosynthesis-specific primers covers all groups of known anoxygenic phototrophs and as such shows promise as a molecular tool for the rapid assessment of natural samples in ecological studies of these organisms.

Res Microbiol, 2001 Apr-May, 152(3-4), 231 - 43
A new family of high-affinity ABC manganese and zinc permeases; Claverys JP; Phylogenetic analysis of 47 extracellular putative metal binding receptors (MBRs) belonging to the newly defined cluster suggests the existence of two subclusters . The question of substrate specificity of the corresponding ATP binding cassette (ABC) permeases is discussed, based on data collected from 19 of them concerning their regulation, metal requirement of permease mutants, metal uptake and metal binding . The proposal that the two subclusters correspond to paralogous metal permeases dedicated primarily to manganese and to zinc transport is made . The question of a direct role of MBRs as adhesins of Gram-positive bacteria is then discussed and the importance of metal permeases for cellular processes and host-bacteria interactions is reviewed.

Retina, 2001, 21(3), 210 - 3
Intraocular vancomycin levels after intravitreal injection in post cataract extraction endophthalmitis; Haider SA et al.; PURPOSE: Intravitreal antibiotics are the mainstay of treatment of endophthalmitis following cataract surgery . The purpose of this study is to determine the range of intraocular vancomycin found after intravitreal therapy and assess the optimum time for repeat injections . METHODS: Aqueous and vitreous vancomycin was assayed at the time of reinjection in 14 patients with endophthalmitis showing a poor clinical response after their primary injection . Nine patients received vancomycin 2 mg and another five received vancomycin 1 mg . In six patients the injection was repeated at 48 hours and in seven at 72 hours . Two patients received three injections . RESULTS: Aqueous vancomycin varied from 8.4 to 170 mg/L and the vitreous vancomycin level ranged from 21.2 to 220 mg/L . CONCLUSIONS: In the current study vitreous vancomycin levels were variable, but well within the therapeutic range for sensitive Gram-positive organisms . At times they exceeded the putative retinotoxic levels (100 mg/L) . Higher aqueous levels were found after an injection of 2 mg than after 1 mg . Vancomycin levels were still very high 3 days after injection of 2 mg where results were available . Assay at the time of repeat injection may provide insight into the adequacy of vitreous levels and guide future therapy.

J Dairy Sci, 2001 Jun, 84(6), 1407 - 12
Influence of subclinical mastitis during early lactation on reproductive parameters; Schrick FN et al.; Our objective was to determine the effects of mastitis during early lactation on the reproductive performance of Jersey cows . From 1986 to 1997, quarter foremilk samples were collected every 4 to 8 wk during lactation, at drying off, near calving, and when clinical mastitis was diagnosed and were evaluated microbiologically to identify causative bacteria . Services per conception, days open, and days to first service were obtained from DHIA records on 752 cows . Cows were separated by mastitis type (clinical, n = 186; subclinical, n = 240; control, uninfected or infected after confirmed pregnancy, n = 326) . Cows were reclassified based on the time of clinical or subclinical mastitis as follows: period 1, before first service (n = 374); period 2, between first service and pregnancy (n = 52); and period 3, after confirmed pregnancy or uninfected (control; n = 326) . Milk production did not differ for any group separations . Reproductive performance did not differ between gram-negative or gram-positive mastitis pathogens . Cows with clinical or subclinical mastitis before first service had increased days to first service (77.3+/-2.7 and 74.8+/-2.7 d), days open (110.0+/-6.9 and 107.7+/-6.9 d), and services per conception (2.1+/-0.2 and 2.1+/-0.2) compared with controls (67.8+/-2.2 d, 85.4+/-5.8 d, 1.6+/-0.2; P < 0.05) . Days to first service were not increased in cows with clinical or subclinical mastitis during period 2 (70.6+/-3.3 and 61.2+/-7.8 d) . However, days open (143.6+/-8.5 d) and services per conception (3.0+/-0.2) were increased (P < 0.05) in cows with clinical mastitis during period 2, but not in cows with subclinical mastitis (90.9+/-20.2 d and 2.1+/-0.5) . Cows initially diagnosed subclinical that became clinical during period 2 exhibited increased days to first service (93.9+/-10.1 d), days open (196.0+/-26.2 d), and services per conception (4.3+/-0.7) compared with control animals (P < 0.05) . In conclusion, subclinical mastitis reduced reproductive performance of lactating cows similar to clinical mastitis . Subclinical mastitis followed by clinical mastitis resulted in the most severe loss in reproductive performance.

J Agric Food Chem, 2001 Jun, 49(6), 2830 - 4
Reaction of B . cereus bacteria and peroxidase enzymes under pressures >400 MPa; Prestamo G et al.; It is known that B . cereus (Gram-positive bacteria) and peroxidase enzymes are resistant to pressures of approximately 400 MPa in fruit and vegetable products among others . The aim of the present work is to have knowledge about their behavior when using pressures >400 MPa without other combined treatments . The results showed that B . cereus was not inactivated at pressures of 1000 MPa for 15 min at 20 degrees C . In peroxidase enzymes the results remained similar and the pressure of 1000 MPa for 30 min was not enough to inactivate them . The apple cell structure at these high-pressure levels revealed that it changed and the cells were less cemented . The treated apple presented a translucent aspect, and some fluids migrated from the inside to the outside of the cell.

Curr Biol, 2001 Apr 3, 11(7), R264 - 7
Bacterial pathogenesis: the answer to virulence is in the pore; Gauthier A et al.; A wide variety of Gram-negative bacterial pathogens use a 'type III' protein secretion system to deliver bacterial virulence factors into host cells . Recent results suggest that Gram-positive pathogens may employ similar methods to deliver virulence factors into host cells.

Int J Syst Evol Microbiol, 2001 May, 51(Pt 3), 937 - 44
Proposal of Mycetocola gen . nov . in the family Microbacteriaceae and three new species, Mycetocola saprophilus sp . nov., Mycetocola tolaasinivorans sp . nov . and Mycetocola lacteus sp . nov., isolated from cultivated mushroom, Pleurotus ostreatus; Tsukamoto T et al.; The taxonomic positions of 10 tolaasin-detoxifying bacteria, which were isolated from the cultivated mushroom Pleurotus ostreatus, were investigated . These strains are Gram-positive, obligately aerobic, non-sporulating and irregular rod-shaped bacteria . They have the following characteristics: the major menaquinone is MK-10, the DNA G+C content ranges from 64 to 65 mol%, the diamino acid in the cell wall is lysine and the muramic acid in the peptidoglycan is an acetyl type . The major fatty acids are anteiso-C15:0 and anteiso-C17:0 . On the basis of morphological, physiological and chemotaxonomic characteristics, together with DNA-DNA reassociation values and 16S rRNA gene sequence comparison data, the new genus Mycetocola gen . nov . is proposed for these bacteria in the family Microbacteriaceae and three new species are also proposed: Mycetocola saprophilus sp . nov . (type strain CM-01T = IFO 16274T = MAFF 211324T = NRRL B-24119T), Mycetocola tolaasinivorans sp . nov . (type strain CM-05T = IFO 16277T = MAFF 211325T = NRRL B-24120T) and Mycetocola lacteus sp . nov . (type strain CM-10T = IFO 16278T = MAFF 211326T = NRRL B-24121T) . The type species of the genus is Mycetocola saprophilus sp . nov.

Int J Syst Evol Microbiol, 2001 May, 51(Pt 3), 933 - 6
Nocardia veterana sp . nov., isolated from human bronchial lavage; Gurtler V et al.; A nocardioform bacterium was isolated from the bronchoscopic lavage of a 78-year-old man with a past history of tuberculous pleurisy, who presented with bilateral upper lobe lesions at Austin and Repatriation Medical Centre, Heidelberg, Australia . The strain was aerobic, Gram-positive, produced beige substrate mycelium and scant white aerial mycelium . It showed chemotaxonomic markers which were consistent with the classification of Nocardia: i.e . meso-diaminopimelic acid, N-glycolylmuramic acid, arabinose and galactose as diagnostic sugars; phospholipids phosphatidylinositol mannosides, phosphatidylinositol, phosphatidylethanolamine and diphosphatidylglycerol; a menaquinone with a cyclic isoprene side chain, MK-8(H4cycl.); a fatty acid pattern composed of unbranched saturated and monounsaturated fatty acids with a considerable amount of tuberculostearic acid; and mycolic acids composed of 54-62 carbon atoms with three principal mycolic acids which were mono- and polyunsaturated, showing a chain length C56, C58 and C60 and accounting for over 70% of the entire pattern . The 16S rDNA sequence showed the highest similarity to the type strain of Nocardia vaccinii; the DNA-DNA similarity of the two strains was 31% . These data, together with distinct physiological traits and molecular biological analyses, as well as chemotaxonomic results, led to the conclusion that the novel isolate represents a new species within the genus Nocardia for which the name Nocardia veterana sp . nov . is proposed . The type strain is M157222T (DSM 44445T = NRRL B-24136T).

Chem Biol, 2001 Jun, 8(6), 593 - 611
Structure-based design and in-parallel synthesis of inhibitors of AmpC beta-lactamase; Tondi D et al.; BACKGROUND: Group I beta-lactamases are a major cause of antibiotic resistance to beta-lactams such as penicillins and cephalosporins . These enzymes are only modestly affected by classic beta-lactam-based inhibitors, such as clavulanic acid . Conversely, small arylboronic acids inhibit these enzymes at sub-micromolar concentrations . Structural studies suggest these inhibitors bind to a well-defined cleft in the group I beta-lactamase AmpC; this cleft binds the ubiquitous R1 side chain of beta-lactams . Intriguingly, much of this cleft is left unoccupied by the small arylboronic acids . RESULTS: To investigate if larger boronic acids might take advantage of this cleft, structure-guided in-parallel synthesis was used to explore new inhibitors of AmpC . Twenty-eight derivatives of the lead compound, 3-aminophenylboronic acid, led to an inhibitor with 80-fold better binding (2; K(i) 83 nM) . Molecular docking suggested orientations for this compound in the R1 cleft . Based on the docking results, 12 derivatives of 2 were synthesized, leading to inhibitors with K(i) values of 60 nM and with improved solubility . Several of these inhibitors reversed the resistance of nosocomial Gram-positive bacteria, though they showed little activity against Gram-negative bacteria . The X-ray crystal structure of compound 2 in complex with AmpC was subsequently determined to 2.1 A resolution . The placement of the proximal two-thirds of the inhibitor in the experimental structure corresponds with the docked structure, but a bond rotation leads to a distinctly different placement of the distal part of the inhibitor . In the experimental structure, the inhibitor interacts with conserved residues in the R1 cleft whose role in recognition has not been previously explored . CONCLUSIONS: Combining structure-based design with in-parallel synthesis allowed for the rapid exploration of inhibitor functionality in the R1 cleft of AmpC . The resulting inhibitors differ considerably from beta-lactams but nevertheless inhibit the enzyme well . The crystal structure of 2 (K(i) 83 nM) in complex with AmpC may guide exploration of a highly conserved, largely unexplored cleft, providing a template for further design against AmpC beta-lactamase.

Crit Care Med, 2001 Jun, 29(6), 1101 - 8
Impact of a rotating empiric antibiotic schedule on infectious mortality in an intensive care unit; Raymond DP et al.; OBJECTIVE: The development of antibiotic-resistant bacteria is associated with significant morbidity and mortality in critically ill patients . We postulated that quarterly rotation of empirical antibiotics could decrease infectious complications from resistant organisms in an intensive care unit (ICU) . DESIGN: Prospective cohort study . SETTING: An ICU at a university medical center . SUBJECTS: All patients admitted to the general, transplant, or trauma surgery services who developed pneumonia, peritonitis, or sepsis of unknown origin . INTERVENTIONS: A 2-yr study consisting of 1 yr of nonprotocol-driven antibiotic use and 1 yr of rotating empirical antibiotic assignment . MEASUREMENTS AND MAIN RESULTS: Over 100 variables were recorded for each infectious episode, including patient characteristics (e.g., Acute Physiology and Chronic Health Evaluation {APACHE} II score, age, comorbidities), infection characteristics (e.g., site, organism), treatment characteristics (e.g., antibiotic, treatment duration) and outcome measures (e.g., mortality, length of stay, antibiotic cost) . Of 1456 consecutive admissions to the ICU, 540 episodes of infection were treated . No differences were noted in age, APACHE II score, race, overall antibiotic utilization or duration of therapy between the 2 yrs of study . Outcome analysis revealed significant reductions in the incidence of antibiotic-resistant Gram-positive coccal infections (7.8 infections/100 admissions vs . 14.6 infections/100 admissions, p <.0001), antibiotic-resistant Gram-negative bacillary infections (2.5 infections/100 admissions vs . 7.7 infections/100 admissions, p <.0001), and mortality associated with infection (2.9 deaths/100 admissions vs . 9.6 deaths/100 admissions, p <.0001) during rotation . Logistic regression identified age (odds ratio {OR}, 1.03; 95% confidence interval {CI}, 1.01-1.06), APACHE II score (OR, 1.06; 95% CI, 1.01-1.13), solid organ transplantation (OR, 9.50; 95% CI, 2.01-52.21), and malignancy (OR, 10.16; 95% CI, 4.11-26.96) as independent predictors of mortality . Antibiotic rotation was an independent predictor of survival (OR 6.27, 95% CI 2.78-14.16) . CONCLUSION: Rotation of empirical antibiotic therapy seems to be a promising method to reduce infectious mortality in an ICU.

J Immunol, 2001 Jun 15, 166(12), 7319 - 26
Characterization of a pattern recognition protein, a masquerade-like protein, in the freshwater crayfish Pacifastacus leniusculus; Lee SY et al.; A multifunctional masquerade-like protein has been isolated, purified, and characterized from hemocytes of the freshwater crayfish, Pacifastacus leniusculus . It was isolated by its Escherichia coli binding property, and it binds to formaldehyde-treated Gram-negative bacteria as well as to yeast, Saccharomyces cerevisiae, whereas it does not bind to formaldehyde-fixed Gram-positive bacteria . The intact masquerade (mas)-like protein is present in crayfish hemocytes as a heterodimer composed of two subunits with molecular masses of 134 and 129 kDa . Under reducing conditions the molecular masses of the intact proteins are not changed . After binding to bacteria or yeast cell walls, the mas-like protein is processed by a proteolytic enzyme . The 134 kDa of the processed protein yields four subunits of 65, 47, 33, and 29 kDa, and the 129-kDa protein results in four subunits of 63, 47, 33, and 29 kDa in 10% SDS-PAGE under reducing conditions . The 33-kDa protein could be purified by immunoaffinity chromatography using an Ab to the C-terminal part of the mas-like protein . This subunit of the mas-like protein has cell adhesion activity, whereas the two intact proteins, 134 and 129 kDa, have binding activity to LPSs, glucans, Gram-negative bacteria, and yeast . E . coli coated with the mas-like protein were more rapidly cleared in crayfish than only E . coli, suggesting this protein is an opsonin . Therefore, the cell adhesion and opsonic activities of the mas-like protein suggest that it plays a role as an innate immune protein.

J Trauma, 2001 May, 50(5), 810 - 6
Early elevation of soluble CD14 may help identify trauma patients at high risk for infection; Carrillo EH et al.; BACKGROUND: Elevated levels of soluble CD14 (sCD14) have been implicated in both gram-positive and gram-negative sepsis, and it has been associated with high mortality in trauma patients who become infected . METHODS: Eleven healthy volunteers and 25 adult trauma patients with multiple injuries and a mean Injury Severity Score of 32 participated . Whole blood was obtained at intervals . Immunohistochemistry was used to quantify membrane CD14 (mCD14), by flow cytometry and plasma levels of sCD14 by enzyme-linked immunosorbent assay . Analysis of variance and Student's T test with Mann-Whitney posttest were used to determine significance at p < 0.05 . RESULTS: On posttrauma day 1, sCD14 was significantly different in the plasma of infected patients compared with normal controls (7.16 +/- 1.87 microg/mL vs . 4.4 +/- 0.92 microg/mL, p < 0.01), but not significantly different from noninfected patients . The percentage of monocytes expressing mCD14 in trauma patients did not differentiate them from normal controls; however, mCD14 receptor density did demonstrate significance in septic trauma patients (n = 15) versus normal controls on posttrauma day 3 (p = 0.0065) . CONCLUSION: On the basis of our data, mCD14 did not differentiate infected and noninfected trauma patients, although trauma in general reduced mCD14 and elevated sCD14 . Interestingly, 100% of patients who exceeded plasma levels of 8 microg/mL of sCD14 on day 1 after injury developed infections . Therefore, early high expressers of sCD14 may be at higher risk for infectious complications after trauma.

Water Sci Technol, 2001, 43(1), 77 - 82
Study of microbial community of brewery-treating granular sludge by denaturing gradient gel electrophoresis of 16S rRNA gene; Chan OC et al.; The microbial community structure of granular sludge from an upflow anaerobic sludge blanket (UASB) reactor treating brewery effluent was studied by denaturing gradient gel electrophoresis (DGGE) . Twelve major bands were observed in the DGGE fingerprint for the Bacteria domain and four bands for the Archaea domain . Of the bacterial bands observed, six were successfully purified and sequenced . Among them, three were related to the gram-positive low G + C group, one to the Delta subclass of the Proteobacteria, one to the Gamma subclass, and one to the Cytophaga group with no close related sequence . The 16S rRNA sequences of the four archaeal bands were closely associated with Methanosaeta concilii and Methanobacterium formicum.

J Int Assoc Physicians AIDS Care, 1996 Apr, 2(4), 35 - 7
Macrolides, azalides, and streptogrammins; Prescott LM; AIDS: The Third International Conference on the Macrolides, Azalides, and Streptogramins was held in Lisbon, Portugal . Conferees were given news on the latest advances in the development of innovative antibiotics belonging to these increasingly important groups of drugs, and learned of their expanding clinical indications . The following areas were emphasized at the conference: multiresistant gram-positive bacteremias in patients with serious underlying infections, azithromycin's effectiveness against acute community-acquired pneumonia, results of clarithromycin plus ethambutol in HIV-infected patients with MAC bacteremia, duodenal ulcers associated with Helicobacter pylori infections, and use of roxithromycin against AIDS-related cryptosporidium diarrhea .

J Mol Microbiol Biotechnol, 2001 Jul, 3(3), 401 - 13
Hierarchical control versus autoregulation of carbohydrate utilization in bacteria; Gunnewijk MG et al.; The involvement of phosphoeno/pyruvate:sugar phosphotransferase (PTS) proteins, like HPr and IIA(Glc), in the regulation of carbohydrate utilization has been well established in Gram-negative and Gram-positive bacteria . The majority of the studies of PTS-mediated regulation have been concerned with the hierarchical control of carbohydrate utilization, which results in the preferential utilization of a particular carbohydrate from a mixture of substrates . The underlying mechanisms of PTS-mediated hierarchical control involve the inhibition of expression of other catabolic enzymes and transporters and/or the allosteric regulation of their activity, which prevents the transcriptional inducer to be formed or taken up into the cell . More recently, it has become clear that PTS components allow also the cell to tune the uptake rate(s) to the carbohydrate availability in the medium and the metabolic capacity of the cell . The different phosphorylated species of HPr play a central role in this autoregulatory control circuit, both at the gene and at the protein level . Our knowledge of hierarchical control and autoregulation of carbohydrate utilization in bacteria is discussed.

J Immunol, 2001 Jun 1, 166(11), 6855 - 60
Bacterial DNA and lipopolysaccharide induce synergistic production of TNF-alpha through a post-transcriptional mechanism; Gao JJ et al.; LPS is well recognized for its potent capacity to activate mouse macrophages to produce TNF-alpha, an important inflammatory mediator in bacterial infection-related diseases such as septic shock . We demonstrate here that while inducing only low levels of TNF-alpha alone, DNA from both Gram-negative and Gram-positive bacteria synergizes with subthreshold concentrations of LPS (0.3 ng/ml) to induce TNF-alpha in the RAW 264.7 macrophage-like cell line . The bacterial DNA effects are mimicked by synthetic CpG-containing oligodeoxynucleotides, but not non-CpG-containing oligodeoxynucleotides . Pretreatment of macrophages with either DNA for 2-8 h inhibits macrophage TNF-alpha production in responses to DNA/LPS . However, when pretreatment was extended to 24 h, DNA/LPS synergy on TNF-alpha is further enhanced . RT-PCR analysis indicates that mRNA levels of the TNF-alpha gene, however, are not synergistically induced by bacterial DNA and LPS . Analyses of the half-life of TNF-alpha mRNA indicate that TNF-alpha message has a longer half-life in bacterial DNA- and LPS-treated macrophages than that in bacterial DNA- or LPS-treated macrophages . These findings indicate that the temporally controlled, synergistic induction of TNF-alpha by bacterial DNA and LPS is not mediated at the transcriptional level . Instead, this synergy may occur via a post-transcriptional mechanism.

Arch Surg, 2001 May, 136(5), 592 - 6
Pancreatic infection in severe pancreatitis: the role of fungus and multiresistant organisms; Gloor B et al.; HYPOTHESIS: Recent controlled clinical studies suggest a positive effect of early antibiotic treatment on late morbidity and mortality in severe acute pancreatitis . However, widespread use of antibiotics may lead to an increased number of fungal infections and multiresistant bacteria, thereby worsening the outcome of the disease . DESIGN: Single-center prospective study . SETTING: University hospital, gastrointestinal surgical service . PATIENTS: One hundred three patients with necrotizing pancreatitis seen consecutively in our service . INTERVENTIONS: In addition to standard treatment, patients with proven necrotizing pancreatitis received a prophylactic intravenous antibiotic treatment . Pancreatic infection was regarded as an indication for surgery . MAIN OUTCOME MEASURES: Pancreatic infection, microbiological findings, drug resistance, fungal infections . RESULTS: Thirty-three patients (32%) had infected necrosis . Gram-negative organisms were isolated from 19 patients (58%), Gram-positive organisms were isolated from 18 patients (55%), fungal organisms were isolated from 8 patients (24%), and multiresistant organisms were isolated from 3 patients (9%) . In 7 patients (21%), the organisms cultured from the pancreatic tissue were resistant to the antibiotics given in for prophylaxis . Infection with multiresistant organisms or organisms resistant to the antibiotic used for prophylaxis, but not with fungal infection or Gram-positive or Gram-negative infection, was correlated with a negative outcome . CONCLUSIONS: Fungal infection under adequate treatment is not associated with a negative outcome . The occurrence of multiresistant organisms seems to be a rare finding (3 of 103 patients) . Antibiotic prophylaxis is effective in preventing infection in necrotizing pancreatitis, but optimal choice and duration of administration of the antibiotic agent(s) need to be carefully determined to avoid the sequelae of multiresistant organisms.

Clin Ther, 2001 Apr, 23(4), 566 - 77
Pharmacokinetics and tolerability of extended-release clarithromycin; Guay DR et al.; BACKGROUND: Clarithromycin is a semisynthetic macrolide that exhibits broad-spectrum activity against gram-positive, gram-negative, and atypical respiratory tract and skin/skin structure pathogens, Mycobacterium species, and Helicobacter pylori . It is indicated for the treatment of a wide variety of respiratory and dermatologic infections in children and adults as well as prophylaxis and treatment of Mycobacterium avium complex infection and peptic ulcers due to H . pylori . OBJECTIVE: In this article, we review the results of 3 studies of the steady-state pharmacokinetic profiles of clarithromycin and 14(R)-hydroxy-clarithromycin after multiple oral once-daily doses of 500-mg extended-release (ER) clarithromycin tablets . We also review the drug tolerability in 2 phase III comparative clinical trials of immediate-release (IR) and ER clarithromycin conducted in adults with acute maxillary sinusitis (AMS) and acute exacerbation of chronic bronchitis (AECB) . METHODS: In the 3 pharmacokinetic studies, multiple-dose regimens of clarithromycin IR (one 250-mg or 500-mg tablet twice daily) and clarithromycin ER (one or two 500-mg tablets once daily), administered to healthy male and female volunteers, were evaluated . The effect of administration in nonfasting versus fasting conditions was assessed as well . Tolerability information was collected from each adult patient enrolled in phase III efficacy studies conducted to support the application for US Food and Drug Administration approval for the treatment of AMS and AECB . Regimens evaluated were 500 mg IR clarithromycin tablets twice daily or 1000 mg (2 x 500 mg) ER clarithromycin tablets once daily for 7 days (AECB) or 14 days (AMS) . RESULTS: Bioavailability of the ER clarithromyin tablet administered with food was equivalent to that of the reference IR tablet, based on area under the plasma concentration-time curve (AUC) for both parent compound and active metabolite . The bioavailability of the ER tablet was 30% lower (based on clarithromycin AUC) when administered under fasting versus nonfasting conditions . Compared with the IR tablet, administration of the ER tablet resulted in significantly lower (P < 0.05) clarithromycin peak plasma concentration (Cmax), delayed time to Cmax, and lower degree of concentration fluctuation, confirming its in vivo extended-release characteristics . The most frequently reported adverse events (AEs) in the phase III clinical trials were diarrhea, abnormal taste, and nausea and were generally mild or moderate . The incidence of AEs was comparable for the 2 formulations . The severity of gastrointestinal AEs was significantly less for the ER formulation than for the IR formulation (P = 0.018), as was the frequency of premature study discontinuation due to gastrointestinal AEs or abnormal taste (P = 0.004) . CONCLUSIONS: The results from the 3 pharmacokinetic studies reviewed demonstrate the bioequivalence of the ER and IR formulations and support the use of this clarithromycin ER formulation in a once-daily dosing regimen in phase III clinical trials . The ER tablet should be taken with food to maximize bioavailability . The results of 2 phase III comparative clinical efficacy and safety trials of clarithromycin ER tablets versus IR tablets in AMS and AECB confirm the good tolerability of the ER formulation.

Antimicrob Agents Chemother, 2001 Jun, 45(6), 1843 - 6
Pharmacokinetics and tissue penetration of linezolid following multiple oral doses; Gee T et al.; The pharmacokinetics of multiple-dose linezolid were determined following administration of five 600-mg oral doses given every 12 h to each of six healthy male volunteers . Concentrations of the drug were determined in plasma and inflammatory blister fluid using high-pressure liquid chromatography . A mean peak concentration in plasma of 18.3 microg/ml (standard deviation {SD}, 6.0) was attained at a mean time of 0.7 h (SD, 0.3) after the final dose . The penetration into the inflammatory fluid was 104% (SD, 20.7) . A mean peak concentration of 16.4 microg/ml (SD, 10.6) was attained in the inflammatory fluid at 3 h (SD, 0.6) after the final dose . The elimination half-life from serum and inflammatory fluid was 4.9 (SD, 1.8) and 5.7 (SD, 1.7) h, respectively . The area under the concentration-time curve in plasma and blister fluid was 140.3 (SD, 73.1) and 155.3 (SD, 80.1) microg x h/ml, respectively . These data suggest that linezolid has good tissue penetration, and we can predict that it will be successful in the treatment of a variety of gram-positive infections.

Przegl Epidemiol, 2000, 54(3-4), 305 - 13
{Microorganisms in clinical material collected from patients at the Public Clinical Hospital Nr 1 in Gdansk from 1997-1999}; Samet A et al.; Occurrence of microorganisms isolated from clinical specimens collected from patients in Clinical Hospital no . 1 in Gdansk in years 1997-1999 was analyzed . In this period there was no change in occurrence of Gram-negative bacteria, that accounted for 44-46% isolates . The number of isolations of Gram-positive bacteria dropped from 45% to 40%, and yeast risen from 5% to 10% . The analysis of blood cultures shows decrease in occurrence of bacteremia caused by Gram-negative bacteria and increase in occurrence bacteremia caused by Gram-positive bacteria and yeast . We observed also that the number of multi-resistant Gram-positive isolates (MRSA, VRE) decreased but there was rise in occurrence of multiresistant Gram-negative isolates (ESBL+, CRPA).

FEMS Microbiol Rev, 2001 May, 25(3), 269 - 84
Cell wall teichoic acids: structural diversity, species specificity in the genus Nocardiopsis, and chemotaxonomic perspective; Naumova IB et al.; Data on the structures of cell wall teichoic acids, the anionic carbohydrate-containing polymers, found in many Gram-positive bacteria have been summarized and the polymers of the actinomycete genus Nocardiopsis have been considered from the taxonomic standpoint . The structures of these polymers or their combinations have been demonstrated to be indicative of each of seven Nocardiopsis species and two subspecies, verified by the DNA-DNA relatedness data, and to correlate well with the grouping of the organisms based on 16S rDNA sequences . As each of the intrageneric taxa discussed is definable by the composition of teichoic acids, the polymers are considered to be valuable taxonomic markers for the Nocardiopsis species and subspecies . The (13)C NMR spectra of the polymers, data on the products of their chemical degradation, and distinguishing constituents of whole cell walls derived from teichoic acids are discussed, which are useful for identification of certain polymers and members of the genus Nocardiopsis at the species and subspecies level in microbiological practice.

J Cardiovasc Pharmacol, 2001 May, 37(5), 548 - 63
Cardiovascular sympathomimetic amine interactions in rats treated with monoamine oxidase inhibitors and the novel oxazolidinone antibiotic linezolid; Humphrey SJ et al.; Linezolid (PNU-100766) is a new gram-positive oxazolidinone antibiotic that is effective at in vitro concentrations < or =4 microg/ml and in vivo doses < or =10 mg/kg . Because linezolid also competitively inhibits human monoamine oxidase-A (MAO-A; Ki = 55 microM), we monitored its effects on the cardiovascular responses to tyramine and amine cold remedies in comparison with standard MAO inhibitors . In anesthetized rats, the pressor response to 16 microg i.v . tyramine was potentiated by the MAO-A inhibitors clorgyline (0.1-1.0 mg/kg i.v.) and moclobemide (5.0-50 mg/kg p.o.), but not by the MAO-B inhibitor selegiline (0.15-15 mg/kg p.o.) . Fifteen milligrams per kilogram intravenous linezolid weakly potentiated i.v . tyramine independent of changes in alpha-adrenoceptor reactivity, but this effect was not enhanced chronically (90-100 mg/kg/day) . In conscious rats, 30 mg/kg/day oral linezolid (8 microg/ml plasma concentration) minimally affected the pressor response to 20 mg/kg oral tyramine, whereas 100 mg/kg/day linezolid (20 microg/ml plasma concentration) moderately potentiated this response similar to 3 mg/kg per day moclobemide . Linezolid's tyramine potentiation was reversible, attenuated by food, and independent of pseudoephedrine, phenylpropanolamine, and dextromethorphan interactions . These studies demonstrate that high-dose linezolid only moderately potentiates the cardiovascular effects of tyramine and validate these models for evaluating such MAO inhibitory interactions.

Biochim Biophys Acta, 2001 Jun 1, 1505(2-3), 185 - 208
A novel scenario for the evolution of haem-copper oxygen reductases; Pereira MM et al.; The increasing sequence information on oxygen reductases of the haem-copper superfamily, together with the available three-dimensional structures, allows a clear identification of their common, functionally important features . Taking into consideration both the overall amino acid sequences of the core subunits and key residues involved in proton transfer, a novel hypothesis for the molecular evolution of these enzymes is proposed . Three main families of oxygen reductases are identified on the basis of common features of the core subunits, constituting three lines of evolution: (i) type A (mitochondrial-like oxidases), (ii) type B (ba3-like oxidases) and (iii) type C (cbb3-type oxidases) . The first group can be further divided into two subfamilies, according to the helix VI residues at the hydrophobic end of one of the proton pathways (the so-called D-channel): (i) type A1, comprising the enzymes with a glutamate residue in the motif -XGHPEV-, and (ii) type A2, enzymes having instead a tyrosine and a serine in the alternative motif -YSHPXV- . This second subfamily of oxidases is shown to be ancestor to the one containing the glutamate residue, which in the Bacteria domain is only present in oxidases from Gram-positive or purple bacteria . It is further proposed that the Archaea domain acquired terminal oxidases by gene transfer from the Gram-positive bacteria, implying that these enzymes were not present in the last common ancestor before the divergence between Archaea and Bacteria . In fact, most oxidases from archaea have a higher amino acid sequence identity and similarity with those from bacteria, mainly from the Gram-positive group, than with oxidases from other archaea . Finally, a possible relation between the dihaemic subunit (FixP) of the cbb3 oxidases and subunit II of caa3 oxidases is discussed . As the families of haem-copper oxidases can also be identified by their subunit II, a parallel evolution of subunits I and II is suggested.

South Med J, 2001 Apr, 94(4), 438 - 40
Rothia dentocariosa endocarditis complicated by multiple intracranial hemorrhages; Ricaurte JC et al.; Rothia dentocariosa is a gram-positive rod found commonly as part of the normal flora of the mouth . It rarely causes clinical disease . Subacute infective endocarditis has been the most commonly reported R dentocariosa infection, and extracardiac complications occur frequently . Solitary intracranial hemorrhages have been reported in two cases . We describe the first case of infective endocarditis complicated by the sequential and unusually prolonged development of multiple new intracranial hemorrhages.

Mol Plant Microbe Interact, 2001 May, 14(5), 639 - 52
Polymorphism of the polyketide synthase gene phID in biocontrol fluorescent pseudomonads producing 2,4-diacetylphloroglucinol and comparison of PhID with plant polyketide synthases; Ramette A et al.; Many biocontrol fluorescent pseudomonads can protect plants from soilborne fungal pathogens through production of the antifungal secondary metabolite 2,4-diacetylphloroglucinol (Phl) . One of the phl biosynthetic genes, phlD, encodes a polyketide synthase similar to plant chalcone synthases . Here, restriction analysis of phlD from 39 Phl+ biocontrol fluorescent pseudomonads yielded seven different banding patterns . The gene was sequenced in seven strains, representing the different restriction patterns . Cluster analysis of phlD restriction data or phlD sequences indicated that phlD polymorphism was high, and two main clusters were obtained when predicted PhlD sequences were compared . When the seven PhlD sequences were studied with those of other procaryotic polyketide synthases (gram-positive bacteria) and plant chalcone synthases, however, Phl+ pseudomonads, gram-positive bacteria, and plants clustered separately . Yet, sequence analysis of active site regions for PhlD and plant chalcone synthases revealed that PhlD can be considered a member of the chalcone synthase family, which may be interpreted as convergent evolution of key enzymes involved in secondary metabolism . For the 39 Phl+ pseudomonads, a relationship was found among phlD restriction patterns, phylogenetic groups defined by 16S rDNA restriction analysis (confirmed by 16S rDNA sequencing), and production levels of Phl in vitro.

Perit Dial Int, 2001 Mar-Apr, 21(2), 154 - 7
Diagnostic value of effluent endotoxin level in gram-negative peritonitis in CAPD patients; Muhammed KO et al.; OBJECTIVE: To evaluate the diagnostic usefulness of effluent endotoxin by Limulus amoebocyte lysate (LAL) assay in gram-negative peritonitis patients on continuous ambulatory peritoneal dialysis (CAPD) therapy . DESIGN: Prospective study with patients serving as their own controls . Standard microbiologic work up and endotoxin analysis of effluents (night dwell) were done during the pre- and posttreatment phases . SETTING: Specimens from three teaching hospitals were processed and tested at a common center . Patients were left for treatment at their respective centers without intervention . PATIENTS: 32 clinical peritonitis and 40 infection-free CAPD patients were studied . RESULTS: 75% (n = 24) of cultures were positive: 41.6% (n = 10) gram-negative and 58.4% (n = 14) gram-positive bacteria . Effluents of pre- and posttreated gram-negative cultures had endotoxin levels of 1.53 +/- 0.169 and 0.214 +/- 0.085 endotoxin units (EU)/mL, respectively (p < 0.0001); pre- and posttreated gram-positive levels of 0.102 +/- 0.06 and 0.122 +/- 0.052 EU/mL, respectively (p > 0.05); pre- and posttreated culture-negative peritonitis levels of 0.110 +/- 6.025 and 0.087 +/- 0.031 EU/mL, respectively (p > 0.05); peritonitis-free effluents contained 0.117 +/- 0.079 EU/mL . The diagnostic specificity and the sensitivity of LAL assay were 100% and 98.2% respectively . CONCLUSION: Where initial microbiological work-up cannot demonstrate a pathogen, effluent endotoxin determined by quantitative LAL assay is a useful marker for diagnosis and management, within safe time limits, of gram-negative peritonitis in CAPD patients.

