Gram negative

Eur J Pharmacol, 1993 Aug 3, 239(1-3), 23 - 30
Nitric oxide, but not interleukin-1, mediates the local blood flow response to lipopolysaccharide in rabbit skin; Pons F et al.; Lipopolysaccharide can mimic many aspects of the inflammatory response to gram-negative bacteria . In rabbit and rat skin, lipopolysaccharide induces neutrophil accumulation and an increase in blood flow . Interleukin-1, tumor necrosis factor-alpha and the vasodilator, nitric oxide (NO) have been implicated in the biological responses to lipopolysaccharide and gram-negative bacteria . In the present study, we characterized the local vascular response to E . coli lipopolysaccharide in rabbit skin and investigated its dependence on de novo protein synthesis, NO, interleukin-1 and neutrophils . The local vascular response to the intradermal injection of lipopolysaccharide was determined by measuring changes in local blood flow with a laser-Doppler flowmeter . Injections of lipopolysaccharide (3, 10 and 30 micrograms/site) induced a dose-related increase in blood flow, with a maximum at 2 h . The response to 10 micrograms/site lipopolysaccharide was abolished by co-injection of actinomycin D (4 x 10(-9) mol/site), reduced by 60% by NG-nitro-L-arginine methyl ester (10(-7) mol/site) and suppressed by 82% in rabbits pretreated with dexamethasone (2 mg/kg, i.v.) . However, the lipopolysaccharide-induced increase in blood flow was not affected by co-injection of recombinant human interleukin-1 receptor antagonist (7.0 x 10(-11) mol/site) or by systemic neutrophil depletion . These results demonstrate that lipopolysaccharide causes delayed vasodilatation of the rabbit cutaneous microvasculature which depends on protein synthesis and involves in part the release of NO . However, this response does not depend on interleukin-1 or neutrophils . The vasodilator effect of lipopolysaccharide may involve the expression of the inducible form of NO synthase in the vessel wall, through a mechanism independent of interleukin-1.

J Laryngol Otol, 1993 Aug, 107(8), 726 - 9
Severe cochlear dysplasia causing recurrent meningitis: a surgical lesson; Stevenson DS et al.; Meningitis may be the sole presenting sign of a cerebrospinal fluid (CSF) fistula of the temporal bone . An eight-year-old boy suffering from recurrent meningitis was found to have bilateral severe cochlear dysplasia . Bilateral tympanotomies were performed, planning to obliterate each vestibule . In the right ear a stapedectomy was performed, resulting in a torrential 'CSF gusher' and difficulty in packing the vestibule . CSF rhinorrhoea requiring revision surgery and two episodes of gram-negative bacterial meningitis complicated the post-operative management, resulting in a prolonged hospital stay . Subsequently, the left ear was managed in a different fashion, leaving the stapes in situ, with grafts placed to seal the oval window niche . We would recommend this alternative procedure in cases of severe cochlear dysplasia, where abnormalities of the vestibule and basal turn of the cochlea mean that performing a stapedectomy to pack the vestibule may result in a severe 'CSF gusher', by opening directly into the subarachnoid space.

Pediatr Nephrol, 1993 Aug, 7(4), 455 - 6
Hypogammaglobulinemia and fatal sepsis in an infant maintained on peritoneal dialysis; Neu AM et al.; Chronic peritoneal dialysis (CPD) is a common form of renal replacement therapy in children . Recent studies suggest that immunological abnormalities, in particular hypogammaglobulinemia, may develop in children and infants on peritoneal dialysis . We report an infant maintained on CPD who died of gram-negative sepsis . At post-mortem examination, he was noted to have severe panhypogammaglobulinemia.

Am J Kidney Dis, 1993 Aug, 22(2), 296 - 9
Gentamicin clearance during hemodialysis: a comparison of high-efficiency cuprammonium rayon and conventional cellulose ester hemodialyzers; Agarwal R et al.; The advent of high-efficiency hemodialyzers has afforded improved efficiency of urea clearance; however, increased clearance of other substances, particularly antibiotics, also may occur, necessitating changes in clinical practice . Accordingly, we compared the efficiency of gentamicin removal using two different hemodialyzers, a conventional saponified cellulose ester (CD 135) and a high-efficiency cuprammonium rayon dialyzer (TAF 175L), in eight hospitalized patients undergoing antibiotic therapy for suspected or proven gram-negative infection . The rate of dialysis, estimated as the ratio of dialyzer urea clearance (K) to urea distribution volume (V) (K/V urea), and the total elimination rate constant (k) of gentamicin were measured during 17 hemodialysis treatments . The K/V urea for the two dialyzers, TAF 175L and CD 135, was 0.390 +/- 0.024 hr-1 and 0.413 +/- 0.129 hr-1 (P = NS), respectively . The TAF 175L hemodialyzer was almost twice as efficient in removing gentamicin as the CD 135: TAF 175, k = 0.263 +/- 0.024 hr-1; CD 135, k = 0.132 +/- 0.027 hr-1 (P < 0.001) . Moreover, the rate of dialysis (K/V urea) was correlated with k of gentamicin for the TAF 175L dialyzer (r2 = 0.50, P < 0.02) but not for the CD 135 dialyzer . We conclude that dialyzer characteristics and the rate of dialysis (K/V urea) should be taken into consideration when determining the dosage of gentamicin in patients on hemodialysis.

Dig Dis Sci, 1993 Aug, 38(8), 1530 - 6
Alterations in rat intestinal transit by morphine promote bacterial translocation; Runkel NS et al.; Translocation of enteric microorganisms from the intestinal tract to extraintestinal sites has been proposed as an early step in the development of gram-negative sepsis . This study examined the role of altered bowel transit in influencing intestinal bacteriostasis and bacterial translocation using morphine as a pharmacologic inhibitor of such transit . In the first experiment, either normal saline (N = 8) or morphine sulfate (20 mg/kg; N = 8) was injected subcutaneously . Two hours later, morphine (7.5 mg/kg) was infused subcutaneously for an additional 22 hr; control animals received saline alone . After completion of this regimen, a volume of 0.2 ml of 2.5 mM FITC dextrans (10,000 daltons) were injected intraduodenally in each group . The bowel was removed 25 min later, divided into 5-cm segments, and the content of dextrans measured . Small bowel propulsion was expressed as the geometric center of the distribution of dextrans throughout the intestine (in percentage length of small bowel) . Gut propulsion was significantly reduced after morphine treatment as compared to controls (32.8 +/- 8.2% vs . 55.8 +/- 4.0%; P < 0.01) . In 16 additional rats, saline or morphine was again administered as described . After 24 hr, samples were obtained from the mesenteric lymph node (MLN) complex, blood, spleen, liver, duodenum, jejunum, ileum, and cecum for standard bacteriology . The bacterial counts increased significantly in each intestinal segment following morphine treatment . Microorganisms translocated to the MLN complex in 5, and to distant sites in four of eight morphine-treated animals, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Surgery, 1993 Aug, 114(2), 140 - 6
Role of bactericidal permeability-increasing protein in the treatment of gram-negative pneumonia; Kelly CJ et al.; BACKGROUND . Gram-negative infections are a major cause of morbidity and death . Bactericidal permeability-increasing protein (BPI) is an endotoxin-neutralizing protein that also exhibits potent bactericidal activity . This study compared the efficacy of a 23 kd recombinant N-terminal fragment of BPI (rBPI23) with that of antiendotoxin antibody E5 in a model of gram-negative sepsis . METHODS . Sixty Swiss-Webster mice (Carworth farm) received an intratracheal inoculation of Escherichia coli (7 x 10(6) colony-forming units) and were randomized to three groups (20 per group) . Starting immediately after inoculation, the groups received either rBPI23 (4 mg/kg intravenously every 2 hours for four doses), E5 (11 mg/kg intravenously every 24 hours for two doses), or an isotype control antibody B55 (11 mg/kg intravenously every 24 hours for two doses) and were followed up for survival . In a second survival study, 40 mice received the same intratracheal inoculation of E . coli and were randomized to two groups . Starting 2 hours after inoculation, the groups received either rBPI23 (4 mg/kg intravenously every 2 hours for four doses) or E5 (8 mg/kg intravenously every 12 hours for four doses) and were followed up for survival . In a third study, mice received an intratracheal inoculation of 3 x 10(6) colony-forming units E . coli, a sublethal dose, and were killed to determine pulmonary and blood clearance of bacteria . RESULTS . rBPI23 conferred significantly greater protection from death than either E5 or B55 when started immediately (95% survival vs 20% and 10%, respectively; p < 0.001) or 2 hours after inoculation (65% survival vs 25% for E5; p < 0.05) . Both pulmonary and vascular clearance of bacteria was enhanced significantly by treatment with rBPI23 . CONCLUSIONS . rBPI23 may be a novel therapeutic agent in the management of gram-negative sepsis.

Crit Care Med, 1993 Aug, 21(8), 1233 - 41
Introduction of new technology into critical care practice: a history of HA-1A human monoclonal antibody against endotoxin; Luce JM; OBJECTIVES: HA-1A, a monoclonal antibody against endotoxin, was thought to be effective in treating patients with Gram-negative sepsis . Because of this possibility, many clinicians felt obligated to use the drug and assumed that its product license application would be approved by the U.S . Food and Drug Administration (FDA) . Nevertheless, the efficacy of HA-1A was not conclusively demonstrated by a first clinical trial . The FDA rejected the product license application and requested a second clinical trial, which was suspended after excess mortality was noted in patients treated with HA-1A . This review of the history of the drug was prepared to provide clinicians and sepsis investigators with information about HA-1A and, by extension, the process by which new technology is introduced into critical care practice . DATA SOURCES: Data used to prepare this review were obtained from the author's personal files as well as the computerized MEDLINE database . STUDY SELECTION: Studies were selected for their relevance to the history of HA-1A and their relevance to the introduction of potentially useful medical technology . DATA EXTRACTION: The author extracted all applicable data . DATA SYNTHESIS: Although the first clinical trial of HA-1A suggested that the drug was effective in treating patients with Gram-negative bacteremia with or without shock, further analysis by the FDA indicated a benefit only for bacteremic patients with shock . Furthermore, the original study design was not followed, leading in part to the FDA's refusal of the product license application . Concern also was raised over the issue of identifying which patients should receive HA-1A and the cost of the drug, which would have put it past the reach of some American hospitals and thereby, would have conflicted with the ethical principle of social justice . Finally, the second trial suggested that HA-1A might be harmful . CONCLUSIONS: Due to the FDA's action, the issues raised about HA-1A, and the results of the two clinical trials, clinicians should not use the drug . The history of HA-1A provides insights about how new technology is and will be introduced into critical care practice.

J Infect Dis, 1993 Aug, 168(2), 473 - 6
Fluid administration, brain edema, and cerebrospinal fluid lactate and glucose concentrations in experimental Escherichia coli meningitis; Tauber MG et al.; The effect of no fluids versus liberal fluid supplementation on brain edema and cerebrospinal fluid (CSF) lactate and glucose concentrations was compared in rabbits with experimental Escherichia coli meningitis . Fluid restriction for the duration of the experiment (19 h) led to a decrease in body weight by approximately 5%, while the high fluid regimen increased body weight by approximately 5% . Infected animals developed brain edema compared with controls, but the fluid regimen had no measurable effect on the degree of edema . In contrast, fluid-restricted animals had significantly higher CSF lactate and lower CSF glucose concentrations than fluid-supplemented animals (lactate, 13.5 +/- 3.5 vs . 10.1 +/- 3.3 mmol/L; glucose, 1.89 +/- 1.39 vs . 4.11 +/- 1.39 mmol/L) . These results fail to support the hypothesis that administration of large amounts of fluid in this model of gram-negative bacterial meningitis aggravates brain edema.

J Bacteriol, 1993 Aug, 175(15), 4911 - 6
Site-directed mutations in the relaxase operon of RP4; Cole SP et al.; Mutations were constructed by site-directed mutagenesis in the relaxase operon of the broad-host-range plasmid RP4 . The mutations were constructed in smaller plasmids, recombined into the 60-kb RP4 plasmid, and tested for their ability to transfer . The relaxase operon contains the transfer genes traJ, traH, and traI, which are involved in nicking at the transfer origin to generate the single strand destined to be transferred to the recipient cell . In the first mutant, the C terminus of TraI was truncated, leaving TraH intact . This mutant decreased transfer by approximately 500-fold in Escherichia coli, and the traI mutation could be complemented by a wild-type copy of traI in trans in the donor . The traI mutation similarly decreased transfer between a variety of gram-negative bacteria . A site-specific mutation was made by the polymerase chain reaction-based unique-site mutagenesis procedure to alter the start site of traH . This mutation had no effect on intraspecific E . coli transfer but reduced transfer by up to sevenfold for some gram-negative bacteria . The traH mutation had no effect on plasmid stability . Thus, neither TraH nor the C terminus of TraI is required for conjugative transfer, but both increase mating efficiency in some hosts.

J Bacteriol, 1993 Aug, 175(15), 4756 - 63
Oar, a 115-kilodalton membrane protein required for development of Myxococcus xanthus; Martinez-Canamero M et al.; Myxococcus xanthus is a developmental gram-negative bacterium which forms multicellular fruiting bodies upon nutrient starvation . This bacterium was found to contain a 115-kDa membrane protein which separated with the inner membrane fraction by sucrose density gradient centrifugation . The gene for this protein was cloned, and its DNA sequence was determined . The deduced amino acid sequence consists of 1,061 residues . This protein contains a putative signal sequence and many short segments, found scattered throughout the entire protein, that have sequence similarities with OmpA, a major outer membrane protein of Escherichia coli . Thus, the gene was designated oar (OmpA-related protein) . A second open reading frame was found 36 bases downstream of the oar termination codon . This open reading frame encodes a protein of 236 residues and contains a putative lipoprotein signal sequence . An aor disruption mutation (delta oar) showed no effect on vegetative growth but caused abnormal morphogenesis during development and reduced myxospore formation . When examined with a light microscope, delta oar cells were unable to aggregate on developmental agar, indicating that Oar is required for cellular adhesiveness during development.

Infect Immun, 1993 Aug, 61(8), 3184 - 9
An interleukin-6-induced acute-phase response does not confer protection against lipopolysaccharide lethality; Bucklin SE et al.; Lipopolysaccharide (LPS), a component of gram-negative bacterial outer cell walls, can stimulate lymphoreticular cells to produce cytokines such as tumor necrosis factor alpha (TNF-alpha), interleukin-1 (IL-1), and IL-6 . One of these proinflammatory cytokines, IL-6, induces hepatic synthesis of a class of proteins termed acute-phase proteins . D-Galactosamine inhibits acute-phase protein synthesis and concurrently sensitizes mice to a lethal dose of LPS approximately 10,000-fold . From these observations, we hypothesized that the acute-phase response may serve as a defense mechanism for protection of the host against the deleterious effects of LPS . To test this hypothesis, murine recombinant IL-6 (mrIL-6) was used to induce an acute-phase response prior to a lethal LPS challenge in both D-galactosamine-treated and normal mice . Induction of the acute-phase response by mrIL-6 was quantitated by measuring the concentrations of fibrinogen and complement component C3, two well-characterized acute-phase proteins, in the circulation . The effect of acute-phase and normal serum on TNF-alpha release by peritoneal macrophages stimulated with LPS in vitro was also examined . The results of these studies confirmed the induction of the acute-phase response by mrIL-6, as reflected in an approximate doubling in circulating levels of fibrinogen and C3 . However, when either D-galactosamine-sensitized or normal mice were challenged with a lethal dose of LPS at various times after mrIL-6 administration, the acute-phase response induced by mrIL-6 did not alter either cumulative lethality or the kinetics of lethality . Additionally, compared with normal serum, acute-phase serum did not affect TNF-alpha release by peritoneal macrophages following LPS-mediated stimulation in vitro . Collectively, these studies would not support a dominant role for an IL-6-mediated acute-phase response as contributing to the resistance of normal mice compared with D-galactosamine-sensitized mice in LPS-induced lethal toxicity.