Clin Diagn Lab Immunol, 2001 May, 8(3), 515 - 21
Peptidoglycan and lipoteichoic acid modify monocyte phenotype in human whole blood; Jorgensen PF et al.; We examined the influence of the gram-positive cell wall products peptidoglycan (PepG) and lipoteichoic acid (LTA), compared to lipopolysaccharide (LPS), on the monocyte expression of receptors involved in antigen presentation (HLA-DR, B7.1, and B7.2), cell adhesion (intercellular adhesion molecule-1 {ICAM-1} and lymphocyte function associated antigen-3 {LFA-3}), phagocytosis (Fc gamma RI), and cell activation (CD14) . We also evaluated possible influences of the immunosuppressive drugs cyclosporine A, tacrolimus, and sirolimus on the expression of these receptors . Pretreatment of whole blood for 4 h with the immunosuppressive drugs did not influence the expression of the surface receptors in normal or stimulated blood . Stimulation with both PepG and LTA caused significant up-regulation of the surface expression of ICAM-1 and HLA-DR on whole blood monocytes, similar to that obtained with LPS, whereas B7.1, B7.2, LFA-3, and Fc gamma RI were not modulated . PepG and LTA also caused increased expression of CD14, whereas LPS down-regulated this molecule . In contrast, we did not detect any significant influence of any of the bacterial products on the plasma concentration of soluble CD14 . We hypothesized that the increased expression of surface CD14 in blood stimulated with PepG would prime for cellular activation by LPS . Indeed, we show that PepG and the partial PepG structure muramyl dipeptide acted in synergy with LPS to cause the release of tumor necrosis factor-alpha . The results suggest that PepG and LPS provoke partly different responses on monocyte phenotype and that CD14 may play different roles in the innate response to gram-positive and gram-negative bacteria.

J Bacteriol, 2001 May, 183(10), 3184 - 92
Role of acid metabolism in Streptomyces coelicolor morphological differentiation and antibiotic biosynthesis; Viollier PH et al.; Studies of citrate synthase (CitA) were carried out to investigate its role in morphological development and biosynthesis of antibiotics in Streptomyces coelicolor . Purification of CitA, the major vegetative enzyme activity, allowed characterization of its kinetic properties . The apparent K(m) values of CitA for acetyl coenzyme A (acetyl-CoA) (32 microM) and oxaloacetate (17 microM) were similar to those of citrate synthases from other gram-positive bacteria and eukaryotes . CitA was not strongly inhibited by various allosteric feedback inhibitors (NAD(+), NADH, ATP, ADP, isocitrate, or alpha-ketoglutarate) . The corresponding gene (citA) was cloned and sequenced, allowing construction of a citA mutant (BZ2) . BZ2 was a glutamate auxotroph, indicating that citA encoded the major citrate synthase allowing flow of acetyl-CoA into the tricarboxylic acid (TCA) cycle . Interruption of aerobic TCA cycle-based metabolism resulted in acidification of the medium and defects in morphological differentiation and antibiotic biosynthesis . These developmental defects of the citA mutant were in part due to a glucose-dependent medium acidification that was also exhibited by some other bald mutants . Unlike other acidogenic bald strains, citA and bldJ mutants were able to produce aerial mycelia and pigments when the medium was buffered sufficiently to maintain neutrality . Extracellular complementation studies suggested that citA defines a new stage of the Streptomyces developmental cascade.

Am J Kidney Dis . 2001 May;37(5):E38.
Antineutrophil cytoplasmic autoantibody specific for proteinase 3 in a patient with shunt nephritis induced by Gemella morbillorum; Nagashima T et al.; A 17-year-old girl had been placed with ventriculoperitoneal, then ventriculoatrial shunts for congenital hydrocephalus since birth . The patient originally was diagnosed as having a lupus-like disease, but later turned out to have shunt nephritis, presenting with fever, proteinuria, pancytopenia, and hypocomplementemia . Antineutrophil cytoplasmic autoantibody specific for proteinase 3 (PR3-ANCA) was detected in her serum . The patient received oral prednisolone and repeated methylprednisolone pulses, with essentially no beneficial effects . A gram-positive coccus, Gemella morbillorum, was recovered from her blood as well as cerebrospinal fluid, and the culture of the shunt catheter established the diagnosis of shunt nephritis . Removal of the shunt catheter improved symptoms dramatically and decreased PR3-ANCA in serum to an undetectable level . Because steroids had no effects and the control of bacterial infection lowered PR3-ANCA levels, the antibody would have been induced by continuous infection with G morbillorum.

Biochem Biophys Res Commun, 2001 Apr 27, 283(1), 195 - 204
Isolation and characterization of the genes encoding a novel oxygenase component of angular dioxygenase from the gram-positive dibenzofuran-degrader Terrabacter sp . strain DBF63; Kasuga K et al.; A gram-positive bacterium Terrabacter sp . strain DBF63 is able to degrade dibenzofuran (DF) via initial dioxygenation by a novel angular dioxygenase . The dbfA1 and dbfA2 genes, which encode the large and small subunits of the dibenzofuran 4,4a-dioxygenase (DFDO), respectively, were isolated by a polymerase chain reaction-based method . DbfA1 and DbfA2 showed moderate homology to the large and small subunits of other ring-hydroxylating dioxygenases (less than 40%), respectively, and some motifs such as the Fe(II) binding site and the {2Fe-2S} cluster ligands were conserved in DbfA1 . DFDO activity was confirmed in Escherichia coli cells containing the cloned dbfA1 and dbfA2 genes with the complementation of nonspecific ferredoxin and ferredoxin reductase component of E . coli . Under this condition, these cells exhibited angular dioxygenation of DF and dibenzo-p-dioxin, and monooxygenation of fluorene, but not angular dioxygenation of carbazole, xanthene, and phenoxathiin . Phylogenetic analysis revealed that DbfA1 formed a branch with recently reported large subunits of polycyclic aromatic hydrocarbon (PAH) dioxygenase from gram-positive bacteria but did not cluster with that of other angular dioxygenases, i.e., DxnA1 from Sphingomonas sp . strain RW1 {Armengaud, J., Happe, B., and Timmis, K . N . J . Bacteriol . 180, 3954-3966, 1998} and CarAa from Pseudomonas sp . strain CA10 {Sato, S., Nam, J.-W., Kasuga, K., Nojiri, H., Yamane, H., and Omori, T . J . Bacteriol . 179, 4850-4858, 1997} .

Vet Rec, 2000 Apr 29, 146(18), 515 - 8
Evaluation of bacteria isolated from infected eyes of captive, non-domestic animals; Williams DL et al.; Bacteria isolated from the eyes of captive species with suspected ocular infections at London Zoo were identified by standard methods . The sensitivity of the organisms to several topical antibiotics was determined by using sensitivity discs, and the minimum inhibitory concentrations of chloramphenicol and fusidic acid were determined . Correlations were evaluated between the results from the antibiotic discs and the minimum inhibitory concentrations and, where possible, between the clinical response to treatment and the results of bacteriological sensitivity tests . Unlike the isolates found in cats and dogs gram-positive cocci accounted for 54 per cent of isolates but almost half of the bacteria isolated were gram-negative organisms.

Int J Syst Evol Microbiol, 2001 Mar, 51(Pt 2), 475 - 9
Aerococcus sanguicola sp . nov., isolated from a human clinical source; Lawson PA et al.; Phenotypic and phylogenetic studies were performed on an unknown, Gram-positive, catalase-negative coccus isolated from human blood . Comparative 16S rRNA gene sequencing demonstrated that the organism represents a new subline within the genus Aerococcus . The unknown bacterium was readily distinguished from the three currently recognized Aerococcus species, Aerococcus christensenii, Aerococcus urinae and Aerococcus viridans, by biochemical tests and electrophoretic analysis of whole-cell proteins . On the basis of phylogenetic and phenotypic evidence, it is proposed that this unknown bacterium from blood be classified as Aerococcus sanguicola sp . nov . The type strain of Aerococcus sanguicola is CCUG 43001T (= CIP 106533T).

Microbiology, 2001 May, 147(Pt 5), 1197 - 212
Genomic analysis of the histidine kinase family in bacteria and archaea; Kim D et al.; Two-component signal transduction systems, consisting of histidine kinase (HK) sensors and DNA-binding response regulators, allow bacteria and archaea to respond to diverse environmental stimuli . HKs possess a conserved domain (H-box region) which contains the site of phosphorylation and an ATP-binding kinase domain . In this study, a genomic approach was taken to analyse the HK family in bacteria and archaea . Based on phylogenetic analysis, differences in the sequence and organization of the H-box and kinase domains, and the predicted secondary structure of the H-box region, five major HK types were identified . Of the 336 HKs analysed, 92% could be assigned to one of the five major HK types . The Type I HKs were found predominantly in bacteria while Type II HKs were not prevalent in bacteria but constituted the major type (13 of 15 HKs) in the archaeon Archaeoglobus fulgidus . Type III HKs were generally more prevalent in Gram-positive bacteria and were the major HK type (14 of 15 HKs) in the archaeon Methanobacterium thermoautotrophicum . Type IV HKs represented a minor type found in bacteria . The fifth HK type was composed of the chemosensor HKs, CheA . Several bacterial genomes contained all five HK types . In contrast, archaeal genomes either contained a specific HK type or lacked HKs altogether . These findings suggest that the different HK types originated in bacteria and that specific HK types were acquired in archaea by horizontal gene transfer.

J Biol Chem, 2001 Jul 13, 276(28), 25680 - 6 Epub 2001 Apr 20.
The role of toll-like receptors (TLRs) in bacteria-induced maturation of murine dendritic cells (DCS) . Peptidoglycan and lipoteichoic acid are inducers of DC maturation and require TLR2; Michelsen KS et al.; Toll-like receptors (TLRs) have been found to be key elements in pathogen recognition by the host immune system . Dendritic cells (DCs) are crucial for both innate immune responses and initiation of acquired immunity . Here we focus on the potential involvement of TLR ligand interaction in DC maturation . TLR2 knockout mice and mice carrying a TLR4 mutation (C3H/HeJ) were investigated for DC maturation induced by peptidoglycan (PGN), lipopolysaccharide (LPS), or lipoteichoic acids (LTAs) . All stimuli induced maturation of murine bone marrow-derived DCs in control mice . TLR2(-)/- mice lacked maturation upon stimulation with PGN, as assessed by expression of major histocompatibility complex class II, CD86, cytokine, and chemokine production, fluorescein isothiocyanate-dextran uptake, and mixed lymphocyte reactions, while being completely responsive to LPS . A similar lack of maturation was observed in C3H/HeJ mice upon stimulation with LPS . DC maturation induced by LTAs from two different types of bacteria was severely impaired in TLR2(-)/-, whereas C3H/HeJ mice responded to LTAs in a manner similar to wild-type mice . We demonstrate that DC maturation is induced by stimuli from Gram-positive microorganisms, such as PGN and LTA, with similar efficiency as by LPS . Finally, we provide evidence that TLR2 and TLR4 interaction with the appropriate ligand is essential for bacteria-induced maturation of DCs.

Clin Chest Med, 2001 Mar, 22(1), 71 - 85
Respiratory infection in the chronically critically ill patient . Ventilator-associated pneumonia and tracheobronchitis; Ahmed QA et al.; The long-term ventilated patient is at high risk for developing nosocomial pneumonia or tracheobronchitis . In general, the frequency of infection increases with the duration of mechanical ventilation, but the risk appears to be greatest in the first week of intubation . Although these types of infection are common and may have morbidity and mortality impact, the daily risk is less in the long-term ventilated patient than in the acutely ill intubated patient . This reduced daily risk may reflect a "survivor effect," with less healthy patients dying early in the hospital stay and not surviving long enough to undergo tracheostomy and long-term ventilation . A number of factors predispose these patients to infection, including host defense impairment and exposure to large numbers of bacteria . This exposure can occur through the airway, and proper care of respiratory therapy devices is essential to minimize the risk for infection . Most infections of the lower respiratory tract are preceded by airway colonization with EGN bacteria and, with improvement in host defenses and nutrition, infection in the face of colonization is less likely . In some patients, colonization can be eliminated . When the long-term ventilated patient does develop infection, it generally involves highly resistant gram-negative or gram-positive organisms and therapy should be prompt and appropriate . Not all such patients respond to systemic antibiotics, and the use of adjunctive aerosol therapy may have benefit for those with either tracheobronchitis or pneumonia, especially if highly resistant pathogens are present.

Rheumatology (Oxford), 2001 Apr, 40(4), 438 - 46
Peptidoglycan from sterile human spleen induces T-cell proliferation and inflammatory mediators in rheumatoid arthritis patients and healthy subjects; Schrijver IA et al.; OBJECTIVES: Peptidoglycan (PG), a component of Gram-positive bacteria, may be involved in rheumatoid arthritis (RA) because of its ability to induce production of proinflammatory cytokines, to induce arthritis in rodents, and its presence in antigen-presenting cells in RA joints . METHODS: In the present study, physiologically relevant PG was able to induce T-cell proliferation in peripheral blood and synovial fluid samples of RA patients, but the magnitude of the response did not differ from that of cells from healthy subjects . In addition, production of cytokines associated with RA (interleukins (IL)-1beta, IL-6, IL-8, IL-10, IL-12 and tumour necrosis factor alpha) and of the matrix metalloproteinase, gelatinase B (MMP-9), was induced in blood and synovial fluid cultures of RA patients . CONCLUSION: The fact that PG, which can be found in synovial tissues of RA patients is able to induce the production of inflammatory mediators supports the hypothesis that PG plays a role in the pathogenesis of RA by influencing the inflammatory microenvironment of the joint.

Intern Med, 2001 Mar, 40(3), 246 - 9
Pulmonary nocardiosis associated with idiopathic thrombocytopenic purpura; Ando T et al.; A 69-year-old woman with idiopathic thrombocytopenic purpura, who was regularly followed and treated with prednisolone and danazol, was admitted to our hospital because of shortness of breath . Chest roentgenogram showed a large amount of left-sided pleural effusion . Gram-positive branching rods, subsequently identified as Nocardia farcinica, were isolated from the fluid . Antibiotic treatment together with pleural drainage with an intercostal catheter resulted in complete remission of pyothorax . Pulmonary nocardiosis is a rare disease, but recognition of the disease in immunocompromised patients and the prompt initiation of appropriate treatments based on isolation of the pathogen can lead to a successful outcome.

Chemotherapy, 2001 May-Jun, 47(3), 226 - 31
Treatment of febrile neutropenia with cefepime monotherapy; Jandula BM et al.; BACKGROUND AND OBJECTIVE: The empirical administration of a broad-spectrum beta-lactam antibiotic, either as monotherapy or in combination with an aminoglycoside, is an essential component of the initial management of patients with fever and severe neutropenia . Multiple antibiotics have been tested for this indication . Cefepime is a fourth-generation cephalosporin with in vitro activity against most gram-negative and many gram-positive bacteria . We have studied the use of this agent as monotherapy in this indication . METHODS: One hundred and twenty-six episodes of febrile neutropenia in 98 adults with hematological malignancies were treated with cefepime monotherapy . Cefepime was given at a dose of 2 g every 8 h i.v . Most episodes (49%) were fever of unexplained origin, while a microbiologically documented and clinically documented infection occurred in 25% episodes each . Seventy-six (61%) episodes occurred after conventional chemotherapy, while 51 (41%) after a hematopoietic stem cell transplantation . Results: Twelve episodes (10%) were not evaluable for response . Among the 114 evaluable episodes, 69 (55% of the initial sample and 61% of those evaluable) responded to cefepime monotherapy, while therapy failed in 45 cases (36% of the initial sample and 39% of those evaluable), including 14 cases who developed breakthrough bacteremia during therapy . There were no deaths due to bacterial infection . At the end of all antibiotic therapy (final outcome) 69 episodes were cured only with monotherapy, 47 were cured with modification of therapy and 10 patients died from an unrelated cause . The only variable that appeared to correlate with response to therapy was the duration of neutropenia, which was longer among patients who failed or developed breakthrough bacteremia than among those who responded to monotherapy . INTERPRETATION AND CONCLUSIONS: Initial empirical antibiotic therapy with cefepime as a single agent in patients with febrile neutropenia and a hematological malignancy is effective, but patients with prolonged neutropenia appear to be at higher risk for failure . However, with appropriate therapeutic changes the risk of dying from a bacterial infection is very low .

J Clin Pharmacol, 2001 Apr, 41(4), 404 - 14
Multiple-dose pharmacokinetics and safety of two regimens of quinupristin/dalfopristin (Synercid) in healthy volunteers; Chevalier P et al.; Quinupristin/dalfopristin (Q/D) is a novel streptogramin antibiotic for the treatment of severe gram-positive infections . The purpose of this open, nonrandomized, parallel-group, phase I trial was to evaluate Q/D pharmacokinetics after single and repeated doses under the two different dosing regimens corresponding to the effective doses and to evaluate tolerability . Two groups of 10 healthy volunteers received multiple 1-hour intravenous infusions of 7.5 mg/kg Q/D either every 8 or 12 hours for 4 or 5 days, respectively . Plasma concentrations of Q, D, and metabolites were determined using high-performance liquid chromatography and selective microbiological assays . The two regimens q8h and q12h lead to the same disposition profile after single and repeated administration . Single-dose data confirmed the high plasma clearances of Q and D (about 0.90 l/h/kg) obtained previously . Unchanged drugs were the main components in plasma, with each of the three metabolites representing about 20% (in terms of the AUC ratio) of the parent drugs . Comparable steady-state concentrations were reached from day 2 of both regimens . A similar moderate increase in Cmax and AUC (about 20%) of parent drugs was observed between the first and last day of treatment . This phenomenon, which was also observed for the metabolites, was not expected considering the short terminal disposition half-lives of the parent drugs and trough plasma concentrations of all components mostly below the limits of quantitation at steady state, whatever the dosing regimen . The clearances of parent drugs at steady state were about 20% lower as compared with that observed following the first drug administration (statistically significant difference) . No trend suggesting a treatment effect on any laboratory parameter, vital signs, or electrocardiographic parameters was identified . However, 80% of subjects reported venous adverse events probably related to treatment.

Pediatr Infect Dis J, 2001 Mar, 20(3), 343 - 9
Empiric use of cefepime in the treatment of lower respiratory tract infections in children; Bradley JS et al.; BACKGROUND: These studies were designed to assess the efficacy and safety of cefepime, a fourth generation cephalosporin, for the treatment of serious infections, including lower respiratory tract infections (LRTI) in children . METHODS: Four clinical trials of cefepime for the treatment of serious bacterial infections enrolled 259 children with LRTI . In 3 trials cefepime was compared with ceftazidime (n = 166), cefotaxime (n = 16) or cefuroxime (n = 12) . One trial was noncomparative (n = 65) . RESULTS: Treatment with cefepime 50 mg/kg/ dose administered every 8 to 12 h produced a satisfactory clinical response (clinical signs of infection resolved or improved with no evidence of recurrent infection at posttreatment follow-up) in 88 to 100% of patients, comparable with comparator therapy . In children from whom a causative pathogen was identified, bacteriologic eradication was comparable between cefepime and comparator therapy . Cefepime was as safe and well-tolerated as comparator therapy . Few treatment-related clinical or laboratory adverse events were noted and were equivalent to comparator in all studies . CONCLUSION: Cefepime is as effective, safe and well-tolerated for the empiric treatment of children with LRTI as comparator agents but offers the advantage of an enhanced spectrum of activity for Gram-positive and Gram-negative pathogens compared with second or third generation cephalosporins.

Crit Care, 2001, 5(2), 53 - 5 Epub 2001 Feb 26.
Superantigen antagonist peptides; Llewelyn M et al.; The production of superantigenic exotoxins by Gram positive bacteria underlies the pathology of toxic shock syndrome . Future treatment strategies for superantigen-mediated diseases are likely to be directed at blocking the three-way interaction between superantigen, T cell receptor and major histocompatibility class II molecule, which inititates an excessive and disordered inflammatory response . In this article, we review the first published data to address one such strategy in the context of other recognised and experimental treatments.

Hum Mutat, 2001 Apr, 17(4), 305 - 16
Fluorescent microsphere-based readout technology for multiplexed human single nucleotide polymorphism analysis and bacterial identification; Ye F et al.; Large-scale human genotyping requires technologies with a minimal number of steps, high accuracy, and the ability to automate at a reasonable cost . In this regard, we have developed a rapid, cost-effective readout method for single nucleotide polymorphism (SNP) genotyping that combines an easily automatable single-tube allele-specific primer extension (ASPE) with an efficient high throughput flow cytometric analysis performed on a Luminex 100 flow cytometer . This robust technique employs an ASPE reaction using PCR-derived target DNA containing the SNP and a pair of synthetic complementary capture probes that differ at their 3' end-nucleotide defining the alleles . Each capture probe has been synthesized to contain a unique 25-nucleotide identifying sequence (ZipCode) at its 5' end . An array of fluorescent microspheres, covalently coupled with complementary ZipCode sequences (cZipCodes), was hybridized to biotin-labeled ASPE reaction products, sequestering them for flow cytometric analysis . ASPE offers both an advantage of streamlining the SNP analysis protocol and an ability to perform multiplex SNP analysis on any mixture of allelic variants . All steps of the assay are simple additions of the solutions, incubations, and washes . This technique was used to assay 15 multiplexed SNPs on human chromosome 12 from 96 patients . Comparison of the microsphere-based ASPE assay results to gel-based oligonucleotide ligation assay (OLA) results showed 99.2% agreement in genotype assignments . In addition, the microsphere-based multiplex SNPs assay system was adapted for the identification of bacterial samples by both ASPE and single base chain extension (SBCE) assays . A series of probes designed for different variable sites of bacterial 16S rDNA permitted multiplex analysis and generated species- or genus-specific patterns . Seventeen bacterial species representing a broad range of gram-negative and gram-positive bacteria were analyzed within 16 variable sites of 16S rDNA sequence . The results were consistent with the published sequences and confirmed by direct DNA sequencing .

Infect Immun, 2001 May, 69(5), 3343 - 9
Fas-mediated apoptosis of neutrophils in sera of patients with infection; Nwakoby IE et al.; In the presence of infection, neutropenia is considered to be a marker of poor prognosis; conversely, neutrophilia may not be a determinant of a better prognosis . Since apoptotic neutrophils are compromised functionally, we evaluated the effect of infection on neutrophil apoptosis . The rate of apoptosis was greater for neutrophils isolated from patients with infection than for healthy controls . Escherichia coli did not directly modulate the rate of neutrophil apoptosis . However, sera from infected patients promoted (P < 0.001) neutrophil apoptosis . Interestingly, the sera of patients with different types of infection (gram negative, gram positive, or culture negative) exerted a more or less identical response on neutrophil apoptosis . Sera of infected patients showed a fivefold greater content of FasL compared to controls . Moreover, anti-FasL antibody partly attenuated the infected-serum-induced neutrophil apoptosis . In in vitro studies, E . coli enhanced monocyte FasL expression . Moreover, conditioned media prepared from activated macrophages from control mice showed enhanced apoptosis of human as well as mouse neutrophils . On the contrary, conditioned media prepared from activated macrophages isolated from FasL-deficient mice induced only a mild degree of neutrophil apoptosis . These results suggest that neutrophils in patients with infection undergo apoptosis at an accelerated rate . Infection not only promoted monocyte expression of FasL but also increased FasL content of the serum . Because the functional status of apoptotic cells is compromised, a significant number of neutrophils may not be participating in the body's defense . Since neutrophils play the most important role in innate immunity, their compromised status in the presence of infection may transfer the host defense burden from an innate response to acquired immunity . The present study provides some insight into the lack of correlation between neutrophilia and the outcome of infection.

J Immunol, 2001 Apr 15, 166(8), 5161 - 7
Induction of cross-tolerance by lipopolysaccharide and highly purified lipoteichoic acid via different Toll-like receptors independent of paracrine mediators; Lehner MD et al.; Exposure of macrophages to LPS induces a state of hyporesponsiveness to subsequent stimulation with LPS termed LPS desensitization or tolerance . To date, it is not known whether similar mechanisms of macrophage refractoriness are induced on contact with components of Gram-positive bacteria . In the present study, we demonstrate that pretreatment with highly purified lipoteichoic acid (LTA) results in suppression of cytokine release on restimulation with LTA in vitro and in vivo in both C3H/HeN and C3H/HeJ mice, but not in macrophages from Toll-like receptor (TLR)-2-deficient mice . Furthermore, desensitization in response to LPS or LTA exposure also inhibits responses to the other stimulus ("cross-tolerance"), suggesting that signaling pathways shared by TLR2 and TLR4 are impaired during tolerance . Finally, we show that LPS- or LTA-induced cross-tolerance is not transferred to hyporesponsive cells cocultured with LPS/LTA-responsive macrophages, showing that soluble mediators do not suffice for tolerance induction in neighboring cells.

Southeast Asian J Trop Med Public Health, 2000 Sep, 31(3), 540 - 6
Peritonitis complicating acute peritoneal dialysis in Northeast Malaysia; Kamaliah MD et al.; A prospective observational study examing the incidence, predisposing factors and microbiological aspects of peritonitis complicating acute intermittent peritoneal dialysis (IPD) was performed in Hospital Universiti Sains Malaysia, a referral hospital situated in Northeast Malaysia . Over a 7- month period, a total of 126 acute IPD treatments were included involving 69 patients . The majority of patients suffered from chronic or end stage renal failure (92.7%) and nearly half (47.8%) have underlying diabetes mellitus . Peritonitis occured in 25 treatment sessions giving a frequency of 19.8% of procedures performed . The mean interval between starting dialysis and the first sign of peritonitis was 3.5 days, with 12% of peritonitis occuring before day 3 of treatment . Frequent catheter manipulation and/or leakages were identified as significant predisposing factors for peritonitis and the risk of peritonitis was increased with longer duration of IPD . Gram-negative infections were seen twice more commonly than gram-positive infections . We recommend the use of cloxacillin in combination with either an aminoglycoside or a cephalosporin as empirical antibiotic coverage until culture reports are available.

Trends Microbiol, 2001 Apr, 9(4), 164 - 8
Processing and export of peptide pheromones and bacteriocins in Gram-negative bacteria; Michiels J et al.; Cell-density-dependent gene expression is widespread in bacteria and is mediated by extracellular communication molecules . Gram-negative bacteria often use N-acyl homoserine lactones, whereas cell-cell signaling in Gram-positive bacteria is accomplished using post-translationally processed peptide pheromones . In many Gram-positive bacteria, export of these peptides requires the activity of a dedicated ATP-binding cassette (ABC) transporter, which cleaves off a typical leader peptide termed the double-glycine leader sequence concomitant with translocation across the membrane . Inspection of bacterial genome sequences has revealed the presence of similar ABC transporters, as well as genes encoding peptides with double-glycine-type leader sequences in Gram-negative bacteria, and it is suggested that the postulated transported peptides could perform a signaling function.

Appl Environ Microbiol, 2001 Apr, 67(4), 1476 - 83
Degradation of phenanthrene and anthracene by cell suspensions of Mycobacterium sp . strain PYR-1; Moody JD et al.; Cultures of Mycobacterium sp . strain PYR-1 were dosed with anthracene or phenanthrene and after 14 days of incubation had degraded 92 and 90% of the added anthracene and phenanthrene, respectively . The metabolites were extracted and identified by UV-visible light absorption, high-pressure liquid chromatography retention times, mass spectrometry, (1)H and (13)C nuclear magnetic resonance spectrometry, and comparison to authentic compounds and literature data . Neutral-pH ethyl acetate extracts from anthracene-incubated cells showed four metabolites, identified as cis-1,2-dihydroxy-1,2-dihydroanthracene, 6,7-benzocoumarin, 1-methoxy-2-hydroxyanthracene, and 9,10-anthraquinone . A novel anthracene ring fission product was isolated from acidified culture media and was identified as 3-(2-carboxyvinyl)naphthalene-2-carboxylic acid . 6,7-Benzocoumarin was also found in that extract . When Mycobacterium sp . strain PYR-1 was grown in the presence of phenanthrene, three neutral metabolites were identified as cis- and trans-9,10-dihydroxy-9,10-dihydrophenanthrene and cis-3,4-dihydroxy-3,4-dihydrophenanthrene . Phenanthrene ring fission products, isolated from acid extracts, were identified as 2,2'-diphenic acid, 1-hydroxynaphthoic acid, and phthalic acid . The data point to the existence, next to already known routes for both gram-negative and gram-positive bacteria, of alternative pathways that might be due to the presence of different dioxygenases or to a relaxed specificity of the same dioxygenase for initial attack on polycyclic aromatic hydrocarbons.

Curr Opin Microbiol, 2001 Apr, 4(2), 172 - 7
Iron and metal regulation in bacteria; Hantke K; In Escherichia coli, the iron regulator Fur is regulated by two oxidative-stress response regulators . The generation of dangerous radicals by oxygen and iron is the basis for this dual regulation, which is also found in eukaryotes . The binding of iron-regulated transcripts to apo-aconitase and results of mRNA half-life studies indicate that there is post-transcriptional iron regulation in bacteria, as in eukaryotes . Fur contains two metal-binding sites, one for Zn2+ and one for Fe2+ . Zinc uptake systems are regulated by members of the Fur protein family, and zinc is a cofactor . DtxR and related proteins constitute another family of iron regulators, first found in Gram-positive organisms with a high GC content . In organisms with Fur-dependent iron regulation, members of the DtxR family regulate manganese transport.

Comb Chem High Throughput Screen, 2001 Apr, 4(2), 171 - 84
Surface display on gram positive bacteria; Hansson M et al.; Heterologous surface display on Gram-positive bacteria was first described almost a decade ago and has since then developed into an active research area . Gram-positive bacterial surface display has today found a range of applications, in immunology, microbiology and biotechnology . Live bacterial vaccine delivery vehicles are being developed through the surface display of selected foreign antigens on the bacterial surfaces . In this field, "second generation" vaccine delivery vehicles are at present being generated by the addition of mucosal targeting signals through co-display of adhesins, in order to achieve targeting of the live bacteria to immunoreactive sites to thereby increase immune responses . Engineered Gram-positive bacteria are further being evaluated as novel microbial biocatalysts with heterologous enzymes immobilized as surface exposed on the bacterial cell surface . A discussion has started whether bacteria can find use as new types of whole-cell diagnostic devices since single-chain antibodies and other variants of tailor-made binding proteins can be displayed on bacteria . Bacteria with increased binding capacity for certain metal ions can be created and potential environmental or biosensor applications for such recombinant bacteria as biosorbents are being discussed . This article explains the basis of Gram-positive bacterial surface display, and discusses current uses and possible future trends of this emerging technology.

Int J Hyg Environ Health, 2001 Mar, 203(3), 245 - 8
Comparison of two sampling methods for the detection of gram-positive and gram-negative bacteria in the environment: moistened swabs versus Rodac plates; Lemmen SW et al.; The objective of this study was to evaluate the moistened swab technique vs . Rodac plates for detecting Gram-positive and Gram-negative bacteria in the inanimate environment . Over a period of 22 months, the environment of 190 patients infected or colonized with MRSA, VRE or multiresistant Gram-negative bacteria was sampled in turn . MRSA and VRE could be detected with either method in 33 out of 54 (61.1%) patient rooms in 174 out of 706 (24.6%) environmental samples . However, multiresistant Gram-negative bacteria were found in 42 out of 136 (30.9%) rooms with a very low frequency of 89 out of 1827 (4.9%) environmental samples (p < 0.0001) . The sensitivity of the swab technique for Gram-positive cocci was 54% (94/174) vs . 69.5% (121/174) for the Rodac plates, ({CI95%}, 47-61% vs . 62-76%, p < 0.05) . In contrast, the sensitivity of the swab technique for Gram-negative bacteria was 74.2% (66/89) vs . 42.7% (38/89) for the Rodac plates, ({CI95%}, 64-83% vs . 32-54%, p < 0.05) . In conclusion, environmental contamination with Gram positive cocci is detected more often than with Gram-negative bacteria . For the detection of Gram-positive cocci, Rodac plates are superior to the swab technique; whereas Gram-negative rods can be detected more often by the swab technique . All these results proved to be statistically significant.

J Biol Chem, 2001 Jun 15, 276(24), 20935 - 44 Epub 2001 Mar 28.
Biosynthesis of nucleotide-activated D-glycero-D-manno-heptose; Kneidinger B et al.; The glycan chain repeats of the S-layer glycoprotein of Aneurinibacillus thermoaerophilus DSM 10155 contain d-glycero-d-manno-heptose, which has also been described as constituent of lipopolysaccharide cores of Gram-negative bacteria . The four genes required for biosynthesis of the nucleotide-activated form GDP-d-glycero-d-manno-heptose were cloned, sequenced, and overexpressed in Escherichia coli, and the corresponding enzymes GmhA, GmhB, GmhC, and GmhD were purified to homogeneity . The isomerase GmhA catalyzed the conversion of d-sedoheptulose 7-phosphate to d-glycero-d-manno-heptose 7-phosphate, and the phosphokinase GmhB added a phosphate group to form d-glycero-d-manno-heptose 1,7-bisphosphate . The phosphatase GmhC removed the phosphate in the C-7 position, and the intermediate d-glycero-alpha-d-manno-heptose 1-phosphate was eventually activated with GTP by the pyrophosphorylase GmhD to yield the final product GDP-d-glycero-alpha-d-manno-heptose . The intermediate and end products were analyzed by high performance liquid chromatography . Nuclear magnetic resonance spectroscopy was used to confirm the structure of these substances . This is the first report of the biosynthesis of GDP-d-glycero-alpha-d-manno-heptose in Gram-positive organisms . In addition, we propose a pathway for biosynthesis of the nucleotide-activated form of l-glycero-d-manno-heptose.

Eur Urol, 2001 Mar, 39(3), 357 - 9
Renal actinomycosis mimicking renal carcinoma; Dieckmann KP et al.; The case of a 52-year-old man is reported who presented with night sweats and slight debilitation . Upon CT scan a left-sided renal mass with centrally liquefied areas was detected . The patient underwent nephrectomy for suspected renal cancer with central necrosis . Histologically, the diagnosis of renal actinomycosis was established based on the detection of sulphur granules . Actinomyces israelii is an anaerobic gram-positive bacterium that may cause localized tumour-like infections mainly in the craniocervical region and exceptionally retroperitoneally . Renal actinomycosis is a rare differential diagnosis of renal masses . As nephrectomy may prove hazardous in these cases, the diagnosis should be attempted pre-operatively by ultrasound-guided aspiration and consecutive antibiotic treatment . In selected cases surgery could be avoided at all.

Surv Ophthalmol, 2001 Mar-Apr, 45(5), 361 - 78
Ocular nocardia infections with special emphasis on the cornea; Sridhar MS et al.; Nocardia are aerobic, gram-positive, nonmotile and branching filamentous bacteria . Corneal infection by Nocardia is rare . Trauma is the most common predisposing factor . Isolated case reports of nocardial infection associated with contact lens wear and laser in situ keratomileusis (LASIK) have been reported . The clinical picture usually consists of superficial patchy infiltrates, which may be arranged in a wreath pattern . Presence of gram-positive, branching, beaded filaments that stain with 1% acid-fast stain (using 1% sulfuric acid, modified Kinyoun's method) in smears of corneal scrapings is suggestive of nocardial infection . Nocardia grow on commonly used media as tiny, white, dry colonies . Available knowledge and clinical experience suggest that although sulfacetamide eyedrops can be tried as the initial drug, trimethoprim-sulfamethoxazole and amikacin are effective drugs . Once therapy is initiated, the infiltrate responds promptly and resolves, forming a corneal scar with or without vascularization, and good visual recovery can be expected.

J Food Prot, 2001 Feb, 64(2), 213 - 9
The effect of nisin on the keeping quality of reduced heat-treated milks; Wirjantoro TI et al.; Milk was subjected to a combination process involving reduced heat treatment (RHT) of 117 degrees C for 2 s and nisin (75 and 150 IU ml(-1)) . The microbial activity and other quality aspects were compared with a RHT control (without nisin) and with a ultrahigh temperature (UHT) milk processed at 142 degrees C for 2 s . Nisin was found to inhibit microbial growth for products stored without refrigeration, and RHT-nisin samples stored at 30 degrees C showed very low spoilage rates during 150 days, although not low enough to satisfy requirements for commercial sterility . RHT-nisin samples could be distinguished from and were preferred to the UHT control . Significant browning occurred during storage at 30 degrees C and above but was less in the RHT-nisin milk samples compared with the UHT milk . In RHT-nisin milk samples stored at 20 and 10 degrees C, no microbial activity could be detected in most samples after storage for 1 year . The effectiveness of this combination of RHT, nisin, and low storage temperatures against gram-positive spore-forming bacteria suggests potential for use of nisin in extended shelf life products.

Vet Microbiol, 2001 Apr 19, 79(4), 351 - 65
Influence of Escherichia coli and Arcanobacterium pyogenes isolated from bovine puerperal uteri on phenotypic and functional properties of neutrophils; Zerbe H et al.; When cows develop endometritis after birth, Escherichia coli and Arcanobacterium pyogenes are usually the most prominent bacteria present in bovine uterine lochial secretions . A . pyogenes alone is rarely found in the course of a disturbed puerperium . This was confirmed in this study, since average and high-grade uterine contaminations were always associated with the presence of both bacteria . The contamination grade was positively correlated with uterine polymorphonuclear granulocyte (PMN) numbers and negatively correlated with blood PMN numbers . Whether E . coli and A . pyogenes affect the phenotype and function of bovine PMN in a similar or differential way was subject to in vitro studies . PMN were tested in the presence of washed bacterial fragments or culture supernatants taken as a source for soluble and/or secreted bacterial products . Fragments and soluble products differed only quantitatively in their effects on PMN . Usually, long-time exposure (24h) of PMN to fragments induced the strongest effects . Accelerated death of granulocytes was only moderately induced by both E . coli and A . pyogenes products . Both E . coli and A . pyogenes products induced the enhanced expression of a membrane molecule detected by mAb IL-A110 and of CD11b . Expression of other surface structures remained largely unchanged (MHC class I, CD11c) . Functional parameters of PMN (phagocytosis; generation of reactive oxygen species, ROS; antibody-independent cellular cytotoxicity, AICC) generally declined after pre-incubation for 24h with products of E . coli or A . pyogenes . Interestingly, soluble products of A . pyogenes stimulated the phagocytosis of PMN . However, co-incubation with E . coli products abrogated this stimulatory effect . The results supply evidence for similar modes of action of the gram-negative E . coli and the gram-positive A . pyogenes on bovine PMN . Alterations in PMN function and phenotype are mainly triggered by direct contact between bacterial fragments and PMN . Inhibition experiments with polymyxin B demonstrated that E . coli-mediated effects were not solely due to the action of lipopolysaccharide . The dominant functional depression of neutrophils by E . coli products strengthens the suggestion that the earlier appearance of E . coli in the uterus may support the co-infection of this organ by A . pyogenes at later times.

Glycoconj J, 2000 May, 17(5), 273 - 82
Synthesis and reactivity of the monosaccharide esters of amino acids as models of teichoic acid fragment; Jeric I et al.; The increasing prevalence of sepsis from gram-positive bacterial pathogens necessitates further evaluation of the basic assumptions about the molecular pathogenesis of septic shock . Since diverse physiological functions of gram-positive bacteria are controlled by the degree of esterification of teichoic acids with D-alanine, we examined the reactivity of monosaccharide esters in which anomerically free or protected D-glucose is linked through its C-6 hydroxy group to either phenylalanyl or tyrosyl residues as models for teichoic acid fragment . We show that the attached sugar moiety induces activation of the amino acid residue . Due to the enhanced reactivity of the NH2 group in the monosaccharide esters studied, the formation of products generated by intramolecular and intermolecular glycation reactions is accelerated resulting in heterogeneous mixture of compounds . These findings suggest that, if similar adducts are formed by glycation of D-alanine in teichoic acid of gram-positive bacteria, they should be examined as potential bioactive ligands or chemical message for infection.

Nat Immunol, 2000 Nov, 1(5), 398 - 401
Pattern recognition receptors TLR4 and CD14 mediate response to respiratory syncytial virus; Kurt-Jones EA et al.; The innate immune system contributes to the earliest phase of the host defense against foreign organisms and has both soluble and cellular pattern recognition receptors for microbial products . Two important members of this receptor group, CD14 and the Toll-like receptor (TLR) pattern recognition receptors, are essential for the innate immune response to components of Gram-negative and Gram-positive bacteria, mycobacteria, spirochetes and yeast . We now find that these receptors function in an antiviral response as well . The innate immune response to the fusion protein of an important respiratory pathogen of humans, respiratory syncytial virus (RSV), was mediated by TLR4 and CD14 . RSV persisted longer in the lungs of infected TLR4-deficient mice compared to normal mice . Thus, a common receptor activation pathway can initiate innate immune responses to both bacterial and viral pathogens.

Gene, 2001 Mar 7, 265(1-2), 103 - 13
Cloning and characterization of biotin biosynthetic genes of Kurthia sp; Kiyasu T et al.; The biotin biosynthesis genes of Kurthia sp., which is an aerobic gram-positive bacterium, were cloned from Kurthia sp . 538-KA26 and characterized . Eleven biotin biosynthetic genes have been identified in Kurthia sp . Kurthia sp . has two genes coding for KAPA synthase, bioF and bioFII, and also has two genes coding for BioH protein, bioH and bioHII . In addition, three genes, orf1, orf2, and orf3, whose functions are unknown, were found in the biotin gene clusters of Kurthia sp . The bioA, bioD, and orf1 genes are arranged in a gene cluster in the order orf1bioDA, and the bioB, bioF, and orf2 genes are arranged in a gene cluster in the order orf2bioFB . These gene clusters proceed to both directions; the face to face promoters and two 40-bp of palindrome sequences exist upstream of the orf1 and orf2 genes . The bioC, bioFII, and bioHII genes are arranged in a gene cluster in the order bioFIIHIIC; a 40-bp of palindrome sequence exists upstream of the bioFII gene . The bioH and orf3 genes are arranged in a gene cluster in the order bioHorf3; a palindrome sequence was not found upstream of the bioH gene . These palindrome sequences are extremely similar to each other, suggesting that the orf1bioDA, orf2bioFB, and bioFIIHIIC gene clusters are regulated by biotin . Kurthia sp . does not have the bioW gene coding pimeloyl-CoA synthase, suggesting that pimeloyl-CoA may be produced by a different pathway than that of gram-positive bacterium B . subtilis or B . sphaericus, further suggesting a modified fatty acid synthesis pathway via acetyl-CoA instead as E . coli has.

Biochimie, 2001 Jan, 83(1), 103 - 8
Enzymology of elongation and constriction of the murein sacculus of Escherichia coli; Holtje JV et al.; Multiple deletions in murein hydrolases revealed that predominantly amidases are responsible for cleavage of the septum during cell division . Endopeptidases and lytic transglycosylases seem also be involved . In the absence of these enzymes E . coli grows normally but forms chains of adhering cells . Surprisingly, mutants lacking up to eight different murein hydrolases still grow with almost unaffected growth rate . Therefore it is speculated that general enlargement of the murein sacculus may differ from cell division by using transferases rather than the two sets of hydrolytic and synthetic enzymes as seems to be the case for the constriction process . A model is presented that describes growth of the murein of both Gram-positive and -negative bacteria by the activity of murein transferases . It is speculated that enzymes exist that catalyze a transpeptidation of the pre-existing murein onto murein precursors or nascent murein by using the chemical energy present in peptide cross-bridges . Such enzymes would at the same time cleave bonds in the murein net and insert new material into the growing sacculus.

Mol Microbiol, 2001 Feb, 39(4), 1036 - 47
Mutational analysis of RsrA, a zinc-binding anti-sigma factor with a thiol-disulphide redox switch; Paget MS et al.; In the Gram-positive bacterium, Streptomyces coelicolor A3(2), expression of the thioredoxin system is modulated by a sigma factor called sigmaR in response to changes in the cytoplasmic thiol-disulphide status, and the activity of sigmaR is controlled post-translationally by an anti-sigma factor, RsrA . In vitro, the anti-sigma factor activity of RsrA, which contains seven cysteines, correlates with its thiol-disulphide redox status . Here, we investigate the function of RsrA in vivo . A constructed rsrA null mutant had very high constitutive levels of disulphide reductase activity and sigmaR-dependent transcription, confirming that RsrA is a negative regulator of sigmaR and a key sensor of thiol-disulphide status . Targeted mutagenesis revealed that three of the seven cysteines in RsrA (C11, C41 and C44) were essential for anti-sigma factor activity and that a mutant RsrA protein containing only these three cysteines was active and still redox sensitive in vivo . We also show that RsrA is a metalloprotein, containing near-stoichiometric amounts of zinc . On the basis of these data, we propose that a thiol-disulphide redox switch is formed between two of C11, C41 and C44, and that all three residues play an essential role in anti-sigma factor activity in their reduced state, perhaps by acting as ligands for zinc . Unexpectedly, rsrA null mutants were blocked in sporulation, probably as a consequence of an increase in the level of free sigmaR.

Nat Immunol, 2000 Aug, 1(2), 113 - 8
Secretion of microbicidal alpha-defensins by intestinal Paneth cells in response to bacteria; Ayabe T et al.; Paneth cells in mouse small intestinal crypts secrete granules rich in microbicidal peptides when exposed to bacteria or bacterial antigens . The dose-dependent secretion occurs within minutes and alpha-defensins, or cryptdins, account for 70% of the released bactericidal peptide activity . Gram-negative bacteria, Gram-positive bacteria, lipopolysaccharide, lipoteichoic acid, lipid A and muramyl dipeptide elicit cryptdin secretion . Live fungi and protozoa, however, do not stimulate degranulation . Thus intestinal Paneth cells contribute to innate immunity by sensing bacteria and bacterial antigens, and discharge microbicidal peptides at effective concentrations accordingly.

Crit Care Med, 2001 Feb, 29(2), 438 - 41
Sclerosing cholangitis and liver cirrhosis after extrabiliary infections: report on three cases; Scheppach W et al.; OBJECTIVE: To describe three unusual cases of sclerosing cholangitis after severe extrahepatic/extrabiliary bacterial infections . DESIGN: Case report, clinical . SETTING: Tertiary care intensive care unit (ICU) . PATIENTS: Three patients admitted to the ICU with infections from Gram-positive bacteria followed by sclerosing cholangitis and secondary biliary cirrhosis . MAIN RESULTS: Three unusual cases of persisting cholestasis that occurred after bacterial infections originating from extrahepatic/extrabiliary foci are described . Endoscopic retrograde cholangiopancreatography and magnetic resonance cholangiopancreatography revealed multiple strictures of the intrahepatic bile ducts as a sign of sclerosing cholangitis . All patients progressed to biliary cirrhosis within months after the onset of cholestasis . CONCLUSION: Infection-associated cholestasis is usually a functional disorder and subsides after effective treatment of the underlying inflammatory focus . In rare cases, however, extrahepatic/extrabiliary infections may lead to sclerosing cholangitis and secondary biliary cirrhosis via unknown mechanisms.

Korean J Intern Med, 2000 Dec, 15(3), 240 - 4
A case of endobronchial actinomycosis; Jin SL et al.; Actinomycosis is an infectious disease caused by certain Actinomyces species . Actinomyces are Gram-positive, non-spore forming organisms characterized by obligate or facultative anaerobic rods that normally inhabit anaerobic niches of the human oral cavity . Cervicofacial, abdominal, pelvic and thoracic infections of Actinomyces are not uncommon, but endobronchial actinomycosis is rarely reported . Endobronchial actinomycosis can be misdiagnosed as unresolving pneumonia, endobronchial lipoma or malignancies . Endobronchial actinomycosis should be included in the differential diagnosis of any endobronchial mass . We report a case of a 43-year-old man who presented with a productive cough and pulmonary consolidation at the right lower lobe on chest radiograph . Fiberoptic bronchoscopy revealed obstruction of the right superior segment of the lower bronchus with an exophytic endobronchial mass . Endobronchial actinomycosis was confirmed by demonstration of sulfur granules in the bronchoscopic biopsy of the mass . Intravenous administration of penicillin G followed by oral amoxacillin/clavulanic acid therapy for 3 months resulted in improving symptoms . Infiltrative consolidation on the chest X-ray was markedly decreased.

J Microbiol Methods, 2001 Apr, 44(3), 193 - 203
Distribution and abundance of Gram-positive bacteria in the environment: development of a group-specific probe; MacGregor BJ et al.; We developed a 16S rRNA-targeted oligonucleotide probe (S-P-GPos-1200-a-A-13) for the Gram-positive bacteria, confirmed its specificity by database searches and hybridization studies, and investigated the effects of humic acids on membrane hybridizations with this probe . S-P-GPos-1200-a-A-13 was used to estimate the abundance of Gram-positive populations in the bovine rumen and Lake Michigan sediments . This probe should be useful for studies of the environmental distribution of Gram-positive bacteria and the detection of uncultured, phylogenetically Gram-positive bacteria with variable or negative Gram staining reactions, and could serve for Gram staining in some diagnostic settings.

Acta Paediatr, 2001 Feb, 90(2), 179 - 83
Detection of bacterial DNA by PCR and reverse hybridization in the 16S rRNA gene with particular reference to neonatal septicemia; Shang S et al.; AIM: The clinical diagnosis of sepsis is difficult, particularly in neonates . It is necessary to develop a rapid and reliable method for detecting bacteria in blood and cerebrospinal fluid (CSF) . Polymerase chain reaction (PCR) and reverse hybridization of the 16S rRNA gene would permit fast and sensitive determination of the presence of bacteria and differentiate gram-positive bacteria from gram-negative ones in clinical specimens . METHODS: We developed a pair of primers according to the gene encoding 16SrRNA found in all bacteria . DNA fragments from different bacterial species and from clinical samples were detected with PCR, and with reverse hybridization using a universal bacterial probe, a gram-positive probe and a gram-negative probe . RESULTS: A 371 bp DNA fragment was amplified from 20 different bacterial species . No signal was observed when human DNA and viruses were used as templates . The sensitivity could be improved to 10(-12) g . All 26 culture-positive clinical samples (22 blood samples and 4 CSF samples) were positive with PCR . The gram-negative and gram-positive probes hybridized to clinical samples and to known bacterial controls, as predicted by Gram's stain characteristics . CONCLUSIONS: Our results suggest that the method of PCR and reverse hybridization is rapid, sensitive and specific in detecting bacterial infections . This finding may be significant in the clinical diagnosis of sepsis in neonates.

Acta Microbiol Immunol Hung, 2001, 48(1), 11 - 5
Fleroxacin uptake in ischaemic limb tissue; Miglioli PA et al.; Antibiotic application to patients with ischaemia of lower limbs may be indicated to avoid or treat infection of soft tissues . Fleroxacin, a fluoroquinolone, active against various Gram-negative and Gram-positive organisms may be used for this purpose . We evaluated the diffusion of fleroxacin into bone, subcutaneous fat, muscle and tendon tissues of lower limb tissue after a 400 mg i.v . dose . Concentrations in ischaemic tissues were similar to those found in non-ischaemic sites . Since the maximum antibiotic levels found were lower than the MICs of various pathogens relevant for infection, we suggest to increase the dose used for this peri-operative prophylaxis to 800 mg.

J Med Microbiol, 2001 Mar, 50(3), 249 - 54
Detection of peptostreptococcus micros DNA in clinical samples by PCR; Riggio MP et al.; Peptostreptococcus micros is a gram-positive anaerobic coccus which, although considered to be a natural commensal of the human oral cavity, is associated with periodontal, endodontal and peritonsillar infections . Identification of the organism has to date relied upon conventional culture methods and biochemical analyses . The purpose of this study was to develop a PCR method for rapid and specific identification of this organism in clinical samples . A pair of primers was selected, each of which was specific at the 3' end for P . micros DNA; they were used in the PCR assay, resulting in a 1074-bp product . The primers were shown to be specific for P . micros DNA as no PCR products were obtained when genomic DNA extracts from a wide range of other Peptostreptococcus species and other oral bacteria were used as templates . The PCR assay was then applied to the identification of P . micros DNA in subgingival plaque samples from adult periodontitis patients and pus samples from subjects with acute dento-alveolar abscesses . Confirmation of specific amplification of P . micros DNA was obtained by digestion of PCR products with the restriction endonuclease RsaI, which gives a unique restriction profile for P . micros, and DNA sequencing . Sixty-eight subgingival plaque samples from 18 patients were analysed, of which 19 (28%) were positive for P . micros DNA; the proportion of patients carrying P . micros DNA in at least one sampled site was 11 (61%) of 18 . Twenty (71%) of 28 pus samples analysed by PCR contained P . micros DNA . These results confirm that P . micros may be involved in the aetiology of acute dentoalveolar abscesses and adult periodontitis . The PCR assay provides a more rapid and reliable alternative to conventional methods for identification of P . micros in clinical samples.

J Clin Microbiol, 2001 Mar, 39(3), 1202 - 3
Case of synovitis potentially caused by Dolosigranulum pigrum; Hall GS et al.; We report a case of synovitis in a 64-year-old man who developed the infection while on steroid therapy for rheumatoid arthritis . Dolosigranulum pigrum, a gram-positive catalase-negative coccus, was isolated from two sets of blood cultures prior to antibiotic therapy . The patient was treated with 4 weeks of appropriate antibiotics, and the synovial inflammation resolved . Although synovial aspirates were never positive for any bacteria or fungi, the timing of positive blood cultures and absence of other pathogens suggest the possible etiology as D . pigrum.

Med Wieku Rozwoj, 2000 Oct-Dec, 4(4), 431 - 9
{Multiresistant bacterial strains in infections of children treated at the National Research Institute of Mother and Child in 1999}; Drejewicz H et al.; The aim of the study was the evaluation of multiresistant strains in infections of children treated at the National Research Institute of Mother and Child (NRIMC) . During 1999, 23145 inoculations and 3873 antibiotic susceptibility tests were done . 2711 multiresistant strains were detected of which 59.2% were Gram-negative rods and 40.8% - Gram-positive cocci . The Gram-negative rods were tested for production of beta-lactamases (ESBL), resistance to aminoglicosides, imipenem and piperacillin/tazobactam . Gram-positive cocci were tested for methicilin and aminoglicosides resistance . Cross-resistance between macrolides, lincosamides and streptogramines was also investigated . Significant participation of multiresistant bacterial strains, mainly Gram-negative, in infections among patients treated in intensive care units of the Institute should be stressed.

Chirurg, 2001 Jan, 72(1), 61 - 71
{Effectiveness of an improved antiseptic in treatment of contaminated soft tissue wounds}; Schmit-Neuerburg KP et al.; A novel antiseptic biguanide has been shown to be more bactericidal and tissue compatible in vitro than other antiseptics . In our controlled, prospective and randomized double-blind study on patients with bacteria-contaminated wound types 2-4, one group (n = 45) was treated with humid cotton swab dressings of 0.2% Lavasept solution compared with Ringer solution (n = 35) . No deterioration of wound healing was observed in either group . Lavasept treatment resulted in faster and significant reduction of gram-positive germs . The tissue compatibility of Lavasept was evaluated as significantly better than Ringer solution.

Khirurgiia (Mosk), 2000, (11), 37 - 40
{Thermal trauma in a combination with respiratory tract burn}; Skuba ND et al.; The detection rate of microbial contamination of burn wounds after flame burn was studied in dynamics to 3 weeks in 56 patients forming 2 similar groups: basic--with thermo-inhalation lesions (TIL) and control--without TIL . General area of skin burns in all the patients was less than 50% of the body surface, on average it was 40%, the area of deep burns was 23% . Cytological and bacteriologic methods were used for study microphlora in the sputum and in bronchoalveolar lavage . There were 186 cases of wound microbial contamination (gram-negative microorganisms--64.4% cases, gram-positive--34%, pathogenic fungi--1.6%) . In 80% cases the microphlora of the wounds was polymicrobial, in 47% cases the microphlora were detected on the second week . Microphlora was detected 2 times more often in the basic group than in the control, corresponded to skin burn area; the greatest exceeding of critical level of wound contamination was revealed on the first 2 weeks . In TIL besides wound contamination, polymicrobial microphlora revealed in tracheobronchial tree, presented on the first 2 weeks with association of gram-positive and gram-negative microorganisms, combining on week 3 with fungal phlora . Early leukocytosis, not reducing during first 3 weeks of the study, due to intoxication by burn products, occurred in TIL . Early lymphopenia in TIL was the sign of resistant immunodeficiency.

J Bacteriol, 2001 Mar, 183(6), 2051 - 8
D-alanylation of lipoteichoic acid: role of the D-alanyl carrier protein in acylation; Kiriukhin MY et al.; The D-alanylation of membrane-associated lipoteichoic acid (LTA) in gram-positive organisms requires the D-alanine-D-alanyl carrier protein ligase (AMP) (Dcl) and the D-alanyl carrier protein (Dcp) . The dlt operon encoding these proteins (dltA and dltC) also includes dltB and dltD . dltB encodes a putative transport system, while dltD encodes a protein which facilitates the binding of Dcp and Dcl for ligation with D-alanine and has thioesterase activity for mischarged D-alanyl-acyl carrier proteins (ACPs) . In previous results it was shown that D-alanyl-Dcp donates its ester residue to membrane-associated LTA (M . P . Heaton and F . C . Neuhaus, J . Bacteriol . 176: 681-690, 1994) . However, all efforts to identify an enzyme which catalyzes this D-alanylation process were unsuccessful . It was discovered that incubation of D-alanyl-Dcp in the presence of LTA resulted in the time-dependent hydrolysis of this D-alanyl thioester . D-Alanyl-ACP in the presence of LTA was not hydrolyzed . When Dcp was incubated with membrane-associated D-alanyl LTA, a time and concentration-dependent formation of D-alanyl-Dcp was found . The addition of NaCl to this reaction inhibited the formation of D-alanyl-Dcp and stimulated the hydrolysis of D-alanyl-Dcp . Since these reactions are specific for the carrier protein (Dcp), it is suggested that Dcp has a unique binding site which interacts with the poly(Gro-P) moiety of LTA . It is this specific interaction that provides the functional specificity for the D-alanylation process . The reversibility of this process provides a mechanism for the transacylation of the D-alanyl ester residues between LTA and wall teichoic acid.

Vet Clin North Am Equine Pract, 2000 Dec, 16(3), 443 - 56, viii
Emergent causes of placentitis and abortion; Donahue JM et al.; The clinical signs, laboratory findings, diagnosis, epidemiology, treatment, and prevention and control of two emerging causes of placentitis and abortion in horses are described in this article . Leptospirosis has been reported as a significant cause of fetal loss in horses in Kentucky, Northern Ireland, and England . Most abortions result from infection by serovars kennewicki or bratislava . Nocardioform placentitis has become the most common cause of placentitis in central Kentucky horses . Nocardioform placentitis is associated with infection by unnamed, gram-positive, filamentous, branching bacteria, and is characterized by distinctive changes in the placental membranes.

Plast Reconstr Surg, 2001 Feb, 107(2), 364 - 73
The use of muscle flaps to treat left ventricular assist device infections; Hutchinson OZ et al.; Left ventricular assist devices have become an important adjunct in the therapeutic armamentarium for patients with end-stage heart failure . Although they may provide a bridge to transplantation, they are prone to certain problems, expecially infection . Because these are life-sustaining devices, changing the device or simple explantation may be a risky, if not impossible, option . Therefore, we evaluated the effectiveness of a surgical alternative, namely, coverage of infected devices with muscle or myocutaneous flaps . Eighty-two consecutive patients who underwent the insertion of 88 left ventricular assist devices at our institution over a 6.5-year period were evaluated . Follow-up was provided for all patients and ranged from 1 to 7.5 years . The duration of ventricular support ranged from 0 to 434 days . All patients who demonstrated clinical evidence of infection were identified . Overall, 54 patients (66 percent) had infections locally at the device site, at distant sites, or systemically during support . Cultured organisms included gram-positive and -negative bacteria, fungi, and viruses . Of the 56 infections in these 54 patients, 21 (38 percent) were device-related, i.e., in the pocket created by the device, in the device itself, or from the driveline . Thus, 24 percent (21 of 88) of all ventricular support devices inserted demonstrated device infection during use . Therapeutic modalities used to combat device-related infection included both nonsurgical management with antibiotics alone and surgical procedures such as device change or relocation, device explant, and flap coverage . Eight of the 20 patients in whom the 21 device-related infections occurred underwent surgical intervention . Four of these eight patients undenwent local flap coverage of their infected left ventricular assist devices . All four patients also had evidence of systemic infection, or "device endocarditis." Coverage was successfully achieved in all cases with pedicled rectus abdominis flaps . There were no perioperative complications . Two patients later underwent successful transplantation; the other two died from causes unrelated to the flap . In conclusion, the treatment of infected left ventricular assist devices currently includes both nonsurgical and surgical alternatives . Of the latter, muscle flaps should be considered a first-line intervention to assist in eradicating infection by providing well-vascularized tissue . Although there were no perioperative complications, the 50 percent mortality rate is consistent with that reported for patients with "device endocarditis." It may be that flap coverage of infected ventricular assist devices, if instituted at an earlier stage in the therapeutic process, could help prevent systemic infection in these patients and, therefore, improve their overall outcome.

J Antibiot (Tokyo), 2000 Nov, 53(11), 1272 - 81
Discovery of RWJ-54428 (MC-02,479), a new cephalosporin active against resistant gram-positive bacteria; Hecker SJ et al.; The discovery of RWJ-54428 (MC-02,479), a new cephalosporin displaying promising activity against sensitive and resistant Gram-positive bacteria, is described . Progressive structural modification from the previously reported 3-phenylthiocephem MC-02,331 afforded an overall increase in potency against MRSA while retaining other key properties such as acceptable solubility and serum binding . Evaluation of the in vitro potency and in vivo efficacy of a series of closely related compounds resulted in selection of RWJ-54428 (MC-02,479) for further studies.

Clin Experiment Ophthalmol, 2000 Dec, 28(6), 437 - 42
Cyanoacrylate glue for corneal perforations: a description of a surgical technique and a review of the literature; Vote BJ et al.; The effective early application of a cyanoacrylate glue corneal patch can aid in the management of small corneal perforations, corneal melts and wound leaks . Their use gives improved visual outcomes with reduced enucleation rates (6% vs 19%) . It may also avoid the need for tectonic penetrating keratoplasty . Cyanoacrylate glue prevents re-epithelialization into the zone of damaged and naked stroma and prevents the development of the critical setting for collagenase production that leads to stromal melting . Cyanoacrylates also have significant bacteriostatic activity against gram-positive organisms . We describe a simple and easily reproducible method of cyanoacrylate corneal patch application, with neglible risk of inadvertent glue complications . It has the further advantage of a smooth corneal surface rather than an irregular surface as often occurs with direct application methods . With corneal application, the major concern is toxicity of cyanoacrylates through direct contact with the corneal endothelium and lens . Fibrin glues may be less toxic; however, they are not as readily available . The longer alkyl chains of currently available cyanoacrylate glues (e.g . Histoacryl) slows degradation significantly, limiting accumulation of histotoxic by-products to amounts that can be effectively eliminated by tissues . Vigilance in monitoring for infection/corneal infiltrate is necessary at all times, especially when the glue has been present for more than 6 weeks . Corneal patching with cyanoacrylate glue is a temporizing procedure only, buying time to allow healing secondary to medical treatment of the underlying condition, or allowing surgery to be elective and under more optimal conditions once inflammation has been reduced and the integrity of the globe restored.

Acta Derm Venereol, 2000 Sep-Oct, 80(5), 321 - 8
Superantigens and their association with dermatological inflammatory diseases: facts and hypotheses; Jappe U; Superantigens have been suggested to play an important role in the pathogenesis of several inflammatory skin diseases as well as systemic diseases such as atopic dermatitis, psoriasis, vasculitis, T-cell lymphoma and autoimmune diseases . Infections often precede the onset and relapse of these diseases, and antibiotic treatment with or without additional glucocorticosteroids and immunoglobulins is occasionally successful . Superantigens are microbial proteins that are able to stimulate up to 20% of the naive T-cell population in a non-specific way . They are produced by gram-positive and -negative bacteria as well as by viruses, parasites and yeasts . The importance of the pathogenic role of superantigens is determined by the potency to induce inflammation by extensive cytokine release after T-cell stimulation and/or T-cell-mediated cytotoxicity and, thereby, tissue damage . Furthermore, superantigens may be able to induce autoimmune processes by stimulation of autoreactive T-cells as well as autoantibody production by stimulation of B-cells . Among the diseases associated with superantigens, atopic dermatitis, guttate and chronic plaque psoriasis, as well as Kawasaki disease, are by far the best-characterized . Nevertheless, conflicting results have been obtained and formal proof of a pathogenic role of superantigens in these diseases is still lacking . The aim of this review is to summarize the data on superantigens in terms of their distribution in microorganisms, their interactions with the adaptive immune system and their contribution to skin pathology.

Genes Immun, 2000 Aug, 1(6), 405 - 7
The common functional C(-159)T polymorphism within the promoter region of the lipopolysaccharide receptor CD14 is not associated with sepsis development or mortality; Hubacek JA et al.; Sepsis is characterised by a systemic inflammatory response to bacterial products during infection, which interestingly both in humans and animal models is gender associated with a higher susceptibility of males than females . The CD14 receptor is involved in activation of cells by lipopolysaccharides released from Gram-negative bacteria and, as recently shown, also by products of Gram-positive bacteria (e.g., peptidoglycans and lipoteichoic acid) . The functional relevance of a C(-159)T CD14 polymorphism recently has been shown based on correlation of the T allele to higher plasma levels of soluble CD14, and higher membrane expression on monocytes . We, therefore, now analysed this CD14 polymorphism in 204 patients with severe sepsis and 247 controls . No significant difference of allele frequencies was observed between sepsis patients and controls neither for males nor females . Mortality also was not associated with the polymorphism studied . This may suggest that other mechanisms for lipopolysaccharide recognition, such as the recently described Toll-like receptors are important for inflammatory cell activation in sepsis.

Avian Dis, 2000 Oct-Dec, 44(4), 901 - 6
In vitro effect of ethylenediaminetetraacetic acid-Tris on the efficacy of hatchery disinfectants; Wooley RE et al.; Solutions of ethylenediaminetetraacetic acid (EDTA)-Tris showed synergistic or additive effects on gram-negative bacteria when combined with hatchery disinfectants consisting of phenol and detergent (Magnaphen-100), quaternary ammonium compound (QAC) and glutaraldehyde (Synergize), QAC (BioSentry 904), and hydrogen peroxide . The gram-positive bacteria reacted less favorably, with reaction mixtures showing all three levels of potentiation (synergistie, additive, and antagonistic) . Combinations of EDTA-Tris and a commercial glutaraldehyde solution (Glutracide), when mixed with the test organisms, showed mostly antagonistic effects . Solutions of EDTA-Tris decreased the concentration of hatchery disinfectants required for bacterial killing by 75% in those situations in which synergistic potentiation occurred . EDTA-Tris is nontoxic to 12-day-old embryos . Serial passage of the test organisms in solutions of EDTA-Tris did not result in the development of resistant forms.

Biodegradation, 2000, 11(1), 49 - 53
Degradation of o-methoxybenzoate by a two-member consortium made up of a gram-positive Arthrobacter strain and a gram-negative Pantotea strain; Gil M et al.; Aromatic carboxylic acids substituted with methoxylated groups are among the most abundant products in "alpechin", the wastes resulting from pressing olives to obtain olive oil . Degradation of o-methoxybenzoate by an stable consortium made of a gram positive bacterium, Arthrobacter oxydans, and gram negative one, Pantotea agglomerans, was shown to mineralize this compound efficiently . The concerted action of both microorganisms was needed for the two first steps in the process, namely, the conversion of o-methoxybenzoate into salycilate, and the hydroxylation of the latter to gentisate . Gentisate was further degraded by the Arthrobacter strain.

Med Sci Monit, 2000 Nov-Dec, 6(6), 1113 - 8
Role of Chlamydia trachomatis in epididymitis . Part I: Direct and serologic diagnosis; Zdrodowska-Stefanow B et al.; The aim of the study was to evaluate the role of Chlamydia trachomatis (C.t.) in the etiopathogenesis of epididymitis . 39 patients with symptoms of acute epididymitis were examined . They were divided into two age groups: 1st--patients aged below 35 (24 patients) and 2nd--patients older than 35 yrs (15 patients) . The patients' urethral swabs were examined and C.t., mycoplasma and other Gram-positive and Gram-negative bacteria were searched for as well as leukocyte count was assessed . Anti-Chlamydia trachomatis antibodies of IgG and IgM classes were assayed in serum . Direct immunofluorescence (antigen detection) and immunoenzymatic (antibodies detection) methods were used in C.t . infection diagnostics . C.t . infection in urethra was found in 30.8 percent of patients suffering from epididymitis, whereas in the control group in 2 percent . C.t . infection was significantly more commonly found in patients below 35 (45.8 percent) as compared with the older men (6.7 percent) . The most common etiological factor in the younger group was C.t., while in the older group it was E . coli . IgG anti-C.t . antibodies were detected in the serum of 51.3 percent of the patients with acute epididymitis, as compared with 5.8 percent of the men in the control group . Specific IgG antibodies were found significantly more often in patients under 35 (66.7 percent) than in the older men (26.7 percent) . Specific IgM antibodies were found in 15.4 percent of the patients, that is in 20.8 percent of the younger patients and in 6.7 percent of the older ones . C.t . is the main etiologic agent of epididymitis in men under 35.

J Immunol, 2001 Mar 1, 166(5), 3599 - 605
IC14, an anti-CD14 antibody, inhibits endotoxin-mediated symptoms and inflammatory responses in humans; Verbon A et al.; CD14 is a receptor for cell wall components of Gram-negative and Gram-positive bacteria that has been implicated in the initiation of the inflammatory response to sepsis . To determine the role of CD14 in LPS-induced effects in humans, 16 healthy subjects received an i.v . injection of LPS (4 ng/kg) preceded (-2 h) by i.v . IC14, a recombinant chimeric mAb against human CD14, at a dose of 1 mg/kg over 1 h, or placebo . In subjects receiving IC14, saturation of CD14 on circulating monocytes and granulocytes was >90% at the time of LPS injection . IC14 attenuated LPS-induced clinical symptoms and strongly inhibited LPS-induced proinflammatory cytokine release, while only delaying the release of the anti-inflammatory cytokines soluble TNF receptor type I and IL-1 receptor antagonist . IC14 also inhibited leukocyte activation, but more modestly reduced endothelial cell activation and the acute phase protein response . The capacity of circulating monocytes and granulocytes to phagocytose Escherichia coli was only marginally reduced after infusion of IC14 . These data provide the first proof of principle that blockade of CD14 is associated with reduced LPS responsiveness in humans in vivo.