Eur J Med, 1993 Aug-Sep, 2(7), 404 - 7
Acid-base and electrolyte abnormalities in febrile patients with bacteraemia; Elisaf M et al.; OBJECTIVES: Very commonly febrile patients with bacteraemia develop a variety of acid-base and electrolyte disturbances which play a significant role in the morbidity and mortality of these patients . This study was undertaken to describe the pathogenetic mechanisms of these abnormalities in febrile patients with bacteraemia . METHODS: Fifteen febrile patients with bacteraemia, aged 24-62 years, were studied . In all patients blood cultures revealed Gram-negative rods . None of them had septic shock, diabetes mellitus, renal or liver failure and none was receiving drugs influencing acid-base balance and electrolyte levels or was a heavy alcohol consumer . RESULTS: Nine patients had respiratory alkalosis, which was possibly due to bacterial toxins, while the remaining 6 had a wide-gap metabolic (lactic) acidosis coexisting with respiratory alkalosis . Hypokalaemia was found in four patients and was mainly due to respiratory alkalosis . However, kaliuria due to hypomagnesaemia contributed to hypokalaemia in 2 patients . Hypomagnesaemia was detected in 3 patients and was attributed to respiratory alkalosis as well as to magnesiuria induced by metabolic acidosis or phosphate depletion . Hypophosphataemia was found in 5 patients who also had respiratory alkalosis and/or phosphaturia due to metabolic acidosis or hypomagnesaemia . Finally, one patient had multifactorial origin hypocalcaemia . CONCLUSION: Febrile patients with bacteraemia develop a number of acid-base and electrolyte disturbances attributed to various pathogenetic mechanisms.

Diagn Microbiol Infect Dis, 1993 Aug-Sep, 17(2), 163 - 6
Central venous catheter infection caused by Moraxella osloensis in a patient receiving home parenteral nutrition; Buchman AL et al.; We report the first case of a central venous catheter infection caused by Moraxella osloensis, which was successfully treated without catheter removal . The isolation, identification, and pathogenesis of this species are discussed . It is recommended that Moraxella isolates be identified to species in order to determine the relative pathogenic and opportunistic roles of the various Moraxella species . Our case also demonstrates that catheter sepsis caused by some Gram-negative organisms may be amenable to systemic antibiotic therapy without the necessity of catheter removal.

Thorax, 1993 Aug, 48(8), 785 - 9
Factors of importance for the long term prognosis after hospital treated pneumonia; Hedlund JU et al.; BACKGROUND--Elderly patients admitted to hospital for community acquired pneumonia have a high risk of recurrence of pneumonia and of death during the years after discharge . In this study potential factors of importance for the long term prognosis after hospital treated pneumonia were retrospectively investigated . METHODS--A total of 241 patients (103 men) with a mean age of 60 (range 18-102) years discharged from hospital after treatment for community acquired pneumonia were studied . After an average follow up period of 31 months, 18 independent variables present during hospital treatment of the initial pneumonia were examined for association with the following end points: recurrence of pneumonia, death from any cause, and death from pneumonia . RESULTS--Age adjusted analysis showed that systemic treatment with corticosteroids correlated significantly with recurrence of pneumonia and with death . The presence of low serum albumin levels on admission or colonisation of the respiratory tract with Gram negative enteric bacteria seemed to be important negative prognostic factors for the outcome during pneumonia recurrences after discharge . CONCLUSIONS--Patients who are admitted to hospital with pneumonia are at risk of subsequent pneumonia and death after discharge . This risk seems to be even higher in patients who are treated with corticosteroids systemically, who have a low serum albumin level on admission, or who become colonised in the respiratory tract with Gram negative enteric bacteria during their hospital stay.

J Rheumatol, 1993 Aug, 20(8), 1388 - 96
Responses to gram negative enteric bacterial antigens by synovial T cells from patients with juvenile chronic arthritis: recognition of heat shock protein HSP60; Life P et al.; OBJECTIVE . To investigate the antigenic specificity of synovial T cells in juvenile chronic arthritis (JCA) . METHODS . Synovial fluid and peripheral blood mononuclear cells from 24 patients with JCA were tested for their proliferative responses to recall antigens, enteric organisms associated with reactive arthritis, influenza A and a recombinant preparation of a mycobacterial heat shock protein, HSP65 . To investigate further recognition of this last antigen, synovial T cells from one B27+ patient with pauciarticular disease were cloned using HSP65 . The specificity of the resultant clones was then examined . RESULTS . Marked synovial T cell responses to enteric organisms and to HSP65 were noted, particularly in HLA-B27+, pauciarticular patients; these were similar to those seen in a B27+ patient with reactive arthritis . Responses to enteric organisms and to HSP65 were significantly correlated, suggesting recognition of an epitope common to these antigen preparations . However, all of the T cell clones obtained using HSP65 proved to recognize E . coli derived antigens contaminating the recombinant HSP65 rather than the mycobacterial antigen; these contaminants included the 60 kDa E . coli HSP, GroEL . The GroEL specific T cells did not respond to heat shocked human cells; this suggests (but does not prove) that they do not crossreact with human HSP60 . CONCLUSION . Synovial T cell recognition of antigens from enteric organisms associated with reactive arthritis is a common feature in pauciarticular JCA . Among the target antigens is the GroEL HSP, but T cells recognizing this antigen do not necessarily crossreact with the homologous human HSP60.

Infect Immun, 1993 Aug, 61(8), 3149 - 56
Endotoxin-mediated endothelial cell injury and activation: role of soluble CD14; Arditi M et al.; Vascular endothelial cell (EC) injury by lipopolysaccharides (LPS) plays a major role in the pathogenesis of gram-negative bacterial sepsis and endotoxic shock . The studies described here were performed to define further the molecular mechanisms involved in the EC responses to LPS . We showed that serum was required for LPS-mediated cytotoxicity for bovine brain microvessel, pulmonary, and aortic ECs and that anti-human CD14 antibodies completely blocked LPS-mediated cytotoxicity for ECs in the presence of human serum . The addition of a recombinant soluble form of human CD14 to serum-free medium restored the LPS-mediated cytotoxicity, whereas the addition of LPS binding protein (LBP), a serum protein that potentiates LPS-induced responses to monocytes, had no effect . A similar dependency on serum or recombinant soluble CD14 (under serum-free conditions) was observed for LPS-induced secretion of interleukin-6 by human umbilical vein ECs . These findings indicate that soluble CD14 is required for LPS-mediated EC responses independently of LPB, suggesting that serum soluble CD14 represents a naturally occurring agonist for EC responses to LPS.

J Immunol, 1993 Jul 15, 151(2), 916 - 21
IFN-gamma involvement in the severity of gram-negative infections in mice; Kohler J et al.; The role of IFN-gamma in the regulation of inflammation leading to gram-negative septic shock is still poorly understood . IFN-gamma blockade has been shown to improve the survival of animals challenged with i.v . bolus injections of LPS and gram-negative bacteria . We have investigated a model of focal Escherichia coli infection leading to peritonitis and septic shock . Mice were challenged i.p . with an inoculum near the LD50 . The addition of rIFN-gamma together with bacteria increased the mortality and the level of blood TNF-alpha and IL-6 . Conversely blockade of IFN-gamma with a neutralizing mAb significantly improved the survival of the mice . This beneficial effect was not associated with a stringent decrease in blood bacterial counts and TNF-alpha and IL-6 levels in survivors . In this model, the protective effect of anti-IFN-gamma mAb contrasted with the ineffectiveness of a neutralizing anti-TNF-alpha mAb . These findings suggest that overproduction of IFN-gamma might have a more detrimental role than overproduction of TNF-alpha during focal gram-negative infections.

Ann Emerg Med, 1993 Jul, 22(7), 1164 - 8
Effect of needle changing and intravenous cannula collection on blood culture contamination rates; Smart D et al.; STUDY OBJECTIVES: We tested the hypotheses that blood culture positivity and contamination rates were not increased by not changing needles between venipuncture and inoculation of blood culture bottles or by taking blood for culture by freshly inserted IV cannulae . DESIGN: A prospective study of blood cultures collected by venipuncture or IV cannulae taken from an emergency department population . Venipuncture samples were randomized into needle change (standard method) or no needle change before inoculation into blood culture bottles . PARTICIPANTS: Nine hundred forty patients requiring blood cultures after assessment in the ED . INTERVENTIONS: A standard disinfection procedure using 0.5% chlorhexidine in 70% alcohol was used . Blood was collected by venipuncture and inoculated with or without needle change . Blood collected by IV cannula was inoculated with a fresh needle applied to the collection syringe . MEASUREMENTS AND MAIN RESULTS: There was no statistically significant difference in contamination rates for blood collected by venipuncture with no needle change (6.4%) compared with needle change (4.2%, P > .30) . No significant difference in contamination rates was noted for blood taken by freshly inserted IV cannulae (4.3%) compared with venipuncture with needle change after sampling (4.2%, P > .90) . Some problems with randomization resulted in unequal numbers in the needle-change (286) versus no-needle-change (141) subgroups, and this may have introduced bias . A higher rate of pathogen growth was observed in blood taken by IV cannula (11.4%) compared with the standard method (6.3%) (P < .025) . A significantly greater rate of Gram-negative sepsis was noted in the IV cannula group (6.6%) compared with direct venipuncture with needle change (1.1%) and no needle change (4.2%, P < .01) . CONCLUSION: The results of this study do not support the practice of changing needles before inoculating blood samples into blood culture bottles . Collection of blood for culture through freshly inserted IV cannulae is associated with a low contamination rate and is an acceptable alternative to direct venipuncture . Sources of bias in this study suggest that further research is needed to determine the optimal technique for collecting blood cultures.

J Gen Microbiol, 1993 Jul, 139 ( Pt 7), 1531 - 5
Phylogeny and phenotypic characterization of the stalk-forming and iron-oxidizing bacterium Gallionella ferruginea; Hallbeck L et al.; The 16S rRNA gene of Gallionella ferruginea was amplified by polymerase chain reaction and sequenced by direct double-stranded sequencing . The phylogenetic analysis placed G . ferruginea in the beta-group of the Proteobacteria, with 90.0% similarity to Nitrosolobus multiformis and 88.6% to Rhodocyclus purpureus . The published phenotypic characteristics of G . ferruginea were compiled and supplemented with growth experiments using ferrous iron, thiosulphate and sulphide as electron donor, and nitrate as nitrogen source . G . ferruginea is a Gram-negative, curved bacterium with one polar flagellum . It grows auto- and mixotrophically with CO2, glucose, fructose and sucrose as carbon sources, ferrous iron as an electron donor and ammonium or nitrate as nitrogen sources . Two G . ferruginea specific oligonucleotide probes are suggested . An iron-oxidizing bacterium without stalk-forming ability, but with the same growth pattern as G . ferruginea, was identified as G . ferruginea by comparison of highly variable parts of the 16S rRNA gene . This indicates that the stalk is not essential for growth.

Curr Opin Oncol, 1993 Jul, 5(4), 625 - 32
Febrile neutropenia; Klastersky J; Severe neutropenia and its related infectious complications remain a permanent threat for patients receiving intensive chemotherapy, especially in the context of bone marrow transplantation . Chemoprophylaxis and use of colony-stimulating factors have altered the severity of the clinical picture in a favorable direction: neutropenia can be shortened, and gram-negative infection can be made less frequent; neither can be yet abolished . Early therapy, eg, empiric combination treatment, remains the cornerstone of our approach to febrile neutropenia; the actual choice of agents is probably less important and should be guided by local epidemiologic conditions . The concepts of empiric therapy also starts to be more widely accepted for the control of fungal and viral infections . Finally, it is fair to recognize that, at the other end of the spectrum of febrile neutropenia, conventional chemotherapy that results in only moderate and short neutropenia can usually be managed without much problem, namely with broad-spectrum monotherapy . Other possible simplified approaches should be investigated under controlled conditions and in patients selected on the basis of favorable prognostic factors.

Pharmacotherapy, 1993 Jul-Aug, 13(4), 330 - 9
The role of Helicobacter pylori in peptic ulcer disease; Partipilo ML et al.; The pathophysiology of peptic ulcer disease (PUD) is often described as an imbalance between aggressive factors such as acid and pepsin and alterations in the mucosal protective mechanisms . Helicobacter pylori is a gram-negative organism that has been identified as a potential causative agent in the pathogenesis of PUD . The exact mechanism by which it contributes to mucosal damage is unknown . It is thought that the organism may disrupt the protective mucous layer, allowing the underlying epithelium to be injured by gastric acid . Significant evidence indicates that H . pylori is a major etiologic factor in type B gastritis . Data confirming its etiologic role in duodenal ulcer (DU) disease is not conclusive; however, eradication of the organism is associated with a reduction in the recurrence of DU . Optimum therapy to eradicate H . pylori has not been established, although several multidrug regimens have been evaluated . Treatment of H . pylori infection should be reserved for individuals in whom conventional therapy for DU is unsuccessful and those whose ulcers relapse during maintenance therapy.

Appl Environ Microbiol, 1993 Jul, 59(7), 2077 - 81
Isolation and characterization of Selenomonas ruminantium strains capable of 2-deoxyribose utilization; Rasmussen MA; Microbes from ruminal contents of cattle were selectively enriched by using 2-deoxyribose (2DR) as a substrate for growth . Bacterial isolates growing on 2DR were gram-negative, curved, motile rods . The isolates grew on a broad range of substrates, including deoxyribose, glucose, ribose, mannitol, and lactate as well as ribonucleosides and deoxyribonucleosides . The strains also grew on rhamnose (6-deoxymannose) but not DNA . Organic acids produced from growth on hexoses and pentoses included acetate, propionate, lactate, and succinate . The isolates were identified as Selenomonas ruminantium subsp . lactilytica on the basis of morphology, substrate specificity, and other biochemical characteristics . Several characterized species of ruminal bacteria were also screened for growth on 2DR, with only one strain (S . ruminantium PC-18) found able to grow on 2DR . Ethanol was produced by 2DR when strains were grown on ribose or 2DR.

Radiographics, 1993 Jul, 13(4), 787 - 96
Imaging of complications of lung transplantation; O'Donovan PB; With the increasing number and improved survival of lung transplant recipients, radiologists should be aware of the imaging features of lung transplants and the associated complications . Reimplantation response, a noncardiogenic pulmonary edema seen 48 hours after transplantation that subsequently resolves, varies in appearance from a mild perihilar haze to a dense consolidation in the perihilar areas and lung bases . A late complication of omentopexy (used to prevent bronchial dehiscence) is herniation of abdominal contents through the diaphragmatic incision into the thorax . Extrabronchial air collections are a radiologic manifestation of anastomotic dehiscence . Stricture formation that compromises the bronchial lumina is usually visible with plain radiography, but computed tomography can aid in the evaluation . Acute rejection is evident radiographically as new or increasing pleural effusions, septal lines, subpleural edema, peribronchial cuffing, and air-space disease, without increase in cardiac size . Radiographic features of chronic rejection include both increased and diminished lung volumes, central and peripheral bronchiectasis, localized air-space disease, partial lobar atelectasis, thin linear irregular areas of increased opacity, pleural thickening, and diminished peripheral lung markings . Infection is frequently seen, especially gram-negative pneumonias, with fewer occurrences of cytomegalovirus infection, candidiasis, and invasive aspergillosis.