ALTEX, 1998, 15(5), 9 - 10
{Development and evaluation of a pyrogen test based on human whole blood}; Hartung T et al.; When cells of the immune system, especially blood monocytes and macrophages, come into contact with pyrogenic (fever-inducing) contaminations, they secrete messenger molecules which initiate an hyperthermic reaction in the organism . Of this group of endogenous pyrogens, most is known about interleukin-1 (IL-1) . A new pyrogen test makes use of this reaction as a system for detection: The substances which are to be screened are incubated with a small volume of blood from a healthy donor . Any pyrogens present induce the production of IL-1 which can be detected by ELISA . This test has a higher sensitivity and is more economical than the conventional pyrogen test in rabbits and furthermore reflects the reaction of the relevant species . In contrast to the customary alternative method, the Limulus amoebocyte lysate test (LAL), this test is not restricted to endotoxins from Gram-negative bacteria and is also not hindered by substances which bind endotoxins, such as blood proteins, to the same extent . Consequently, more than 50 non-endotoxin pyrogens have already been traced by this test . The whole blood test is even superior to the LAL in regard to the detection of endotoxins: in a comparison of about 60 endotoxins, there was a correlation of the potency of the individual endotoxins between the whole blood test and the pyrogen test in rabbits, but neither test correlated with the LAL test . In some cases, endotoxins with equal effects in the LAL test differed in potency in the human blood model by a factor of 10 000 . A method has been developed by which cryopreserved blood can be put to use in the test . In this way, blood donations from a donor can be pre-tested so that uniform material may be employed in the test . This test opens up entirely new perspectives on pyrogen testing for Gram-positive or fungal pyrogens as well as in medicinal products . In addition, it could fill the dangerous security gap which might result from the limitations of testing medications and blood products with LAL only . The project was supported by ZEBET, D-Berlin, BMBF, D-Bonn, and set, D-Mainz.

Arch Surg, 2001 Feb, 136(2), 229 - 34
Increased resource use associated with catheter-related bloodstream infection in the surgical intensive care unit; Dimick JB et al.; HYPOTHESIS: Catheter-related bloodstream infection (CRBSI) in critically ill surgical patients with prolonged intensive care unit (ICU) stays is associated with a significant increase in health care resource use . DESIGN: Prospective cohort study . SETTING: Surgical ICU at a large tertiary care center . PATIENTS: Critically ill surgical patients (N = 260) with projected surgical ICU length of stay greater than 3 days . INTERVENTIONS: Central venous catheters were cultured for clinical suspicion of infection . MAIN OUTCOME MEASURES: Increases in total hospital cost, ICU cost, hospital days, and ICU days attributable to CRBSI were estimated using multiple linear regression after adjusting for demographic factors and severity of illness (APACHE III {Apache Physiology and Chronic Health Evaluation III} score) . RESULTS: The incidence of CRBSI per 1000 catheter-days was 3.6 episodes (95% confidence interval {CI}, 2.1-5.8 episodes) . Microbiologic isolates were Gram-positive bacteria in 75%, Gram-negative bacteria in 20%, and yeast in 5% . After adjusting for demographic factors and severity of disease, CRBSI was associated with an increase of $56 167 (95% CI, $11 523-$165 735; P =.001) (in 1998 dollars) in total hospital cost, an increase of $71 443 (95% CI, $11 960-$195 628; P<.001) in ICU cost, a 22-day increase in hospital length of stay, and a 20-day increase in ICU length of stay . CONCLUSIONS: For critically ill surgical patients, CRBSI is associated with a profound increase in resource use . Prevention, early diagnosis, and intervention for CRBSI might result in cost savings in this high-risk population.

Biochem Soc Trans, 2000 Dec, 28(6), 806 - 10
Molecular evolution of lycopene cyclases involved in the formation of carotenoids with ionone end groups; Krubasik P et al.; A survey is given of the lycopene cyclase genes present in bacteria, fungi and plants where two completely unrelated types exist . One is the classical monomeric bacterial beta-cyclase gene, crtY, which may be an ancestor of crtL, the gene for a beta-cyclase in cyanobacteria . From crtL a line of evolution can be drawn to plant beta- and epsilon-cyclase genes and to the gene of capsanthin/capsorubin synthase . In Gram-positive bacteria two genes crtYc and crtYd are present . They encode two proteins which have to interact as a heterodimer for lycopene beta-cyclization . From this type of lycopene cyclase gene the fungal lycopene cyclase/phytoene synthase fusion gene evolved.

J Appl Microbiol, 2001 Feb, 90(2), 237 - 47
Isolation and characterization of a novel hydrocarbon-degrading, Gram-positive bacterium, isolated from intertidal beach sediment, and description of Planococcus alkanoclasticus sp . nov; Engelhardt MA et al.; AIMS: Characterization of a bacterial isolate (strain MAE2) from intertidal beach sediment capable of degrading linear and branched alkanes . METHODS AND RESULTS: A Gram-positive, aerobic, heterotrophic bacterium (strain MAE2), that was capable of extensive degradation of alkanes in crude oil but had a limited capacity for the utilization of other organic compounds, was isolated from intertidal beach sediment . MAE2 had an obligate requirement for NaCl but could not tolerate high salt concentrations . It was capable of degrading branched and n-alkanes in crude oil from C11 to C33, but was unable to degrade aromatic hydrocarbons . Comparative 16S rRNA sequence analysis placed the isolate with members of the genus Planococcus . That finding was corroborated by chemotaxonomic and physiological data . The fatty acid composition of strain MAE2 was very similar to the type species of the genus Planococcus, P . citreus (NCIMB 1493T) and P . kocurii (NCIMB 629T), and was dominated by branched acids, mainly a15:0 . However, the 16S rRNA of strain MAE2 had less than 97% sequence identity with the type strains of P . citreus (NCIMB 1439T), P . kocurii (NCIMB 629T) and two Planococcus spp . (strain MB6-16 and strain ICO24) isolated from Antarctic sea ice . This indicated that strain MAE2 represented a separate species from these planococci . Morphologically, the isolate resembled P . okeanokoites (NCIMB 561T) and P . mcmeekinii S23F2 (ATCC 700539T) . The cellular fatty acid composition of P . okeanokoites and P . mcmeekinii was considerably different from strain MAE2, and the mol % G + C content of P . mcmeekinii was far lower than that of MAE2 . CONCLUSION: On the basis of phenotypic and genotypic data, it is proposed that strain MAE2 is a new species of Planococcus, Planococcus alkanoclasticus sp . nov., for which the type strain is P . alkanoclasticus MAE2 (NCIMB 13489T) . SIGNIFICANCE AND IMPACT OF THE STUDY: Planococcus species are abundant members of the bacterial community in a variety of marine environments, including some in sensitive Antarctic ecosystems . The occurrence of hydrocarbon-degrading Planococcus spp . is potentially of importance in controlling the impact of hydrocarbon contamination in sensitive marine environments.

J Appl Microbiol, 2001 Feb, 90(2), 180 - 9
Filter-based PNA in situ hybridization for rapid detection, identification and enumeration of specific micro-organisms; Perry-O'Keefe H et al.; AIMS: A method for rapid and simultaneous detection, identification and enumeration of specific micro-organisms using Peptide Nucleic Acid (PNA) probes is presented . METHODS AND RESULTS: The method is based on a membrane filtration technique . The membrane filter was incubated for a short period of time . The microcolonies were analysed by in situ hybridization, using peroxidase-labelled PNA probes targeting a species-specific rRNA sequence, and visualized by a chemiluminescent reaction . Microcolonies were observed as small spots of light on film, thereby providing simultaneous detection, identification and enumeration . The method showed 95-100% correlation to standard plate counts along with definitive identification due to the specificity of the probe . CONCLUSION: Using the same protocol, results were generated approximately three times faster than culture methods for Gram-positive and -negative bacterial species and yeast species . SIGNIFICANCE AND IMPACT OF THE STUDY: The method is an improvement on the current membrane filtration technique, providing rapid determination of the level of specific pathogens, spoilage or indicator micro-organisms.

J Appl Microbiol, 2001 Feb, 90(2), 163 - 71
Evaluation of microbial proteolysis in meat products by capillary electrophoresis; Martin A et al.; AIMS: The available methods for evaluating proteolysis in meat products, particularly the contribution of micro-organisms, are expensive, time-consuming and require an unacceptable sample size . To minimize these problems, two capillary electrophoresis-based methods have been developed . METHODS AND RESULTS: Six Gram-positive, catalase-positive cocci, four moulds and three yeasts, isolated from dry-cured ham, were tested on sterile pork slices . Using the Capillary Gel Electrophoresis (CGE) method, changes in sarcoplasmic and myofibrillar proteins due to endogenous and microbial enzymes were detected . The Capillary Zone Electrophoresis (CZE) analysis allowed evaluation of bulk changes by micro-organisms in soluble nitrogen compounds . CONCLUSION: CGE analysis of myofibrillar proteins and CZE determination of soluble nitrogen compounds have proved to be valuable tools for evaluating proteolytic activity of endogenous and microbial origin . SIGNIFICANCE AND IMPACT OF THE STUDY: The CGE and CZE methods developed can be used for a rapid and sensitive analysis of proteolysis in meat products.

Clin Exp Immunol, 2001 Jan, 123(1), 140 - 6
Reduced systemic IgG levels against peptidoglycan in rheumatoid arthritis (RA) patients; Schrijver IA et al.; The gut flora is believed to play a role in the pathogenesis of RA . Peptidoglycan, a major cell wall component of Gram-positive bacteria, is a candidate antigen because of its capability to trigger production of proinflammatory cytokines, to induce arthritis in rodents, and because of its presence in antigen-presenting cells in RA joints . We investigated whether the systemic and local antibody levels against a peptidoglycan-polysaccharide (PG-PS) are related to the presence and disease activity of RA . Significantly lower levels of systemic IgG directed against PG-PS were found in healthy females compared with healthy males, and systemic IgA levels specific for PG-PS were negatively correlated with age . Levels of systemic IgG directed against PG-PS were significantly reduced in RA patients compared with sex- and age-matched healthy controls . Local (synovial fluid) levels of IgG did not correlate with disease activity whereas synovial fluid levels of IgA correlated positively with disease activity . These data suggest that IgG in healthy people mediates protection against spreading of PG to non-mucosal sites.

Acta Ophthalmol Scand, 2001 Feb, 79(1), 97 - 9
Foreign body orbital cyst; Yazdanfard Y et al.; PURPOSE: To present the clinical and histopathological characteristics of a retained orbital foreign body . METHODS: A 34-year-old male was hit in his left eye by a flower stake . At first examination there was a massive haematoma of the left orbit and eyelids, reduced movement of the left eye and a small conjunctival lesion beneath the upper lid . There were no signs of deeper orbital or ocular lesions . During the subsequent months the haematoma disappeared, but increased proptosis and impaired eye movements with diplopia were noticed . One year after the injury, ultrasound and CT-scan revealed an ovoid cystic tumour behind the globe . During surgery the content of the cyst appeared purulent and from the centre of the cyst a foreign body measuring 25 x 13 x 1 mm was removed . RESULTS: Microscopy of the foreign body showed the typical structure of a decidual leaf . The superficial cells were empty, whereas the centrally located cells contained remnants of cytoplasm . Confined to the border between the empty and the filled cells, a band of cells containing groups of gram positive cocci was noted . Staining for immunoglobulins revealed traces of IgG in the superficial empty cells only . CONCLUSION: The high degree of preservation of the leaf and the survival of the cocci may be due to a barrier function of the intact plant cell walls with their high content of cellulose.

Hum Immunol, 2001 Jan, 62(1), 50 - 63
Interactions of bacterial lipopolysaccharide and peptidoglycan with a 70 kDa and an 80 kDa protein on the cell surface of CD14+ and CD14- cells; Triantafilou K et al.; Bacterial cell wall components, lipopolysaccharide (LPS), lipoteichoic acid (LTA), and peptidoglycan (PGN) are known to stimulate cells of the immune, inflammatory and vascular systems contributing to septic shock . CD14 has been identified as the main LPS receptor, a process that is accelerated by the serum protein LPS-binding protein (LBP) . CD14 has also been found to bind LTA and PGN from the cell wall of gram positive bacteria . Recently, toll-like receptor proteins TLR-2 and TLR-4 have been shown to be required for LPS and LTA-induced intracellular signalling . Although CD14 functions as either a glycosylphosphatidylinositol (GPI)-anchored molecule that does not transverse the cell membrane or as a soluble serum protein, the mechanisms by which the CD14-LPS/LTA complex interacts with the TLRs remains to be elucidated . We have looked directly for cell surface protein(s) that bind LPS or LTA in a CD14-dependent manner . Using biochemical approaches we have identified two proteins of molecular weight 70 kDa (LAP-1) and 80 kDa (LAP-2) that can be precipitated from both CD14(+) and CD14(-) cells with LPS- or LTA-specific antibodies . Binding of LPS and LTA to LAP-1 and -2 required serum . While soluble CD14 (sCD14) was sufficient to allow precipitation of these two proteins from CD14(-) cells, serum could not be replaced by purified sCD14 and/or LBP when mCD14-expressing cells were used.

J Immunol, 2001 Feb 1, 166(3), 2018 - 24
Bacterial lipopolysaccharide and IFN-gamma induce Toll-like receptor 2 and Toll-like receptor 4 expression in human endothelial cells: role of NF-kappa B activation; Faure E et al.; Toll-like receptor (TLR) 4 has been identified as the primary receptor for enteric LPS, whereas TLR2 has been implicated as the receptor for Gram-positive and fungal cell wall components and for bacterial, mycobacterial, and spirochetal lipoproteins . Vascular endothelial cell (EC) activation or injury by microbial cell wall components such as LPS is of critical importance in the development of sepsis and septic shock . We have previously shown that EC express predominantly TLR4, and have very little TLR2 . These cells respond vigorously to LPS via TLR4, but are unresponsive to lipoproteins and other TLR2 ligands . Here we show that LPS, TNF-alpha, or IFN-gamma induce TLR2 expression in both human dermal microvessel EC and HUVEC . Furthermore, LPS and IFN-gamma act synergistically to induce TLR2 expression in EC, and LPS-induced TLR2 expression is NF-kappaB dependent . LPS and IFN-gamma also up-regulate TLR4 mRNA expression in EC . These data indicate that TLR2 and TLR4 expression in ECs is regulated by inflammatory molecules such as LPS, TNF-alpha, or IFN-gamma . TLR2 and TLR4 molecules may render EC responsive to TLR2 ligands and may help to explain the synergy between LPS and lipoproteins, and between LPS and IFN-gamma, in inducing shock associated with Gram-negative sepsis.

J Immunol, 2001 Feb 1, 166(3), 1938 - 44
MD-2 enables Toll-like receptor 2 (TLR2)-mediated responses to lipopolysaccharide and enhances TLR2-mediated responses to Gram-positive and Gram-negative bacteria and their cell wall components; Dziarski R et al.; MD-2 is associated with Toll-like receptor 4 (TLR4) on the cell surface and enables TLR4 to respond to LPS . We tested whether MD-2 enhances or enables the responses of both TLR2 and TLR4 to Gram-negative and Gram-positive bacteria and their components . TLR2 without MD-2 did not efficiently respond to highly purified LPS and LPS partial structures . MD-2 enabled TLR2 to respond to nonactivating protein-free LPS, LPS mutants, or lipid A and enhanced TLR2-mediated responses to both Gram-negative and Gram-positive bacteria and their LPS, peptidoglycan, and lipoteichoic acid components . MD-2 enabled TLR4 to respond to a wide variety of LPS partial structures, Gram-negative bacteria, and Gram-positive lipoteichoic acid, but not to Gram-positive bacteria, peptidoglycan, and lipopeptide . MD-2 physically associated with TLR2, but this association was weaker than with TLR4 . MD-2 enhanced expression of both TLR2 and TLR4, and TLR2 and TLR4 enhanced expression of MD-2 . Thus, MD-2 enables both TLR4 and TLR2 to respond with high sensitivity to a broad range of LPS structures and to lipoteichoic acid, and, moreover, MD-2 enhances the responses of TLR2 to Gram-positive bacteria and peptidoglycan, to which the TLR4-MD-2 complex is unresponsive.

Appl Environ Microbiol, 2001 Feb, 67(2), 880 - 7
Evaluation of PCR-generated chimeras, mutations, and heteroduplexes with 16S rRNA gene-based cloning; Qiu X et al.; To evaluate PCR-generated artifacts (i.e., chimeras, mutations, and heteroduplexes) with the 16S ribosomal DNA (rDNA)-based cloning approach, a model community of four species was constructed from alpha, beta, and gamma subdivisions of the division Proteobacteria as well as gram-positive bacterium, all of which could be distinguished by HhaI restriction digestion patterns . The overall PCR artifacts were significantly different among the three Taq DNA polymerases examined: 20% for Z-Taq, with the highest processitivity; 15% for LA-Taq, with the highest fidelity and intermediate processitivity; and 7% for the conventionally used DNA polymerase, AmpliTaq . In contrast to the theoretical prediction, the frequency of chimeras for both Z-Taq (8.7%) and LA-Taq (6.2%) was higher than that for AmpliTaq (2.5%) . The frequencies of chimeras and of heteroduplexes for Z-Taq were almost three times higher than those of AmpliTaq . The total PCR artifacts increased as PCR cycles and template concentrations increased and decreased as elongation time increased . Generally the frequency of chimeras was lower than that of mutations but higher than that of heteroduplexes . The total PCR artifacts as well as the frequency of heteroduplexes increased as the species diversity increased . PCR artifacts were significantly reduced by using AmpliTaq and fewer PCR cycles (fewer than 20 cycles), and the heteroduplexes could be effectively removed from PCR products prior to cloning by polyacrylamide gel purification or T7 endonuclease I digestion . Based upon these results, an optimal approach is proposed to minimize PCR artifacts in 16S rDNA-based microbial community studies.

Appl Environ Microbiol, 2001 Feb, 67(2), 591 - 7
Development of a gene cloning and inactivation system for halorespiring Desulfitobacterium dehalogenans; Smidt H et al.; Efficient host-vector systems have been developed for the versatile, strictly anaerobic, halo- and fumarate-respiring gram-positive bacterium Desulfitobacterium dehalogenans . An electroporation-based transformation procedure resulting in approximately 10(3) to 10(4) transformants per microg of the cloning vector pIL253 was developed and validated . The broad-host-range vector pG+host9 was shown to replicate at a permissive temperature of 30 degrees C, whereas the replicon was not functional at 40 degrees C . The D . dehalogenans frdCAB operon, predicted to encode a fumarate reductase, was cloned, characterized, and targeted for insertional inactivation by pG+host9 carrying a 0.6-kb internal frdA fragment . Single-crossover integration at the frdA locus occurred at a frequency of 3.3 x 10(-4) per cell and resulted in partially impaired fumarate reductase activity . The gene cloning and inactivation systems described here provide a solid basis for the further elucidation of the halorespiratory network in D . dehalogenans and allow for its further exploitation as a dedicated degrader.

Crit Care Med, 2000 Mar, 28(3), 638 - 42
Phase II multicenter clinical study of the platelet-activating factor receptor antagonist BB-882 in the treatment of sepsis; Vincent JL et al.; OBJECTIVE: To evaluate the safety and efficacy of the platelet-activating factor receptor antagonist BB-882 in the treatment of patients with sepsis . DESIGN: Double-blind, placebo-controlled, randomized, multi-centered study . SETTING: Thirty-four European intensive care units . PATIENTS: One hundred fifty-two patients with clinical suspicion of infection and a mean APACHE II score between 15 and 35 in the 24 hrs before entry into the trial . INTERVENTIONS: Patients received either a loading dose of 4 mg of BB-882 on the first day, followed by an intravenous infusion of 96 mg/24 hrs for up to 120 hrs, or placebo . MEASUREMENTS: Hemodynamic, respiratory and oxygen transport variables, blood lactate concentrations, interleukin-6, interleukin-8, tumor necrosis factor (TNF)-alpha, soluble TNF receptor concentrations, organ failure score, 28-day mortality rate, Acute Physiology And Chronic Health Evaluation (APACHE) II score within 24 hrs of entry . RESULTS: Sixty-nine patients (42 male, 27 female) received placebo and 83 (59 male, 24 female) received BB-882 . Patients ranged in age from 16 to 89 yrs (mean, 60 yrs) . No important differences existed between the two groups in terms of gender distribution, age, or initial APACHE II score . Sepsis was identified as Gram-positive in 49 patients, Gram-negative in 40, mixed in 37, and unknown in 26 . No important differences were shown in hemodynamic, respiratory, or oxygen transport variables between groups during the study . Organ failure scores were similar in the two groups throughout the study . Cytokine concentrations were not significantly different in the two groups . Within 28 days of entering the study, 75 patients died, including 31 (45%) in the placebo group and 44 (53%) in the treatment group, p = .32 . The median time to death in the placebo group was 6.0 days, and in the treatment group, it was 4.5 days (p = .30) . CONCLUSION: Treatment of sepsis with the platelet-activating factor antagonist BB-882 offers no advantage over placebo on survival, hemodynamic status, respiratory function, or organ failure scores.

J Biol Chem, 2001 Mar 30, 276(13), 9631 - 9 Epub 2000 Dec 22.
Biochemical properties and cDNa cloning of two new lectins from the plasma of Tachypleus tridentatus: Tachypleus plasma lectin 1 and 2+; Chen SC et al.; A Sepharose CL-4B-binding protein, Tachypleus plasma lectin 1 (TPL-1), and a lipopolysaccharide (LPS)-binding protein, Tachypleus plasma lectin-2 (TPL-2), have been isolated from the plasma of Tachypleus tridentatus and biochemically characterized . Each protein is coded by a homologous family of multigenes . TPL-1 binds to Sepharose CL-4B and was eluted with buffer containing 0.4 m GlcNAc . The deduced amino acid sequence of TPL-1 consisted of 232 amino acids with an N-glycosylation site, Asn-Gly-Ser at residues 74-76 . It shares a 65% sequence identity and similar internal repeats of about 20 amino acid motifs with tachylectin-1 . Tachylectin-1 was identified as a lipopolysaccharide-agarose binding nonglycosylated protein from the amebocytes of T . tridentatus . TPL-2 was eluted from the LPS-Sepharose CL-4B affinity column in buffer containing 0.4 m GlcNAc and 2 m KCl . The deduced amino acid sequence of TPL-2 consisted of 128 amino acids with an N-glycosylation site, Asn-Cys-Thr, at positions 3-5 . It shares an 80% sequence identity with tachylectin-3, isolated from the amebocytes of T . tridentatus . TPL-2 purified by LPS-affinity column from the plasma predominantly exists as a dimer of a glycoprotein with an apparent molecular mass of 36 kDa . Tachylectin-3 is an intracellular nonglycosylated protein that also exists as a dimer in solution with an apparent molecular mass of 29 kDa . It recognizes Gram-negative bacteria through the 0-antigen of LPS . Western blot analyses showed that, in the plasma, TPL-1 and TPL-2 exist predominantly as oligomers with molecular masses above 60 kDa . They both bind to Gram-positive and Gram-negative bacteria, and this binding is inhibited by GlcNAc . Possible binding site of TPL-1 and TPL-2 to the bacteria could be at the NAc moiety of GlcNAc-MurNAc of the peptidoglycan . The physiological function of TPL-1 and TPL-2 is most likely related to their ability to form a cluster of interlocking molecules to immobilize and entrap invading organisms.

Int J Syst Evol Microbiol, 2000 Nov, 50 Pt 6, 2037 - 41
Characterization of a Gemella-like organism isolated from an abscess of a rabbit: description of Gemella cunicula sp . nov; Hoyles L et al.; An unknown Gram-positive, catalase-negative, ovoid-shaped bacterium isolated from the submandibular abscess of a rabbit was subjected to a polyphasic taxonomic analysis . Comparative 16S rRNA gene sequencing demonstrated the unknown coccus represents a new subline within the genus Gemella . The unknown isolate was readily distinguished from other recognized members of the genus Gemella, namely Gemella haemolysans, Gemella bergeri, Gemella morbillorum, Gemella palaticanis and Gemella sanguinis, by biochemical tests and electrophoretic analysis of whole-cell proteins . Based on both phylogenetic and phenotypic evidence, it is proposed that the unknown bacterium is classified in the genus Gemella as Gemella cuniculi sp . nov . The type strain is CCUG 42726T.

Appl Microbiol Biotechnol, 2000 Dec, 54(6), 799 - 807
Identification of Amycolatopsis sp . strain HR167 genes, involved in the bioconversion of ferulic acid to vanillin; Achterholt S et al.; The gene loci ech, encoding enoyl-CoA hydratase/aldolase, and fcs, encoding an unusual feruloyl-CoA synthetase, which are involved in the bioconversion of ferulic acid to vanillin by the gram-positive bacterium Amycolatopsis sp . strain HR167, were localized on a 4,000 bp PstI fragment (P40) . The nucleotide sequence of P40 was determined, revealing open reading frames of 864 bp and 1,476 bp, representing ech and fcs, respectively . The deduced amino acid sequences of ech exhibited 62% amino acid identity to the enoyl-CoA hydratase/aldolase from Pseudomonas sp . strain HR199 and the enoyl-CoA hydratase/lyase from P . fluorescens strain AN103 . The deduced amino acid sequences of fcs exhibited up to 37% amino acid identity to long-chain fatty acid coenzymeA ligases but no significant similarity to the feruloyl-CoA synthetase of Pseudomonas sp . strain HR199 . Fragment P40 was cloned in pBluescript SK- and fcs and ech were expressed in Escherichia coli . Recombinant strains were able to transform ferulic acid to vanillin . In crude extracts of these recombinant strains, feruloyl-CoA synthetase and enoyl-CoA hydratase/aldolase activities were detected by photometric assay and high-performance liquid chromatography . The obtained data suggest that ferulic acid degradation in the gram-positive Amycolatopsis sp . strain HR167 proceeds via a pathway similar to that recently described for the gram-negative P . fluorescens strain AN103 and Pseudomonas sp . strain HR199.

Pediatr Infect Dis J, 2000 Dec, 19(12), 1178 - 84
Single dose pharmacokinetics of linezolid in infants and children; Kearns GL et al.; BACKGROUND: Linezolid is an oxazolidinone antibiotic with excellent in vitro activity against a number of Gram-positive organisms including antibiotic-resistant isolates . The safety and pharmacokinetics of intravenously administered linezolid were evaluated in children and adolescents to examine the potential for developmental dependence on its disposition characteristics . METHODS: Fifty-eight children (3 months to 16 years old) participated in this study; 44 received a single 1.5-mg/kg dose and 14 received a single 10-mg/kg dose of linezolid administered by intravenous infusion . Repeated blood samples (n = 10 in children > or = 12 months; n = 8 in children 3 to 12 months) were obtained during 24 h after drug administration, and linezolid was quantitated from plasma by high performance liquid chromatography with mass spectrometry detection . Plasma concentration vs . time data were evaluated with a model independent approach . RESULTS: Linezolid was well-tolerated by all subjects . The disposition of linezolid appears to be age-dependent . A significant although weak correlation between age and total body clearance was observed . The mean (+/- SD) values for elimination half-life, total clearance and apparent volume of distribution were 3.0 +/- 1.1 h, 0.34 +/- 0.15 liter/h/kg and 0.73 +/- 0.18 liter/kg, respectively . Estimates of total body clearance and volume of distribution were significantly greater in children than historical values of adult data . As such maximum achievable linezolid plasma concentrations were slightly lower in children, and concentrations 12 h after a single 10-mg/kg dose were below the MIC90 for selected pathogens with in vitro susceptibility to the drug . CONCLUSION: Based on these data a linezolid dose of 10 mg/kg given two to three times daily would appear appropriate for use in pediatric therapeutic clinical trials of this agent.

J Am Soc Mass Spectrom, 2001 Jan, 12(1), 49 - 54
Exploring the limits of bacterial identification by intact cell-mass spectrometry; Evason DJ et al.; The limits of intact cell-mass spectrometry (ICM-MS) were tested with regard to the minimum number of bacterial cells detectable and its power to discriminate mixed-bacterial cultures . The technique is a surface analysis tool, as is supported by evidence showing that mass fingerprints correspond to material desorbed directly from the cell wall . The brief exposure to solvents, which occurs during sample preparation, does not extract internal cellular material . Spectra were collected over the m/z range of 500 to 10,000 . The UV absorbing matrices used were found to be highly specific to bacterial gram type: alpha-cyano-4-hydroxycinnamic acid for gram-negative bacteria and 5-chloro-2-mercaptobenzothiazole for gram-positive bacteria . This specificity allows mixed cultures of different gram types to be differentiated by ICM-MS . The minimum number of cells that could reliably give spectra of sufficient data was 10(4) cells (10(7) cells/mL).

Infection, 2000 Nov-Dec, 28(6), 367 - 73
Clinical infections and bloodstream isolates associated with fever in patients undergoing chemotherapy for acute myeloid leukemia; Madani TA; BACKGROUND: Patients with acute myeloid leukemia (AML) are at high risk for infections.The aim of this study was to identify the sources of fever and the type of pathogens that cause bloodstream infection in patients with AML undergoing cytotoxic chemotherapy and antibiotic prophylaxis . PATIENTS AND METHODS: The source of fever and the type of pathogens causing bloodstream infection were identified for 129 febrile episodes experienced by 42 patients with AML receiving cytotoxic chemotherapy and antibiotic prophylaxis . RESULTS: A source of fever was identified in 81% of all febrile episodes . Mucositis (21.7%), pneumonia (13.2%), central venous catheter infection (12.4%), neutropenic enterocolitis (9.3%) and invasive fungal disease (9.3%) were the most common sources of fever . Of 16 central venous catheter infections, seven (43.8%) were not associated with local signs . 49 febrile episodes (37.9%) were associated with bloodstream infections, of which 14 (28.6%) were polymicrobic and seven (14.3%) had an undefined source of infection . Bloodstream infection was commonly associated with cellulitis (60%), mucositis (57.1%), central venous catheter infection (55.6%), neutropenic enterocolitis (41.7%) and invasive fungal disease (41.7%) . Gram-positive microorganisms were the most common blood isolates (75.8%) . Gram-negative bacteremic infections occurred in eight episodes (12.1%) experienced by patients who were not receiving ciprofloxacin prophylaxes at the time of bacteremia . Noninfectious sources of fever accounted for 23 (17.8%) of the 129 febrile episodes . CONCLUSION: Although the spectrum of pathogens that cause infection in this group of patients has shifted from gram-negative to gram-positive bacteria, the most common sources of infection remain the same as previously described and they mainly involve integumental surfaces.

Am J Kidney Dis, 2001 Jan, 37(1), 49 - 55
Initial treatment of peritoneal dialysis peritonitis without vancomycin with a once-daily cefazolin-based regimen; Goldberg L et al.; To reduce the use of vancomycin, the current recommendations of the International Society of Peritoneal Dialysis (PD) for the initial treatment of peritonitis complicating PD are to administer intraperitoneal (IP) cefazolin or cephalothin in every PD fluid bag, together with once-daily gentamicin . In view of the inherent impracticalities of this regimen, we studied the efficacy of once-daily cefazolin (1.5 g) IP with gentamicin IP as initial treatment for primary (nonrecurrent) PD peritonitis . This regimen has been used in all episodes of peritonitis not associated with tunnel or exit-site infections or fluid leaks . Sixty-nine episodes in 61 patients were analyzed (44 patients, continuous ambulatory PD; 22 patients, automated PD; and 3 patients, hospital-based intermittent PD), of which 38 episodes (55%) were gram-positive infections, 6 episodes (9%) were gram-negative infections, and 18 episodes (26%) had negative culture results . Four patients died within 4 weeks of infection (none considered attributable to inadequate treatment of their peritonitis) . Ten catheters (14.5%) required removal to clear the infection; 7 catheters were in patients with gram-negative infections . The relapse rate within 4 weeks of ceasing antibiotic therapy was 8.9% . Compared with the results of 40 episodes of peritonitis treated initially with our previous IP vancomycin and gentamicin regimen, successful treatment (no death, catheter removal, or recurrence) was achieved in 52 of 69 episodes in the cefazolin group (75.4%) versus 23 of 40 episodes in the vancomycin group (57.5%; P: = 0.058) . In conclusion, once-daily IP cefazolin and gentamicin for the initial treatment of PD peritonitis is at least as effective as a vancomycin-based regimen and is well tolerated.

Pediatr Res, 2001 Jan, 49(1), 81 - 3
Ontogeny of Toll-like receptors Tlr2 and Tlr4 in mice; Harju K et al.; Toll-like receptors (TLR:) have recently been linked to the immunostimulatory function of microbial toxins in human and mice . TLR: signals activation of nuclear factor kappaB that leads to the production of a number of proinflammatory mediators . TLR:4 mediates the endotoxin-induced inflammatory response, whereas TLR:2 may be involved in the response to yeast and Gram-positive bacterial products . To better understand age-related changes in acute inflammatory response, we studied the ontogeny of TLR:2 and TLR:4 mRNA in murine fetal lung, liver, and placenta by quantitative reverse transcriptase-PCR . Different expression patterns were seen between the tissues and between the TLR: This is in accordance with the evidence that there are differences in the receptors for different microbial toxins and that the response is organ specific . We additionally show that the expression of TLR: was dependent on the stage of differentiation . In the liver, the levels of Tlr2 and Tlr4 were high regardless of the age . In the lung, Tlr2 and Tlr4 expression levels were barely detectable in immature fetus (d 14-15) . Tlr2 and Tlr4 were increased several-fold during prenatal development and further increased after birth . The present results support the finding of a deficient inflammatory response of the immature lung to microbial toxins.

Appl Environ Microbiol, 2001 Jan, 67(1), 190 - 7
Phylogenetic specificity and reproducibility and new method for analysis of terminal restriction fragment profiles of 16S rRNA genes from bacterial communities; Dunbar J et al.; Terminal restriction fragment (TRF) analysis of 16S rRNA genes is an increasingly popular method for rapid comparison of microbial communities, but analysis of the data is still in a developmental stage . We assessed the phylogenetic resolution and reproducibility of TRF profiles in order to evaluate the limitations of the method, and we developed an essential analysis technique to improve the interpretation of TRF data . The theoretical phylogenetic resolution of TRF profiles was determined based on the specificity of TRFs predicted from 3,908 16S rRNA gene sequences . With sequences from the Proteobacteria or gram-positive division, as much as 73% of the TRFs were phylogenetically specific (representing strains from at most two genera) . However, the fraction decreased when sequences from the two divisions were combined . The data show that phylogenetic inference will be most effective if TRF profiles represent only a single bacterial division or smaller group . The analytical precision of the TRF method was assessed by comparing nine replicate profiles of a single soil DNA sample . Despite meticulous care in producing the replicates, numerous small, irreproducible peaks were observed . As many as 85% of the 169 distinct TRFs found among the profiles were irreproducible (i.e., not present in all nine replicates) . Substantial variation also occurred in the height of synonymous peaks . To make comparisons of microbial communities more reliable, we developed an analytical procedure that reduces variation and extracts a reproducible subset of data from replicate TRF profiles . The procedure can also be used with other DNA fingerprinting techniques for microbial communities or microbial genomes.