Parasitology, 1993 Jul, 107 ( Pt 1), 49 - 53
Endotoxins and the pathogenesis of Trypanosoma brucei brucei infection in mice; Alafiatayo RA et al.; The involvement of endotoxins in Trypanosoma brucei brucei infection in CD-1 mice was investigated by the Limulus amoebocyte lysate (LAL) test . At 7 days post-infection mean serum endotoxin level was elevated by 2.5 times (36.4 pg/ml cf . control 14.25 pg/ml, P < 0.001) and a similar increase was maintained throughout the infection (survival 28-35 days) . Purified disrupted parasites contained significant endotoxin activity (mean value 280 pg/mg protein) . The mouse infections were also associated with progressive Gram-negative bacteraemia (present in 4 out of 5 infected animals by day 28 p.i.) . The increased endotoxin levels may be due to parasite products, the products of intercurrent bacterial infections, other unidentified sources (e.g . from the gut), or a combination of these . It is concluded that the raised endotoxins may be important contributive factors in the pathogenesis of experimental murine trypanosomiasis.

Infect Control Hosp Epidemiol, 1993 Jul, 14(7), 383 - 9
A prospective randomized trial comparing manual and automated endoscope disinfection methods; Fraser VJ et al.; OBJECTIVE: To compare the efficacy of endoscope disinfection using automated and manual systems . DESIGN: Prospective randomized trial . SETTING: A 1,000-bed tertiary care referral center . METHODS: All endoscopes underwent a three-stage decontamination process including brushing and cleaning with water and detergent, manual or automated disinfection with 2% glutaraldehyde, and 70% alcohol rinse with forced air drying . Cultures were obtained from endoscopes from both groups before and after alcohol rinse and then after overnight storage . RESULTS: Cultures from 8/30 (27%) automated and 11/30 (37%) manually disinfected (P = 0.58) endoscopes grew gram-negative bacteria and/or nontuberculous mycobacteria before the alcohol rinse . After alcohol rinse, 3 (10%) of 30 automated and 8 (27%) of 30 manually disinfected endoscopes remained contaminated (P = 0.28) . Manually disinfected endoscopes were contaminated more frequently with coliform bacteria, whereas endoscopes undergoing automated disinfection were more frequently contaminated with nontuberculous mycobacteria, but the differences were not statistically significant . After alcohol rinse and forced air drying, there was no difference in contamination rates between freshly disinfected endoscopes and those stored overnight (7/30 (23%) versus 4/30 (13%), P = 0.50) . Colonoscopes and duodenoscopes were contaminated more often than gastroscopes (P = 0.00001) . CONCLUSION: The persistent endoscope contamination after manual and automated disinfection indicates the importance of developing more reliable and effective disinfection methods.

J Clin Microbiol, 1993 Jul, 31(7), 1936 - 9
Use of immunoelectron microscopy to demonstrate Francisella tularensis; Geisbert TW et al.; Three immunoelectron microscopy (IEM) methods were employed to show laboratory-cultivated Francisella tularensis . By the IEM assays, F . tularensis was distinguished from four antigenically distinct gram-negative bacteria . IEM should be a valuable tool for confirming presumptive isolates of F . tularensis and may potentially be useful for demonstrating other medically important bacteria.

J Leukoc Biol, 1993 Jul, 54(1), 81 - 8
Lipopolysaccharide-induced suppression of erythrocyte binding and phagocytosis via Fc gamma RI, Fc gamma RII, Fc gamma RIII, and CR3 receptors in murine macrophages; Sundaram R et al.; In this study we have investigated the ability of lipopolysaccharide (LPS) to suppress binding and phagocytosis of erythrocytes via various receptors on mouse macrophages . Thioglycollate-elicited peritoneal macrophages were treated in vitro with LPS and the ability to bind and phagocytose radiolabeled sheep red blood cells was determined . We show that LPS can directly suppress phagocytosis of immunoglobulin G-opsonized and nonopsonized sheep red blood cells (SRBCs) by inflammatory macrophages . Suppression was dose dependent and was observed after 4 h of exposure . This effect lasted for at least 24 h following the removal of LPS . LPS suppressed the binding, rate, and absolute level of phagocytosis via Fc receptors . Phagocytosis via all Fc receptors (Fc gamma RI, Fc gamma RII, and Fc gamma RIII) was suppressed by LPS . Furthermore, suppression was not limited to Fc receptor-mediated phagocytosis because binding and uptake of C3bi-opsonized SRBCs to CR3 receptors was also decreased following LPS treatment . LPS did not exert its effects via the production of interleukin-1 (IL-1), IL-6, tumor necrosis factor alpha, or interferon alpha/beta, because antibodies to these cytokines did not abrogate the effect . The ability of LPS to suppress binding and phagocytosis of microorganisms may contribute to the toxic effects of LPS during gram-negative sepsis by preventing or delaying elimination of bacteria by host macrophages.

Ann Surg, 1993 Jul, 218(1), 79 - 90
Comparison of peripheral blood leukocyte kinetics after live Escherichia coli, endotoxin, or interleukin-1 alpha administration . Studies using a novel interleukin-1 receptor antagonist; Hawes AS et al.; OBJECTIVE: This study was undertaken to evaluate whether hematologic and immunologic effects observed after bacteremia and endotoxemia in the host could be replicated by administration of recombinant human interleukin-1 alpha (IL-1 alpha) in a primate model . Furthermore, to determine whether endogenously produced interleukin-1 (IL-1) contributes to the changes observed during endotoxemia or gram-negative septic shock, a specific IL-1 receptor antagonist (IL-1 ra) was administered . SUMMARY BACKGROUND DATA: The lipopolysaccharide (LPS) component of the outer membrane of gram-negative bacteria initiates a constellation of metabolic and immunologic host responses . IL-1, a macrophage-derived cytokine, acts as a key mediator in the host response to infection and inflammation . METHODS: Baboons were randomly assigned to receive either recombinant human IL-1 alpha, LPS, or live Escherichia coli both with or without concomitant administration of IL-1ra . Blood was collected hourly and analyzed using flow cytometric techniques . RESULTS: Both endotoxemia and live E . coli bacteremia induced an acute granulocytopenia; however, the granulocytopenia gradually resolved in the endotoxemic group, but was sustained in the bacteremic group . An early lymphopenia and monocytopenia was elicited by LPS or E . coli and persisted throughout the experiment . Recombinant human IL-1 alpha induced the following: (1) an early, transient decline in granulocytes followed by a sustained granulocytosis; (2) a lymphopenia; and (3) a transient monocytopenia followed by a gradual return to baseline . Although IL-1ra had no effect on leukocyte kinetics with either live E . coli or LPS, the IL-1ra significantly abrogated the monocytopenia seen with recombinant human IL-1 alpha administration alone . CONCLUSIONS: These results suggest that administration of recombinant human IL-1 alpha can replicate some of the characteristic patterns of hematologic change associated with bacteremia and endotoxemia . However, an endogenous IL-1 response is not required for these changes to occur . Rather, the data suggest that other inflammatory mediators induced by endotoxemia or gram-negative bacteremia, such as tumor necrosis factor-alpha (TNF alpha), may be involved.

Hosp Pract (Off Ed), 1993 Jul, 28 Suppl 2, 36 - 9; discussion 60-1
Outpatient parenteral antibiotic therapy . Management of serious infections . Part II: Amenable infections and models for delivery . Osteomyelitis; Tice AD; Osteomyelitis is one of the most common and well-established indications for outpatient parenteral antibiotic therapy . Because patients are usually otherwise healthy and therapy is prolonged (four to six weeks), this infection is especially suited to outpatient management . While most gram-negative infections in adults can be treated with an oral quinolone, others usually require IV therapy.

Hepatology, 1993 Jul, 18(1), 173 - 8
Gram-negative bacterial lipopolysaccharide impairs hyaluronan clearance in vivo and its uptake by the isolated, perfused rat liver; Deaciuc IV et al.; The purpose of this investigation was to examine the effect of gram-negative bacterial lipopolysaccharide on hyaluronan concentration in blood plasma, hyaluronan removal from the blood and hyaluronan uptake by isolated, perfused rat liver . Intravenous administration of Escherichia coli lipopolysaccharide to rats markedly increased plasma hyaluronan concentration in a dose-dependent manner . One day after lipopolysaccharide challenge (0.1 or 1.0 mg per 100 gm body wt), plasma hyaluronan levels were 570.7 +/- 66.8 ng x ml-1 and 1,951.0 +/- 120.3 ng x ml-1, respectively, as compared with 94.2 +/- 12.2 ng x ml-1 in the time-matched control animals . Removal of intravenously injected hyaluronan (30 micrograms per 100 gm body wt) was suppressed 32% by lipopolysaccharide administration (100 micrograms per 100 gm body wt) . At the same dose, lipopolysaccharide induced a severe inhibition (60% to 80%) of hyaluronan uptake by perfused livers isolated 3 or 24 hr after lipopolysaccharide administration . The inhibitory effect of lipopolysaccharide on hyaluronan uptake by the isolated, perfused liver was not abolished by pretreatment with either antibodies to tumor necrosis factor-alpha IgG or indomethacin, an inhibitor of the cyclooxygenase pathway . Continuous intravenous infusion of recombinant murine tumor necrosis factor-alpha for 18 to 20 hr did not affect plasma hyaluronan concentration . These data suggest that neither tumor necrosis factor-alpha, an early cytokine induced by lipopolysaccharide, nor prostaglandins are involved in the mechanism of lipopolysaccharide-induced inhibition of hyaluronan uptake by the perfused rat liver.(ABSTRACT TRUNCATED AT 250 WORDS)

J Bacteriol, 1993 Jul, 175(13), 4225 - 34
Molecular cloning and sequence analysis of the gene encoding OmpL1, a transmembrane outer membrane protein of pathogenic Leptospira spp; Haake DA et al.; Pathogenic Leptospira spp . are spirochetes that have a low transmembrane outer membrane protein content relative to that of enteric gram-negative bacteria . In a previous study we identified a 31-kDa surface protein that was present in strains of Leptospira alstoni in amounts which correlated with the outer membrane particle density observed by freeze fracture electron microscopy (D . A . Haake, E . M . Walker, D . R . Blanco, C . A . Bolin, J . N . Miller, and M . A . Lovett, Infect . Immun . 59:1131-1140, 1991) . The N-terminal amino acid sequence was used to design a pair of oligonucleotides which were utilized to screen a lambda ZAP II library containing EcoRI fragments of L . alstoni DNA . A 2.5-kb DNA fragment which contained the entire structural ompL1 gene was identified . The structural gene deduced from the sequence of this DNA fragment would encode a 320-amino-acid polypeptide with a 24-amino-acid leader peptide and a leader peptidase I cleavage site . Processing of OmpL1 results in a mature protein with a predicted molecular mass of 31,113 Da . Secondary-structure prediction identified repeated stretches of amphipathic beta-sheets typical of outer membrane protein membrane-spanning sequences . A topological model of OmpL1 containing 10 transmembrane segments is suggested . A recombinant OmpL1 fusion protein was expressed in Escherichia coli in order to immunize rabbits with the purified protein . Upon Triton X-114 extraction of L . alstoni and phase separation, anti-OmpL1 antiserum recognized a single band on immunoblots of the hydrophobic detergent fraction which was not present in the hydrophilic aqueous fraction . Immunoelectron microscopy with anti-OmpL1 antiserum demonstrates binding to the surface of intact L . alstoni . DNA hybridization studies indicate that the ompL1 gene is present in a single copy in all pathogenic Leptospira species that have been tested and is absent in nonpathogenic Leptospira species . OmpL1 may be the first spirochetal transmembrane outer membrane protein for which the structural gene has been cloned and sequenced.

Am J Pathol, 1993 Jul, 143(1), 76 - 84
Murine tissue factor gene expression in vivo . Tissue and cell specificity and regulation by lipopolysaccharide; Mackman N et al.; Regulation of tissue factor (TF) gene expression was studied in vivo employing a murine model system . In untreated mice, TF mRNA was detected in brain, lung, kidney, and heart by Northern blot analysis . After administration of lipopolysaccharide, steady-state levels of TF mRNA were unchanged in brain, decreased in heart, and increased in both kidney and lung . In the brain, Bergmann glia within the Purkinje cell layer of the cerebellum and neuroglia within the cerebral cortex expressed TF mRNA by in situ hybridization . Epidermal cells of the skin and tongue also expressed TF mRNA . At present, we have not identified the cell type(s) in the kidney and lung responsible for increased TF gene expression . These results demonstrate tissue- and cell-specific TF gene expression in vivo . Lipopolysaccharide-mediated increases in TF expression in the kidney and lung may promote fibrin deposition in these organs during Gram-negative sepsis.

West Afr J Med, 1993 Jul-Sep, 12(3), 180 - 4
Acute encephalopathy, hypertension and gram negative sepsis in sickle cell disease; Akpede GO et al.; The case histories of two patients with sickle cell disease and gram negative sepsis complicated by encephalopathy and hypertension is presented . The first patient had 2 episodes of "hypertensive encephalopathy" before control of her blood pressure was achieved while the second patient had only one . The occurrence, though apparently rare, can have serious implications . Possible mechanisms are discussed and the need to monitor the blood pressure of children with sickle cell disease is stressed.

ASAIO J, 1993 Jul-Sep, 39(3), M773 - 7
Production of platelet activating factor by human neutrophils after backfiltration of endotoxin contaminated dialysate; David S et al.; Lipopolysaccharide (LPS) from gram negative bacteria has been shown to prime human polymorphonuclear neutrophil (PMN) production of platelet activating factor (PAF) . PAF is a lipid mediator of inflammation and endotoxic shock and is also involved in leukocyte activation occurring during hemodialysis; PAF induces leukopenia, degranulation of lysosomal granules, and adherence to hemodialysis membranes . Transmembrane passage of LPS has also been shown to occur . To evaluate the relevance of transmembrane passage of LPS on the priming of PMN derived production of PAF, we designed in vitro studies using an experimental circuit equipped with different membranes (Cuprophan, polysulfone, polymethylmethacrylate, polyamide) and recirculation of purified human PMNs . At different time intervals, PMNs were stimulated with FMLP (10 microM) after back-filtration of sterile and LPS contaminated dialysate . The results of these studies suggest that priming of PMN derived production of PAF was related to the percent of backfiltered LPS, and they emphasize the need for careful assessment of microbiologic quality to improve biocompatibility.

Cytokine, 1993 Jul, 5(4), 348 - 53
Antilipid a monoclonal antibody HA-1A: immune complex clearance of endotoxin reduces TNF-alpha, IL-1 beta and IL-6 production; Katsikis P et al.; HA-1A is a human monoclonal IgM antibody which recognizes the lipid A component of lipopolysaccharide (LPS) . This antibody has reduced mortality in the septic shock syndrome resulting from Gram negative bacteria, in which many of the manifestations are considered to be due to cellular activation and secretion of cytokines, most notably TNF-alpha . However HA-1A does not directly neutralize LPS effectively in vitro, and studies reported to date have not defined its mechanism of action . Here we demonstrate that HA-1A, which in the presence of complement promotes immune adherence, may inhibit LPS action by facilitating its sequestration on red blood cells and clearance to an extent that cytokine production is reduced . Incubation of LPS at clinically significant (pg/ml) does with HA-1A at therapeutic levels (e.g . 10 micrograms/ml) and complement resulted in LPS association with erythrocyte CR1 receptors . This reduced the ability of the residual, free LPS by 50-70% to induce the secretion of TNF-alpha, IL-1 beta and IL-6 from normal blood mononuclear cells . This mechanism is likely to be operative in vivo, and could account for the protective effect of HA-1A, and its reduction of TNF-alpha production in vivo.

Infection, 1993 Jul-Aug, 21(4), 245 - 7
The effect of clindamycin gel insert in periodontal pockets, as observed on smears and cultures; Sauvetre E et al.; This study is aimed at the evaluation of a 1% clindamycin hydrochloride containing gel on the microbial flora of periodontal pockets deeper than 5 mm . In order to achieve that purpose, 20 patients with pocketing in the premolar-molar regions were selected . Active and placebo gel were inserted once during the first 2 weeks of this experimental study . Microbial samplings were performed 1, 2, 4 and 12 weeks after the experiment started . The samples were submitted to microscopic examination and also to culture . Changes in the microbial content of the periodontal pockets treated by subgingival scaling and clindamycin 1% gel were significant, compared with those obtained with subgingival scaling and placebo gel, particularly with respect to anaerobic black-pigmented bacteria and the motile gram-negative flora . However, after 3 months, most of the treated cases were recolonized by the same initial species, though never at pre-clindamycin levels . In the light of this study, it will be concluded that the use of a small amount of clindamycin hydrochloride inserted into a periodontal pocket, once a week for 2 weeks as a complement to periodontal subgingival scaling, is beneficial in the treatment of adult periodontitis, by eliminating more effectively the microbial pocket colonization.

Eur J Biochem, 1993 Jun 15, 214(3), 685 - 93
Outer-membrane porins from gram-negative bacteria stimulate platelet-activating-factor biosynthesis by cultured human endothelial cells; Tufano MA et al.; Porins are a family of hydrophobic proteins located in the outer membrane of the cell wall in Gram-negative bacteria . The effect of porins on the biosynthesis of platelet-activating factor (PAF) by cultured human umbilical-cord-vein-derived endothelial cells (HUVEC) was investigated . The results demonstrate that porins were able to induce a dose-dependent synthesis of PAF in HUVEC . PAF, synthesized after stimulation with porins, was mainly cell associated and the synthesis peaked at 15 min, decreasing rapidly thereafter . Experiments with radiolabeled precursors demonstrated that PAF, a 1-O-alkyl-2-acetyl-sn-glyceryl-3-phosphorylcholine, was synthesized via the remodeling pathway involving the acetylation of 1-O-alkyl-2-lyso-sn-glyceryl-3-phosphorylcholine (2-lysoPAF) generated from 1-O-alkyl-2-acyl-sn-glyceryl-3-phosphorylcholine by phospholipase-A2 activity . The activation of phospholipase A2 in HUVEC stimulated by porins was detected by observing the mobilization of {14C}arachidonic acid . In addition, the activity of acetyl-CoA:1-alkyl-sn-glycero-3-phosphorylcholine 2-O-acetyltransferase was transiently increased in porin-stimulated HUVEC and, after incubation with {3H}CoASAc or {3H}acetate, the {3H}acetyl group was incorporated into newly synthesized PAF . Porins, by forming transmembrane channels, induced a sustained influx of extracellular 45Ca2+ into the cytosol . The activation of PAF synthesis by porins depended on this influx rather than on intracellular calcium mobilization, since PAF synthesis did not occur in the absence of extracellular Ca2+.

J Immunol, 1993 Jun 15, 150(12), 5556 - 65
Neutrophil CD14: biochemical properties and role in the secretion of tumor necrosis factor-alpha in response to lipopolysaccharide; Haziot A et al.; CD14 is a myeloid cell differentiation Ag expressed primarily by monocytes and macrophages . CD14 has recently been shown to function as a receptor for a complex of LPS and LPS binding protein (LBP), an acute phase serum protein also present in normal serum in trace amounts . In the presence of LBP, LPS strongly activates monocytes via CD14 as measured by TNF secretion . This pathway of monocyte activation is thought to be a major contributor to the symptoms of endotoxin shock . Another major cell type involved in the response to Gram-negative infection is the neutrophil . Recent studies have shown that neutrophils also express CD14 and suggest that they can respond to LPS through a similar pathway . However, the biochemical nature of neutrophil CD14 has not previously been described . In this report, we have analyzed several biochemical characteristics of neutrophil CD14 . We show that CD14 is actively synthesized by neutrophils as a glycosylphosphatidyl-inositol-anchored protein, indistinguishable in size from monocyte CD14 . Furthermore, neutrophils, like monocytes, shed a smaller soluble form of CD14 into culture supernatants . In addition, like monocytes, neutrophils respond to LPS/LBP complexes via CD14 by releasing TNF-alpha . The described properties and function of neutrophil CD14 suggest that it may directly participate in the acute inflammatory response and in endotoxin shock.

Schweiz Med Wochenschr, 1993 Jun 5, 123(22), 1165 - 8
{Cervical abscess caused by Capnocytophaga ochracea}; Henzen C et al.; Capnocytophaga is a gram-negative capnophilic bacterium which is part of the normal oral flora of humans (C . ochracea, C . gingivalis, C . sputigena) and mammals such as canines, cats, and rodents (C . animorsus and C . cynodegmi) . Its role in the pathogenesis of periodontal disease is not well defined, and normally it represents an opportunistic germ of low pathogenicity . Threatening and fulminant infections have been observed in immunodeficient patients, and lately in immunocompetent hosts . We describe an otherwise healthy woman who developed a cervical abscess due to C . ochracea . Recurrent aphthous lesions are suspected to be the port of entrance for the germs . Bacteriological, clinical, epidemiological, and therapeutic aspects of Capnocytophaga infection are discussed.

Transplantation, 1993 Jun, 55(6), 1250 - 6
Lectin-dependent cell-mediated cytotoxicity and natural killer function in rejecting and infected lung allografts; Nguyen DM et al.; Differentiation between rejection and infection of lung allografts remains difficult . The effects of these two pathologic entities on the cytolytic activity of bronchoalveolar lavage (BAL) and PBL were investigated . Left lung allotransplantation was performed on 16 mongrel dogs of which 12 were available for complete studies . All animals received CA, AZA, and PRED for 2 weeks . Four grafts developed left lower lobe Gram negative pneumonia . The eight remaining recipients progressed gradually to severe rejection after acute reduction of immunosuppression . Cytolytic activity of blood and left lung BAL lymphocytes was quantitated by the natural killer (NK) and lectin-dependent cell-mediated cytotoxicity (LDCMC) assays . Two additional groups serving as controls were either given a 10-day course of immunosuppressants or had right lower lobe pneumonia induced by transbronchial inoculation of gram negative bacteria . Immunosuppressed control animals showed significant depression of PBL and BAL lymphocyte LDCMC and NK activity . Similarly, BAL lymphocytes expressed very low LDCMC in normal allografts (2.8 +/- 0.8%) . Once rejection developed and progressed, LDCMC became significantly higher (15.6 +/- 2.2 and 52.7 +/- 2.8% in mild and severe rejection, respectively) . There was no detectable NK activity in rejecting lung allografts . BAL lymphocytes from infected allografts, on the other hand, showed an elevation of both NK and LDCMC activity (9.1 +/- 1.1 and 14.6 +/- 1.0%, respectively) . Similarly, bacterial pneumonia in control animals manifested an increase in NK and LDCMC activity in lung and blood . PBL lymphocytes of lung allograft recipients, however, had increased NK and LDCMC activity in both rejection and infection . LDCMC/NK activity ratio (LM/NK index) of lung lymphocytes was significantly higher in rejecting allografts (11.2 +/- 1.0 and 12.4 +/- 1.6 for mild and severe rejection, respectively) than in infected ones (1.2 +/- 0.3, P < 0.0001) . It appears, from this study, that rejection of the lung allograft results in alterations in BAL lymphocyte phenotypes and functions that differ from those associated with bacterial infection . Such differences may be useful in distinguishing episodes of acute allograft rejection from bacterial infection.

J Clin Invest, 1993 Jun, 91(6), 2850 - 60
Tissue factor pathway inhibitor reduces mortality from Escherichia coli septic shock; Creasey AA et al.; This study was designed to test the hypothesis that tissue factor pathway inhibitor (TFPI) plays a significant role in vivo in regulating coagulation that results from exposure of blood to tissue factor after vascular injury as in the case of gram negative sepsis . Highly purified recombinant TFPI (6 mg/kg) was administered either 30 min or 4 h after the start of a lethal intravenous Escherichia coli infusion in baboons . Early posttreatment of TFPI resulted in (a) permanent seven-day survivors (5/5) with significant improvement in quality of life, while the mean survival time for the controls (5/5) was 39.9 h (no survivors); and (b) significant attenuations of the coagulation response and various measures of cell injury, with significant reductions in pathology observed in E . coli sepsis target organs, including kidneys, adrenals, and lungs . TFPI administration did not affect the reduction in mean systemic arterial pressure, the increases in respiration and heart rate, or temperature changes associated with the bacterial infusion . TFPI treated E . coli infected baboons had significantly lower IL-6 levels than their phosphate buffered saline-treated controls, however tumor necrosis factor levels were similarly elevated in both groups . In contrast to the earlier 30-min treatment, the administration of TFPI at 4 h, i.e., 240 min, after the start of bacterial infusion resulted in prolongation of survival time, with 40% survival rate (2/5) and some attenuation of the coagulopathic response, especially in animals in which fibrinogen levels were above 10% of normal at the time of TFPI administration . Results provide evidence for the significance of tissue factor and tissue factor pathway inhibitor in bacterial sepsis, and suggest a role for blood coagulation in the regulation of the inflammatory response.

Biophys J, 1993 Jun, 64(6), 1691 - 700
Deformations in the cytoplasmic membrane of Escherichia coli direct the synthesis of peptidoglycan . The hernia model; Norris V et al.; To explain the growth of the Gram-negative envelope and in particular how it could be strengthened where it is weakest, we propose in the hernia model that local weakening of the peptidoglycan sacculus allows turgor pressure to cause the envelope to bulge outwards in a hernia; the consequent local alteration in the radius of curvature of the cytoplasmic membrane causes local alterations in phospholipid structure and composition that determine both the synthesis and hydrolysis of peptidoglycan . This proposal is supported by evidence that phospholipid composition determines the activity of phospho-N-acetylmuramic acid pentapeptide translocase, UDP-N-acetylglucosamine:N-acetylmuramic acid-(pentapeptide)-P-P-bactoprenyl-N-acetylglucosamine transferase, bactoprenyl phosphate phosphokinase, and N-acetylmuramyl-L-alanine amidase . We also propose that the shape of Escherichia coli is maintained by contractile proteins acting at the hernia . Given the universal importance of membranes, these proposals have implications for the determination of shape in eukaryotic cells.

J Clin Pharmacol, 1993 Jun, 33(6), 562 - 7
Determination of gentamicin pharmacokinetics by bioelectrical impedance in critically ill adults; Zarowitz BJ et al.; This investigation compares the accuracy of calculating gentamicin pharmacokinetic parameters by a noninvasive body composition technique (bioelectrical impedance analysis; BIA) with an empiric method, against the two-point method as the criterion standard . A prospective concurrent open label design was used . The 32 medical and surgical intensive care unit beds at Henry Ford Hospital, a not-for-profit, university-affiliated teaching hospital, served as the setting . Twenty critical ill adults, Therapeutic Index Scoring System (TISS) = 4, who required gentamicin as part of their normal course of therapy for gram-negative bacillary infections, were evaluated . Gentamicin Vd and k were calculated by three methods . After measurement of body composition parameters by BIA, previously derived gentamicin dosing equations were used to predict gentamicin volume of distribution (Vd) and elimination rate constant (k) (BIA method) . Empiric estimates of these parameters (Vd = 0.3L/kg and k derived from creatinine clearance) were compared with the BIA parameters against a criterion standard Vd and k determined from a two-point sampling of gentamicin serum concentrations . Measurements of BIA parameters and gentamicin serum concentrations were made in duplicate with coefficients of variation, < or = 2% and < or = 3%, respectively . The BIA and empiric methods produced resultant pharmacokinetic parameters (Vd and k) not different than those measured by the two-point method . There were no statistically significant differences in mean error (bias), or mean squared error (precision) for both Vd and k assessed by the empiric or BIA methods.(ABSTRACT TRUNCATED AT 250 WORDS)

Anasthesiol Intensivmed Notfallmed Schmerzther, 1993 Jun, 28(4), 258 - 60
{Septic shock with acute abdomen in idiopathic hemochromatosis}; Sydow M et al.; The case of a patient with abdominal crisis and shock without any discernible origin who died 36 hours after hospital admission despite maximal therapy is described . Gram-negative sepsis, peritonitis and haemochromatosis with hepatic siderosis was the post-mortem diagnosis . We consider spontaneous peritonitis arising from translocation of normal intestinal flora (E . coli) through the intact wall of the gut combined with the impaired ability of the reticulo-endothelial system to remove endotoxin to be the causative factors . It is unknown whether the adrenal insufficiency due to siderophilic adrenal hypophysis and adrenal glands contributed to the fulminant course of the disease . Undiagnosed liver cirrhosis and especially haemochromatosis should therefore be included in the differential diagnostic considerations in patients presenting with these symptoms, and in whom no obvious cause for a septic focus can be found.

Cesk Epidemiol Mikrobiol Imunol, 1993 Jun, 42(2), 87 - 92
{Lyme borreliosis: review of present knowledge}; Schwarzova K; The author reviews hitherto assembled knowledge on a bacterial disease, Lyme borreliosis transmitted by ticks . Initial information on Lyme borreliosis appeared at the beginning of the 20th century . In Czechoslovakia attention to the disease was paid since cca 1960 . The infection occurs as a rule in the summer months during the period when ticks are parasitic and at that time the causal agent of the disease is transmitted to hosts . Information on the prevalence and incidence of Lyme borreliosis in Europe is not complete and so far we do not possess a standardized diagnostic method for assessment of circulating antibodies in the patients' serum and cerebrospinal fluid . The infectious disease is caused by the gram-negative spirochete Borrelia burgdorferi . The Borrelia cell has a similar morphological structure as cells of other gram-negative bacteria . Chemical analysis of the external membrane of B . burgdorferi revealed the presence of 46% proteins, 51% lipids and 3% carbohydrates . The typical shape of borrelias indicates marked ondulation of 8-14 periplasmatic flagellae along the cell body . Borrelias can be cultivated in vitro in modified Barbour-Stoenner-Kelly medium at an optimal temperature of 30-37 degrees C . The change of morphology during cultivation is typical for B . burgdorferi . Clinically Lyme borreliosis is manifested in two stages . A typical manifestation of the early stage is a skin lesion--erythema migrans . The later stage is characterized above all by relapsing arthritis, CNS infection and chronic acrodermatitis chronica atrophicans . The disease is treated by administration of a number of antibiotics either by the oral route or by injection.(ABSTRACT TRUNCATED AT 250 WORDS)

Changgeng Yi Xue Za Zhi, 1993 Jun, 16(2), 93 - 8
Ultrasonography and fine needle aspiration cytology of acute suppurative thyroiditis; Lin JD et al.; Although acute suppurative thyroiditis is a rare disorder, from January . 1985 to December 1991, we observed 11 cases of acute suppurative thyroiditis in our hospital . All patients underwent thyroid ultrasonography and fine needle aspiration biopsy and cytologic evaluation of the specimens . In eight cases, pus cultures were positive, four of them grew Gram negative bacteria . Ultrasonographic finding showed either local or diffuse low to a very low echo density on both lobes . However, a cystic lesion was also found . Polymorphonuclear cells were the main elements in the smears from the thyroid aspirates . In conclusion, thyroid ultrasound with fine needle aspiration and cytology can assist in confirming the early diagnosis of acute suppurative thyroiditis.

Clin Infect Dis, 1993 Jun, 16 Suppl 4, S203 - 10
Bacterial mediators in periodontal disease; Loesche WJ; Periodontal disease is the general description given to the inflammatory response of the gingiva and underlying connective tissue to bacterial accumulations (dental plaque) on the teeth . A limited number of cultivable species are usually associated with periodontal disease . The majority of putative periodontal pathogens are gram-negative anaerobic rods . Some of the characteristics of Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, and Treponema denticola will be discussed, given their prominence in the literature . These organisms share the ability to penetrate the gingival epithelium, such that their endotoxins, immunologically active compounds, and cytotoxic enzymes and molecules are presented directly to the host's inflammatory cells . This ability may be what distinguishes these gram-negative species from the plethora of other gram-negative species that inhabit the subgingival plaque . In addition, these organisms tend to be selected for in disease-associated plaques, suggesting that their nutritional needs are met when the gingival crevicular fluid contains a variety of inflammatory mediators and products of tissue breakdown . A . actinomycetemcomitans produces a leukotoxin, and the immunologic response of the host to this antigen may explain the unique pattern of tooth involvement in localized juvenile periodontitis . Both P . gingivalis and T . denticola have a trypsin-like enzyme that could be a virulence factor, primarily because this enzyme(s) may allow these organisms to grow in the presence of the inflammatory response of the host.