J Dairy Sci, 2000 Dec, 83(12), 2753 - 8
Influence of fluorescence of bacteria stained with acridine orange on the enumeration of microorganisms in raw milk; Rapposch S et al.; The staining of gram-positive and gram-negative cultures with acridine orange in metabolically active and inactive states was investigated using a Bactoscan, direct epifluorescent filter technique (DEFT), and standard plate count as the reference method . The evaluation of the bacterial cultures in the Bactoscan revealed a linear relationship between Bactoscan counts (pulses) and the quantity of pure culture suspension used . But the proper detection of bacteria with the fluorescence optic methods was dependent on the type of microorganism and the physiological state of the cells . The Bactoscan and DEFT underestimated the bacterial counts of gram-negative cultures as compared with standard plate counting . When stained with acridine orange, metabolically active bacteria showed more orange fluorescence and a lower percentage of green fluorescent cells as compared with inactive bacteria . Bactoscan pulse height analysis (PHA) diagrams, graphs of the detected pulses and their intensity, showed low pulses of inactive bacteria . Many of these weak pulses were eliminated from counting because of their faint fluorescent staining . In contrast, PHA diagrams of metabolically active microorganisms showed bright staining and, therefore, high pulses . A complete count of these bacteria was possible . These investigations point out that discrepancies between the fluorescence optical counting methods and the standard plate count depend strongly on the staining of the cultures with acridine orange and, therefore, on the type of microorganism and the metabolic state of the cells measured.

J Biol Chem, 2001 Mar 30, 276(13), 10234 - 46 Epub 2000 Dec 20.
The RepE initiator is a double-stranded and single-stranded DNA-binding protein that forms an atypical open complex at the onset of replication of plasmid pAMbeta 1 from Gram-positive bacteria; Le Chatelier E et al.; The RepE protein of the broad host range pAMbeta1 plasmid from Gram-positive bacteria is absolutely required for replication . To elucidate its role, we purified the protein to near homogeneity and analyzed its interactions with different nucleic acids using gel retardation assays and footprinting experiments . We show that RepE is monomeric in solution and binds specifically, rapidly, and durably to the origin at a unique double-stranded binding site immediately upstream from the initiation site of DNA replication . The binding induces only a weak bend (31 degrees ) . Unexpectedly, RepE also binds nonspecifically to single-stranded DNA with a 2-4-fold greater affinity than for double-stranded origin . On a supercoiled plasmid, RepE binding to the double-stranded origin leads to the denaturation of the AT-rich sequence immediately downstream from the binding site to form an open complex . This open complex is atypical since (i) its formation requires neither multiple RepE binding sites on the double-stranded origin nor strong bending of the origin, (ii) it occurs in the absence of any cofactors (only RepE and supercoiling are required), and (iii) its melted region serves as a substrate for RepE binding . These original properties together with the fact that pAMbeta1 replication depends on a transcription step through the origin on DNA polymerase I to initiate replication and on a primosome to load the replisome suggest that the main function of RepE is to assist primer generation at the origin.

Ann Plast Surg, 2000 Dec, 45(6), 629 - 32
The effects of hyperglycemia on skin graft survival in the burn patient; Mowlavi A et al.; The authors elected to determine the relative effects of hyperglycemia and/or elevated wound Gram-positive bacterial counts on success of skin graft survival in 74 burn patients . Results of serum glucose and quantitative wound biopsies on the day of admission and on postoperative day 4 were charted . Cases were separated into the following groups for analysis: normoglycemia plus normal bacterial counts, elevated bacterial counts only, hyperglycemia only, and hyperglycemia plus elevated bacterial counts . Successful graft "take" was defined as survival of 80% to 100% of the grafted area as assessed on postoperative day 4 . Significant results included decreased incidence of graft take for groups with hyperglycemia only (62.5%), elevated bacterial counts only (63.3%), as well as hyperglycemia plus elevated bacterial counts (54.5%) when compared with the group with normoglycemia plus normal bacterial counts (92.8%; p = 0.020, p = 0.042, p = 0.012 respectively) for physiological parameters measured on postoperative day 4 only . Additionally, incidence of graft take was reassessed and found to be decreased significantly in groups with hyperglycemia (60.0%) vs . groups with normoglycemia (84.6%), regardless of Gram-positive bacterial counts (p = 0.034).

Clin Orthop, 2000 Dec, (381), 91 - 100
Assessment of patient selection criteria for treatment of the infected hip arthroplasty; Hanssen AD et al.; In a group of 37 infected hip arthroplasties (36 patients) treated consecutively between January 1997 and October 1999, the feasibility of published patient selection criteria for direct exchange arthroplasty was tested . These criteria include the requirement of a healthy patient with good soft tissues, minimal femoral bone loss, and an organism identified preoperatively as an antibiotic sensitive gram-positive organism . After assignment of the selection criteria, only four patients (four hips) (11%) were deemed potential candidates for a direct exchange procedure . Infected arthroplasties excluded from a primary exchange included 14 patients (15 hips) with gram-negative or methicillin-resistant gram-positive organisms obtained from preoperative joint aspirations, 10 patients (10 hips) with moderate or severe femoral bone loss, four patients (four hips) who required a proximal femoral osteotomy for component removal, two patients (two hips) with poor health status, and two patients (two hips) with poor soft tissues . Because of the increasing emergence of antibiotic-resistant bacteria and an increased prevalence of revision arthroplasties with associated bone loss, the feasibility of published selection criteria for direct exchange are limited . A philosophy of delayed reconstruction for the treatment of the infected hip arthroplasty seems most appropriate in the current era of patient treatment.

Paediatr Anaesth, 2001 Jan, 11(1), 93 - 8
Postoperative infectious complications in paediatric liver transplantation: a study of 48 transplants; Bouchut JC et al.; We studied, retrospectively, postoperative infectious complications following paediatric liver transplantation at a single university centre . The objectives were to characterize the epidemiology of infection and to determine the associated risk factors during the early postoperative period, either the first postoperative month or the entire duration of paediatric intensive care unit (PICU) stay . Forty-eight liver transplants were performed on 46 patients . Sixty-three infections occurred in 32 patients who underwent 34 liver transplantations (1.36 infection/patient); 47 were bacterial, 6 fungal and 10 viral . The most common sites of infection were bloodstream (36.5%) and abdomen (30%) . Gram-positive bacteria (78%) predominated over gram-negative bacteria (22%) . Initial analysis revealed infection risk factors to be age <1 year, body weight <10 kg, extrahepatic biliary atresia, intraoperative transfusion > 160 ml x kg(-1), mechanical ventilation > 8 days and PICU stay > 19 days . After stratified analysis, the main risk factor for infection was low body weight of the recipient.

J Infect, 2000 Nov, 41(3), 221 - 6
Use of polymerase chain reaction (PCR) and DNA probe hybridization to determine the Gram reaction of the infecting bacterium in the intraocular fluids of patients with endophthalmitis; Anand AR et al.; OBJECTIVES: To evaluate polymerase chain reaction (PCR) combined with DNA probe hybridization to determine the Gram reaction of the bacterium in intraocular specimens from patients with infectious endophthalmitis . METHODS: Fifty-seven intraocular specimens - 17 aqueous humor (AH) and 40 vitreous fluid (VF) - from 55 patients with clinically diagnosed infectious endophthalmitis and 25 control intraocular specimens from non-infectious ocular disorders (10 AH and 15 VF) were evaluated by microscopy, culture and PCR-DNA probe hybridization to detect the Gram reaction of the bacterium . RESULTS: PCR-DNA probe hybridization was specific and sensitive to detect 30 fg of both gram-positive and gram-negative bacterial DNA . None of the controls showed bacteria by microscopy, culture or PCR . Of the 57 intraocular specimens, conventional microbiological methods could detect a bacterial aetiology in 32 (56.1%), while PCR-DNA probe hybridization could detect 52 (91.2%) specimens . This difference was statistically significant (P= 0.003) . In bacteriologically positive specimens, there was absolute correlation of the Gram reaction between the results of smear and culture methods and PCR-DNA probe hybridization . Of the 25 bacteriologically negative specimens, 20 (80%) were positive by PCR-DNA probe hybridization, of which seven (35%) were gram-positive, 12 (60%) gram-negative and one (5%) positive by both . Results of PCR on AH and VF were not significantly different . CONCLUSION: PCR and DNA probe hybridization to determine the Gram reaction of the bacterium in intraocular fluids is a specific and sensitive method in the diagnosis of bacterial endophthalmitis . AH is an ideal specimen for PCR, since its collection is a simple and safe office procedure .

Rapid Commun Mass Spectrom, 2000, 14(23), 2220 - 9
On-probe sample pretreatment for the detection of proteins above 15 KDa from whole cell bacteria by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry; Madonna AJ et al.; A rapid methodology is described for the enhancement of the signal-to-base-line (S/B) ratio of high molecular weight protein signals from whole cell bacteria analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) . The procedure involves depositing growing bacteria colonies from culture dishes directly onto the MALDI probe followed by treatment of the sample spot with a 2 microL aliquot of 40% ethanol prior to the addition of a ferulic acid matrix solution (12.5 mg dissolved in 17% formic acid/33% acetonitrile/50% H(2)O) . Protein signals of more than 20 kDa were routinely produced from both Gram positive and Gram negative bacteria prepared in this manner . Moreover, a substantial number of intense protein signals were also produced in the more 'conventional' fingerprint region extending from 4 to 20 kDa . This approach is rapid, easy to implement into existing methodologies, and does not require any special hardware .

Gene, 2000 Nov 27, 258(1-2), 55 - 62
Genetic and biochemical characterization of a protein phosphatase with dual substrate specificity in Streptomyces coelicolor A3(2); Umeyama T et al.; A gene encoding a protein phosphatase (SppA) with a phosphoesterase motif, which was predicted by the genome project of the Gram-positive bacterium Streptomyces coelicolor A3(2), was cloned by PCR in pET32a(+) and expressed in Escherichia coli . SppA fused to thioredoxin (TRX-SppA) showed distinct heat-stable phosphatase activity toward p-nitrophenyl phosphate with optimal pH 8.0 and optimal temperature 55 degrees C . Mn2+ greatly enhanced enzyme activity, as is found with other protein Ser/Thr phosphatases . TRX-SppA was not inhibited by sodium orthovanadate or okadaic acid, both of which are known to be specific inhibitors of protein phosphatases . TRX-SppA showed phosphatase activity toward not only phosphoThr (pThr) and pTyr but also oligopeptides containing pSer, pThr, and pTyr, indicating that SppA is a protein phosphatase with dual substrate specificity . Disruption of the chromosomal sppA gene resulted in severe impairment of vegetative growth . All of these observations show that SppA, a protein phosphatase with dual specificity, plays an important, but not essential, role in vegetative growth of S . coelicolor A3(2) . The presence of a single copy of sppA in all the 13 Streptomyces species examined, as determined by Southern hybridization, suggests a common role of SppA in general in Streptomyces species.

Acta Clin Belg, 2000 Sep-Oct, 55(5), 279 - 84
Penicillin or vancomycin plus ceftazidime in febrile neutropenic patients after stem cell transplantation; Kargar Samani K et al.; The response of gram-positive cocci to third generation cephalosporin therapy in febrile neutropenic patients is not optimal . We evaluated the safety and efficacy of ceftazidime plus penicillin (C + P) and compared it to ceftazidime plus vancomycin (C + V) in febrile neutropenic patients . The study includes 64 patients admitted to the Department of Haematology . Thirty-six patients were treated with C + V and 28 patients with C + P . Control of infection was observed in 78% of the patients in the C + V group and in 57% of the patients in C + P group (p = 0.5) . Infection was the cause of the death of 1 patient in each group . We conclude that the combination of C + P has the same activity as the combination of C + V in febrile neutropenic patients . The morbidity and mortality were identical in both groups but cost effectiveness was in favour of the C + P group.

Przegl Lek, 2000, 57(7-8), 374 - 7
{Tissue factor and tissue factor pathway inhibitor in peritonitis treated with dialysis}; Malyszko J et al.; TFPI-tissue factor pathway inhibitor appears to play a primary role in regulating TF-induced coagulation . During CAPD procoagulant and anticoagulant activities of the mesothelium are balanced under normal conditions . The aim of the work was to assess TF and TFPI concentrations during peritonitis in CAPD patients . The study was performed in 9 CAPD subjects with peritonitis and 14 clinically stable CAPD patients . TF, total, free and truncated TFPI, thrombomodulin concentrations were measured in plasma and dialysate; C-reactive protein and tumor necrosis factor-TNF alpha were assayed in serum . In 8 patients with S . aureus peritonitis TF and TFPI were not found in dialysate, but were detectable in a case with E . coli peritonitis . C-reactive protein and TNF alpha significantly elevated at the beginning of peritonitis, fell sharply after the therapy . Further studies are needed to establish whether the kind of bacterial peritonitis (Gram-positive or negative) may affect TF and TFPI in plasma and dialysate in CAPD patients.

Int Microbiol, 1998 Dec, 1(4), 259 - 64
Induction of microbial secondary metabolism; Demain AL; Precursors often stimulate production of secondary metabolites either by increasing the amount of a limiting precursor, by inducing a biosynthetic enzyme (synthase) or both . These are usually amino acids but other small molecules also function as inducers . The most well-known are the auto-inducers which include gamma-butyrolactones (butanolides) of the actinomycetes, N-acylhomoserine lactones of Gram-negative bacteria, oligopeptides of Gram-positive bacteria, and B-factor (3'-{1-butylphosphoryl} adenosine) of Amycolatopsis mediterranei . The actinomycete butanolides exert their effects via receptor proteins which normally repress chemical and morphological differentiation (secondary metabolism and differentiation into aerial mycelia and spores respectively) but, when complexed with the butanolide, can no longer function . Homoserine lactones of Gram-negative bacteria function at high cell density and are structurally related to the butanolides . They turn on plant and animal virulence, light emission, plasmid transfer, and production of pigments, cyanide and beta-lactam antibiotics . They are made by enzymes homologous to Lux1, excreted by the cell, enter other cells at high density, bind to a LuxR homologue, the complex then binding to DNA upstream of genes controlled by "quorum sensing" and turning on their expression . Quorum sensing also operates in the case of the peptide pheromones of the Gram-positive bacteria . Here, secretion is accomplished by an ATP binding casette (ABC transporter), the secreted pheromone being recognized by a sensor component of a two-component signal transduction system . The pheromone often induces its own synthesis as well as those proteins involved in protein/peptide antibiotic (including bacteriocins and lantibiotics) production, virulence and genetic competence . The B-factor of A . mediterranei is an inducer of ansamycin (rifamycin) formation.

J Vet Pharmacol Ther, 2000 Oct, 23(5), 273 - 80
Comparison of plasma pharmacokinetics and bioequivalence of ceftiofur sodium in cattle after a single intramuscular or subcutaneous injection; Brown SA et al.; Ceftiofur sodium, a broad-spectrum cephalosporin, is active against gram-positive and gram-negative pathogens of veterinary importance . This study was designed to compare the bioequivalence of the sodium salt in cattle after a single intramuscular (i.m.) or subcutaneous dose (s.c.) of 2.2 mg ceftiofur equivalents/kg body weight . The criteria used to evaluate bioequivalence were (1) the area under the curve from time of injection to the limit of quantitation (LOQ) of the assay (AUC0-LOQ), and (2) time concentrations remained above 0.2 microg/mL (t>0.2) . Twelve crossbred beef cattle were enrolled in a three-period, two-treatment crossover trial, with a minimum 2-week washout period between doses of 2.2 mg ceftiofur equivalents/kg . Blood samples were collected serially for up to 72 h post-injection . Plasma samples were then analyzed using a validated assay that measures ceftiofur, and all desfuroylceftiofur-related metabolites, by high-performance liquid chromatography (HPLC) as the stable derivative, desfuroylceftiofur acetamide . A maximum plasma concentration (Cmax) of 13.9+/-3.55 microg/mL was observed from 0 . 67-2.0 h after i.m . administration, whereas a Cmax of 13.6+/-3.85 microg/mL was observed from 0.67-3.0 h after s.c . administration . The AUC0-LOQ was 108+/-35.0 microg . h/mL after i.m . dosing, compared with 105+/-29.8 microg . h/mL after s.c . dosing . The pre-established criterion for equivalence of the AUC0-LOQ for the i.m . and s.c . routes of administration was satisfied . The t>0.2 was 49.2+/-8.55 h after i.m . administration, compared with 47.0+/-9.40 h after s.c . administration . The pre-established criterion for equivalence of the t>0.2 for i.m . and s.c . administration was satisfied . The equivalence of AUC0-LOQ and t>0.2 for i.m . and s.c . administration of 2.2 mg ceftiofur equivalents (CE)/kg doses of ceftiofur sodium suggest similar therapeutic efficacy and systemic safety for the two routes of administration.

Vet Pathol, 2000 Nov, 37(6), 650 - 2
Pyogranulomatous meningoencephalitis due to Actinomyces sp . in a dog; Couto SS et al.; Actinomyces sp . are commensal, filamentous, gram-positive, acid-fast-negative bacteria that can cause pyogranulomatous inflammation in animals and humans . Central nervous system (CNS) disease is a rare presentation of actinomycosis and is usually due to extension from infected wounds or seeding from distant sites . A dog with progressive, poorly localized neurologic signs had primary CNS actinomycosis without history or evidence of previous trauma or other organ involvement . Histologically, there was a severe pyogranulomatous meningoencephalitis with intralesional filamentous bacteria that were also visible on cytology of the cerebral spinal fluid (CSF) postmortem . Actinomyces sp . was cultured postmortem from the CSF, confirming the diagnosis . This case demonstrates that Actinomyces sp . can be a causative agent of primary CNS disease in dogs.

Life Sci, 2000 Oct 13, 67(21), 2639 - 45
Effects of nitric oxide synthase inhibitors on the febrile response to lipopolysaccharide and muramyl dipeptide in guinea pigs; Kamerman P et al.; We investigated the effect of N-nitro-L-arginine methyl ester (L-NAME), an unspecific nitric oxide synthase (NOS) inhibitor, and aminoguanidine, a relatively selective inhibitor of the inducible NOS enzyme, on both gram-negative lipopolysaccharide (LPS) and gram-positive muramyl dipeptide (MDP) fever in guinea pigs . Intraperitoneal injection of either 10 mg/kg L-NAME or 25 mg/kg aminoguanidine inhibited the febrile response to an intramuscular injection of 50 microg/kg MDP . However, LPS fever (20 microg/kg) was inhibited only by L-NAME . The development of LPS fever may therefore occur independently of the synthesis of nitric oxide by the inducible NOS enzyme, while MDP fever may involve synthesis of nitric oxide by both the inducible and the constitutively expressed NOS enzymes.

FEMS Microbiol Ecol, 2000 Dec 1, 34(2), 103 - 111
Identification and characterization of humic substances-degrading bacterial isolates from an estuarine environment; Esham EC et al.; Bacterial isolates were obtained from enrichment cultures containing humic substances extracted from estuarine water using an XAD-8 resin . Eighteen isolates were chosen for phylogenetic and physiological characterization based on numerical importance in serial dilutions of the enrichment culture and unique colony morphology . Partial sequences of the 16S rRNA genes indicated that six of the isolates were associated with the alpha subclass of Proteobacteria, three with the gamma-Proteobacteria, and nine with the Gram-positive bacteria . Ten isolates degraded at least one (and up to six) selected aromatic single-ring compounds . Six isolates showed ability to degrade {(14)C}humic substances derived from the dominant salt marsh grass in the estuary from which they were isolated (Spartina alterniflora), mineralizing 0.4-1.1% of the humic substances over 4 weeks . A mixture of all 18 isolates did not degrade humic substances significantly faster than any of the individual strains, however, and no isolate degraded humic substances to the same extent as the natural marine bacterial community (3.0%) . Similar studies with a radiolabeled synthetic lignin ({beta-(14)C}dehydropolymerisate) showed measurable levels of degradation by all 18 bacteria (3.0-8.8% in 4 weeks), but mineralization levels were again lower than that observed for the natural marine bacterial community (28.2%) . Metabolic capabilities of the 18 isolates were highly variable and generally did not map to phylogenetic affiliation.

J Antibiot (Tokyo), 2000 Sep, 53(9), 928 - 33
Xanthoepocin, a new antibiotic from Penicillium simplicissimum IFO5762; Igarashi Y et al.; A new antifungal antibiotic xanthoepocin was isolated from the culture broth of Penicillium simplicissimum IFO5762 . Xanthoepocin was obtained from the culture fluid by solvent extraction and chromatographic purification . It showed antibiotic activity against Gram-positive bacteria and yeasts.

FEMS Microbiol Ecol, 2000 Sep 1, 33(3), 241 - 249
Polyphasic microbial community analysis of petroleum hydrocarbon-contaminated soils from two northern Canadian communities; Juck D et al.; The cold-adapted bacterial communities in petroleum hydrocarbon-contaminated and non-impacted soils from two northern Canadian environments, Kuujjuaq, Que., and Alert, Nunavut, were analyzed using a polyphasic approach . Denaturing gradient gel electrophoresis (DGGE) separation of 16S rDNA PCR fragments from soil total community DNA revealed a high level of bacterial diversity, as estimated by the total number of bands visualized . Dendrogram analysis clustered the sample sites on the basis of geographical location . Comparison of the overall microbial molecular diversity suggested that in the Kuujjuaq sites, contamination negatively impacted diversity whereas in the Alert samples, diversity was maintained or increased as compared to uncontaminated controls . Extraction and sequencing analysis of selected 16S rDNA bands demonstrated a range of similarity of 86-100% to reference organisms, with 63.6% of the bands representing high G+C Gram-positive organisms in the order Actinomycetales and 36.4% in the class Proteobacteria . Community level physiological profiles generated using Biolog GN plates were analyzed by cluster analysis . Based on substrate oxidation rates, the samples clustered into groups similar to those of the DGGE dendrograms, i.e . separation based upon geographic origin . The coinciding results reached using culture-independent and -dependent analyses reinforces the conclusion that geographical origin of the samples, rather than petroleum contamination level, was more important in determining species diversity within these cold-adapted bacterial communities.

Clin Infect Dis, 2000 Dec, 31(6), 1357 - 61 Epub 2000 Nov 29.
Population-based study of the risk and short-term prognosis for bacteremia in patients with liver cirrhosis; Thulstrup AM et al.; We examined the risk of bacteremia in patients with liver cirrhosis (compared with the risk for all Danish citizens >20 years of age who were living in North Jutland County, Denmark), as well as the type of bacteremia and the 30-day case-fatality rate . We used the Danish National Registry of Patients to identify 1339 patients with liver cirrhosis, and we used the North Jutland County Bacteremia Database to identify episodes of bacteremia . We observed 117 cases of bacteremia in patients with liver cirrhosis (11.0 cases were expected), which yielded a standardized incidence ratio of 10.5 (95% confidence interval {CI}, 8.8-12.7) . Sixty-two cases of bacteremia were nosocomial infections . There were 53 cases of gram-positive bacteremia, 55 cases of gram-negative bacteremia, and 8 cases of polymicrobial bacteremia (1 case of candidemia was excluded from the analysis) . The most common cause of death was bleeding from gastroesophageal varices; the second most common cause of death was infection in the respiratory system . The 30-day case-fatality rate was 0.53 (95% CI, 0.39-0.73) . Patients with liver cirrhosis had an increased risk of bacteremia and a poor prognosis.

Proc Natl Acad Sci U S A, 2000 Dec 5, 97(25), 13766 - 71
The repertoire for pattern recognition of pathogens by the innate immune system is defined by cooperation between toll-like receptors; Ozinsky A et al.; Toll-like receptors (TLRs) have been shown to participate in the recognition of pathogens by the innate immune system, but it is not clear how a restricted family of receptors has the capacity to recognize the wide spectrum of TLR stimuli known to exist . We report here that two members of the TLR family, TLR2 and TLR6, together coordinate macrophage activation by Gram-positive bacteria and the yeast cell-wall particle, zymosan . TLR6 and TLR2 both are recruited to the macrophage phagosome, where they recognize peptidoglycan, a Gram-positive pathogen component . By contrast, TLR2 recognizes another component, bacterial lipopeptide, without TLR6 . The requirement for TLR cooperation is supported by the finding that TLR2 needs a partner to activate tumor necrosis factor-alpha production in macrophages . Dimerization of the cytoplasmic domain of TLR2 does not induce tumor necrosis factor-alpha production in macrophages, whereas similar dimerization of the TLR4 cytoplasmic domain does . We show that the cytoplasmic domain of TLR2 can form functional pairs with TLR6 or TLR1, and this interaction leads to cytokine induction . Thus, the cytoplasmic tails of TLRs are not functionally equivalent, with certain TLRs requiring assembly into heteromeric complexes, whereas others are active as homomeric complexes . Finally, we show that TLR6, TLR2, and TLR1 are recruited to macrophage phagosomes that contain IgG-coated erythrocytes that do not display microbial components . The data suggest that TLRs sample the contents of the phagosome independent of the nature of the contents, and can establish a combinatorial repertoire to discriminate among the large number of pathogen-associated molecular patterns found in nature.

Med Klin (Munich), 2000 Oct 15, 95(10), 568 - 77
{Infectiology update . II: Diagnosis and therapy}; Salzberger B et al.; In this article, relevant recent changes in the diagnosis and therapy of infectious diseases are reviewed . Several new diagnostic tools for the quantification of viral DNA and RNA have been developed in the last years (PCR-based and related methods) . A number of new antiviral agents has been licensed for the treatment of different viral infections . The clinical management of viral infections has dramatically changed with these tools and agents . Especially morbidity and mortality of HIV-infected patients have both been reduced by antiretroviral combination therapy . With rising rates of antibiotic resistance in gram-positive bacteria, therapy of infections with these agents has become increasingly difficult . A number of new antibiotic agents for the treatment of these infections have been developed and licensed.

J Mol Biol, 2000 Dec 1, 304(3), 311 - 21
Relationship between exopolysaccharide production and protein-tyrosine phosphorylation in gram-negative bacteria; Vincent C et al.; The phosphorylation of proteins at tyrosine residues is known to play a key role in the control of numerous fundamental processes in animal systems . In contrast, the biological significance of protein-tyrosine phosphorylation in bacteria, which has only been recognised recently, is still unclear . Here, we have analysed the role in Escherichia coli cells of an autophosphorylating protein-tyrosine kinase, Wzc, and a phosphotyrosine-protein phosphatase, Wzb, by performing knock-out experiments on the corresponding genes, wzc and wzb, and looking at the metabolic consequences induced . The results demonstrate that the phosphorylation of Wzc, as regulated by Wzb, is directly connected with the production of a particular capsular polysaccharide, colanic acid . Thus, when Wzc is phosphorylated on tyrosine, no colanic acid is synthesised by bacteria, but when dephosphorylated by Wzb, colanic acid is produced . This process is rather specific to the pair of proteins Wzc/Wzb . Indeed, a much lesser effect, if any, on colanic acid synthesis is observed when knock-out experiments are performed on another pair of genes, etk and etp, which also encode respectively a protein-tyrosine kinase, Etk, and a phosphotyrosine-protein phosphatase, Etp, in E . coli . In addition, the analysis of the phosphorylation reaction at the molecular level reveals differences between Gram-negative and Gram-positive bacteria, namely in the number of protein components required for this reaction to occur .

Intensive Care Med, 2000 Sep, 26(9), 1350 - 4
The use of the arterial line as a source for blood cultures; Levin PD et al.; OBJECTIVE: To determine the reliability of blood cultures obtained through indwelling arterial lines as compared to that of blood cultures obtained by venipuncture . DESIGN: A prospective observational study . SETTING: Six-bed mixed medical surgical intensive care unit (ICU) of a 550-bed university-affiliated medical center . MEASUREMENTS: During a 3-month period blood culture sets, when clinically indicated, were drawn in parallel from indwelling arterial catheters and one-time venipuncture and the results compared . Each blood sample consisted of 15 ml and was distributed equally between three blood culture bottles: aerobic, anaerobic and one aerobic resin-containing bottle . Blood culture results from the two sources were compared according to preset definitions . MAIN RESULTS: During the study period 90 parallel blood culture sets (540 bottles) were obtained from 36 patients . Forty-three (16%) venipuncture bottles were positive versus 88 (32%) arterial line culture bottles (p < 0.001) . Of the parallel sets, 83% yielded equivalent results - either both sterile or both growing the same organism . Amongst the discordant sets, the arterial line cultures grew 37 gram-positive and 18 gram-negative isolates not found in venipuncture sets (i.e . 50% of 109 arterial line isolates), while only two gram-positive isolates were solely grown in venipuncture cultures (4% of all 55 venipuncture isolates, p < 0.001) . On clinical correlation, all the gram-positive organisms in the discordant cultures were found not to reflect bacteremia, while five of the 18 gram-negative isolates (28%) grown only in arterial line cultures probably did reflect ongoing bacteremia . CONCLUSION: The results of blood cultures taken from the arterial line are frequently equivalent to those taken from venipuncture . When discordant, the growth of gram-positive bacteria almost certainly reflects contamination or arterial line colonization, whereas the growth of gram-negative bacteria may have to be considered as reflecting bacteremia.

Ann Thorac Surg, 2000 Oct, 70(4), 1404 - 5
Diagnosis and management of left ventricular assist device valve-endocarditis: LVAD valve replacement; de Jonge KC et al.; A 57-year-old man who has been wearing a Novacor N100 left ventricular assist device (LVAD) for more than 3 years suffered from LVAD endocarditis . Only by immunoscintigraphic methods was it possible to localize the septic focus . After successful exchange of in- and outflow tract valves, the infection was eradicated . Microscopic investigation confirmed the scintigraphic findings: Gram-positive bacteria were found . The valves showed no gross degenerative lesions after more than 1,100 days of implantation . The patient is now doing well.

Nature, 2000 Nov 2, 408(6808), 111 - 5
Structural basis for signal transduction by the Toll/interleukin-1 receptor domains; Xu Y et al.; Toll-like receptors (TLRs) and the interleukin-1 receptor superfamily (IL-1Rs) are integral to both innate and adaptive immunity for host defence . These receptors share a conserved cytoplasmic domain, known as the TIR domain . A single-point mutation in the TIR domain of murine TLR4 (Pro712His, the Lps(d) mutation) abolishes the host immune response to lipopolysaccharide (LPS), and mutation of the equivalent residue in TLR2, Pro681His, disrupts signal transduction in response to stimulation by yeast and gram-positive bacteria . Here we report the crystal structures of the TIR domains of human TLR1 and TLR2 and of the Pro681His mutant of TLR2 . The structures have a large conserved surface patch that also contains the site of the Lps(d) mutation . Mutagenesis and functional studies confirm that residues in this surface patch are crucial for receptor signalling . The Lps(d) mutation does not disturb the structure of the TIR domain itself . Instead, structural and functional studies indicate that the conserved surface patch may mediate interactions with the down-stream MyD88 adapter molecule, and that the Lps(d) mutation may abolish receptor signalling by disrupting this recruitment.

Biochim Biophys Acta, 2001 Nov 15, 1524(1), 17 - 26
Bacterial peptidoglycan binds to tubulin; Dziarski R et al.; A search for cellular binding proteins for peptidoglycan (PGN), a CD14- and TLR2-dependent macrophage activator from Gram-positive bacteria, using PGN-affinity chromatography and N-terminal micro-sequencing, revealed that tubulin was a major PGN-binding protein in mouse macrophages . Tubulin also co-eluted with PGN from anti-PGN vancomycin affinity column and bound to PGN coupled to agarose . Tubulin-PGN binding was preferential under the conditions that promote tubulin polymerization, required macromolecular PGN, was competitively inhibited by soluble PGN and tubulin, did not require microtubule-associated proteins, and had an affinity of 100-150 nM . By contrast, binding of tubulin to lipopolysaccharide (LPS) had 2-3 times lower affinity, faster kinetics of binding, and showed positive cooperativity . PGN enhanced tubulin polymerization in the presence of 4 M glycerol, but in the absence of glycerol, both PGN and LPS decreased microtubule polymerization . These results indicate that tubulin is a major PGN-binding protein and that PGN modulates tubulin polymerization.

J Mol Microbiol Biotechnol, 2000 Oct, 2(4), 425 - 32
Antibiotic selective markers and spirochete genetics; Hardham JM et al.; Until very recently, the pathogenic spirochetes have been refractory to genetic manipulation . This has been due, in part, to difficulties with in vitro growth and the genetic distance that spirochetes are from typical Gram-negative and Gram-positive organisms . Insertional mutagenesis and other genetic techniques are now possible in some of the pathogenic spirochetes such as Borrelia burgdorferi, Brachyspira (Serpulina) hyodysenteriae, Leptospira sp., and Treponema denticola . However, organisms such as Treponema pallidum, which cannot be grown in vitro, are still not amenable to genetic manipulation . These recent advances have paved the way for more detailed genetic studies of transcriptional regulation, protein function, protein localization, metabolic capabilities, motility, and pathogenic nature of this group of spirochetes . This review article will discuss the current repertoire of antibiotic markers that are useful for spirochetal genetic manipulation . Further advances in selectable markers and shuttle vectors will allow researchers to complete Koch's molecular hypothesis for various virulence genes of the pathogenic spirochetes and increase the overall understanding of these challenging bacteria.

Mol Microbiol, 2000 Oct, 38(2), 335 - 47
The CtsR regulator of stress response is active as a dimer and specifically degraded in vivo at 37 degrees C; Derre I et al.; CtsR (class three stress gene repressor) negatively regulates the expression of class III heat shock genes (clpP, clpE and the clpC operon) by binding to a directly repeated heptanucleotide operator sequence (A/GGTCAAA NAN A/GGTCAAA) . CtsR-dependent genes are expressed at a low level at 37 degrees C and are strongly induced under heat shock conditions . We performed a structure/function analysis of the CtsR protein, which is highly conserved among low G+C Gram-positive bacteria . Random chemical mutagenesis, in vitro cross-linking, in vivo co-expression of wild-type and mutant forms of CtsR and the construction of chimeric proteins with the DNA-binding domain of the lambda CI repressor allowed us to identify three different functional domains within CtsR: a helix-turn-helix DNA-binding domain, a dimerization domain and a putative heat-sensing domain . We provide evidence suggesting that CtsR is active as a dimer . Transcriptional analysis of a clpP'-bgaB fusion and/or Western blotting experiments using antibodies directed against the CtsR protein indicate that ClpP and ClpX are involved in CtsR degradation at 37 degrees C . This in turn leads to a low steady-state level of CtsR within the cell, as CtsR negatively autoregulates its own synthesis . This is the first example of degradation of a repressor of stress response genes by the Clp ATP-dependent protease.

Ann Surg, 2000 Nov, 232(5), 619 - 26
Acute necrotizing pancreatitis: treatment strategy according to the status of infection; Buchler MW et al.; OBJECTIVE: To determine benefits of conservative versus surgical treatment in patients with necrotizing pancreatitis . SUMMARY BACKGROUND DATA: Infection of pancreatic necrosis is the most important risk factor contributing to death in severe acute pancreatitis, and it is generally accepted that infected pancreatic necrosis should be managed surgically . In contrast, the management of sterile pancreatic necrosis accompanied by organ failure is controversial . Recent clinical experience has provided evidence that conservative management of sterile pancreatic necrosis including early antibiotic administration seems promising . METHODS: A prospective single-center trial evaluated the role of nonsurgical management including early antibiotic treatment in patients with necrotizing pancreatitis . Pancreatic infection, if confirmed by fine-needle aspiration, was considered an indication for surgery, whereas patients without signs of pancreatic infection were treated without surgery . RESULTS: Between January 1994 and June 1999, 204 consecutive patients with acute pancreatitis were recruited . Eighty-six (42%) had necrotizing disease, of whom 57 (66%) had sterile and 29 (34%) infected necrosis . Patients with infected necrosis had more organ failures and a greater extent of necrosis compared with those with sterile necrosis . When early antibiotic treatment was used in all patients with necrotizing pancreatitis (imipenem/cilastatin), the characteristics of pancreatic infection changed to predominantly gram-positive and fungal infections . Fine-needle aspiration showed a sensitivity of 96% for detecting pancreatic infection . The death rate was 1.8% (1/56) in patients with sterile necrosis managed without surgery versus 24% (7/29) in patients with infected necrosis (P <.01) . Two patients whose infected necrosis could not be diagnosed in a timely fashion died while receiving nonsurgical treatment . Thus, an intent-to-treat analysis (nonsurgical vs . surgical treatment) revealed a death rate of 5% (3/58) with conservative management versus 21% (6/28) with surgery . CONCLUSIONS: These results support nonsurgical management, including early antibiotic treatment, in patients with sterile pancreatic necrosis . Patients with infected necrosis still represent a high-risk group in severe acute pancreatitis, and for them surgical treatment seems preferable.