Tokai J Exp Clin Med, 1993 Jun, 18(1-2), 71 - 80
Results and prognostic factors in the radical sinus operation for chronic sinusitis; Iida M et al.; Radical surgical procedures of the maxillary sinus with or without involvement of the ethmoid, sphenoid and frontal sinuses were performed in 382 adult patients in the Department of Otorhinolaryngology . Tokai University Hospital in the 8-year period from 1984 to 1991 . Of 382 patients, the postoperative symptomatology was studied in 189 patients by mcans of questionnaires . The postoperative results were finally analyzed in 63 patients whose subjective symptoms and objective findings were completely recorded . The follow-up period was over 3 months and subjective postoperative improvement was achieved in 83% and objective postoperative improvement in 75%; the overall improvement rate was 73% . In an analysis of results of the radical surgical operation, it appeared that poor results were closely related to the following factors: (1) maxillary sinus mucosa with cellular infiltration or edematous changes; (2) multiplicity of bacterial species including Gram-negative rods isolated from maxillary sinus secretion; (3) poor mucociliary clearance function; and (4) unsatisfactory postoperative treatment.

J Immunol, 1993 Jun 1, 150(11), 5033 - 40
Differences in cytokine response and induction of nitric oxide synthase in endotoxin-resistant and endotoxin-sensitive mice after intravenous gram-negative infection; Evans TJ et al.; Previous reports have suggested that the endotoxin-resistant C3H/HeJ strain of mouse is more susceptible to infection than is the endotoxin-sensitive parent strain, C3H/HeN, although they have never been compared in an i.v . model of sepsis . We therefore have used these mouse strains in an i.v . model of Gram-negative sepsis to compare their sensitivities to infection, their cytokine responses, and the levels of induction of the enzyme nitric oxide synthase assayed in their livers . By using i.v . infection with Escherichia coli we have found that both strains are approximately equally sensitive to this organism, despite the C3H/HeJ mice having a markedly attenuated TNF-alpha response . IFN-gamma levels after infection were identical in the two strains; the levels of nitric oxide synthase induced in their livers were about fourfold greater in the C3H/HeJ mice . This difference could not be explained by differences in bacterial load . These experiments suggest that factors other than TNF-alpha are important in determining outcome from Gram-negative sepsis and that TNF-alpha is not a major factor in the induction of hepatic nitric oxide synthase after infection in vivo.

FEBS Lett, 1993 May 17, 322(3), 231 - 4
Pentoxifylline inhibits the expression of tissue factor mRNA in endotoxin-activated human monocytes; Ollivier V et al.; Tissue factor (TF) is a transmembrane glycoprotein which, in association with factor VII(a), is the main activator of coagulation . In illnesses such as Gram-negative endotoxemia, circulating monocytes synthesize and express substantial TF activity, resulting in extensive disseminated intravascular coagulation . We investigated the way in which TF is suppressed by pentoxifylline (PTX), and found that PTX down-regulates immunologic TF expression and specific mRNA production in response to LPS . Since TF mRNA stability is not altered, this effect appears to take place at the transcriptional level.

Gene, 1993 May 15, 127(1), 127 - 31
Sequence and transcriptional analysis of the nourseothricin acetyltransferase-encoding gene nat1 from Streptomyces noursei; Krugel H et al.; We have determined the nucleotide (nt) sequence of nat1, a gene encoding nourseothricin (Nc) acetyltransferase (AT) from Streptomyces noursei, and its transcriptional start point (tsp) . The nt sequence upstream from the coding region is completely different from that of the stat gene (encoding streptothricin AT) from Streptomyces lavendulae {S . Horinouchi, K . Furuya, M . Nishiyama, H . Suzuki and T . Beppu, J . Bacteriol . 169 (1987) 1929-1937}, even though the nt sequences of the two genes and the deduced amino acid (aa) sequences of the two enzymes show a high degree of similarity . Another stat gene, derived from a Gram-negative plasmid, showed only deduced aa similarity, but not nt sequence similarity, to the above two . A database search for related aa sequences did not reveal any clear-cut homologies to other types of protein . A multiple aa sequence alignment of several ATs is presented.

JAMA, 1993 May 5, 269(17), 2221 - 7
A controlled trial of HA-1A in a canine model of gram-negative septic shock; Quezado ZM et al.; OBJECTIVE--To investigate the therapeutic efficacy and microbiological and physiological effects of a human IgM monoclonal antibody (HA-1A) directed against the lipid A component of endotoxin in a canine model of sepsis that simulates the cardiovascular abnormalities of human septic shock . DESIGN--Blinded, placebo-controlled 28-day trial . INTERVENTIONS--Purpose-bred beagles were implanted with an intraperitoneal clot infected with Escherichia coli O111:B4 . At clot placement, animals received HA-1A (10 mg.kg-1), control human IgM antibody (10 mg.kg-1), or control human serum albumin intravenously . All animals were given antibiotic and fluid therapy . MEASURES--Survival and microbiological and physiological events . RESULTS--Only two (15%) of 13 animals in the HA-1A group, compared with eight (57%) of 14 control animals (combined control human IgM antibody and control human serum albumin groups) (P = .05), survived 28 days . At 24 hours, the HA-1A group had lower mean arterial pressure (P = .04) and cardiac index (P = .004) and higher lactate levels (P = .05) compared with the combined-controls group . In addition, these parameters in the HA-1A group were significantly more predictive of death . The HA-1A and combined-controls groups had similar significant increases in the level of endotoxemia and bacteremia . Studies of toxic effects showed no harmful effects of control human IgM antibody in infected animals or HA-1A in non-infected animals . CONCLUSION--In a canine model of E coli sepsis, HA-1A did not alter levels of bacteremia or endotoxemia and actually decreased survival . If these data are relevant to human septic shock, HA-1A therapy should be limited until the conditions under which this monoclonal antibody has beneficial or deleterious effects are more completely defined.

Biochemistry, 1993 May 4, 32(17), 4579 - 86
Effect of pH on solubility and ionic state of lipopolysaccharide obtained from the deep rough mutant of Escherichia coli; Din ZZ et al.; The dissociation of the highly aggregated form of lipopolysaccharide (LPS) from Gram-negative bacteria to the monomeric (or soluble) form is though to be the initial step in the activation of responding cells (macrophages, B-cells, neutrophils, monocytes, and endothelial cells) by LPS . This process is presently not adequately understood . Using the equilibrium dialysis apparatus and a highly purified and well-characterized radiolabeled deep rough chemotype LPS ({14C}ReLPS) from Escherichia coli D31m4, we have examined the effect of pH on its solubility (CT) and ionic states in aqueous media . The solubility range of {14C}ReLPS suspended in 50 mM Tris-HCl-100 mM KCl buffer (or 50 mM MES-100 mM KCl buffer at pH 6.5) was determined to be from (2.91 +/- 0.01) x 10(-8) to (4.55 +/- 0.07) x 10(-8) M over a pH range of 6.50-8.20, respectively . These experimental data satisfactorily fitted the curve generated by the solubility equation CT = S0(1 + K5/{H+})/({H+}/K4' + 1), where S0 is the concentration of the tetraanionic ReLPS, K5 is the dissociation constant of the tetraanionic ReLPS in solution, and K4' is the dissociation constant of the trianionic ReLPS at the surface of the solid particles in suspension . The increase in solubility of ReLPS with increase in pH from 7.00 to 8.20 is primarily caused by the formation of the pentaanionic form from the tetraanions . The pK5 (primarily the second dissociation of the 1-phosphate) of ReLPS was determined to be 8.58 from experimental data.(ABSTRACT TRUNCATED AT 250 WORDS)

Toxicol Lett, 1993 May, 68(1-2), 251 - 63
Ozone- and endotoxin-induced mucous cell metaplasias in rat airway epithelium: novel animal models to study toxicant-induced epithelial transformation in airways; Harkema JR et al.; Mucous (goblet) cell proliferation and hypersecretion of airway mucus are important characteristics of human respiratory disorders, especially chronic bronchitis and cystic fibrosis . These changes in secretory patterns also occur in animals experimentally exposed to chemical irritants such as ozone (O3), sulfur dioxide (SO2), and cigarette smoke . The cellular and molecular mechanisms involved in irritant-induced mucous cell metaplasia (MCM; transformation of airway epithelium, normally devoid of mucous cells, to a secretory epithelium containing numerous mucous cells) are still unclear . We used two experimental models of toxicant-induced MCM in rat airways to study the cellular and molecular changes that occur during the development of this respiratory tract lesion . MCM can be induced in the nasal transitional epithelium of rats by repeated exposure to ambient levels of ozone . In addition, MCM can be induced in the tracheobronchial airways of rats repeatedly exposed to endotoxin, a lipopolysaccharide-protein molecule found in the outer walls of Gram-negative bacteria . The pathogenesis of ozone- or endotoxin-induced MCM has been partially characterized using a variety of morphometric and histochemical techniques . Toxicant-induced changes in the numbers and types of airway epithelial cells have been estimated using morphometric methods designed for estimating the abundance of cell populations . Nasal pulmonary airway tissues are also processed for light microscopy and stained with Alcian Blue (pH 2.5)/Periodic Acid Schiff (AB/PAS) for detection of acidic and neutral mucosubstances (the specific glycoprotein product of mucous cells), respectively, within the tissue . Computerized image analysis is used to quantitate the amount of the stained mucous product within the airway epithelium . To better characterize the molecular and cellular events in the pathogenesis of ozone- or endotoxin-induced MCM in the rat airway epithelium, we are conducting studies to determine when, and in which epithelial cells, the mucin gene is expressed after exposure to the toxicant . In these studies, rats undergo single or repeated exposures to ozone or endotoxin and are then sacrificed immediately or a few days after the end of the exposures . Airway tissues are microdissected from specific regions of the exposed respiratory tract, and changes in mucin core polypeptide mRNA are evaluated by Northern analysis using human and rat mucin cDNA . In future studies using in situ hybridization, we will establish when, and in which epithelial cells, the expression of high molecular weight airway mucin is initiated in response to ozone or endotoxin.(ABSTRACT TRUNCATED AT 400 WORDS)

J Clin Microbiol, 1993 May, 31(5), 1122 - 6
Occurrence of bacterial endosymbionts in Acanthamoeba spp . isolated from corneal and environmental specimens and contact lenses; Fritsche TR et al.; Free-living and parasitic protozoa are known to harbor a variety of endosymbiotic bacteria, although the roles such endosymbionts play in host survival, infectivity, and invasiveness are unclear . We have identified the presence of intracellular bacteria in 14 of 57 (24%) axenically grown Acanthamoeba isolates examined . These organisms are gram negative and non-acid fast, and they cannot be cultured by routine methodologies, although electron microscopy reveals evidence for multiplication within the amoebic cytoplasm . Examination for Legionella spp . with culture and nucleic acid probes has proven unsuccessful . We conclude that these bacteria are endosymbionts which have an obligate need to multiply within their amoebic hosts . Rod-shaped bacteria were identified in 5 of 23 clinical Acanthamoeba isolates (3 of 19 corneal isolates and 2 of 4 contact lens isolates), 4 of 25 environmental Acanthamoeba isolates, and 2 of 9 American Type Culture Collection Acanthamoeba isolates (ATCC 30868 and ATCC 30871) previously unrecognized as having endosymbionts . Coccus-shaped bacteria were present in one clinical (corneal) isolate and two environmental isolates . There was no statistical difference (P > 0.8) between the numbers of endosymbiont strains originating from clinical (26% positive) and environmental (24% positive) amoebic isolates, suggesting that the presence alone of these bacteria does not enhance amoebic infectivity . Rods and cocci were found in both clinical and environmental isolates from different geographical areas (Seattle, Wash., and Portland, Oreg.), demonstrating their widespread occurrence in nature . Our findings suggest that endosymbiosis occurs commonly among members of the family Acanthamoebidae and that the endosymbionts comprise a diverse taxonomic assemblage . The role such endosymbionts may play in pathogenesis remains unknown, although a variety of exogenous bacteria have been implicated in the development of amoebic keratitis, warranting further evaluation.

Cell Immunol, 1993 May, 148(2), 397 - 407
Iron augments macrophage-mediated killing of Brucella abortus alone and in conjunction with interferon-gamma; Jiang X et al.; Brucella abortus are Gram negative facultative intracellular bacteria, which survive and replicate in host macrophages . We have recently demonstrated that activation of macrophages with interferon-gamma increases their anti-brucella activities but does not result in elimination of intracellular brucellae . Here we demonstrate that iron-loaded macrophages have an enhanced capacity to kill or prevent replication of intracellular brucellae . Iron added bound to transferrin or as a salt, iron-nitrilotriacetate, can mediate the effect . Macrophages supplemented with iron-loaded transferrin in addition to activation with interferon-gamma can frequently eliminate the intracellular organisms by 48 hr after infection . The effect is apparent following phagocytosis of either nonopsonized or antibody-opsonized brucellae, and with both attenuated and virulent strains of B . abortus . The killing can be blocked by the hydroxyl radical scavengers mannitol and thiourea . This is consistent with the Haber-Weiss reaction, in which iron catalyzes the generation of hydroxyl radicals from hydrogen peroxide.

Infect Immun, 1993 May, 61(5), 1756 - 63
Reactivity of the human antiendotoxin immunoglobulin M monoclonal antibody HA-1A with lipopolysaccharides from rough and smooth gram-negative organisms; Mascelli MA et al.; Clinical data suggest that the human immunoglobulin M antiendotoxin antibody HA-1A reduced mortality in patients diagnosed with gram-negative bacteremia and bacteremia with shock . Previous studies have demonstrated that HA-1A binds to the lipid A domain of lipopolysaccharide (LPS) . The present study evaluated the ability of HA-1A to interact with LPs isolated from various strains of gram-negative bacteria by using liquid-phase rate nephelometry and solid-phase immunoblotting assays . HA-1A formed immune complexes in solution with LPSs isolated from both rough and smooth gram-negative organisms . Western blot (immunoblot) analysis of these LPS preparations revealed that HA-1A bound to LPS isolated from rough gram-negative organisms and to a rough LPS-like component present in smooth LPS . HA-1A also bound to LPS-protein complexes found in certain commercial rough LPS preparations . Preincubation of HA-1A with lipid A completely blocked subsequent binding of HA-1A to LPS in both liquid- and solid-phase assay formats, suggesting that the interaction of HA-1A with LPS is through the lipid A domain . Evidence that the binding of HA-1A to LPS was mediated through the antigen-combining (Fv) region of the antibody was provided by the finding that a murine anti-idiotypic antibody to HA-1A inhibited binding . These findings suggested that the broad antiendotoxin reactivity exhibited by HA-1A appeared to be due to the ability of HA-1A to bind to the conserved lipid A moiety of LPSs derived from both smooth- and rough-phenotype gram-negative bacterial strains.

Infect Immun, 1993 May, 61(5), 1715 - 21
Cytocidal effects of Escherichia coli hemolysin on human T lymphocytes; Jonas D et al.; Escherichia coli hemolysin is the prototype of a large family of pore-forming toxins produced by gram-negative organisms . Besides its known cytotoxic activities against granulocytes, monocytes, endothelial cells, and renal epithelial cells, we now demonstrate that the toxin potently kills human T lymphocytes . Evidence based on different and independent approaches indicates that lymphocidal activity is due to formation of transmembrane pores . Additionally, cells prestimulated with phytohemagglutinin respond to low doses of E . coli hemolysin with DNA fragmentation similar to that observed in cells undergoing programmed cell death . Kinetic considerations lead us to conclude that DNA degradation may, however, represent an epiphenomenon . Killing of T cells is another means through which E . coli hemolysin could directly impair host defense.