Intern Med, 2000 Nov, 39(11), 901 - 9
Current practice of management of bacteremic sepsis: a study in a tertiary care teaching hospital in Japan; Aoki Y et al.; OBJECTIVE: To investigate how patients with bacteremic sepsis are managed in a tertiary care teaching hospital . PATIENTS AND METHODS: Prospective observational study on patients with bacteremic sepsis . Clinical and microbiological characteristics of bacteremic sepsis were analyzed in relation to prognosis . Severity of the illness was quantitatively analyzed by the APACHE (Acute Physiology, Age, Chronic Health Evaluation) III scoring system . Also investigated was how closely physicians paid attention to acute physiological alterations in patients . RESULTS: The 28-day mortalities in fifty hemodynamically stable patients and in twenty-three septic shock patients were 26% and 52%, respectively (p=0.028) . Gram-positive organisms accounted for 54% of all organisms, with the mortality and incidence of septic shock being the same as with Gram-negative infections . The mean APACHE III score was 42.9 in survivors, and 76.5 in non-survivors (p < 0.001) . Although serum levels of C-reactive protein and acute physiology score (APS) was significantly higher in non-survivors than in survivors, the correlation with APACHE III score was more prominent in APS . The number of vital signs recorded was 1.67 in physicians and 3.6 in nurses (p < 0.001) . CONCLUSIONS: The present study proved that the APACHE III score accurately discriminates between survivors and non-survivors of patients with sepsis . By addressing the need for an objective evaluation of severity of illness, it strongly recommends that physicians should be made aware of physiologically defined sepsis and that they should pay closer attention to patients' physiological alterations to identify the development of sepsis in critically ill patients.

Brain Res, 2000 Nov 10, 883(1), 157 - 63
Muramyl dipeptide potentiates cytokine-induced activation of inducible nitric oxide synthase in rat astrocytes; Trajkovic V et al.; We investigated the influence of muramyl dipeptide (MDP), a cell wall component of Gram-positive bacteria, on cytokine-induced nitric oxide (NO) production in rat primary astrocytes . MDP alone did not induce NO release in astrocyte cultures . However, MDP increased astrocyte NO production and subsequent nitrite accumulation triggered by IFN-gamma . IFN-gamma-activated expression of mRNA for inducible NO synthase (iNOS) and iNOS transcription factor interferon regulatory factor-1 (IRF-1) was markedly enhanced in astrocytes treated with MDP . The potentiating effect of MDP on IFN-gamma-induced NO production in astrocytes was completely blocked with protein tyrosine kinase (PTK) inhibitor genistein or mitogen activated protein kinase (MAPK) inhibitor PD98059 . In contrast, protein kinase C (PKC) inhibitor calphostin C did not affect ability of MDP to augment IFN-gamma-triggered astrocyte NO synthesis . These results suggest that MDP synergizes with IFN-gamma in the induction of iNOS gene in astrocytes through mechanisms involving PTK and MAPK, but not PKC activation . Finally, MDP also augmented NO production and iNOS mRNA expression in astrocytes treated with IL-1beta.

Clin Diagn Lab Immunol, 2000 Nov, 7(6), 889 - 92
Procalcitonin does not discriminate infection from inflammation after allogeneic bone marrow transplantation; Blijlevens NM et al.; Procalcitonin (PCT) is an early marker of bacterial infection but little is known about its value in neutropenic allogeneic bone marrow transplant (BMT) recipients . We collected plasma from 12 recipients of T-cell-depleted HLA-matched related BMT recipients who had been treated preemptively with meropenem from the day after BMT for at least 15 days . PCT and C-reactive protein (CRP) concentrations were determined on BMT days 1, 5, 8, 12, and 15, and their relationship to inflammatory events (IE), including mucositis, microbiologically and clinically defined infections, acute graft-versus-host disease (GVDH), and unexplained fever, was then determined . The PCT concentrations were all low and never exceeded 4 microg/liter, unlike CRP concentrations, which spanned the full range up to 350 mg/liter . All patients had mucositis, and there was no significant difference between PCT concentrations associated with mucositis alone and those associated with an additional IE on BMT days 1 to 12 . However, on BMT day 15, the mean concentrations of PCT were 0.37 +/- 0.05 microg/liter for the 10 patients that had an additional IE, compared with 0.11 +/- 0.03 microg/liter for the 2 patients with mucositis only (P = 0.012), and GVHD rather than infection was involved in six cases . PCT was also not a sensitive marker of gram-positive bacteremia or pulmonary aspergillosis . Thus, PCT is of little value in discriminating infections from other inflammatory complications that occur following allogeneic BMT.

Expert Opin Investig Drugs, 2000 Oct, 9(10), 2351 - 69
Inhibitors of bacterial two-component signalling systems; Macielag MJ et al.; Bacterial two-component regulatory systems (TCS) play a pivotal role in the process of infection . These signal transduction systems enable bacterial pathogens to mount an adaptive response and cope with diverse environmental stresses, including nutrient deprivation, antibiotic onslaught and phagocytosis . Interest in these systems as novel bacterial targets has been rekindled by the recent discovery of several essential systems in important Gram-positive and Gram-negative pathogens . Several series of TCS inhibitors derived from broad screening approaches have been reported in the literature, however, most appear to suffer from poor selectivity, excessive protein binding and/or limited bioavailability . Consequently, pharmaceutical chemists have turned to alternate strategies, such as the design of substrate-based inhibitors, the generation of combinatorial libraries and the isolation of natural products, to identify inhibitors with more desirable properties . Recent structural studies of the histidine protein kinase and response regulator proteins that constitute TCS may provide a foundation for a structure-based design approach to TCS inhibitors.

J Clin Microbiol, 2000 Nov, 38(11), 4254 - 5
Meningoencephalitis associated with Globicatella sanguinis infection in lambs; Vela AI et al.; Thirty lambs displayed symptoms of meningoencephalitis . An unusual gram-positive coccus was isolated in pure culture from the blood and brain samples from one of the affected animals, and phenotypic and phylogenetic characterization showed this to be Globicatella sanguinis . This is the first report of the isolation of G . sanguinis in pure culture from an animal infection.

Scand J Infect Dis, 2000, 32(5), 511 - 4
Plasmapheresis as part of the treatment for septic shock; Hjorth V et al.; Plasmapheresis is one of the methods which has been tried in the effort to influence the course of severe sepsis with septic shock and to improve survival rates . This is a retrospective study of 17 consecutive patients with septic shock who were treated with acute plasmapheresis . Nine out of 16 patients with verified etiology suffered from infections with Gram-positive bacteria . Three (18%) of the treated patients died . The estimated mortality rate based on the patients' initial APACHE II scores was 62% . No serious side-effects of treatment were observed . This study indicates that plasmapheresis is a safe treatment for patients with septic shock and has a positive effect on survival . Prospective, randomized studies would be valuable to determine the role of plasmapheresis in the management of patients with septic shock.

Am J Kidney Dis, 2000 Nov, 36(5), 1000 - 8
Polymicrobial peritonitis in continuous ambulatory peritoneal dialysis patients; Kim GC et al.; We retrospectively evaluated 232 continuous ambulatory peritoneal dialysis (CAPD) patients entering our program from January 1, 1987, to December 31, 1997, for polymicrobial peritonitis . Polymicrobial peritonitis occurred in 16% of the patients (polymicrobial-peritonitis group), whereas 52% of the patients had peritonitis episodes with only a single organism (single-organism group), and 32% of the patients had no episode of peritonitis . Polymicrobial peritonitis accounted for 8% of the 554 peritonitis episodes, occurred after 23 +/- 20 months on peritoneal dialysis (PD), and was preceded by greater than three episodes of peritonitis in 73% of the patients . Peritonitis rates were greater in the polymicrobial-peritonitis group compared with patients in the single-organism group (1.8 versus 1.2 episodes/patient-year; P: < 0.001) . The majority of polymicrobial infections involved gram-negative and/or fungal pathogens, but in 21% of the episodes, only gram-positive organisms were identified . An intra-abdominal process was identified in only 7% of the patients . Catheter loss overall was greatest in the polymicrobial-peritonitis group (65% versus single-organism group, 30% versus patients without peritonitis, 5%; P < 0.001), but only 33% of the polymicrobial infections resulted in catheter loss . At last follow-up, 70% of the patients in the polymicrobial-peritonitis group had permanently transferred to hemodialysis compared with 25% from the single-organism group and 15% from the no-peritonitis group (P < 0.001) . In conclusion, polymicrobial peritonitis is an infrequent but serious complication of CAPD that occurs late in the course of PD and is often preceded by recurrent episodes of peritonitis . Polymicrobial peritonitis is rarely the result of a catastrophic intra-abdominal process, and although the majority of patients can be successfully treated without catheter removal, the long-term prognosis is poor, with a high rate of transfer to hemodialysis.

J Bacteriol, 2000 Nov, 182(22), 6292 - 301
Maltose and maltodextrin transport in the thermoacidophilic gram-positive bacterium Alicyclobacillus acidocaldarius is mediated by a high-affinity transport system that includes a maltose binding protein tolerant to low pH; Hulsmann A et al.; We have studied the uptake of maltose in the thermoacidophilic gram-positive bacterium Alicyclobacillus acidocaldarius, which grows best at 57 degrees C and pH 3.5 . Under these conditions, accumulation of {(14)C}maltose was observed in cells grown with maltose but not in those grown with glucose . At lower temperatures or higher pH values, the transport rates substantially decreased . Uptake of radiolabeled maltose was inhibited by maltotetraose, acarbose, and cyclodextrins but not by lactose, sucrose, or trehalose . The kinetic parameters (K(m) of 0.91 +/- 0.06 microM and V(max) ranging from 0.6 to 3.7 nmol/min/mg of protein) are consistent with a binding protein-dependent ATP binding cassette (ABC) transporter . A corresponding binding protein (MalE) that interacts with maltose with high affinity (K(d) of 1.5 microM) was purified from the culture supernatant of maltose-grown cells . Immunoelectron microscopy revealed distribution of the protein throughout the cell wall . The malE gene was cloned and sequenced . Five additional open reading frames, encoding components of a maltose transport system (MalF and MalG), a putative transcriptional regulator (MalR), a cyclodextrinase (CdaA), and an alpha-glucosidase (GlcA), were identified downstream of malE . The malE gene lacking the DNA sequence that encodes the signal sequence was expressed in Escherichia coli . The purified wild-type and recombinant proteins bind maltose with high affinity over a wide pH range (2.5 to 7) and up to 80 degrees C . Recombinant MalE cross-reacted with an antiserum raised against the wild-type protein, thereby indicating that the latter is the product of the malE gene . The MalE protein might be well suited as a model to study tolerance of proteins to low pH.

J Biotechnol, 2000 Oct 13, 83(3), 211 - 7
Gentle lysis of mucous producing cold-adapted bacteria by surfactant treatment combined with mechanical disruption; Nandakumar R et al.; A procedure for the enhanced lysis of mucous producing psychrotrophic gram positive bacteria for subsequent enzyme studies is described . An initial washing of bacterial cells with Tween 80 was found to improve the degree of cell disruption in subsequent sonication or grinding with glass beads, resulting in about 20-200% increase in total soluble protein content . However, in terms of lactate dehydrogenase (LDH) activity present in the lysate, pretreatment with Tween 80 was more effective in combination with grinding, especially in the highly mucous producing strain GY11 . The type of surfactant used in the pretreatment procedure before grinding strongly influenced the percentage lysis of tested strains, both in terms of released soluble protein and enzyme activity . Zymograms of LDH and glutamate dehydrogenase (GDH) activity present in the lysates also very well supported the results obtained by total protein and enzyme activity measurements.

FEMS Microbiol Ecol, 2000 Oct 1, 34(1), 73 - 80
Detection of Desulfotomaculum in an Italian rice paddy soil by 16S ribosomal nucleic acid analyses; Stubner S et al.; Two specific primers were developed for the amplification of 16S rRNA genes of Desulfotomaculum lineage 1 to detect members of the genus Desulfotomaculum in rice field soil . The combination of both primers in PCR allowed the specific amplification and cloning of ten 16S rDNA sequences of this group from rice paddy soil DNA extracts . The phylogenetic analysis showed that these sequences formed a deeply branching cluster within Desulfotomaculum lineage 1, together with two sequences from the database and two sequences from a hydrocarbon-contaminated aquifer . Dissimilarity values to validly described species, including recently isolated strains of Desulfotomaculum from rice paddy microcosms, were higher than 12% . Within the new cluster the cloned sequences formed three separate groups which were each represented by at least two sequences with identities of >/=99% while one sequence represented an additional group . The sequences should represent sulfate-reducing organisms because they clearly fell into the physiologically coherent group of Gram-positive sulfate reducers . The relative abundance of bacteria of the Desulfotomaculum lineage 1 in rice paddy soil and root samples was estimated with rRNA dot blot hybridizations of extracted RNA . The relative RNA content of Desulfotomaculum lineage 1 was 0.55% in the bulk soil and 1% in the rice root samples, respectively, of the total 16S rRNA content (probe Eub338) . Hybridization of rRNA with a probe targeting the new cluster represented by the cloned sequences confirmed the high abundance of 16S rRNA sequences from this cluster in the rice paddy field samples . Another hybridization probe detecting Desulfotomaculum acetoxidans and two closely related Desulfotomaculum isolates from rice paddy soil indicated that these bacteria were less abundant.

J Biol Chem, 2001 Jan 26, 276(4), 2361 - 71 Epub 2000 Oct 26.
Phosphorylation of Wzc, a tyrosine autokinase, is essential for assembly of group 1 capsular polysaccharides in Escherichia coli; Wugeditsch T et al.; Wzc proteins are tyrosine autokinases . They are found in some important bacterial pathogens of humans and livestock as well as plant-associated bacteria, and are often encoded within gene clusters determining synthesis and assembly of capsular and extracellular polysaccharides . Autophosphorylation of Wzc(cps) is essential for assembly of the serotype K30 group 1 capsule in Escherichia coli O9a:K30, although a genetically unlinked Wzc(cps)-homologue (Etk) can also participate with low efficiency . While autophosphorylation of Wzc(cps) is required for assembly of high molecular weight K30 capsular polysaccharide, it is not essential for either the synthesis of the K30 repeat units or for activity of the K30 polymerase enzyme . Paradoxically, the cognate phosphotyrosine protein phosphatase for Wzc(cps), Wzb(cps), is also required for capsule expression . The tyrosine-rich domain at the C terminus of Wzc(cps) was identified as the site of phosphorylation and autophosphorylation of Wzc requires a functional Walker A motif . Intermolecular transphosphorylation of Wzc(cps) was detected in strains expressing a combination of mutant Wzc(cps) derivatives . The N- and C-terminal domains of Wzc(cps) were expressed independently to mimic the situation found naturally in Gram-positive bacteria . In this format, both domains were required for phosphorylation of the Wzc(cps) C terminus, and for capsule assembly . Regulation by a post-translational phosphorylation event represents a new dimension in the assembly of bacterial cell-surface polysaccharides.

Adv Perit Dial, 2000, 16, 276 - 9
Prevalence of microbial colonization in removed peritoneal catheters: a prospective study; Rodriguez-Carmona A et al.; We conducted a prospective bacteriologic study of 89 peritoneal catheters removed from 77 peritoneal dialysis patients . Reasons for catheter removal included severe peritonitis (n = 36, Group A), persistent exit-site infection (n = 29, Group B), and dormant, seemingly uninfected catheters (n = 22, Group C) . We studied the external cuff (EC) and internal cuff (IC) as well as the catheter tip . In Group A, microbial growth was seen in 86.1% of ECs, 66.7% of ICs, and 67.6% of tips . In cases of positive isolation, concordance was 91.7% IC versus EC, 84.2% IC versus tip, and 80.0% EC versus tip . The peritonitis agent was recovered from 61.1% of ECs, 50.0% of ICs, and 55.6% of tips . In Group B, colonization was seen in 72.4% of ECs, 44.8% of ICs, and 31.0% of tips . When an isolation was obtained from both EC and IC, concordance was 81.8% . The exit-site infection agent was recovered from 69% of ECs and 24% of ICs . In Group C, microbial growth was observed in 77.3% of ECs, 45.5% of ICs, and 31.8% of tips . Gram-positive bacteria predominated, with the same bacteria colonizing EC and IC in 66.7% of cases . In conclusion, removed peritoneal catheters present a high prevalence of extensive microbial colonization, even in the absence of overt infection.

Adv Perit Dial, 2000, 16, 243 - 7
The influence of seasonal factors on the incidence of peritonitis in continuous ambulatory peritoneal dialysis in the temperate zone; Kim MJ et al.; Many factors contribute to the development of continuous ambulatory peritoneal dialysis (CAPD) peritonitis . The role of climate in CAPD peritonitis has received relatively little attention . We retrospectively analyzed the incidence of CAPD peritonitis according to temperature and relative humidity . Data from 80 patients were examined . The monthly mean temperature and relative humidity during study period varied between -3.4 degrees C and 25.4 degrees C and between 61% and 81%, respectively . In 1123 patient-months, 53 cases of peritonitis occurred . The occurrence of peritonitis paralleled temperature and relative humidity, being the highest (0.180 episodes/patient-month) in July (mean temperature, 24.6 degrees C; relative humidity, 81%) and lowest (0.013 episodes/patient-month) in November (mean temperature, 6.6 degrees C; relative humidity, 66%) . Significant correlations were seen between the monthly frequency of CAPD peritonitis and temperature (r = 0.53, p < 0.05) and relative humidity (r = 0.59, p < 0.05) . The incidence was higher in the warm season (months with a mean temperature > or = 15 degrees C, that is, May-September) than in the cold season (months with a mean temperature < 15 degrees C, that is, October-April), at 0.074 episodes/patient-month versus 0.024 episodes/patient-month, p < 0.05 . We also found a tendency for gram-negative peritonitis to occur uniformly throughout the year, but for gram-positive peritonitis to increase during hot and humid months, especially the rainy month of July . Gram-positive organisms caused 50% of peritonitis from March to August, but just 17.7% from September to February . Gram-negative organisms caused 7.3% and 29.4% of peritonitis during the same periods (p < 0.05) . The results indicate a clear seasonal change in the rate of CAPD peritonitis and in the causative micro-organisms . The observation that CAPD peritonitis increases in the season of high temperature and high humidity suggests the influence of climate on CAPD peritonitis in the temperate zone.

Biochem Soc Trans, 2000 Oct, 28(5), 563 - 6
Toll-like receptor family and signalling pathway; Muzio M et al.; Toll is a Drosophila gene essential for ontogenesis and anti-microbial resistance . Several orthologues of Toll have been identified and cloned in vertebrates, namely Toll-like receptors (TLRs) . Human TLRs are a growing family of molecules involved in innate immunity . TLRs are characterized structurally by a cytoplasmic Toll/interleukin-1 receptor (TIR) domain and by extracellular leucine-rich repeats . TLRs characterized so far activate the MyD88/interleukin-1 receptor-associated kinase (IRAK) signalling pathway . Genetic, gene-transfer and dominant-negative approaches have involved TLR family members (TLR2 and TLR4) in Gram-positive and Gram-negative bacteria recognition and signalling . Accumulating evidence suggests that TLR2 is also involved in signalling-receptor complexes that recognize components of yeast and mycobacteria . However, the definitive roles of other TLRs are still lacking . A systematic approach has been used to determine whether different human leucocyte populations selectively or specifically express TLR mRNA . Based on expression pattern, TLR can be classified as ubiquitous (TLR1), restricted (TLR2, TLR4 and TLR5) and specific (TLR3) . Expression and regulation of distinct but overlapping ligand-recognition patterns may underlie the existence of a large, seemingly redundant TLR family . Alternatively, the expression of a TLR in a single cell type may indicate a specific role for this molecule in a restricted setting.

Biochem Soc Trans, 2000 Oct, 28(5), 557 - 63
The Toll/interleukin-1 receptor domain: a molecular switch for inflammation and host defence; O'Neill L; The pro-inflammatory cytokine interleukin-1 (IL-1) signals via the Type-I IL-1 receptor (IL-1RI), inducing an increase in the expression of many genes with roles in immunity and inflammation . The signalling pathways involve two adapter proteins, MyD88 and Tollip, which via two IL-1 receptor-associated kinases (IRAK and IRAK-2) activate transcription factors such as nuclear factor-kappa B and protein kinases such as p38 mitogen-activated protein kinase . A role for the low-molecular-mass G-proteins Rac, Ras and Rap in these processes has also been indicated . IL-1RI is the founder of a diverse superfamily of receptors, which all share a cytosolic domain, termed the Toll/IL-1 receptor (TIR) domain . The superfamily can be divided broadly into three subgroups . The first of these is most similar to IL-1RI and includes the receptor for IL-18 and the Th2 cell regulator T1/ST2 . The second subgroup is most similar to the Drosophila melanagaster protein Toll and includes Toll-like receptor 2 (TLR2), which is required for host defence against Gram-positive bacteria and fungi, and TLR4, which is required for lipopolysaccharide responsiveness, and thus is involved in host defence against Gram-negative bacteria . There are also a number of TLRs in plants and insects, all involved in host defence . The third subgroup contains non-receptor proteins which possess a TIR domain and are cytosolic . MyD88 is a member, and it presumably complexes with IL-1RI via a TIR-TIR interaction . The other two members are proteins encoded by the vaccinia virus, A46R and A52R, which block TIR-dependent signalling . This receptor superfamily therefore appears to play a central role in inflammation and host defence against infection, pointing to the TIR domain as a critical molecular player in the innate immune response.

Biochem Soc Trans, 2000 Oct, 28(5), 551 - 6
Toll-like receptors: lessons from knockout mice; Akira S; The Toll signalling pathway, which is required for establishment of dorsoventral polarity in Drosophila embryos, plays an important role in the response to microbial infections . Recently, Toll-like receptors (TLRs) have also been identified in mammals . TLR4 has been shown to function as the transmembrane component of the lipopolysaccharide receptor, while TLR2 recognizes peptidoglycans from Gram-positive bacteria, lipoproteins and yeast . Although various microbial cell-wall components are recognized by different receptors, all of these responses are abrogated in MyD88-deficient cells . These results show that different TLRs recognize different microbial cell-wall components, and that MyD88 is an essential signalling molecule shared among interleukin-1 receptor/Toll family members.

Arthritis Rheum, 2000 Oct, 43(10), 2160 - 8
Antigen-presenting cells containing bacterial peptidoglycan in synovial tissues of rheumatoid arthritis patients coexpress costimulatory molecules and cytokines; Schrijver IA et al.; OBJECTIVE: Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by intimal lining hyperplasia and massive infiltration of the synovial sublining by antigen-presenting cells (APCs), lymphocytes, and plasma cells . Peptidoglycan (PG), a major cell wall component of gram-positive bacteria, which is abundantly expressed in all mucosa, is believed to be involved in the pathogenesis of RA because of its ability to induce the production of proinflammatory cytokines as well as to induce arthritis in rodents . While PG has been detected in APCs in RA joints, little is known about the role of these cells in RA . In this study, the presence and immune competence of PG-containing cells in synovial tissues from 14 RA and 14 osteoarthritis (OA) patients were analyzed in situ . METHODS: Using immunohistochemistry, we examined the coexpression of phenotypic markers, costimulatory molecules, and cytokines by PG-containing cells . RESULTS: PG was present in higher numbers in RA than in OA synovial tissues, although the difference was not significant . PG-containing cells were mainly macrophages, but some mature dendritic cells also contained PG . A high percentage of PG-containing cells in both RA and OA synovial tissues coexpressed HLA-DR . CD40, CD80, and CD86 expression by PG-containing cells was higher in RA than in OA tissues . Furthermore, PG-containing cells coexpressed cytokines, which modulate inflammatory reactions, in particular, tumor necrosis factor alpha and interleukins 6 and 10 . CONCLUSION: The results suggest that PG-containing cells may contribute to inflammation within the microenvironment of the joint in RA patients.

Antimicrob Agents Chemother, 2000 Nov, 44(11), 2948 - 53
Once-daily dosing in dogs optimizes daptomycin safety; Oleson FB Jr et al.; Daptomycin is a novel lipopeptide antibiotic with potent bactericidal activity against most clinically important gram-positive bacteria, including resistant strains . Daptomycin has been shown to have an effect on skeletal muscle . To guide the clinical dosing regimen with the potential for the least effect on skeletal muscle, two studies were conducted with dogs to compare the effects of repeated intravenous administration every 24 h versus every 8 h for 20 days . The data suggest that skeletal-muscle effects were more closely related to the dosing interval than to either the maximum concentration of the drug in plasma or the area under the concentration-time curve . Both increases in serum creatine phosphokinase activity and the incidence of myopathy observed at 25 mg/kg of body weight every 8 h were greater than those observed at 75 mg/kg every 24 h despite the lower maximum concentration of drug in plasma . Similarly, the effects observed at 25 mg/kg every 8 h were greater than those observed at 75 mg/kg every 24 h at approximately the same area under the concentration-time curve from 0 to 24 h . Once-daily administration appeared to minimize the potential for daptomycin-related skeletal-muscle effects, possibly by allowing for more time between doses for repair of subclinical effects . Thus, these studies with dogs suggest that once-daily dosing of daptomycin in humans should have the potential to minimize skeletal-muscle effects . In fact, interim results of ongoing clinical trials, which have focused on once-daily dosing, appear to be consistent with this conclusion.

Int J Syst Evol Microbiol, 2000 Sep, 50 Pt 5, 1899 - 903
Janibacter brevis sp . nov., a new trichloroethylene-degrading bacterium isolated from polluted environments; Imamura Y et al.; Two strains that were strongly able to degrade trichloroethylene were isolated from contaminated environmental samples . The isolates are non-motile, aerobic, non-spore-forming, gram-positive coccoid to short rods with cell-wall peptidoglycan containing meso-diaminopimelic acid . The major type of menaquinone is MK-8(H4) . The polar lipids were diphosphatidylglycerol, phosphatidylglycerol and phosphatidylinositol . Mycolic acids were absent . The G+C content of the DNA was 72 mol% . Furthermore, comparison of 16S rDNA sequences shows that the isolates clearly belong to the genus Janibacter . The isolates differ from Janibacter limosus at the species level, as shown by the value for DNA-DNA hybridization . It is therefore proposed that the strains be assigned to the genus Janibacter as Janibacter brevis sp . nov.

J Emerg Med, 2000 Oct, 19(3), 217 - 23
Bedside measurement of D-dimer in the identification of bacteremia in the emergency department; Quick G et al.; The objective of this pilot study was to determine the clinical utility of the SimpliRed D-dimer bedside assay to identify patients with bacteremia in a university hospital Emergency Department . We tested 265 patients and compared blood culture results with a novel D-dimer semiquantitative whole blood assay . Bacteremia was confirmed in 25/262 patients . Sensitivity of D-dimer assay was 66.7% for Gram-positive bacteremia and 61.5% for Gram-negative bacteremia with negative predictive value of 98% for Gram-positive and 96% for Gram-negative bacteremia patients . Measurement of D-dimer appears to be of value in identifying patients at low risk for bacteremia and can be accomplished rapidly using a whole blood semiquantitative bedside assay . Although increases in D-dimer are not detected in all patients subsequently documented to have bacteremia on a single sampling, the results of this and other earlier studies suggest assay of D-dimer is useful in rapid differentiation of patients with bacteremia from those who have no bacteremia using blood culture positivity as the standard for bacteremia.

Diagn Microbiol Infect Dis, 2000 Sep, 38(1), 1 - 5
Evaluation of antiseptic-impregnated central venous catheters for prevention of catheter-related infection in intensive care unit patients; Sheng WH et al.; Central venous catheterization represents a significant medical advancement, particularly in the treatment of critical ill . However, there is a high risk of central venous catheters-related infection . A novel antiseptic central venous catheter, made of polyurethane and impregnated with chlorhexidine and silver sulfadiazine, was developed to reduce the risk of catheters-related infection . In this study, we did a randomized clinical study to determine the efficacy by using antiseptic catheters for the prevention of central venous catheters-related infection in the intensive care units . A total of 204 patients with 235 central venous catheters were studied at the surgical intensive care units at National Taiwan University Hospital between November 1998 and June 1999 . Participants received either a standard triple-lumen polyurethane catheter or an antiseptic catheter (Arrow International, Reading, Pennsylvania, USA) . Both were indistinguishable from each other . Compared to standard polyurethane catheters, antiseptic catheters were less likely to be colonized by microorganisms when they were cultured at the removal (8.0 versus 20.0 colonized catheters per 100 catheters; relative risk 0.34 {95% CI, 0.15 to 0.74}; p<0.01) . There was no significant differences between both groups in catheter-related infections (0.9 versus 4.9 infections per 100 catheters; relative risk 0.17 {95% CI, 0.03 to 1.15}; p = 0.07) . Gram-positive cocci and fungi were more likely to colonize in the standard polyurethane catheters (p = 0.06 and 0.04, compared to antiseptic catheters respectively) . Two of our cases in the control group died directly due to catheter-related candidemia . No adverse reactions such as hypersensitivity or leukopenia were found in the antiseptic catheter group . Our study showed that central venous catheters with antiseptic coating were safe and had less risk of colonization of bacteria and fungi than standard catheters in the critically ill patients.

FEMS Immunol Med Microbiol, 2000 Oct, 29(2), 101 - 6
Effect of Alloiococcus otitidis and three pathogens of otitis media in production of interleukin-12 by human monocyte cell line; Himi T et al.; Alloiococcus otitidis is detected in middle ear effusion of otitis media with effusion (OME) . Only a limited number of studies are available concerning the immunological profile of A . otitidis . We have studied the ability of A . otitidis and three other representative pathogens of otitis media to stimulate the production of interleukin-12 (IL-12) from a monocytic cell line THP-1 . Viable A . otitidis induced the production of IL-12 in THP-1 cells but IL-12 production was reduced if glutaraldehyde-fixed bacteria were used as stimulants . When viable bacteria were physically separated from THP-1 cells during the stimulation period, remarkable reductions of IL-12 secretion were shown after challenge with gram-positive bacteria A . otitidis and S . pneumoniae . When stimulated with soluble extracts of A . otitidis, THP-1 secreted IL-12 in a dose-dependent manner . The subfraction with a molecular mass over 100 kDa showed a strong ability to induce IL-12 production . Our results show that A . otitidis has immunostimulatory capacity with regard to IL-12 production . We also show that soluble antigen(s) of A . otitidis can modulate the immune response in OME.

FEMS Microbiol Lett, 2000 Oct 15, 191(2), 191 - 7
Structural and functional analysis of the riboflavin synthesis genes encoding GTP cyclohydrolase II (ribA), DHBP synthase (ribBA), riboflavin synthase (ribC), and riboflavin deaminase/reductase (ribD) from Helicobacter pylori strain P1; Fassbinder F et al.; The functions of the riboflavin synthesis gene homologues ribA, ribBA, ribC, and ribD from Helicobacter pylori strain P1 were confirmed by complementation of defined Escherichia coli mutant strains . The H . pylori ribBA gene, which is similar to bifunctional ribBA genes of Gram-positive bacteria, fully complemented the ribB mutation and partially restored growth in a ribC mutant . However, ribBA did not complement the ribA mutation in E . coli, thus explaining the presence of the additional separate copy of the ribA gene in the H . pylori chromosome . In E . coli exclusively ribA conferred hemolytic activity and gave rise to production of molecules with fluorescence characteristics similar to flavins, as observed earlier . The E . coli hemolysin ClyA was not involved in causing the hemolytic phenotype . No riboflavin synthesis genes on plasmids conferred iron uptake functions to a siderophore-deficient mutant of E . coli . Marker exchange mutagenesis of the genes in H . pylori was not successful indicating that riboflavin synthesis is essential for basic metabolic functions of the gastric pathogen.

Microbes Infect, 2000 Sep, 2(11), 1373 - 81
Nocardia brasiliensis: from microbe to human and experimental infections; Salinas-Carmona MC; Nocardia brasiliensis is a Gram-positive bacterium that lives as a saprophyte in soil . In this article the physical properties, chemical composition and taxonomic position of this species is reviewed . Human infections and an experimental model of actinomycetoma in BALB/c mice as well as the host-immune response is described.

J Clin Microbiol, 2000 Oct, 38(10), 3755 - 8
Evaluation of a PCR primer based on the isocitrate dehydrogenase gene for detection of Helicobacter pylori in feces; Argyros FC et al.; In order to improve detection and identification of Helicobacter pylori in highly contaminated samples, we evaluated new specific primers based on the DNA base sequence within the isocitrate dehydrogenase (icd) gene to amplify a 1,200-bp DNA segment . The specificity of the icd primer was tested against DNA derived from various bacteria, including 7 Helicobacter species and a panel of 1 gram-variable, 2 gram-positive, and 16 gram-negative bacteria, as well as DNA from houseflies and feces from H . pylori-negative patients . The primers permitted the detection of all clinical H . pylori isolates tested, but no reactions were observed with negative controls . Several procedures for DNA extraction from feces were evaluated using PCR with icd primers . The lower limits of detection of H . pylori DNA from two different sources containing the same number of H . pylori organisms, a pure culture and feces spiked with H . pylori, were established for each extraction method tested . The results were 8.0 x 10(3) CFU/ml for cultures of pure H . pylori, and 8.0 x 10(6) CFU/ml for H . pylori from feces, using the phenol-chloroform method; 8.0 x 10(2) and 7.0 x 10(3) CFU/ml, respectively, for a glass matrix and chaotropic solution protocol; 8.0 x 10(2) and 7.0 x 10(3) CFU/ml, respectively, for the QIAamp tissue kit; and 5.0 x 10(2) and 5.0 x 10(3) CFU/ml, respectively, for the XTRAX DNA extraction kit . We conclude that the use of the icd gene as a primer for PCR represents a specific and sensitive assay for detection of H . pylori in highly contaminated samples.

J Infect Dis, 2000 Nov, 182(5), 1549 - 52 Epub 2000 Oct 03.
Detection of circulating lipopolysaccharide-bound monocytes in children with gram-negative sepsis; Takeshita S et al.; The possibility that gram-negative sepsis can be diagnosed by detection of lipopolysaccharide (LPS) bound on the surface of monocytes in the circulation of patients with gram-negative sepsis was investigated . Peripheral monocytes were analyzed by flow cytometer and an anti-LPS monoclonal antibody in 3 groups: children with gram-negative sepsis, children with gram-positive sepsis, and healthy children . LPS-bound monocytes were found in all patients with gram-negative sepsis but not in children with gram-positive sepsis or in healthy children . Therefore, the flow cytometry method developed for this study may be a novel method for diagnosing gram-negative sepsis.

Zygote, 2000 Aug, 8(3), 257 - 65
Enzymes responsible for the bactericidal effect in extracts of vitelline and fertilisation envelopes of rainbow trout eggs; Kudo S; Extracts from both the vitelline envelope (VE) and fertilisation envelopes (FE) of rainbow trout eggs have the ability to exert a bactericidal effect on Gram-positive and -negative bacteria . The effect may be due to the presence of phospholipase D (PLD), lysozyme, proteinase and DNases, as the extracts contain these enzyme activities . The intensity of chorionic PLD and lysozyme activities in the VE extract was maintained in the FE without any alteration in activity even after transformation in the course of the cortical reaction, as components of a fundamental architecture of the envelope . Both extracts also contain different types of proteinase activities . Treatment with VE or FE extract seriously damaged the outer membrane of Gram-negative bacteria and the plasma membrane of Gram-positive and -negative bacteria at the ultrastructural level . Chorionic DNases probably degrade DNA of bacterial cells killed by virtue of the action of PLD and/or lysozyme and contribute to the transmigration of nucleosides and/or nucleotides produced by degrading bacterial DNA after degradation of bacterial components by the actions of the chorionic PLD, lysozyme and proteinase . These results suggest that the bactericidal process manifested by the VE or FE extract may start with the action of PLD and/or lysozyme against bacteria and be completed by subsequent degradation of constitutive proteins and DNA by the action of proteinases and DNases, respectively . Thus the VE and FE are able to protect the egg itself and the embryo, respectively, from bacterial infection in the internal or external environments.

Appl Environ Microbiol, 2000 Oct, 66(10), 4486 - 96
Development of a vital fluorescent staining method for monitoring bacterial transport in subsurface environments; Fuller ME et al.; Previous bacterial transport studies have utilized fluorophores which have been shown to adversely affect the physiology of stained cells . This research was undertaken to identify alternative fluorescent stains that do not adversely affect the transport or viability of bacteria . Initial work was performed with a groundwater isolate, Comamonas sp . strain DA001 . Potential compounds were first screened to determine staining efficiencies and adverse side effects . 5-(And 6-)-carboxyfluorescein diacetate, succinimidyl ester (CFDA/SE) efficiently stained DA001 without causing undesirable effects on cell adhesion or viability . Members of many other gram-negative and gram-positive bacterial genera were also effectively stained with CFDA/SE . More than 95% of CFDA/SE-stained Comamonas sp . strain DA001 cells incubated in artificial groundwater (under no-growth conditions) remained fluorescent for at least 28 days as determined by epifluorescent microscopy and flow cytometry . No differences in the survival and culturability of CFDA/SE-stained and unstained DA001 cells in groundwater or saturated sediment microcosms were detected . The bright, yellow-green cells were readily distinguished from autofluorescing sediment particles by epifluorescence microscopy . A high throughput method using microplate spectrofluorometry was developed, which had a detection limit of mid-10(5) CFDA-stained cells/ml; the detection limit for flow cytometry was on the order of 1,000 cells/ml . The results of laboratory-scale bacterial transport experiments performed with intact sediment cores and nondividing DA001 cells revealed good agreement between the aqueous cell concentrations determined by the microplate assay and those determined by other enumeration methods . This research indicates that CFDA/SE is very efficient for labeling cells for bacterial transport experiments and that it may be useful for other microbial ecology research as well.

Biochem Pharmacol, 2000 Nov 1, 60(9), 1367 - 79
Novel mechanisms of DNA topoisomerase II inhibition by pyranonaphthoquinone derivatives-eleutherin, alpha lapachone, and beta lapachone; Krishnan P et al.; Pyranonaphthoquinones have diverse biological activities against Gram-positive bacteria, fungi, and mycoplasms, and, recently, there has also been an increasing interest in their anti-cancer activity . This study includes three derivatives: eleutherin (compound 1), beta lapachone (compound 2), and its structural isomer, alpha lapachone (compound 3) . The mechanism of topoisomerase II inhibition by the three derivatives was examined systematically with respect to the steps of the catalytic cycle of the enzyme . Etoposide, the prototypical enzyme poison, was used as a control and in combination with compounds 1-3 to localize their mechanism of action . The study revealed that eleutherin (1) and beta lapachone (2) inhibited topoisomerase II by inducing religation and dissociation of the enzyme from DNA in the presence of ATP . Whereas compound 2 was an "irreversible" inhibitor of topoisomerase II, compound 1 merely slowed the catalytic cycle of the enzyme . alpha Lapachone (3), on the other hand, inhibited initial non-covalent binding of topoisomerase II to DNA and, in addition, induced religation of DNA breaks (even in pre-established ternary complexes) before dissociating the enzyme from DNA . Compound 3 was an "irreversible" inhibitor of topoisomerase II . The diverse and unique mechanisms of topoisomerase II inhibition by pyranonaphthoquinone derivatives reveal novel ways to target the enzyme with potential for anti-cancer drug design.

Avian Dis, 2000 Jul-Sep, 44(3), 721 - 4
An outbreak of erysipelas in 2-day-old poults; Hollifield JL et al.; Systemic erysipelas infection was seen in 2-to-4-day-old poults from three separate ranches owned by the same company . The affected poults were all from the same breeder source; poults from other breeder sources were seemingly unaffected . Mortality increased on days 2 and 3, ranging from 2% to 8.5% . Birds submitted were dehydrated and very weak, with one half of the poults submitted having died during transport to the lab . Gross lesions included swollen, congested livers and spleens, as well as hemorrhagic breast muscle in one case . Toes were swollen and reddish-purple in color . The poults had been toe-trimmed during hatchery processing using a commercial microwave . Histologically, periportal inflammation with heterophilic infiltration in the liver was noted . Spleens showed hyalinization of arteries, lymphoid depletion, and necrosis . Toe joints showed purulent synovitis and cellulitis . Gram stains done on impression smears of liver and spleen showed rare to moderate numbers of small gram-positive rods . Erysipelothrix rhusiopathiae was isolated from 18 of 22 livers cultured, five of six toe joints cultured, and from the yolk sac in two birds.

J Pathol, 2000 Oct, 192(2), 263 - 72
Role of macrophage scavenger receptor in endotoxin shock; Kobayashi Y et al.; Lipopolysaccharide (LPS) is known to bind to several receptors on macrophages, including CD14 and macrophage scavenger receptor class A types I and II (MSR-A), and stimulates macrophages to release various inflammatory mediators . MSR-A recognizes a broad range of polyanionic ligands such as chemically modified lipoproteins, LPS of Gram-negative bacteria, and lipoteichoic acid of Gram-positive bacteria, suggesting a role in host defence . In this study, mice lacking MSR-A were used to elucidate the role of MSR-A in endotoxin shock . Peritoneal macrophages from MSR-A-deficient (MSR-A(-/-)) mice bound less remarkably to LPS than those from wild-type (MSR-A(+/+)) mice and the binding activity of MSR-A(+/+) macrophages to LPS was reduced by the addition of an anti-MSR-A antibody . Clearance of LPS in serum was retarded in MSR-A(-/-) mice after intraperitoneal administration of LPS . LPS-induced expression of cytokines in the liver was similar in MSR-A(+/+) and MSR-A(-/-) mice, but levels of interleukin (IL)-1beta expression and serum IL-1beta were lower in MSR-A(-/-) mice . Administration of large doses of LPS resulted in a higher mortality of MSR-A(+/+) mice and pretreatment with an IL-1 receptor antagonist reduced the mortality . Thus, MSR-A-mediated macrophage activation plays a negative role in protecting mice from endotoxin shock by enhancing IL-1beta production by macrophages .

J Bacteriol, 2000 Oct, 182(20), 5683 - 91
Transcriptional regulation of the cpr gene cluster in ortho-chlorophenol-respiring Desulfitobacterium dehalogenans; Smidt H et al.; To characterize the expression and possible regulation of reductive dehalogenation in halorespiring bacteria, a 11.5-kb genomic fragment containing the o-chlorophenol reductive dehalogenase-encoding cprBA genes of the gram-positive bacterium Desulfitobacterium dehalogenans was subjected to detailed molecular characterization . Sequence analysis revealed the presence of eight designated genes with the order cprTKZEBACD and with the same polarity except for cprT . The deduced cprC and cprK gene products belong to the NirI/NosR and CRP-FNR families of transcription regulatory proteins, respectively . CprD and CprE are predicted to be molecular chaperones of the GroEL type, whereas cprT may encode a homologue of the trigger factor folding catalysts . Northern blot analysis, reverse transcriptase PCR, and primer extension analysis were used to elucidate the transcriptional organization and regulation of the cpr gene cluster . Results indicated halorespiration-specific transcriptional induction of the monocistronic cprT gene and the biscistronic cprBA and cprZE genes . Occasional read-through at cprC gives rise to a tetracistronic cprBACD transcript . Transcription of cprBA was induced 15-fold upon addition of the o-chlorophenolic substrate 3-chloro-4-hydroxyphenylacetic acid within 30 min with concomitant induction of dehalogenation activity . Putative regulatory protein binding motifs that to some extent resemble the FNR box were identified in the cprT-cprK and cprK-cprZ intergenic regions and the promoter at cprB, suggesting a role for FNR-like CprK in the control of expression of the cprTKZEBACD genes.

Ginekol Pol, 2000 Jun, 71(6), 542 - 9
{The influence of severe infections on the natural killer cells in neonates with vary gestational age}; Mazur B et al.; In 27 infected neonates (16 boys and 11 girls) with birth weight ranged from 1200 to 4500 g (mean 2670 +/- 827 g) and gestational age from 31 to 41 weeks (mean 36.7 +/- 4.1), among them 11 (41%) with birth asphyxia, between 1 and 7 day of life flow cytometric immunophenotyping of the relative sizes of natural killer cells (CD3-/CD16-56+) using monoclonal antibodies of Becton Dickinson were made . The following infections were diagnosed in the neonates: bacterial septicemia (gram-positive in 13 cases, gram-negative in 5 cases), pneumonia in 6 cases and purulent meningitis in 3 cases . Control group consisted of full-term, healthy, eutrophic neonates born in Military Medical Academy in Lodz between 1994-95 years, in which immunological studies were made with the same method and dr Banasik, agreed for using her results for statistical analysis . It was stated, that in infected full-term neonates the mean relative size of NK cells was 10.7 +/- 5.1% and in prematures 12.8 +/- 7.2% and did'n differ significantly from the control . Kind of infection had no significant influence on the mean relative size of these cells, because in septic neonates was 11.7 +/- 5.7% and in other infected babies 11.6 +/- 7.2% . It was found that infected neonates without respiratory insufficiency and shock had significantly higher (13.2 +/- 6.4%) relative size of NK cells than healthy . The lowest value (7.0 +/- 4.8%) of NK cells during infections in dead neonates were noted (4 cases, among them 2 septic with shock and 2 pneumonic prematures with intraventricular haemorrhages) . CONCLUSION: The severity and bad results of the treatment in neonatal infections may be due to deficit of NK cells.

J Bacteriol, 2000 Oct, 182(19), 5470 - 8
WhiA, a protein of unknown function conserved among gram-positive bacteria, is essential for sporulation in Streptomyces coelicolor A3(2); Ainsa JA et al.; The whiA sporulation gene of Streptomyces coelicolor A3(2), which plays a key role in switching aerial hyphae away from continued extension growth and toward sporulation septation, was cloned by complementation of whiA mutants . DNA sequencing of the wild-type allele and five whiA mutations verified that whiA is a gene encoding a protein with homologues in all gram-positive bacteria whose genome sequence is known, whether of high or low G+C content . No function has been attributed to any of these WhiA-like proteins . In most cases, as in S . coelicolor, the whiA-like gene is downstream of other conserved genes in an operon-like cluster . Phenotypic analysis of a constructed disruption mutant confirmed that whiA is essential for sporulation . whiA is transcribed from at least two promoters, the most downstream of which is located within the preceding gene and is strongly up-regulated when colonies are undergoing sporulation . The up-regulation depends on a functional whiA gene, suggesting positive autoregulation, although it is not known whether this is direct or indirect . Unlike the promoters of some other sporulation-regulatory genes, the whiA promoter does not depend on the sporulation-specific sigma factor encoded by whiG.

Clin Infect Dis, 2000 Aug, 31 Suppl 2, S24 - 8
Mechanisms of action and resistance of older and newer fluoroquinolones; Hooper DC; The fluoroquinolones interact with 2 bacterial targets, the related enzymes DNA gyrase and topoisomerase IV, both of which are involved in DNA replication . Quinolones form complexes of these enzymes with DNA, complexes that block movement of the DNA-replication fork and thereby inhibit DNA replication . Many older quinolones differ in their relative activities against gyrase and topoisomerase IV in a bacterial cell, having greater potency against gyrase than against topoisomerase IV in many gram-negative bacteria and greater potency against topoisomerase IV than against gyrase in many gram-positive bacteria . Several newer quinolones appear to have more closely balanced activity against these enzymes . Resistance to fluoroquinolones occurs as a result of mutational amino acid substitutions in the subunits of the more sensitive (or primary-target) enzyme within the cell . If, however, both enzymes are similarly susceptible to a fluoroquinolone, then the level of resistance caused by a primary-target mutation may be low and may be limited by the sensitivity of the secondary target . Fluoroquinolones also differ in the extent to which common bacterial multidrug efflux pumps affect their activity, with some compounds being unaffected by resistance mechanisms because of overexpression of such pumps . Newer fluoroquinolone interaction with dual targets and avoidance of efflux-resistance mechanisms may each contribute to the lower frequencies of selection of resistant mutants in the laboratory.

Laryngoscope, 2000 Sep, 110(9), 1511 - 5
An animal model for the study of genetic predisposition in the pathogenesis of middle ear inflammation; Clark JM et al.; OBJECTIVES: Chronic otitis media with effusion (COME) is the most prevalent inflammatory disease in children and is associated with numerous adverse long-term sequelae . Many factors have been associated with an increased risk of developing COME, one of which may be a genetic predisposition to the disease . To study the role that genetics play in the pathogenesis of COME, we used an animal model to compare the middle ear inflammatory responses in two different strains of rats (Lewis and Fisher) . METHODS: In earlier studies, we demonstrated that exposure of the middle ear to endotoxin caused early extensive exudation and, later, goblet cell hyperplasia and mucin hypersecretion . In the present study, the animals were divided into six groups . In each group the animals were given transtympanic injection with gram-positive bacterial cell wall product (peptidoglycan-polysaccharide {PG-PS}) . The middle ear bullae were studied at 1 week and 3 weeks after infection, and after systemic reinfection . Comparisons were made of the quantity of mucin exudate by enzyme-linked immunosorbent assay and by histological evaluation of the middle ear epithelial thickness . RESULTS: Our data demonstrate a statistically significant difference in middle ear inflammation and effusion formation between the two genetically different strains of rats . CONCLUSIONS: These data support the hypothesis that the middle ear response to PG-PS may be genetically determined and therefore suggest that genetic predisposition may play a role in the pathogenesis of COME.

Nucleic Acids Res, 2000 Sep 15, 28(18), 3417 - 32
SURVEY AND SUMMARY: holliday junction resolvases and related nucleases: identification of new families, phyletic distribution and evolutionary trajectories; Aravind L et al.; Holliday junction resolvases (HJRs) are key enzymes of DNA recombination . A detailed computer analysis of the structural and evolutionary relationships of HJRs and related nucleases suggests that the HJR function has evolved independently from at least four distinct structural folds, namely RNase H, endonuclease, endonuclease VII-colicin E and RusA . The endonuclease fold, whose structural prototypes are the phage lambda exonuclease, the very short patch repair nuclease (Vsr) and type II restriction enzymes, is shown to encompass by far a greater diversity of nucleases than previously suspected . This fold unifies archaeal HJRs, repair nucleases such as RecB and Vsr, restriction enzymes and a variety of predicted nucleases whose specific activities remain to be determined . Within the RNase H fold a new family of predicted HJRs, which is nearly ubiquitous in bacteria, was discovered, in addition to the previously characterized RuvC family . The proteins of this family, typified by Escherichia coli YqgF, are likely to function as an alternative to RuvC in most bacteria, but could be the principal HJRs in low-GC Gram-positive bacteria and AQUIFEX: Endonuclease VII of phage T4 is shown to serve as a structural template for many nucleases, including MCR:A and other type II restriction enzymes . Together with colicin E7, endonuclease VII defines a distinct metal-dependent nuclease fold . As a result of this analysis, the principal HJRs are now known or confidently predicted for all bacteria and archaea whose genomes have been completely sequenced, with many species encoding multiple potential HJRs . Horizontal gene transfer, lineage-specific gene loss and gene family expansion, and non-orthologous gene displacement seem to have been major forces in the evolution of HJRs and related nucleases . A remarkable case of displacement is seen in the Lyme disease spirochete Borrelia burgdorferi, which does not possess any of the typical HJRs, but instead encodes, in its chromosome and each of the linear plasmids, members of the lambda exonuclease family predicted to function as HJRs . The diversity of HJRs and related nucleases in bacteria and archaea contrasts with their near absence in eukaryotes . The few detected eukaryotic representatives of the endonuclease fold and the RNase H fold have probably been acquired from bacteria via horizontal gene transfer . The identity of the principal HJR(s) involved in recombination in eukaryotes remains uncertain; this function could be performed by topoisomerase IB or by a novel, so far undetected, class of enzymes . Likely HJRs and related nucleases were identified in the genomes of numerous bacterial and eukaryotic DNA viruses . Gene flow between viral and cellular genomes has probably played a major role in the evolution of this class of enzymes . This analysis resulted in the prediction of numerous previously unnoticed nucleases, some of which are likely to be new restriction enzymes.

Front Biosci, 2000 Sep 01, 5, D837 - 65
Proteasomes in the archaea: from structure to function; Maupin-Furlow JA et al.; Survival of cells is critically dependent on their ability to rapidly adapt to changes in the natural environment no matter how 'extreme'the habitat . An interplay between protein folding and hydrolysis is emerging as a central mechanism for stress survival and proper cell function . In eucaryotic cells, most proteins destined for destruction are covalently modified by the ubiquitin-system and then degraded in an energy-dependent mechanism by the 26S proteasome, a multicatalytic protease . The 26S proteasome is composed of a 20S proteolytic core and 19S cap (PA700) regulator which includes six AAA+ ATPase subunits . Related AAA+ proteins and 20S proteasomes are found in the archaea and Gram positive actinomycetes . In general, 20S proteasomes form a barrel-shaped nanocompartment with narrow openings which isolate rather non-specific proteolytic active-sites to the interior of the cylinder and away from interaction with cytosolic proteins . The proteasome-associated AAA+ proteins are predicted to form ring-like structures which unfold substrate proteins for entry into the central proteolytic 20S chamber resulting in an energy-dependent and processive destruction of the protein . Detailed biochemical and biophysical analysis as well as identification of proteasomes in archaea with developed genetic tools are providing a foundation for understanding the biological role of the proteasome in these unusual organisms.

FEMS Microbiol Ecol, 2000 Aug 1, 33(2), 111 - 120
Diverse, yet-to-be-cultured members of the Rubrobacter subdivision of the Actinobacteria are widespread in Australian arid soils; Holmes AJ et al.; Phylogenetic analyses of ribosomal RNA gene sequences (rDNAs) retrieved from an Australian desert soil sample (Sturt National Park) revealed the presence of a number of clones which branched deeply from the high GC Gram-positive division line of descent . The most abundant group of these clones were related to Rubrobacter . An oligonucleotide probe was designed to have broad specificity to Rubrobacter and relatives . This probe was used to interrogate eight rDNA libraries representing four distinct land forms within the Australian arid zone . Relative abundance of Rubrobacter-relatives in these samples ranged from 2.6 to 10.2% . Clones from these libraries were selected for sequence analysis on the basis of a heteroduplex mobility assay to maximise the diversity represented in the sample . Phylogenetic analyses of these rDNA clones and Rubrobacter-related clones reported in the literature show strong support for three distinct groups . Database-searching revealed 'Rubrobacteria' were relatively abundant in a number of published soil rDNA libraries but absent from others . A PCR assay for group-1 'Rubrobacteria' was used to test for their presence in 21 environmental samples . Only marine and arid-zone soil samples gave positive PCR results . Taken together these results indicate 'Rubrobacteria' are a widespread group of variable abundance and diversity.

Ophthalmology, 2000 Sep, 107(9), 1769 - 75
Corneal microsporidiosis . Report of case, including electron microscopic observations; Font RL et al.; OBJECTIVE: To report a case of corneal stromal infection caused by a protozoon of the genus MICROSPORIDIA:, including clinical, histopathologic, and electron microscopic observations . DESIGN: Case report . METHODS: Light and electron microscopy studies were performed on keratectomy specimens from a 67-year-old immunocompetent man who had a unilateral chronic stromal keratitis that was refractory to medical treatment . Initial corneal biopsy followed by lamellar and penetrating keratoplasty were performed on the patient . All the specimens were studied histopathologically . RESULTS: Light microscopy of the corneal biopsy and the subsequent keratectomy specimens demonstrated myriad small, round to oval microsporidial organisms measuring 3.5 to 5.0 micrometer in length that stained positively with the periodic acid-Schiff, Grocott-methenamine silver, and acid-fast methods and were gram positive . Electron microscopic observations demonstrated viable blastospores that had a thin osmiophilic outer cell wall and contained 11 to 13 coils of the filament . The light and electron microscopic features, the tinctorial characteristics, and the selective corneal stromal involvement are consistent with microsporidial keratitis . CONCLUSIONS: Microsporidiosis should be considered in the differential diagnosis of a culture-negative stromal keratitis refractory to medical treatment . The diagnosis can be easily established based on the morphologic features of the protozoa in the keratectomy specimens . No effective medical treatment for the stromal disease is available . Full-thickness keratoplasty is suggested because, in our patient, lamellar keratoplasty did not preclude recurrence of the disease.

Microbes Infect, 2000 Jul, 2(8), 933 - 43
The role of the interleukin-1/Toll-like receptor superfamily in inflammation and host defence; Fitzgerald KA et al.; The IL-1 receptor/Toll-like receptor superfamily comprises a diverse family of cell surface receptors defined by a characteristic conserved sequence in their cytosolic regions, termed the Toll/IL-1 receptor domain, which function in inflammation and host defence against microbial pathogens . Members include receptors for the proinflammatory cytokines IL-1 and IL-18 and Toll-like receptors 2 and 4, which are involved in host responses to Gram-positive and Gram-negative bacteria, respectively . Signalling pathways activated by these receptors are conserved and the superfamily represents a pan-genomic system involved in the host response to infection and injury.

Org Lett, 2000 Aug 10, 2(16), 2467 - 70
Total synthesis of cristatic acid; Furstner A et al.; The first total synthesis of cristatic acid 1, an antibiotic endowed with considerable activity against Gram-positive bacteria, hemolytic properties, and significant cytotoxicity, is described . Key to success are the formation of its 2,4-disubstituted furan moiety via a palladium-catalyzed alkylation of vinylepoxide 10 derived from sulfonium salt 8 and the use of SEM ethers as the protecting groups for the phenolic OH functions.

J Biol Chem, 2000 Dec 1, 275(48), 37373 - 81
Immulectin-2, a lipopolysaccharide-specific lectin from an insect, Manduca sexta, is induced in response to gram-negative bacteria; Yu XQ et al.; A lipopolysaccharide-specific lectin, immulectin-2, was isolated from plasma of the tobacco hornworm, Manduca sexta . Immulectin-2 has specificity for xylose, glucose, lipopolysaccharide, and mannan . A cDNA clone encoding immulectin-2 was isolated from an Escherichia coli-induced M . sexta larval fat body cDNA library . The cDNA is 1253 base pairs long, with an open reading frame of 981 base pairs, encoding a 327-residue polypeptide . Immulectin-2 is a member of the C-type lectin superfamily . It consists of two carbohydrate recognition domains, which is similar to the organization of M . sexta immulectin-1 . Immulectin-2 was present at a constitutively low level in plasma of control larvae and increased 3-4-fold after injection of Gram-negative bacteria or lipopolysaccharide . Immulectin-2 mRNA was detected in fat body of control larvae, and its level increased dramatically after injection of E . coli . The concentration of immulectin-2 in plasma did not change significantly after injection of Gram-positive bacteria or yeast, even though its mRNA level was increased by these treatments . Compared with immulectin-1, immulectin-2 has a more restricted specificity for binding to Gram-negative bacteria . Immulectin-2 at low physiological concentrations agglutinated E . coli in a calcium-dependent manner . It also bound to immobilized lipopolysaccharide from E . coli . Binding of immulectin-2 to lipopolysaccharide stimulated phenol oxidase activation in plasma . The properties of immulectin-2 are consistent with its function as a pattern recognition receptor for detection and defense against Gram-negative bacterial infection in M . sexta.

J Heart Valve Dis, 2000 Jul, 9(4), 544 - 51
The Quattro valve and active infective endocarditis of the mitral valve; Middlemost SJ et al.; BACKGROUND AND AIM OF THE STUDY: Even today, infective endocarditis remains a therapeutic challenge . Active endocarditis at the time of valve implantation is an important risk factor for the development of prosthetic valve infection . This study reports results following implantation of the Quattro valve, a stentless chordally supported quadrileaflet mitral valve made from bovine pericardium . METHODS: The Quattro valve was implanted in seven patients (four females, three males; mean age 34 years) requiring isolated mitral valve replacement for active bacterial endocarditis . All had congestive heart failure; two were in cardiogenic shock . The diagnosis of active endocarditis was based on clinical and echocardiographic findings, together with macroscopic evidence of acute infection at surgery, blood culture or histopathological evidence of valve infection . Postoperatively, all patients received at least four weeks of parenteral antibiotic therapy . RESULTS: Congestive heart failure (and large pedunculated vegetations and mobile septic left atrial thrombi in two patients) prompted early surgical intervention . Patients underwent surgery at a mean of 7 days (range: 1-16 days) after admission . Endocarditis was caused by Gram-positive cocci in all patients except one . At a mean follow up of 15 months (range: 6-24 months) all patients were alive and symptomatically improved . To date, all remain free of prosthetic valve endocarditis, reoperation and thromboembolism . CONCLUSION: The Quattro valve can be implanted safely in patients with acute bacterial endocarditis . The results also reflect the benefit of early surgical intervention in patients with infective endocarditis complicated by congestive heart failure, with or without large vegetations.

J Food Prot, 2000 Aug, 63(8), 1100 - 6
Bactericidal properties of ozone and its potential application as a terminal disinfectant; Moore G et al.; The efficacy of ozone as a terminal disinfectant was evaluated under laboratory conditions . Different microorganisms of importance to the food industry were inoculated onto stainless steel squares and incubated at various temperatures and relative humidities for up to 4 h . Survival of microorganisms from these controls was compared with identically incubated squares exposed to ozone . Exposure of the contaminated surfaces to ozone (2 ppm for 4 h) resulted in a reduction in microbial viability that ranged, depending on organism type, from 7.56 to 2.41 log values . For all the microorganisms tested, this loss in viability was significantly greater (P < 0.05) than that observed in the absence of ozone . Gram-negative bacteria were more sensitive to ozone than gram-positive organisms; bacteria were more sensitive than the yeast strain tested . Exposure to ozone (2 ppm for 4 h) in the presence of ultra-high temperature (UHT) milk resulted in a reduction in bacterial viability that ranged from 5.64 to 1.65 log values . In most cases, this reduction was significantly less (P < 0.05) than that achieved in the absence of organic material, although still significantly greater (P < 0.05) than that observed in the absence of ozone . The presence of a meat-based broth reduced the effectiveness of ozone to a greater extent, although the number of surviving gram-negative organisms was still significantly less (P < 0.05) than in the absence of ozone . Less than 1 log unit of yeast cells was destroyed when exposed to ozone in the presence of UHT milk or meat-based broth . Results of this investigation suggest that if applied after adequate cleaning ozone could be used as an effective disinfectant.

Microbiol Immunol, 2000, 44(6), 511 - 7
Interleukin-8 secretion of human epithelial and monocytic cell lines induced by middle ear pathogens; Kita H et al.; Otitis media with effusion (OME) is one of the most common diseases in children . Alloiococcus otitidis, a new gram-positive bacterial species, was isolated from the middle ear fluid of children with OME; however, the pathogenic role of this bacteria is yet unknown . In this study, the ability of cultured epithelial cell lines (Hep-2 and Hela) and monocytic cell lines (THP-1 and U 937) to secrete chemokine interleukin-8 (IL-8) in response to the A . otitidis organism and three bacterial organisms mainly detected from middle ear fluid in OME, and bacterial cell components was investigated . When stimulated with four viable bacterial cells, epithelial cells and monocytes secreted IL-8 in a time-dependent manner . The monocytes produced significantly higher levels of IL-8 than the epithelial cells . Compared with that by viable bacterial cells, IL-8 secretion by stimulated epithelial cells and monocytes was reduced when the bacteria were heated and treated with glutaraldehyde . With bacterial stimulations, cell treatment of interferon-gamma caused monocytes to increase the induction of IL-8 production, however, the induction of monocyte differentiation caused monocytes to reduce the induction of IL-8 production . Furthermore, epithelial cells and monocytes stimulated by four viable bacterial organisms physically separated from cultured cells reduced the induction of IL-8 compared with directly stimulated cells, and monocytes stimulated with soluble extracts prepared from A . otitidis organisms produced IL-8 in a dose-dependent manner . These results suggest that part of the IL-8 stimulation of the A . otitidis organism may exist in a diffusable factor released by the bacteria or soluble components of the bacteria itself.

J Mol Microbiol Biotechnol, 1999 Aug, 1(1), 107 - 25
The RND permease superfamily: an ancient, ubiquitous and diverse family that includes human disease and development proteins; Tseng TT et al.; A previous report identified and classified a small family of gram-negative bacterial drug and heavy metal efflux permeases, now commonly referred to as the RND family (TC no . 2.6) . We here show that this family is actually a ubiquitous superfamily with representation in all major kingdoms . We report phylogenetic analyses that define seven families within the RND superfamily as follows: (1) the heavy metal efflux (HME) family (gram negative bacteria), (2) the hydrophobe/amphiphile efflux-1 (HAE1) family (gram negative bacteria), (3) the nodulation factor exporter (NFE) family (gram negative bacteria), (4) the SecDF protein-secretion accessory protein (SecDF) family (gram negative and gram positive bacteria as well as archaea), (5) the hydrophobe/amphiphile efflux-2 (HAE2) family (gram positive bacteria), (6) the eukaryotic sterol homeostasis (ESH) family, and (7) the hydrophobe/amphiphile efflux-3 (HAE3) family (archaea and spirochetes) . Functionally uncharacterized proteins were identified that are members of the RND superfamily but fall outside of these seven families . Some of the eukaryotic homologues function as enzymes and receptors instead of (or in addition to) transporters . The sizes and topological patterns exhibited by members of all seven families are shown to be strikingly similar, and statistical analyses establish common descent . Multiple alignments of proteins within each family allow derivation of family-specific signature sequences . Structural, functional, mechanistic and evolutionary implication of the reported results are discussed.

Ned Tijdschr Geneeskd, 2000 Jul 29, 144(31), 1494 - 7
{Septic arthritis in two young children caused by unusual gram-negative pathogens}; Bruijn J et al.; Two children, a girl aged 2 years and a boy aged 10 months, were moderately ill with signs of inflammation of the left and the right knee, respectively . Both had had pharyngitis, and the boy also had paronychia of the right foot . The Gram preparation of synovial fluid showed Gram-positive cocci in the girl, while Kingella kingae was cultured . In the boy, a Moraxella was cultured from the synovial fluid using an aerobic blood culture system . Both recovered without sequelae after adequate antibiotic treatment . The micro-organisms cultured were Gram-negative bacteria, which are rarely seen in septic arthritis and are difficult to demonstrate . In young children, septic arthritis often presents with mild symptoms and inconclusive laboratory findings . Even if the Gram preparation of the synovial fluid shows no micro-organisms, unusual pathogens may be isolated by means of an aerobic blood culture system.

Annu Rev Biophys Biomol Struct, 2000, 29, 265 - 89
The structural biology of molecular recognition by vancomycin; Loll PJ et al.; Vancomycin is the archetype among naturally occurring compounds known as glycopeptide antibiotics . Because it is a vital therapeutic agent used world-wide for the treatment of infections with gram-positive bacteria, emerging bacterial resistance to vancomycin is a major public health threat . Recent investigations into the mechanisms of action of glycopeptide antibiotics are driven by a need to understand their detailed mechanism of action so that new agents can be developed to overcome resistance . These investigations have revealed that glycopeptide antibiotics exhibit a rich array of complex cooperative phenomena when they bind target ligands, making them valuable model systems for the study of molecular recognition.

J Refract Surg, 2000 Jul-Aug, 16(4), 467 - 9
Nocardia keratitis after traumatic detachment of a laser in situ keratomileusis flap; Kim EK et al.; PURPOSE: Nocardia are gram-positive bacteria existing ubiquitously in the environment; they can cause keratitis . Nocardia asteroides keratitis occurred in the interface between the stromal bed and flap after traumatic detachment of the flap 4 months after an initially uncomplicated laser in situ keratomileusis (LASIK) procedure . METHODS: Nocardia asteroides keratitis was confirmed by culture . Therapy included topical and oral trimethoprim-sulfamethoxazole . RESULTS: Thirteen months after the trauma, the patient's spectacle-corrected visual acuity was 20/20 with a manifest refraction of -2.25 -1.00 x 30 degrees . CONCLUSIONS: The immediate steps of management consisting of surgically lifting the corneal flap, rapid microbial identification, and proper treatment with specific antibiotics resulted in the successful treatment of Nocardia asteroides keratitis in a traumatized eye after LASIK.

Int J Syst Evol Microbiol, 2000 Jul, 50 Pt 4, 1553 - 61
Arthrobacter flavus sp . nov., a psychrophilic bacterium isolated from a pond in McMurdo Dry Valley, Antarctica; Reddy GS et al.; CMS 19YT, a psychrophilic bacterium, was isolated from a cyanobacterial mat sample from a pond in Antarctica and was characterized taxonomically . The bacterium was aerobic, gram-positive, non-spore-forming, non-motile, exhibited a rod-coccus growth cycle and produced a yellow pigment that was insoluble in water but soluble in methanol . No growth factors were required and it was able to grow between 5 and 30 degrees C, between pH 6 and pH 9 and tolerated up to 11.5% NaCl . The cell wall peptidoglycan was Lys-Thr-Ala3 (the A3alpha variant) and the major menaquinone was MK-9(H2) . The G+C content of the DNA was 64+/-2 mol% . The 16S rDNA analysis indicated that CMS 19YT is closely related to group I Arthrobacter species and showed highest sequence similarity (97.91%) with Arthrobacter agilis . Furthermore, DNA-DNA . hybridization studies also indicated 77% homology between CMS 19YT and A . agilis . It differed from A . agilis, however, in that it was psychrophilic, non-motile, yellow in colour, exhibited a rod-coccus growth cycle, had a higher degree of tolerance to NaCl and was oxidase- and urease-negative and lipase-positive . In addition, it had a distinct fatty acid composition compared to that of A . agilis: the predominant fatty acids were C15:0, anteiso-C15:0, C16:0, iso-C16:0, C17:0, anteiso-C17:0 and C18:0 . It is proposed, therefore, that CMS 19YT should be placed in the genus Arthrobacter as a new species, i.e . Arthrobacter flavus sp . nov . The type strain of A . flavus is CMS 19YT (= MTCC 3476T).