Infect Immun, 1993 May, 61(5), 1630 - 5
Endotoxin-induced tumor necrosis factor alpha synthesis in murine embryo fibroblasts; Havell EA et al.; Murine embryo fibroblasts (MEF) were found to secrete tumor necrosis factor (TNF) in response to stimulation with endotoxin . Endotoxin-induced TNF production by MEF was inhibited by cycloheximide . However, reversal of the effect of this inhibitor on protein synthesis results in TNF being secreted in amounts equivalent to those produced by endotoxin-induced MEF not treated with cycloheximide . Actinomycin D treatment of MEF blocked the production of endotoxin-induced TNF . Maximal production of TNF required MEF gene transcription during the first 6 h of incubation with endotoxin . To determine whether endotoxin-induced TNF alpha (TNF-alpha) and/or TNF beta were produced by MEF, cDNA was synthesized from the total RNA isolated from endotoxin-induced MEF and amplified by the polymerase chain reaction in the presence of oligonucleotide primers specific for each cytokine . On the basis of the polymerase chain reaction analysis, it was determined that TNF-alpha mRNA levels were increased in endotoxin-induced MEF . Thus, production of TNF-alpha by fibroblasts in response to the endotoxin component of bacterial cell walls is likely to contribute to the expression of TNF-mediated effects occurring in fibroblast-rich tissues infected with gram-negative bacteria.

Am J Crit Care, 1993 May, 2(3), 224 - 35; quiz 236-7
Epidemiology, pathophysiology and clinical presentation of gram-negative sepsis; Hazinski MF et al.; OBJECTIVE: To review the epidemiology and pathophysiology of gram-negative sepsis and the new consensus terminology describing the clinical signs of sepsis . DATA SOURCES: Review of the medical literature and compiled data from animal and clinical trials . PARTICIPANTS: Members of the Society of Critical Care Medicine, American College of Chest Physicians and American Association of Critical-Care Nurses with expertise on the subject of sepsis and its complications . RESULTS: Preconference and general sessions were offered at the National Teaching Institutes of the American Association of Critical-Care Nurses, with the goal of clarifying the epidemiology, risk factors and pathophysiology of gram-negative sepsis . In addition, current terminology and new (1992) consensus terminology describing the clinical signs of sepsis were presented . Special emphasis was placed on the role of the healthcare provider in the prevention and recognition of sepsis and the role of the septic mediators in the septic cascade . CONCLUSIONS: If the incidence of sepsis is to be reduced, the healthcare provider must be aware of the risk factors for sepsis and methods of reducing nosocomial infections . A thorough understanding of the role of mediators and consensus terminology used to describe sepsis, severe sepsis, septic shock and multiple organ dysfunction syndrome is necessary to recognize early or progressive signs of sepsis and to initiate state-of-the-art therapy.

J Gen Microbiol, 1993 May, 139 ( Pt 5), 1105 - 14
The melA gene is essential for melanin biosynthesis in the marine bacterium Shewanella colwelliana; Fuqua WC et al.; The surface-adhering, Gram-negative marine bacterium Shewanella colwelliana synthesizes a red-brow melanin in the late stage of exponential growth in laboratory culture . Previous studies identified a single gene, melA, from S . colwelliana that could impart the ability to produce melanin to an E . coli host . However, these studies did not demonstrate a requirement for melA during melanization in S . colwelliana . In this paper, genetic analyses, using a broad host range conjugation system to generate specific lesions, reveal that melA null mutants fail to synthesize pigment . The wild-type melA gene provided in trans on a low copy number plasmid complemented these null mutations, as well as a spontaneous pigment variant, to wild-type melanin synthesis . Polyclonal antibodies, raised against a MelA-LacZ fusion protein, were used to confirm the presence of the melA gene product in wild-type S . colwelliana and verify its absence in the non-pigmented mutants . In addition, detection of the MelA protein over the course of growth in batch culture revealed a constant steady-state level of MelA protein, suggesting that the timing of melanization and the quantity of melanin synthesized is not controlled at the level of melA expression.

J Antimicrob Chemother, 1993 May, 31 Suppl D, 61 - 70
Pharmacodynamics of antibiotics in the therapy of meningitis: infection model observations; Schmidt T et al.; Detailed studies of pharmacodynamic principles relevant to the therapy of bacterial meningitis are difficult to perform in man, while the rabbit model of bacterial meningitis has proved to be extremely valuable and has led to insights that appear relevant for the treatment of humans . Most importantly in the light of the restricted penetration of antibiotics into the CSF, animal studies have shown that in meningitis there is a dose-response curve between the CSF concentrations achieved by antibiotics and their bactericidal activity . This appears to be true for all classes of antibiotics thus far examined, including the beta-lactams, which do not show such a dose-response behaviour in other infections . Only CSF concentrations that exceed the MBC of the infecting organism by at least 10-30-fold achieve consistent and rapid bactericidal activity . Such rapid bactericidal activity is a requirement for successful therapy with beta-lactams and can be impaired with certain antibiotics by the specific conditions in infected CSF (protein content; acidic pH; slow-growing bacteria) . However, rapid antibiotic killing of the infecting organisms may not be without adverse effects either . Some antibiotics, particularly beta-lactams lead to the brisk liberation of bacterial cell wall components (e.g . endotoxin, in the case of Gram-negative organisms) which have an inflammatory effect on the host and can lead to a temporary deterioration of the disease . Dexamethasone, when administered with the antibiotic, can prevent some of the adverse effects of rapid bacterial lysis.

Circ Shock, 1993 May, 40(1), 53 - 60
Lipopolysaccharide alters suckling rat liver glycogenolysis; Goto M et al.; Gram-negative sepsis/septic shock in the newborn continues to be a major medical problem, causing high mortality . Hyperglycemia followed by hypoglycemia is a common symptom in endotoxic shock . However, the mechanism of newborn glucoregulatory response to endotoxin has not been well understood . Paradoxically, monocyte-phagocytes can contribute to shock by overwhelming secretion of cytokines and also host defense by detoxifying endotoxin . Since monocyte-phagocyte function is immature in the newborn, this study was performed to evaluate Kupffer cell's role in liver glycogenolysis during endotoxic shock . Endotoxin (LPS) induced hyperglycemia in 10-day-old rats, and increased net glucose output in the isolated perfused liver . 1) Cytarabine decreased Kupffer cell function (decreased hepatic colloid carbon uptake) and blunted LPS-increased liver net glucose output in the Cytarabine + LPS-treated group (104 +/- 4 vs . 146 +/- 3 micrograms/min/g wet liver in the LPS-treated group: P < .001) . 2) Indomethacin (IND) suppressed LPS-induced liver net glucose output in the LPS + IND-treated group (133 +/- 5 vs . 146 +/- 3 micrograms/min/g wet liver, P < .05) . Thus, prostaglandins were suggested to contribute to glycogenolysis in the 10-day-old rat liver . 3) Phorbol 12-myristate 13-acetate (PMA) increased liver net glucose output (166 +/- 4 micrograms/min/g wet liver), and H-7, a protein kinase C inhibitor, blunted PMA-induced liver glucose output (140 +/- 2 micrograms/min/g wet liver, P < .05) . H-7 enhanced LPS-induced liver net glucose output (196 +/- 9 micrograms/min/g wet liver, P < .01) . Therefore, protein kinase C may not be the dominant cell signaling system for LPS stimulation in suckling rat Kupffer cells.

J Dairy Res, 1993 May, 60(2), 223 - 8
Routine limulus amoebocyte lysate (LAL) test for endotoxin determination in milk using a Toxinometer ET-201; Mottar J et al.; A rapid method of performing the Limulus amoebocyte lysate (LAL) test in milk is proposed using the Toxinometer ET-201 . This instrument measured the increase in turbidity due to the interaction between the endotoxins of the Gram-negative bacteria and the LAL reagent, monitored the ratio Rt of the sequential to the initial transmission at 12 s intervals and quantified endotoxins by determination of the reaction time Tr required to obtain a 5% decrease in Rt . There was a good correlation between the toxinometrically determined endotoxin concentrations and the number of Gram-negative bacteria (SD, 0.18 log(plate count units)), and the repeatability (CV, 6-10%) was high . The assay may be useful for screening raw materials for UHT milk production, as the endotoxin content of the raw material is related to the rest proteinase activity in the UHT milk.

Lancet, 1993 May 1, 341(8853), 1133 - 5
Reappraisal of the role of endotoxin in the sepsis syndrome; Hurley JC; There is strong evidence to implicate endotoxin released from gram negative bacteria in the pathogenesis of the sepsis syndrome and related conditions, but equally compelling data bring the role of endotoxin into doubt . Reappraisal of endotoxin and its release from gram negative bacteria suggests that it is not directly responsible for the complications of sepsis syndrome . Rather, release of endotoxin is a marker for the transition of gram negative organisms to cell-wall-deficient forms (L-forms) that may persist undetected despite antibiotic therapy directed against the parental form . This transition has two consequences in compromised patients: L-forms cause organ failure, and they serve as a sanctuary from which cell-wall-intact revertants may arise.

New Horiz, 1993 May, 1(2), 312 - 23
Disseminated intravascular coagulation: pathophysiology, diagnosis, and treatment; ten Cate H et al.; Disseminated intravascular coagulation is a frequent finding in critically ill patients, and may be diagnosed in the majority of patients with Gram-negative sepsis . Tissue damage may result from intravascular thrombosis, and disseminated intravascular coagulation is an underestimated causal factor in the pathogenesis of organ failure in sepsis . The diagnosis of disseminated intravascular coagulation is difficult, as the initial coagulation process that leads to thrombosis is counteracted by fibrinolytic responses, that in the context of ongoing consumption of clotting factors may result in an overwhelming bleeding tendency . Hence, depending on underlying disease, and the time at which the patient is evaluated, different laboratory results may be obtained . The treatment of disseminated intravascular coagulation is even more challenging than its diagnosis . No well-designed, controlled clinical trials exist that form a basis for rational treatment decisions . Treatment frequently needs to be individualized, and rapid adjustments may be necessitated by the course of the disease . Nonetheless, we believe that recent insights in the pathophysiology of disseminated intravascular coagulation, in particular concerning the role of the extrinsic coagulation pathway, provide ground for some optimism concerning future therapeutic options.

Trends Microbiol, 1993 May, 1(2), 50 - 5
A novel secretion apparatus for the assembly of adhesive bacterial pili; Jacob-Dubuisson F et al.; The biogenesis of most types of bacterial pili requires two specialized proteins: a chaperone that caps the pilus subunits in the periplasm, and an outer membrane usher that receives the subunits and serves as an assembly platform . This secretion and assembly machinery is proposed to be a novel export apparatus found widely in Gram-negative pathogens.

J Struct Biol, 1993 May-Jun, 110(3), 180 - 7
Demonstration of the glycocalyces associated with three oral gram-negative bacterial species using a modern acrylic resin technique; Barber PM et al.; The complex highly hydrated chemical composition of the bacterial glycocalyx renders it difficult to preserve and visualize at the ultrastructural level . Polyanionic stains such as ruthenium red help to maintain some structural integrity, and other more modern approaches include antibody stabilization, lectins, and the addition of lysine to the primary fixative . It has been suggested that the glycocalyx of certain disease-associated organisms may play a role in the pathogenesis of some microbially based diseases such as periodontitis . New, more adequate, modern methodologies are therefore required for the further study of this structure . In the present study a cold dehydration process in conjunction with LR white acrylic resin has been employed to study the glycocalyces of three oral gram-negative bacterial species reported to be periodontopathogens . When compared with organisms processed conventionally and with ruthenium red, the organisms processed by the cold dehydration and LR white method demonstrated a fibrous matrix that was not seen in the other specimens . These results indicate that a combination of reduced dehydration temperature and cold acrylic resin embedding provides the best methodology for the visualization of the fine structure of the bacterial glycocalyx . This approach may be particularly useful in the study of organisms within specific disease-associated environments such as the periodontal pocket.

J Clin Microbiol, 1993 May, 31(5), 1027 - 9
Rejection criteria for endotracheal aspirates from adults; Morris AJ et al.; Although criteria have been established to assess the quality of sputum specimens, no criteria for assessing the quality of endotracheal suction aspirates (ETSA) exist . Therefore, we compared the Gram stain (GS) and culture results for 504 consecutive ETSA specimens . Results recorded for GS included the numbers of squamous epithelial cells (SEC) and polymorphonuclear leukocytes (PML) per low-power field (LPF) (magnification, x100) as well as the quantities and types of organisms per high-power field (HPF) (magnification, x1,000) . Culture results were quantitated by organism . Only 15% of ETSA specimens tested by GS contained > 10 SEC per LPF, and 21, 20, and 59% had < or = 10, 11 to 24, and > or = 25 PML per LPF, respectively . For 40% of ETSA specimens, no organisms were visible by GS . Of these specimens, 40% were sterile, 48% grew normal oropharyngeal flora (NF) only, 5% grew 1+ NF (i.e., > 10 colonies in the first quadrant) and 1+ gram-negative rods (GNR), and 7% grew < or = 1+ GNR either alone or in mixed culture . The mean numbers of organisms recovered from ETSA with < or = 10 SEC per LPF and > 10 SEC per LPF were 2.35 and 4.05, respectively . We therefore recommend that ETSA specimens that show no organisms by GS be rejected, in addition to those with > 10 SEC per LPF . Application of these rejection criteria enabled us to reject 847 (41%) of 2,068 ETSA specimens over a 6-month period . This represents a saving of approximately $66,000/year in unnecessary laboratory charges to patients.

J Clin Invest, 1993 May, 91(5), 2076 - 83
Mechanisms of stimulation of interleukin-1 beta and tumor necrosis factor-alpha by Mycobacterium tuberculosis components; Zhang Y et al.; The granulomatous immune response in tuberculosis is characterized by delayed hypersensitivity and is mediated by various cytokines released by the stimulated mononuclear phagocytes, including tumor necrosis factor-alpha (TNF alpha) and IL-1 beta . We have demonstrated that Mycobacterium tuberculosis cell wall component lipoarabinomannan (LAM), mycobacterial heat shock protein-65 kD, and M . tuberculosis culture filtrate, devoid of LPS as assessed by the Amebocyte Lysate assay, stimulate the production of TNF alpha and IL-1 beta proteins and mRNA from mononuclear phagocytes (THP-1 cells) . The effect of LAM on the release of these cytokines was specific, as only LAM stimulation was inhibited by anti-LAM monoclonal antibody . Interestingly, we found that LAM and Gram-negative bacterial cell wall-associated endotoxin LPS may share a similar mechanism in their stimulatory action as demonstrated by inhibition of TNF alpha and IL-1 beta release by monoclonal antibodies to CD14 . Anti-CD14 monoclonal antibody MY4 inhibited both TNF alpha and IL-1 beta release with LAM and LPS but no effect was observed with other mycobacterial proteins . An isotype antibody control did not inhibit release of cytokines under the same experimental conditions . M . tuberculosis and its components upregulated IL-1 beta and TNF alpha mRNAs in THP-1 cells . Nuclear run-on assay for IL-1 beta demonstrated that LAM increased the transcription rate . The induction of IL-1 beta was regulated at the transcriptional level, in which these stimuli acted through cis-acting element(s) on the 5' flanking region of the IL-1 beta genomic DNA . M . tuberculosis cell wall component LAM acts similarly to LPS in activating mononuclear phagocyte cytokine TNF alpha and IL-1 beta release through CD14 and synthesis at the transcriptional level; both cytokines are key participants in the host immune response to tuberculosis.

J Immunol, 1993 Apr 15, 150(8 Pt 1), 3397 - 403
Mice treated with a leumedin or antibody to Mac-1 to inhibit leukocyte sequestration survive endotoxin challenge; Burch RM et al.; Endotoxin challenge causes metabolic dysfunction mediated by TNF, and sequestration of leukocytes . NPC 15669, N-carboxy-L-leucine, N-{2,7-dimethylfluoren-9-yl)methyl} ester, inhibits leukocyte recruitment into inflammatory lesions in animals, and inhibits endotoxin-induced neutropenia and lymphopenia in mice . This study was carried out to determine whether the ability of NPC 15669 to inhibit leukocyte sequestration is sufficient to promote survival after endotoxin challenge . To inhibit leukocyte sequestration directly, mice were treated with anti-CD11a (LFA-1) or anti-CD11b (Mac-1) before endotoxin challenge . Anti-CD11b partly inhibited neutropenia and lymphopenia in response to challenge with LPS, but anti--CD11a had little effect on leukopenia . At doses of 100 and 1000 micrograms/kg, anti-CD11b increased survival to endotoxin challenge from 0 to 20 and 40%, respectively, whereas anti-CD11a was without effect . These observations, coupled with the finding that NPC 15669 does not inhibit endotoxin-induced TNF release suggest that inhibition of leukocyte sequestration can increase survival after endotoxin challenge, and that NPC 15669 or antibodies to Mac-1 may represent effective therapies for gram-negative sepsis and shock.

JAMA, 1993 Apr 14, 269(14), 1829 - 35
Anticytokine strategies in the treatment of the systemic inflammatory response syndrome; Dinarello CA et al.; The systemic inflammatory response syndrome (SIRS) is an acute illness characterized by generalized activation of the endothelium . The most severe form of the syndrome is found in patients with shock due to gram-negative sepsis . We examined both animal and limited human data for the contribution of cytokines to this syndrome . Cytokines are endogenously produced proteins of small molecular weight and multiple biological effects . The cytokines interleukin 1 (IL-1) and tumor necrosis factor (TNF), as well as interferon-gamma and interleukin 8, are discussed . Laboratory investigations suggest that these cytokines play a critical role in SIRS by promoting the biochemical and clinical characteristics of SIRS . The biochemical changes induced by TNF and IL-1 include increased synthesis of nitric oxide, prostaglandins, platelet-activating factor, and endothelial cell adhesion molecules . Specific blockade of TNF using neutralizing antibodies or soluble receptors to TNF in animal models of SIRS reduces mortality and severity of disease . Similar results have been observed blocking IL-1 using soluble IL-1 receptors or IL-1 receptor antagonists . Preliminary clinical studies suggest that blockade may be useful in treating human SIRS . The various strategies for blocking IL-1 and TNF are presented; in addition, their mechanism(s) of action and safety in humans are discussed . We conclude that based on animal studies and preliminary clinical trials, strategies to block IL-1 or TNF may benefit patients with the syndrome, although thorough clinical trials have not been completed.

Anaesth Intensive Care, 1993 Apr, 21(2), 172 - 3
Aminoglycoside volume of distribution and illness severity in critically ill septic patients; Marik PE; The volume of distribution of amikacin and the APACHE II score were determined in 42 critically ill patients being treated for a gram-negative infection . The mean volume of distribution (Vdt) was 0.41 +/- 0.12 l/kg with a wide range (normal of 0.25 l/kg) . There was a good relationship between the Vdt and illness severity as measured by the APACHE II score (r = 0.70; P < 0.001) . Critically ill patients should receive larger loading doses of aminoglycosides in order to achieve therapeutic blood levels . The aminoglycoside Vdt may be useful in determining the degree of capillary leak and tissue oedema that accompanies sepsis.

Infusionsther Transfusionsmed, 1993 Apr, 20 Suppl 1, 16 - 9; discussion 20
{Detection of endotoxin in plasma: specificity and value for development and prognosis of infection}; Urbaschek R et al.; A number of problems may be involved in the detection of endotoxin in plasma of patients using LAL (Limulus amebocyte lysate) . When collecting blood or processing samples, contamination with endotoxin or its adsorption to material must be avoided . In our laboratory a kinetic LAL microtiter assay was developed that takes into account plasma-related interferences with the LAL endotoxin reaction by performing an internal standardization in each sample . Negative results do not absolutely exclude the involvement of endotoxins in the underlying disease . High levels of endotoxins do not necessarily reflect the severeness of the clinical status of the patient . Due to nonendotoxin-specific reactions with some complete lysates, false-positive levels may result, e.g., following immunoglobulin therapy . In spite of these limitations, the LAL test remains a valuable tool in the evaluation of gram-negative infections.

Am J Vet Res, 1993 Apr, 54(4), 576 - 9
Pharmacokinetics of cefotaxime in neonatal pony foals; Gardner SY et al.; Serum concentrations of cefotaxime and desacetylcefotaxime were measured in 1-week-old pony foals after IV administration of a single dose of cefotaxime . The cefotaxime disposition data conformed to a two-compartment model with elimination half-life of 0.60 hour . The combined cefotaxime and desacetylcefotaxime data was best described by a four-compartment model . The apparent half-life describing the disappearance of desacetylcefotaxime was 1.69 hours . Dosage of 40 mg/kg of body weight given IV every 4 to 6 hours for neonatal foals with gram-negative septicemia and every 2 hours for foals with meningitis is recommended for further study.

J Am Vet Med Assoc, 1993 Apr 1, 202(7), 1137 - 42
Cholelithiasis in dogs: 29 cases (1980-1990); Kirpensteijn J et al.; Medical records of 29 dogs with cholelithiasis were reviewed . Aged female small-breed dogs were overrepresented . Mean age was 9.5 years, and mean weight was 12 kg . Vomiting, anorexia, weakness, polyuria/polydipsia, weight loss, icterus, fever, and signs of abdominal pain were the most common clinical signs . Leukocytosis, neutrophilia with left shift, monocytosis, high activity of serum hepatic enzymes, hypoalbuminemia, and high concentrations of serum total bilirubin were common . Radiopaque choleliths were evident on abdominal radiography of 13 of 27 dogs . Microbial culturing of bile isolated organisms in 15 of 20 dogs . Gram-negative bacteria were most common . Surgery was performed in 22 dogs . Four dogs were treated medically, and 3 dogs were euthanatized without treatment . Surgical treatment consisted of cholecystectomy in 11 dogs, choledochotomy in 5 dogs, cholecystotomy in 4 dogs, and cholecystojejunostomy in 1 dog . Sphincter of Oddiotomy was performed in 1 dog . Five dogs had concurrent generalized peritonitis attributable to bile . Multiple choleliths were detected in most of the dogs . Choleliths were located in the gallbladder in 20 dogs and in the bile ducts in 14 dogs . The most common abnormalities of the gallbladder, identified histologically, were chronic cholecystitis, mucosal hyperplasia, and pericholecystic inflammation . The most common abnormalities of the liver were cholestasis, hepatocellular degeneration, and periportal fibrosis . Survival rate of dogs that underwent cholecystectomy tended to be higher (86%) than that of dogs treated via cholecystotomy (50%) or cholecystectomy in combination with choledochotomy (33%) . Dogs that underwent medical treatment, abdominal exploratory, cholecystojenunostomy, choledochotomy, and sphincter of Oddiotomy died or were euthanatized because of redevelopment of clinical signs associated with cholelithiasis.(ABSTRACT TRUNCATED AT 250 WORDS)

EMBO J, 1993 Apr, 12(4), 1265 - 75
Growth phase dependence of the activation of a bacterial gene for carotenoid synthesis by blue light; Fontes M et al.; Myxococcus xanthus responds to blue light by producing carotenoid pigments . A mutation at a gene named carC is known to block the metabolism of phytoene, a carotenoid precursor, and this gene has now been cloned and sequenced . We show here that gene carC, which is homologous to phytoene dehydrogenase genes from other organisms, is tightly regulated by light through a mechanism that operates only when the cells have reached the stationary phase or are starved of a carbon source . A genetic element that mediates the effect of the growth phase has been identified . Gene carC is integrated with another unlinked carotenogenic gene in a single 'light regulon' controlled by common trans-acting genetic elements . A potential -35 site for the binding of sigma factors has been found upstream of the carC transcriptional start . However, the -10 region shows no similarity with analogous sites at promoters of other Gram-negative bacteria.

J Bacteriol, 1993 Apr, 175(7), 2157 - 61
Heat shock-induced axenic growth of Bdellovibrio bacteriovorus; Gordon RF et al.; The bdellovibrios are obligately predatory bacteria that attack other gram-negative bacteria . They grow only in the periplasmic space of prey unless they mutate to forms that can grow axenically . A culture medium that promoted enhanced growth of prey-independent bdellovibrios was developed . The ability of this medium to support the growth of prey-dependent bdellovibrios was tested under transcription-altering conditions . This approach tested the hypothesis that the inability to grow prey-dependent bdellovibrios in artificial media was rooted in both nutritional and transcriptional signal deficiencies . It was assumed that nutritional deficiencies had been resolved and that empirically applied artificial signals may evoke the expression of genes required for axenic growth of bdellovibrios . Prey-dependent bdellovibrios could be grown in PPYE medium (0.1% proteose peptone 3 and 0.03% Bacto yeast extract adjusted to pH 7.0 and supplemented with 3 mM MgCl2 and 2 mM CaCl2 after autoclaving) after heat shock, and subsequent rounds of growth occurred after additional heat shocks . Heat shock may have generated or simulated signals normally derived from prey.

J Infect Dis, 1993 Apr, 167(4), 865 - 75
Human anti-endotoxin antibody HA-1A mediates complement-dependent binding of Escherichia coli J5 lipopolysaccharide to complement receptor type 1 of human erythrocytes and neutrophils; Krieger JI et al.; HA-1A has been shown clinically to decrease mortality in septic patients with gram-negative bacteremia . In this study, the ability of HA-1A to augment the serum complement-dependent immune adherence of 125I-labeled Escherichia coli J5 lipopolysaccharide (LPS) to human erythrocytes (RBC) and polymorphonuclear leukocytes (PMNL) was evaluated . In vitro studies indicated three things: HA-1A mediates immune adherence of 125I-J5 LPS to human RBC and PMNL in a dose-dependent manner; under these conditions, high concentrations of LPS (400 ng/mL) could be specifically bound . Immune adherence occurs via the classical complement pathway as demonstrated by its calcium dependence; HA-1A-J5 LPS-C' immune complexes bound to CR1 on human RBC and PMNL . PMNL binding and internalization of immune complexes was demonstrated by trypsin stripping of externally bound immune complexes . These studies support the proposal that HA-1A can lower the bioavailability of endotoxin by mediating binding and potential clearance of LPS via human RBC through the reticuloendothelial system or via direct internalization by peripheral blood PMNL.

J Histochem Cytochem, 1993 Apr, 41(4), 601 - 8
Binding of bacterial endotoxins to the macrophage surface: visualization by fracture-flip and immunocytochemistry; Risco C et al.; Endotoxins (lipopolysaccharides, LPS) are surface components of gram-negative bacteria that stimulate macrophage activation and cause endotoxic shock . How LPS is recognized by host cells is still an open question, but it is generally accepted that many effects of endotoxins follow the overproduction of cytokines by macrophages . In the present study, we used fracture-flip and immunolabeling to study the morphology of isolated commercial LPS (C-LPS), the endotoxin release from the bacterial wall in presence of serum (S-LPS), and the distribution of these two endotoxins on the macrophage surface . Cells treated with C-LPS exhibited large LPS aggregates bound to smooth and particulate areas of the membrane and to microvilli . In contrast, macrophages incubated with S-LPS showed a uniform monodispersed labeling over the free surface of the membrane . Our results show that fracture-flip provides high-resolution images of the binding of ligands to the cell surface . They also suggest the importance of using highly dispersed LPS suspensions when the mechanisms of cell activation and damage by endotoxins are studied.

Avian Dis, 1993 Apr-Jun, 37(2), 628 - 34
Pharmacokinetic properties of gentamicin and amikacin in the cockatiel; Ramsay EC et al.; Gentamicin and amikacin are commonly used in veterinary medicine to treat a variety of gram-negative bacterial infections . The present study evaluates the pharmacokinetics of gentamicin sulfate and amikacin sulfate in the cockatiel (Nymphicus hollandicus), a small (approximate body weight = 100 g) psittacine bird, utilizing treatment regimens developed in larger parrot species . Serum antibiotic concentrations were determined in cockatiels following twice-daily intramuscular treatment with 5 mg gentamicin/kg body weight and 15 mg amikacin/kg body weight . In the present study, peak values of gentamicin were 4.6 +/- 1.45 micrograms/ml, and trough values were 0.17 +/- 0.04 micrograms/ml . Amikacin administration resulted in peak values of 27.3 +/- 6.9 micrograms/ml and trough concentrations of 0.9 +/- 0.3 micrograms/ml . Based on the present study, an appropriate intramuscular dose regimen for gentamicin in cockatiels is 5 to 10 mg/kg body weight either two or three times per day . An intramuscular amikacin dosage of 15 to 20 mg/kg body weight either two or three times per day is recommended for treatment of infections caused by susceptible bacteria.

Immun Infekt, 1993 Apr, 21(2), 40 - 4
{The role of cytokines in endotoxic shock and in endotoxin hypersensitivity}; Freudenberg MA et al.; Endotoxins (lipopolysaccharides, LPS) are constituents of the outer membrane of gram-negative bacteria . The application of purified LPS to experimental animals leads to the development of many pathophysiological activities which are also seen during infection with gram-negative microorganisms . One important prerequisite for the development of endotoxin shock is the interaction of LPS with macrophages, the activation of which leads to the release of cytokines, in particular of TNF alpha and IL-1, which mediate the toxic activity of LPS . The interaction of LPS with target cells and the subsequent initiation of endotoxin shock may be modified by plasma proteins that are capable of binding LPS . The most important LPS-binding proteins known so far are high density lipoprotein (HDL), low density lipoprotein (LDL), LPS-binding protein (LBP) and specific LPS antibodies . Native LPS released from bacteria may be associated partly with bacterial proteins (e.g . OmpA) which may also modify its interaction with host targets . The sensitivity of the organism to LPS is genetically determined, but may be altered under different experimental conditions . Hypersensitivity to LPS may be achieved by treatment with live (infection) or killed microorganisms, growing tumors, hepatotoxic agents or treatment with proteins to which the organism has been immunologically primed . The state of hypersensitivity is characterized by an increased ability of hypersensitive animals to produce cytokines on LPS challenge, as well as by an increased susceptibility to the toxic activity of TNF alpha . Recently it could be shown that interferon gamma (IFN gamma) is a central mediator in the development of the hypersensitivity to LPS induced by infection.

Immun Infekt, 1993 Apr, 21(2), 26 - 35
{Bacterial endotoxins: relationship between chemical structure and biological effect}; Rietschel ET et al.; Gram-negative bacteria carry on their surface endotoxins, which are essential for bacterial growth and survival . If released from the bacterial cell, endotoxins induce in higher organisms a great variety of pathophysiological effects . Chemically, endotoxins constitute lipopolysaccharides (LPS), the lipid component (termed lipid A) of which is responsible for the induction of endotoxin effects . The structural and conformational parameters, endowing lipid A with its potent bioactivity, have been well characterized . The toxic effects of endotoxins are initiated by the specific interaction of lipid A with macrophages/monocytes resulting in the production of peptide or lipid mediators . This interaction is governed by a unique (toxic) conformation of lipid A on the one hand, and by specific cellular receptors on the other . The interaction and subsequent mediator production can be specifically and antagonistically inhibited by lipid A partial structures . A recently developed monoclonal anti-LPS-antibody cross-reacts with endotoxins of various bacterial origin, and it cross-protects against harmful endotoxin effects such as pyrogenicity and lethality.