Kidney Int Suppl, 2000 Aug, 76, S148 - 55
Adsorption in sepsis; Ronco C et al.; The pathophysiology of sepsis offers a highly complicated scenario . In sepsis, endotoxin or other gram-positive-derived products induce a complex and dynamic cellular response, giving rise to several mediators known to be relevant in the pathogenesis of septic shock such as specific mediators, substances responsible for up- or down-regulation of cytokine receptors and cytokine antagonists, inactivators of translational or transductional pathways, and precursor molecules . In this review, we delve into some new concepts stemming up from the use of sorbents in continuous plasma filtration . Nonspecific simultaneous removal of several mediators of the inflammatory cascade have led to improved outcomes in animal models of septic shock and to improved hemodynamics in a pilot clinical study . It seems of great importance to explore all possible treatment techniques that may have a direct impact on circulating mediators of sepsis and that also may interfere with the imbalance between proinflammatory and anti-inflammatory substances in the critically ill patient with multiple organ failure . In this view, the application of sorbents appears to open new and interesting therapeutic options . The search for innovative treatments specifically targeted to the special needs of the critically ill patients seems therefore more important than the attempt to adjust concepts and technologies that are normally applied to patients with chronic renal failure.

J Biotechnol, 2000 Jul 28, 81(1), 63 - 72
Molecular cloning of a gene encoding the thermoactive levansucrase from Rrahnella aquatilis and its growth phase-dependent expression in Eescherichia coli; Seo JW et al.; A levansucrase gene (lsrA) from Rahnella aquatilis ATCC33071 was isolated from a genomic library and the nucleotide sequence of the lsrA structural gene was determined . lsrA is composed of 1248 bp and encodes 415 amino acid residues with a calculated molecular mass of 45.9 kDa . Although the amino acid sequence of lsrA gene showed good conservation with the sequences of reported levansucrases and of the conserved regions thought to be implicated in the enzyme activity, comparison of the deduced amino acid sequences certified the dissimilarity of the proteins from Gram-negative and Gram-positive bacteria . The lsrA gene was expressed from its own promoter in Escherichia coli in an active form . The lsrA expression in E . coli-pRL1CPR was affected by the growth phase of cells: it was repressed in the early phase of growth, but was significantly stimulated during the entrance of cells into the late phase of growth . The growth-phase-dependent fashion of lsrA expression was altered in a constitutive-like fashion by the deletion of an upstream region of lsrA (pNd137), suggesting that the growth-phase dependent expression of lsrA was mediated by the deleted upstream region.

Rev Latinoam Microbiol, 1996 Jul-Dec, 38(3-4), 185 - 91
Antibiotic 26-deoxylaidlomycin isolated from Streptomyces sp . Ar386 from Brazilian soil; Ujikawa K et al.; An actinomycete strain (Ar386) was isolated from the soil of the Araraquara region, SP, Brazil . The strain, named Streptomyces jacareensis, formed irregular rayed, rugose, grayish-white mycelium with sinuous, branched hyphae carrying rare isolated spores; assimilated glucose, galactose, inositol, ribose, maltose, sucrose, melibiose and starch but not mannitol, rhamnose, arabinose, xylose, lactose and raffinose; and contained LL-diaminopimelic acid in its cell wall . An antibiotic active against Gram-positive bacteria, which was characterized as being 26-deoxylaidlomycin and which may have application against poultry coccidiosis, was isolated from cultures of the strain . This was the first isolation of this antibiotic from a microorganism of the genus Streptomyces and also the first isolation of this antibiotic in Brazil.

Mol Microbiol, 2000 Aug, 37(3), 477 - 84
Plasmid rolling-circle replication: recent developments; Khan SA; It is now well established that a large majority of small, multicopy plasmids of Gram-positive bacteria use the rolling-circle (RC) mechanism for their replication . Furthermore, the host range of RC plasmids now includes Gram-negative organisms as well as archaea . RC plasmids can be broadly classified into at least five families, individual members of which are spread among widely different bacteria . There is significant homology in the basic replicons of plasmids belonging to a particular family, and there is compelling evidence that such plasmids have evolved from common ancestors . Major advances have recently been made in our understanding of plasmid RC replication, including the characterization of the biochemical activities of the plasmid initiator proteins and their interaction with the double-strand origin, the domain structure of the initiator proteins and the molecular basis for the function of single-strand origins in plasmid lagging strand synthesis . Over the past several years, there has been a 'renaissance' in studies on RC replication as a result of the discovery that many plasmids replicate by this mechanism, and studies in the next few years are likely to reveal new and novel mechanisms used by RC plasmids for their regulated replication.

Syst Appl Microbiol, 2000 Jun, 23(2), 219 - 29
New LL-diaminopimelic acid-containing actinomycetes from hypersaline, heliothermal and meromictic Antarctic Ekho Lake: Nocardioides aquaticus sp . nov . and Friedmanniella {correction of Friedmannielly} lacustris sp . nov; Lawson PA et al.; Two Gram-positive, non-motile and aerobic bacteria were isolated from a water sample of the hypersaline Ekho Lake, Antarctica . The cocci or short rods grew well on oligotrophic PYGV agar of pH 7.5 and at 26 degrees C . Strains EL-17KT and EL-17AT both required thiamine and biotin, strain EL-17AT also required nicotinic acid . Carbon sources utilized by both strains were acetate, pyruvate, alpha-D-glucose, glutamate and (weakly) citrate, but succinate, malate or butyrate were utilized only by EL-17KT . Gelatin, starch and DNA were hydrolyzed, NH, was formed from peptone, and nitrate was reduced aerobically by both strains . The isolates had the same temperature tolerance for growth in the range tested (below 3 to above 33.5 degrees C) and pH range (<5.5 to >9.5) and were sensitive to chloramphenicol and penicillin G . Their cell walls contained LL-diaminopimelic acid and had a single glycine residue as interpeptide bridge . Strain EL-17AT contained glycine at position 1 of the peptide subunit (peptidoglycan type A 3gamma') . Isolates EL-17KT and EL-17AT differed in their maximum NaCl tolerance, which was 15% or 6-8%, respectively . The major fatty acid of EL-17KT was C18:1 and that of EL-17AT was ai-C15:0 . The major respiratory quinones of EL-17KT and EL-17AT were MK-8(H4) and MK-9(H4), respectively . The former isolate had 69 mol% G+C, the latter had 73 mol% G+C . Comparative 16S rRNA gene sequencing revealed phylogenetic relationships of isolate EL-17KT with the genus Nocardioides, with N . pyridinolyticus and N . plantarum as the closest relatives . Phenotypic and genotypic characteristics support the description of a new species, Nocardioides aquaticus sp . nov., with EL-17KT as the type strain (= DSM 11439T) . Isolate EL-17AT is related to the genus Friedmanniella, with E antarctica and E spumicola as the closest relatives . The differentiating characteristics support the description of a new species, Friedmanniella lacustris sp . nov., with EL-17AT as the type strain (= DSM 11465T).

Ann Med Interne (Paris), 2000 Jun, 151(4), 243 - 7
{Abdominal actinomycosis . Report of three cases and review of the literature}; Kacem C et al.; Actinomyces are Gram positive, anaerobic, filamentous bacteria, saprophytes of the oral cavity and intestinal tract of humans . They rarely cause chronic suppurative infections . Abdominal abscess can masquerade as a malignant process and lead to a surgical intervention with resection . We report three patients with abdominal actinomycosis; the first affected the liver with a favorable outcome on medical treatment, the second of abdomino-pelvic localisation was related to an intra-uterine device and the last affecting the transverse colon simulated a malignant process and required an hemicolectomy . These observations illustrates the difficulties of the diagnosis of this rare and unrecognized disease.

Harefuah, 1998 May 1, 134(9), 695 - 7, 750
{Necrotizing soft tissue infection}; Rath E et al.; The dramatic course of necrotizing soft tissue infection represents a medical emergency, since it is limb- and life-threatening . Most necrotizing soft tissue infections are caused by mixed aerobic and anaerobic Gram-negative and Gram-positive organisms . Most case have been reported in immuno-compromised hosts after penetrating trauma or surgery . We describe a unique series of cases of necrotizing soft tissue infection . The mainstay of treatment is early and daily debridement of devitalized tissue and broad-spectrum antibiotics . Hyperbaric oxygen therapy should be considered.

Rev Lat Am Enfermagem, 2000 Jan, 8(1), 57 - 65
{The effect of sugar on the process of cicatrization of infected surgical incisions}; Haddad M do C et al.; This is an experimental research based on microbiological analysis . Our aim was to evaluate the influence of sugar on the healing process of infected surgical wounds in 25 patients . On the dehiscences, there have been identified gram positive and gram negative bacteria, and fungi as well . Every patient was administered systemic antibiotics and had the wounds treated locally with crystal sugar three times a day . Statistics showed, by Person's correlation, that sugar did not influence on the healing process of infected surgical wounds in undernourished, overweight and elderly individuals.

Acta Otolaryngol Suppl, 2000, 543, 122 - 6
Acute mastoidectomy in a Danish county from 1977 to 1997--operative findings and long-term results; Petersen CG et al.; Data from patients undergoing acute mastoidectomy were examined retrospectively to evaluate if the nature of acute mastoiditis (AM) treated surgically has changed during the last 20 years (1977-97) . Moreover, a prevalence study was conducted to clarify the otological and audiological course following acute mastoidectomy . Patients with cholesteatoma and intracranial complications were excluded . Thus, 79 patients with a median age of 16 months were included . Thirty-seven percent had a history of middle ear disease, and the mean duration from onset of symptoms to admission was 9 days . Well-being was affected in 46%, and 82% had fever . The clinical picture was dominated by auricular protrusion (77%) and pathological tympanic membrane (94%) . Postauricular oedema, hyperaemia and tenderness were demonstrated in 89%, 78% and 49% of cases, respectively . Peroperatively, purulent middle ear effusion was recognized in 92%, subperiosteal abscess in 66% and pus in the mastoid in 90% . Specimens revealed growth of pathogens in 58%, predominantly Gram-positive bacteria . The observation period was 1-20 years . The findings in operated ears were not significantly different from the contralateral non-operated ears concerning incidence of otitis media, hearing and ear canal volume . Conclusively, acute mastoidectomy is a safe and effective treatment to eliminate infection . The operation can be done with negligible risk and does not leave long-term sequelae.

J Comp Pathol, 2000 Jul, 123(1), 72 - 6
Actinomycosis of dogs caused by Actinomyces hordeovulneris; Pelle G et al.; Actinomyces hordeovulneris was isolated from the lesions of chronic pyogranulomatous pleuritis and pericarditis of one of three dogs showing similar symptoms . The parietal pleura and the pericardium were thickened and covered with fine short threads of angiofibroblastic tissue . About 500-1000 ml of reddish purulent exudate in the thorax of all three dogs contained large numbers of rice-grain-sized, soft, yellowish-white granules ("sulphur granules") . These granules had a central core of branching filaments of gram-positive bacteria embedded in thick granulation tissue . The parietal pleura, the mediastinal pleura and the pericardium were infiltrated mainly with neutrophils, and to a lesser extent with lymphocytes and plasma cells . A small number of eosinophils and giant cells was also observed . Large numbers of pyogranulomas embedded in the granulation tissue were composed of a core of necrotized granulation tissue, mixed with clusters of gram-positive branching bacteria, surrounded by an area of intact and degenerating neutrophils and lymphocytes . Bacteria were detected in the lesions by Brown-Brenn staining and were isolated from one of the affected animals . The isolated bacteria were identified as A . hordeovulneris . This was the first isolation of A . hordeovulneris in Hungary .

J Hepatol, 2000 Jul, 33(1), 9 - 18
Amplification and sequence analysis of partial bacterial 16S ribosomal RNA gene in gallbladder bile from patients with primary biliary cirrhosis; Hiramatsu K et al.; BACKGROUND/AIMS: The etiopathogenesis of bile duct lesion in primary biliary cirrhosis is unknown, though the participation of bacteria and/or their components and products is suspected . In this study, we tried to detect and identify bacteria in the bile of patients with primary biliary cirrhosis by polymerase chain reaction using universal bacterial primers of the 16S ribosomal RNA gene . METHODS: Gallbladder bile samples from 15 patients with primary biliary cirrhosis, 5 with primary sclerosing cholangitis, 5 with hepatitis C virus-related liver cirrhosis, 11 with cholecystolithiasis, and from 12 normal adult gallbladders were used . In addition to the culture study, partial bacterial 16S ribosomal RNA gene was amplified by polymerase chain reaction (PCR) taking advantage of universal primers that can amplify the gene of almost all bacterial species, and the amplicons were cloned and sequenced . Sequence homology with specific bacterial species was analyzed by database research . Bacterial contamination at every step of the bile sampling, DNA extraction and PCR study was avoided . Furthermore, to confirm whether bacterial DNA is detectable in liver explants, the same analysis was performed using 10 liver explants of patients with primary biliary cirrhosis . RESULTS: In primary biliary cirrhosis, 75% (p<0.0001) of 100 clones were identified as so-called gram-positive cocci while these cocci were positive in only 5% in cholecystolithiasis (p<0.0001) . In cholecystolithiasis gram-negative rods were predominant instead . One bacterial species detected in a normal adult was not related to those detected in primary biliary cirrhosis and cholecystolithiasis patients . No bacterial DNA was detected by PCR amplification in 10 liver explants of patients with primary biliary cirrhosis . CONCLUSIONS: The present results raise several possible roles of gram-positive bacteria in bile in the etiopathogenesis of primary biliary cirrhosis . However, these results could also reflect an epiphenomenon due to decreased bile flow in the patients with primary biliary cirrhosis at an advanced stage.

Biochem J, 2000 Aug 1, 349 Pt 3, 717 - 28
Structure-function relationships in novel peptide dodecamerswith broad-spectrum bactericidal and endotoxin-neutralizing activities; Mayo KH et al.; A series of designed peptide 33-mers (betapep peptides) areknown to be bactericidal {Mayo, Haseman, Ilyina and Gray (1998)Biochim . Biophys . Acta 1425, 81-92} . Here dodecapeptides (SC-1-SC-8), which 'walk through' the sequence ofbetapep-25, were investigated for their ability to kill Gram-negativeand -positive bacteria and to neutralize endotoxin . SC-4 (KLFKRHLKWKI I-NH(2); the -NH(2) at the right of each sequenceindicates amidation of the C-terminal carboxylate group) is the mosteffective, more so than betapep-25, at killing Gram-negative bacteriawith nanomolar LD(50) values . Against Gram-positive bacteria,SC-4 also shows good activity with submicromolar LD(50)values . Leakage studies indicate rapid bacterial membrane permeability,with t(1/2) valuesof 10-15 min . SC-4 in the micromolar range also effectivelyneutralizes endotoxin and is not haemolytic below 10(-4)M . For all SC peptides, CD and NMR data indicate the presence of both 3(10)- and alpha-helix . For SC-4, nuclear-Overhauser-effect-based computational modelling yields an amphipathic helix with K1, K4,R5, and K8 arrayed on the same face (K is lysine, R is arginine).Activity differences among SC peptides and single-site variants of SC-4allow some structure-function relationships to be deduced.Relative to other known bactericidal peptides in the linear peptide,helix-forming category, SC-4 is the most potent broad-spectrumantibacterial identified to date . The present study contributes to thedevelopment of agents involved in combating the ever-recurring problemof drug-resistant micro-organisms.

J Am Acad Dermatol, 2000 Aug, 43(2 Pt 2), 333 - 6
Cutaneous Mycobacterium chelonae in a liver transplant patient; Nathan DL et al.; A 51-year-old woman with an orthotopic liver transplant on tacrolimus (SKF 506) and prednisone presented with an erythematous ulcerated nodule on the knee . No preceding trauma was noted . A skin biopsy specimen demonstrated beaded gram-positive, acid-fast rods and the skin culture grew Mycobacterium chelonae (formerly M chelonae subsp chelonae ) . This report describes the first case in a liver transplant patient of cutaneous Mycobacterium chelonae under the current method of designating atypical Mycobacterium species.

J Bone Joint Surg Am, 2000 Jul, 82-A(7), 1050 - 1
Paternalism; Capozzi JD et al.; J . S . is a sixty-five-year-old man who was treated at another hospital with arthroscopic debridement of an infection at the site of a right total knee replacement and was placed on long-term intravenous antibiotics . He signed out of that hospital against medical advice . One month later, he presented at our hospital with recurrent sepsis of his knee . Knee aspiration yielded frank pus with a white blood-cell count of 80,000 cells per cubic millimeter . Gram-staining demonstrated gram-positive cocci . The patient was placed on intravenous antibiotics . The patient appeared cachectic, reporting a sixty-pound (27.2-kilogram) weight loss over the past year . A metastatic workup, including a chest radiograph, an abdominal sonogram, prostate-specific antigen, a complete blood-cell count, erythrocyte sedimentation rate, and a purified-protein-derivative skin test, was negative; however, an occult neoplasm could not be excluded . The patient displayed episodes of confusion, disorientation, and argumentative behavior . Medical and psychiatric consults did not determine whether this behavior was due to previous substance abuse or a primary psychiatric disorder . Nevertheless, psychiatrists at our institution determined that the patient lacked decisional capacity . Attempts were made to salvage the knee replacement, and the patient underwent an extensive surgical debridement of the knee with insertion of drains . He was placed on intravenous antibiotics . The plan was for the patient to be managed with long-term oral suppressive antibiotics . After treatment, the patient was transferred to a skilled-nursing facility . Psychiatrists at the nursing facility deemed the patient to have decisional capacity, and the patient was permitted to leave the facility . He was discharged without antibiotics . Several weeks later, he presented at our hospital with a grossly purulent knee . The orthopaedic options were reviewed with the patient and his brother . Removal of the components was recommended . The patient did not want to "lose" his knee replacement, and he refused surgical intervention . We did not believe that the infection could be either controlled or eradicated with the components in place.

Eur J Surg Suppl, 1999, (584), 33 - 8
The overall concept: immunoglobulins as adjuvants for mediastinitis--does it make sense?
Werdan K.
Supportive intravenous immunoglobulin (ivIg) treatment may reduce the incidence and severity of infections in a well-defined group of patients in intensive care, including those who have had cardiac surgery . Sternal wound infections and mediastinitis after cardiac surgery are to be investigated in the ATMI study with Pentaglobin . In this article I focus on optimising ivIg dosage, on likely mechanisms, on reasons why ivIg might improve a primarily gram-positive infection, and on the use of serial scoring as an endpoint in a trial of immunoglobulin.

J Clin Periodontol, 2000 Jun, 27(6), 437 - 46
PCR monitoring for tetracycline resistance genes in subgingival plaque following site-specific periodontal therapy . A preliminary report; Manch-Citron JN et al.; BACKGROUND: The selection of antibiotic resistance genes during antibiotic therapy is a critical problem complicated by the transmission of resistance genes to previously sensitive strains via conjugative plasmids and transposons and by the transfer of resistance genes between gram-positive and gram-negative bacteria . The purpose of this investigation was to monitor the presence of selected tetracycline resistance genes in subgingival plaque during site specific tetracycline fiber therapy in 10 patients with adult periodontitis . METHOD: The polymerase chain reaction (PCR) was used in separate tests for the presence of 3 tetracycline resistance genes (tetM, tetO and tetQ) in DNA purified from subgingival plaque samples . Samples were collected at baseline, i.e., immediately prior to treatment, and at 2 weeks, and 1, 3, and 6 months post-fiber placement . The baseline and 6-month samples were also subjected to DNA hybridization tests for the presence of 8 putative periodontal pathogenic bacteria . RESULTS: PCR analysis for the tetM resistance gene showed little or no change in 5 patients and a decrease in detectability in the remaining 5 patients over the 6 months following tetracycline fiber placement . The results for tetO and tetQ were variable showing either no change in detectability from baseline through the 6-month sampling interval or a slight increase in detectability over time in 4 of the 10 patients . DNA hybridization analysis showed reductions to unmeasurable levels of the putative periodontal pathogenic bacteria in all but 2 of the 10 patients . CONCLUSIONS: These results complement earlier studies of tet resistance and demonstrate the efficacy of PCR monitoring for the appearance of specific resistance genes during and after antibiotic therapy.

Microbiology, 2000 Jul, 146 ( Pt 7), 1525 - 33
Extracytoplasmic proteins of Mycobacterium tuberculosis - mature secreted proteins often start with aspartic acid and proline; Wiker HG et al.; A surrogate expression system, based on fusions to the phoA bacterial reporter gene, was used to identify Mycobacterium tuberculosis genes that encode exported proteins and the promoter regions required for their expression in the heterologous host Mycobacterium smegmatis . To assess these results in the context of the complete M . tuberculosis genome sequence, the corresponding genes were identified and computational algorithms were employed to identify signal peptide (SP), transmembrane domain and membrane lipoprotein attachment motifs . This information was used to predict the subset of M . tuberculosis genes that encode exported proteins . Of the 34 genes identified by the phoA method, 22 were classified to encode potential soluble secreted proteins . Among these, 14 genes may encode novel secreted proteins . Six of the remaining 12 genes were predicted to encode membrane lipoproteins and an additional six to encode integral membrane proteins . Published observations of proteins proven to be secreted into M . tuberculosis culture filtrates were reviewed to further characterize the mycobacterial SP motif . It was concluded that mycobacterial SPs are comparable in size to Gram-positive SPs, but certain features are different . In particular, arginine was the predominant N-terminally positively charged amino acid in contrast to lysine in the Gram-positives . The hydrophobic transmembrane segment of the SP was dominated by alanine, in contrast to leucine . At the C-terminal end of the SPs, the (-3, -1) rule (AXA motif) holds, with alanine as the dominant amino acid in both positions, being most dominant in the (-1) position . A high proportion of mature sequences start with aspartic acid in the (+1) position and proline in the (+2) position - the DP motif . The authors propose that the DP sequence serves as a sorting signal, following translocation and cleavage by signal peptidase I . Alternatively, the DP motif may be part of the recognition site for the signal peptidase.

Z Gastroenterol, 2000 May, 38(5), 375 - 9
{Actinomycosis of the pelvis with an indwelling IUD}; Kurz R et al.; Infection with actinomycosis israeli (an anaerobic, gram-positive bacterium) presents as chronic inflammation with tendency to fibrosis and suppuration with formation of external sinuses . Cervicofacial, thoracic and abdominal forms of the disease made up 95% of cases of actinomycosis . A 53-year-old woman was admitted to the hospital because of a pelvic mass which was thought to be malignant . A laparotomy was performed and the histologic examination showed actinomycosis . The patient first received penicillin followed by tetracyclin and the pelvic mass shrunk . One year later no more mass was detectable . We think that the IUP in place over years is the source for this infection.

Ann Fr Anesth Reanim, 2000 May, 19(5), 375 - 81
{Meningitis after locoregional spinal anesthesia}; Gorce P et al.; OBJECTIVE: Meningitis is a severe and an uncommon complication of both spinal and epidural anaesthesia . This review summarizes the knowledge on epidemiology, clinical and microbiological diagnosis and the ways to prevent them . DATA SOURCES: Articles published in English and French language since 1989 has been collected on Medline database, using "meningitis", "spinal anaesthesia" and "epidural anaesthesia" as keywords . DATA SYNTHESIS: Bacterial meningitis are usually in relation with Gram positive bacterias which is a clue for an exogenous contamination . Another unusual ways of contamination are blood circulating bacterias and spreading of local infection . Aseptic meningitis has been described, in relation to introduction of irritant agents in subarachnoid space . Lumbar puncture must be done each time meningitis suspected so that it can assert the diagnosis and guide antibiotherapy . Easy hygienic guidelines has been widely published to prevent meningitis . Usually, antibiotherapy alone is sufficient to treat meningitis but with an unjustified cost and sometimes severe persistent neurologic sequelae . CONCLUSION: The unexpected appearance of meningitis during the wearing-off of a spinal anesthesia is exceptional; the possibility of death or serious sequela must be taken into account . The sources of contamination are quite frequently exogenous, the germs coming most often from the patient's cutaneous flora or the anesthetist's ENT flora . Prevention of this risk involves a rigorous respect for cutaneous disinfection and hygiene procedures . The anesthetist's medico-legal responsibilities will be called upon in case of exogenous contamination.

Plasmid, 2000 Jul, 44(1), 66 - 84
IS1294, a DNA element that transposes by RC transposition; Tavakoli N et al.; IS1294, found on the ColD-like resistance plasmid pUB2380, is IS91-like . It is an active 1.7-kb insertion sequence that lacks terminal inverted repeats, displays insertion-site specificity, and does not generate direct repeats of the target site . The element has one large open reading frame, tnp(1294), encoding a transposase of 351 amino acids, related to members of the REP family of replication proteins used by RC-plasmids of gram-positive bacteria . IS1294 transposes using rolling-circle replication, initiated at one end of the element, oriIS, and terminated at the other, terIS . oriIS and terIS are highly conserved among like IS elements . oriIS resembles the leading strand replication origins of RC-plasmids; terIS resembles a rho-independent transcription terminator . IS1294 mediates not only its own transposition, but also sequences adjacent to terIS . A transposition model for IS1294 and related elements, involving rolling-circle replication and single-strand DNA intermediates, is presented .

J Pharmacol Exp Ther, 2000 Jul, 294(1), 280 - 6
Effect of lipoteichoic acid on dermal vascular permeability in mice; Wada K et al.; Lipoteichoic acid (LTA), the cell wall component of Gram-positive bacteria, has been shown to cause inflammatory responses comparable to lipopolysaccharide (LPS) of Gram-negative bacteria . This study examined the activity of LTA to induce dermal microvascular permeability changes in mice . Vascular permeability was assessed by extravasation of Pontamine sky blue . Subcutaneous injection of LTA (200-400 microg/site) in mice that were preinjected i.v . with the dye increased local dye leakage in the skin at 1 to 3 h . The LTA-induced dye leakage was inhibited by indomethacin, valeryl salicylate, diphenhydramine, and a platelet-activating factor antagonist but not by inhibitors of nitric-oxide synthase, cyclooxygenase-2, or guanylate cyclase or by antibodies against tumor necrosis factor-alpha or interleukin-1alpha . LTA induced comparable increases in dye leakage in inducible nitric-oxide synthase-deficient mice and wild-type controls . Pretreatment of normal mice with i.v . LTA did not confer tolerance to LTA- or LPS-induced dye leakage . In contrast, systemic LPS administration induced tolerance against subsequent challenge with LPS but not LTA . Serum corticosterone levels, which were suggested to induce tolerance, were not increased by LTA pretreatment but were increased by LPS . Thus, LTA increases dermal microvascular permeability in mice . Among the inflammatory mediators, eicosanoids, platelet-activating factor, and histamine mediate the effect of both LTA and LPS, whereas nitric oxide, tumor necrosis factor-alpha, and interleukin-1alpha may not play a major role in LTA-induced dye leakage . The difference between LTA and LPS to stimulate corticosterone may partially explain the failure of LTA to induce tolerance against vascular dye leakage.

J Antibiot (Tokyo), 2000 Apr, 53(4), 329 - 36
New biologically active rubiginones from Streptomyces sp; Puder C et al.; Four new polyketides, named rubiginone D2 (2), 4-O-acetyl-rubiginone D2 (3), rubiginone H (6) and rubiginone I (7) were isolated from the cultures of Streptomyces sp . (strain Go N1/5) . Their structures were established by a detailed spectroscopic analysis . The absolute configuration of 3 was determined by derivatization with chiral acids (Helmchen's method) . The rubiginones inhibit the growth of some Gram-positive bacteria and are cytostatically active against different tumor cell lines.

J Intraven Nurs, 1999 Nov-Dec, 22(6), 336 - 42
Vancomycin peak serum concentration monitoring; Tam VH et al.; Vancomycin is a glycopeptide antibiotic commonly used in the treatment of gram-positive bacterial infections . With the rising prevalence of resistant gram-positive bacterial infections in healthcare institutions and the advent of attempts to curb escalating medical cost, vancomycin is being used with increasing frequency in the homecare setting for infection management . In view of its variability in pharmacokinetics and the potential for toxicities, serum concentrations often are measured . This article reviews the pharmacology of vancomycin and suggests a practical approach to serum concentration monitoring in homecare.

Anat Rec, 2000 Jul 1, 259(3), 276 - 87
Transmission electron microscope study of bacterial morphotypes on the anterior dorsal surface of human tongues; Arora HK et al.; The human tongue has been the subject of many cytological and histological studies . When a literature search disclosed no reports of the ultrastructure of the morphotypes of bacteria residing on the tongue's surface, a transmission electron microscope study of ultrathin sections of bacteria obtained by scraping eight human tongues was undertaken . The scrapings from the anterior dorsal tongue surfaces, processed conventionally for electron microscope study, revealed 33-35 different bacterial morphotypes . Several of the morphotypes were unique to a tongue . Morphotype differences were also related to donor characteristics such as smoking, tongue site, location in centrifuge pellet, diet, and medications . The predominant morphotypes were Gram-positive cocci . These preliminary findings suggest that the microbiota of the human tongue and variations in that microbiota, related to physical condition, lifestyle, medications, and dietary preferences, merit more attention from anatomists .

SADJ, 1999 Jun, 54(6), 253 - 6
External bacterial contamination of local anaesthetic cartridges; Basson NJ et al.; In this study external bacterial contamination of local anaesthetic cartridges from newly opened and open containers was examined . Colonies of mainly Gram-positive cocci and Gram-positive rods were grown from cartridges from both the freshly opened and open containers . However, the total number of colonies grown from open containers was significantly higher than that from freshly opened ones . It was concluded that where local anaesthetic cartridges are bulk-packed in containers, strict infection control measures are to be instituted in clinical practice . We suggest that these include keeping containers tightly capped, removing cartridges only when needed, using forceps to handle cartridges and swabbing cartridges with alcohol prior to loading into syringes.

Antimicrob Agents Chemother, 2000 Jul, 44(7), 1778 - 82
Stenotrophomonas maltophilia D457R contains a cluster of genes from gram-positive bacteria involved in antibiotic and heavy metal resistance; Alonso A et al.; A cluster of genes involved in antibiotic and heavy metal resistance has been characterized from a clinical isolate of the gram-negative bacterium Stenotrophomonas maltophilia . These genes include a macrolide phosphotransferase (mphBM) and a cadmium efflux determinant (cadA), together with the gene cadC coding for its transcriptional regulator . The cadC cadA region is flanked by a truncated IS257 sequence and a region coding for a bin3 invertase . Despite their presence in a gram-negative bacterium, these genetic elements share a common gram-positive origin . The possible origin of these determinants as a remnant composite transposon as well as the role of gene transfer between gram-positive and gram-negative bacteria for the acquisition of antibiotic resistance determinants in chronic, mixed infections is discussed.

J Bacteriol, 2000 Jul, 182(13), 3649 - 54
Characterization of the fructosyltransferase gene of Actinomyces naeslundii WVU45; Bergeron LJ et al.; Oral actinomycetes produce fructosyltransferase (FTF) enzymes which convert sucrose into polymers of D-fructose, known as levans, and these polymers are thought to contribute to the persistence and virulence of the organisms . A gene encoding FTF was isolated from Actinomyces naeslundii WVU45; the deduced amino acid sequence showed significant similarity to known levansucrases of gram-negative environmental isolates but was less similar to FTFs from gram-positive bacteria . A transcriptional start site was mapped by primer extension 70 bp 5' from the putative start codon . Promoter fusions to a chloramphenicol acetyltransferase gene were used to confirm that there was a functional promoter driving ftf expression and to show that sequences located 86 to 218 bp upstream of the transcription initiation site were required for optimal ftf expression . Quantitative slot blot analysis against total RNA from cells grown on different sugars or from different growth phases revealed that ftf was constitutively transcribed . Thus, the A . naeslundii FTF is more similar in primary sequence and the regulation of expression to levansucrases of gram-negative bacteria than gram-positive bacteria.

Arterioscler Thromb Vasc Biol, 2000 Jun, 20(6), 1521 - 8
Lipoteichoic acid induces delayed protection in the rat heart: A comparison with endotoxin; Zacharowski K et al.; Classic ischemic preconditioning transiently (30 to 120 minutes) protects the myocardium against subsequent lethal ischemia/reperfusion injury . After dissipation of this acute protection, a second window of protection (SWOP) appears 12 to 24 hours later; this SWOP lasts up to 3 days . Several triggers induce a SWOP, including brief repetitive cycles of coronary artery occlusion, rapid ventricular pacing, stimulation of adenosine A(1) receptors, and administration of wall fragments of Gram-negative bacteria, such as lipopolysaccharide (LPS) . The aim of this study was to investigate whether lipoteichoic acid (LTA), a cell wall fragment of Gram-positive bacteria, can induce a SWOP in a rat model of left anterior descending coronary artery (LAD) occlusion (25 minutes) and reperfusion (2 hours) . Thus, 166 male Wistar rats were pretreated (2 to 24 hours) with saline, LTA (1 mg/kg IP), or LPS (1 mg/kg IP) and subjected to LAD occlusion/reperfusion . Pretreatment with LTA or LPS for 16 hours led to a substantial, approximately 65%, reduction in infarct size and a reduction in the release of cardiac troponin T into the plasma . The dose of LTA used had no toxic effect (on any of the parameters studied), whereas the same dose of LPS caused a time-dependent activation of the coagulation system and liver injury . By use of RNase protection assays, it was determined that LPS caused a time-dependent induction of tumor necrosis factor-alpha, interleukin-1beta, and manganese superoxide dismutase mRNA content in the heart, whereas LTA failed to induce manganese superoxide dismutase . LPS also caused an upregulation of the expression of intercellular adhesion molecule-1 and P-selectin, whereas LTA downregulated these molecules and attenuated the accumulation of polymorphonuclear granulocytes caused by myocardial ischemia/reperfusion . This study demonstrates for the first time that pretreatment with LTA at 8 to 24 hours before myocardial ischemia significantly reduces (1) infarct size, (2) cardiac troponin T, and (3) the histological signs of tissue injury in rats subjected to LAD occlusion and reperfusion . The mechanism(s) underlying the observed cardioprotective effects of LTA warrants further investigation but is likely to be related to its ability to inhibit the interactions between the coronary vascular endothelium and polymorphonuclear granulocytes . Therefore, LTA represents a novel and promising agent capable of enhancing myocardial tolerance to ischemia/reperfusion injury.

What Is Biofilter?, What Is Antibiotic?, What Is Cell Biology?, What Is Anthrax?, What Is Genetics?, o, Microbes, c, Bacterium, s, Microorganism, r, Microorganisms, r, Microbe, o, Escherichia coli, i, Cell suspensions, n, Gram negative, i, Pseudomonas aeruginosa, a, Antimicrobials, s, Bacillus subtilis, e, Bactericidal, r, Escherichia coli, e, Candida albicans, n, Escherichia coli, n, Streptomycin, r, Enterobacters, i, Yeasts, i, Clostridia, o, Escherichia coli, o, Antibiotics, a, Bacteria, r, Microbiological, i, Escherichia coli, o, Bacteria, c, Streptococci

Scientific Publications - Work Done by Microbiology Reader Bioscreen C
Agricultural Microbiology Anaerobic Microbiology Antimicrobial Susceptibility Artificial Atmosphere Bioassay of Antibiotics Biofilm Microbiology Bioreactor Technology Biotechnology Cell Biology Clinical Microbiology Environmental Microbiology Experiments with Yeast	Fermentation Food Microbiology Functional Genomics Gene Technology Growth Media Development Growth Rate and Lag Time Industrial Microbiology Medical/Pharmaceutical Field Microbiological Assay Microbiological Research Microbiology of Cosmetics go to a specific theme...	Military Microbiology Molecular Microbiology Mutagenicity and Genotoxicity Oral Microbiology Patents Postantibiotic Studies Soil Microbiology Spore Microbiology Veterinary Microbiology Waste/Wastewater Treatment Water Microbiology Wine Microbiology

� 2005 Transgalactic Ltd (manufacturer of Bioscreen C software) | Privacy Statement | P.O. Box 1393, 00101 Helsinki, Finland, phone: +358 9 85172920, fax: +358 9 8749481, e-mail: microbiology@bionewsonline.com

Last modified: May 25, 2005