J Surg Res, 1993 Apr, 54(4), 342 - 8
Anti-lipopolysaccharide monoclonal antibodies inhibit macrophage TNF messenger RNA synthesis in vitro; Battafarano RJ et al.; Gram-negative bacterial lipopolysaccharide (LPS, endotoxin) directly stimulates macrophages to produce tumor necrosis factor (TNF) . TNF, in turn, produces a constellation of adverse effects that includes hypotension, systemic acidosis, arterial hypoxemia, and death . Transcription of the TNF gene occurs within minutes of LPS stimulation and appears to be a critical control point in the synthesis and secretion of TNF protein by macrophages . We hypothesized that murine monoclonal antibody (mAb) 8G9 directed against Escherichia coli 0111:B4 LPS would provide protective capacity against an E . coli 0111:B4 bacterial challenge in vivo and would concurrently inhibit LPS-induced synthesis of TNF mRNA and secretion of TNF protein in vitro . E . coli 0111:B4 LPS was used to stimulate a macrophage-derived cell line (RAW 264.7) to produce TNF in the presence or absence of mAb 8G9 . Media alone and LPS without 8G9 mAb served as controls against which the effect of 8G9 mAb was compared . Total cellular RNA was purified and analyzed by a Northern blotting technique utilizing a radiolabeled cDNA probe specific for TNF mRNA . TNF mRNA levels from each sample were quantitated by autoradiograph densitometry . Pretreatment with mAb 8G9 provided protective capacity against an intraperitoneal E . coli 0111:B4 bacterial challenge in vivo when compared with saline pretreatment alone (22% versus 90% mortality respectively, P < 0.05) . Preincubation of LPS with mAb 8G9 resulted in a significant inhibition of LPS-induced TNF mRNA synthesis (63 +/- 20%, P < 0.01) and TNF protein secretion (88 +/- 10%, P < 0.001) in vitro.(ABSTRACT TRUNCATED AT 250 WORDS)

Immunobiology, 1993 Apr, 187(3-5), 464 - 77
Immunotherapy of endotoxemia and septicemia; Baumgartner JD et al.; Neutralization of endotoxin (lipopolysaccharide, LPS) would be of considerable benefit in the treatment of Gram-negative sepsis . Administration of anti-LPS antibodies is an old approach which has been renewed by improvements in monoclonal antibody technology . The antibodies directed at the conserved core region of LPS or at the lipid A which have been studied in humans are discussed in this review . Some of these antibodies appeared to be protective in animal models or in clinical trials, but discrepant results have been reported and the mechanism of the postulated protection was not clarified . The polyclonal antibody preparations have given variable results in patients . The clinical studies of anti-lipid A monoclonal antibodies seemed promising because both antibodies appeared to protect subsets of patients . However, the studies gave discrepant results concerning the type of patients reported to benefit from the administration of these antibodies . One of these antibodies, E5, appeared to improve the survival of patients with Gram-negative sepsis provided they were not in shock, but a second trial failed to confirm this . The other antibody, HA-1A, appeared to protect patients with Gram-negative sepsis who were in refractory shock, but only when they were bacteremic . This antibody was recently released on the market in some european countries . However, the FDA agency decided that a confirmatory study should be done before it could consider to approve HA-1A because a careful reanalysis suggested that the observed differences were only of marginal statistical significance . Therefore, this type of treatment has not yet clearly been shown to benefit patients . More studies are needed to delineate the role of core LPS antibodies in the management of Gram-negative sepsis.

Immunobiology, 1993 Apr, 187(3-5), 430 - 46
Lipopolysaccharide, lipid A, and liposomes containing lipid A as immunologic adjuvants; Alving CR; Numerous studies have demonstrated that most or all of the potent adjuvant activity of Gram-negative bacterial endotoxin resides in the lipid A moiety of lipopolysaccharide (LPS) . Synthetic analogues of lipid A have provided insights into structure-activity relationships . Several cellular mechanisms of LPS and lipid A adjuvant activities have been identified . Activation of macrophages by LPS or lipid A results in cytokine secretions that enhance the immune response . LPS and lipid A cause recruitment of antigen-presenting cells, particularly macrophages . Liposomes containing lipid A serve as an in vivo adjuvant to recruit increased numbers of macrophages . Liposomal lipid A that has been phagocytized by cultured macrophages also serves as an "intracellular adjuvant" to cause increased immunologic presentation of liposomal antigen by the macrophages to specific T lymphocytes . Lipid A can abolish suppressor T cell activity, resulting in increased immune responses to polysaccharide antigens . Upon combination of lipid A or lipid A analogues with nonionic block polymers, modulation of murine antibody isotypes can be achieved with antibodies against a variety of antigens in vivo . Liposomes containing monophosphoryl lipid A (MPL) have been utilized in a phase I clinical trial of a proposed malaria vaccine in humans . The liposomal malaria vaccine resulted in very high levels of antibodies against the malarial antigen, and despite the presence of huge amounts of MPL (up to 2.2 mg), the liposomal lipid A was nonpyrogenic and safe for use in humans . Lipid A and lipid A analogues, and liposomes or other carriers containing lipid A, have shown considerable promise both as adjuvants for immunization of animals and for human vaccines.

Immunobiology, 1993 Apr, 187(3-5), 403 - 16
Response of man to endotoxin; Martich GD et al.; Endotoxin, a cell wall component of Gram-negative bacteria, plays a central role in the pathogenesis of septic shock . By administering small doses of intravenous endotoxin to humans, a variety of acute inflammatory responses are induced which are qualitatively similar to those that occur during the early stages of septic shock . Within hours of the administration of intravenous endotoxin to human volunteers, changes occur in systemic hemodynamics, ventricular function, pulmonary gas exchange and permeability . In conjunction with these changes in organ function, a wide variety of inflammatory mediators are released which appear to contribute to these responses . These include the release of proinflammatory cytokines (e.g . tumor necrosis factor-alpha, IL-1 beta, IL-6, IL-8), activation of the fibrinolytic system, kallikrein-kinin generation and phospholipase A2 release . Phagocytic leukocytes are primed for enhanced inflammatory responses following endotoxin administration . Counter-regulatory responses are initiated in parallel and may serve to limit some of the end-organ responses by the inflammatory mediators . This human model provides a unique opportunity to extend previous concepts of acute inflammation and to evaluate the earliest responses activated after exposure to an important bacterial component . Defining the pathways and responses initiated during acute human endotoxemia may allow a better understanding of host responses that are critical to the development of organ dysfunction and shock due to severe infections.

Immunobiology, 1993 Apr, 187(3-5), 346 - 56
Mechanisms of endotoxin shock and endotoxin hypersensitivity; Galanos C et al.; Endotoxins (lipopolysaccharide, LPS) are biologically active substances present in Gram-negative bacteria . Injection of purified LPS into experimental animals leads to the development of many biological activities that can lead to shock with lethal outcome . The biological activities of LPS are not direct effects of the LPS molecule since LPS usually expresses no direct cytotoxic activity . The toxic and other biological properties of LPS are caused indirectly through the action of endogenous mediators that are formed following interaction of LPS with cellular targets, macrophages occupying a key position in the development of endotoxin shock . The interaction of LPS with macrophages may proceed directly leading to activation of these cells, with subsequent synthesis and secretion of a number of endogenous mediators which initiate the different biological activities of LPS . Tumor necrosis factor alpha (TNF-alpha), a macrophage derived cytokine, is a primary mediator of the lethal action of endotoxin . Sensitivity to LPS is genetically determined, varying considerably among different species . The sensitivity of normal animals (mice) to endotoxin may be enhanced considerably under different experimental conditions that include treatment with live (infection) or killed Gram-negative and -positive bacteria . Sensitization to endotoxin proceeds in all LPS-responder strains investigated and in the LPS-resistant mice of the strain C3H/HeJ . It does not proceed in a second LPS-resistant strain, C57BL/10ScCr . The absence of sensitization in the latter mice was found to be due to an impaired IFN-gamma production . IFN-gamma could be identified as the mediator of endotoxin hypersensitivity induced by bacteria.

Immunobiology, 1993 Apr, 187(3-5), 169 - 90
The chemical structure of bacterial endotoxin in relation to bioactivity; Rietschel ET et al.; Lipopolysaccharides (LPS) constitute the O-antigens and endotoxins of Gram-negative bacteria . Whereas both the polysaccharide and lipid portion of LPS contribute to the pathogenic potential of this class of bacteria, it is the lipid component (lipid A) which determines the endotoxic properties of LPS . The primary structure of lipid A of various bacterial origin has been elucidated and Escherichia coli lipid A has been chemically synthesized . The biological analysis of synthetic lipid A partial structures proved that the expression of endotoxic activity depends on a unique structural arrangement and conformation . Such analyses have furthermore provided insight into the determinants required for lipid A binding to and activation of human target cells . Present research efforts aim at the molecular characterization of the specificity, modulation and biomedical consequences of the interaction of lipid A with host cells.

West J Med, 1993 Apr, 158(4), 393 - 9
Monoclonal antibodies to endotoxin in the management of sepsis; Fang KC; New monoclonal antibodies directed against the lipid A moiety of the endotoxin present in gram-negative bacteria have been developed to improve the clinical outcome in patients with sepsis . Two studies of monoclonal antibodies HA-1A and E5 retrospectively identified specific patient subgroups showing benefit with therapy . I analyze and summarize the new sepsis nomenclature, the structure of endotoxin, the data implicating endotoxin as a causative agent in septic patients' morbidity and mortality, and specific data from the 2 clinical studies of monoclonal antibody therapy.

S Afr Med J, 1993 Apr, 83(4), 253 - 6
Abdominal complications in black and Indian children with nephrotic syndrome; Adhikari M et al.; Abdominal complications were detected and investigated in 19 (10%) of 191 children with nephrotic syndrome who experienced 35 episodes of these complications . Fourteen children were Indian with steroid-responsive nephrotic syndrome, and 5 were black, of whom 4 had membranous nephropathy and 1 focal proliferative nephritis . All had clinical features of peritonitis and hypovolaemia was frequently present . Eleven of the 35 episodes were culture-proven peritonitis (5 due to Pneumococcus, 6 due to Gram-negative bacteria) and in 24 the cultures were negative . Hypovolaemia occurred in 6 of the former group and 5 of the latter . The occurrence of these episodes bore no temporal relationship to steroid and cyclophosphamide treatment . Sixty-nine per cent of the complications appeared within the first 3 years of onset of the nephrotic syndrome and 8 of 19 patients experienced multiple episodes . In this study, hypovolaemia always occurred in the context of clinically detected peritonitis and not as a separate complication, suggesting infection together with fluid and protein losses as likely pathogenetic mechanisms.

Mol Microbiol, 1993 Apr, 8(2), 379 - 88
A sequence-specific function for the N-terminal signal-like sequence of the TonB protein; Karlsson M et al.; TonB is a proline-rich protein which provides a functional link between the inner and outer membranes of Gram-negative bacteria . TonB is anchored to the inner membrane via an N-terminal signal-like sequence and spans the periplasm, interacting with transport receptors in the outer membrane . We have investigated the role of the N-terminal signal-like peptide in TonB function . Replacement of the N-terminal sequence with heterologous sequences indicates that it has at least three distinct roles in TonB function: (i) to facilitate translocation of TonB across the cytoplasmic membrane; (ii) to anchor TonB to the cytoplasmic membrane; (iii) a sequence-specific functional interaction with the ExbBD proteins.

Chest, 1993 Apr, 103(4), 1107 - 12
Elevated concentrations of cross-linked fibrin degradation products in plasma . An early marker of gram-negative bacteremia; Deitcher SR et al.; PURPOSE: Because coagulant and fibrinolytic activity are increased by endotoxin, we hypothesized that concentrations of cross-linked fibrin degradation products, sensitive markers of plasmin activity, would be increased in patients with Gram-negative bacteremia . PATIENTS AND METHODS: Cross-linked fibrin degradation products were measured with an enzyme-linked immunosorbent assay (ELISA) based on a monoclonal antibody specific for cross-linked fibrin degradation products, and with a novel, semiquantitative hemagglutination assay based on the same antibody coupled to FAB' fragments from a monoclonal antibody against a universal red blood cell antigen . Blood samples were obtained from 100 consecutive emergency department patients evaluated for sepsis . RESULTS: Blood cultures were positive in 21 percent of the patients, 75 percent (12/21) with Gram-negative organisms . Elevations of cross-linked fibrin degradation products (normal < 200 ng/ml) were measured by ELISA in all patients (12/12) with Gram-negative bacteremia . The hemagglutination assay was positive in ten of these . Detectable increases in fibrinolytic activity occurred in all patients with Gram-negative bacteremia, even in the absence of clinical signs or laboratory evidence of sepsis or disseminated intravascular coagulation . The ELISA and hemagglutination assay had negative predictive values for Gram-negative bacteremia of 100 percent and 96 percent, respectively . CONCLUSION: Elevation of cross-linked fibrin degradation products consistent with increased fibrinolytic activity occurs in all patients with Gram-negative bacteremia . Accordingly, the lack of elevation may be clinically useful in identifying patients less likely to have Gram-negative bacteremia.

J Formos Med Assoc, 1993 Apr, 92(4), 317 - 23
Gram-negative bacillary meningitis in adults; Hsu GJ et al.; To evaluate the clinical aspects of gram-negative bacillary meningitis (GNBM), we reviewed 41 adult patients with bacteriologically proven gram-negative bacillary meningitis, seen from 1985 to 1990 . Thirty-two patients had post-neurosurgical GNBM and nine patients had spontaneous GNBM . Spontaneous GNBM appeared to have a sudden onset, a relatively fulminant course, and was caused most often by Escherichia coli . Post-neurosurgical GNBM, however, had a more insidious onset, a more protracted course, and was more often caused by nosocomial organisms which were resistant to multiple antibiotics . The overall mortality was 39% . Patients treated with combined aminoglycoside therapy had a lower mortality rate than those treated with intravenous aminoglycoside (17% vs 48%) . The use of third-generation cephalosporins has made a significant therapeutic advance in the treatment of GNBM, with a lower mortality of 21% . We recommend treatment of GNBM with third-generation cephalosporins and aminoglycosides . If aminoglycosides are to be employed, it is suggested that they be administered both intravenously and directly into the central nervous system.

Mol Microbiol, 1993 Apr, 8(2), 389 - 96
ExbB acts as a chaperone-like protein to stabilize TonB in the cytoplasm; Karlsson M et al.; The TonB protein is required to transduce energy from the cytoplasmic membrane to outer membrane transport proteins of Gram-negative bacteria . Two accessory proteins, ExbB and ExbD, are required for TonB function and it has been suggested that TonB and ExbBD form a complex in the membrane . In this paper we demonstrate that there are two spatially distinct, functional interactions between ExbBD and TonB . First, there is an interaction between ExbBD and the N-terminal signal-like peptide of TonB, probably the formation of a stable complex in the membrane . Second, ExbB interacts with TonB in the cytoplasm . This interaction involves the domain of TonB that is normally periplasmic . Thus, this is a transient interaction which occurs during the synthesis and/or localization of TonB, implying a chaperone-like role for ExbB . The transmembrane topology of ExbB was shown to be consistent with this role.

J Clin Invest, 1993 Apr, 91(4), 1459 - 68
Tumor necrosis factor-alpha blockade prevents neutrophil CD18 receptor upregulation and attenuates acute lung injury in porcine sepsis without inhibition of neutrophil oxygen radical generation; Windsor AC et al.; Tumor necrosis factor (TNF alpha), both by direct action and by trafficking cells of the immune system, is implicated in cardiopulmonary derangements and PMN-mediated microvascular injury associated with gram-negative sepsis . We examined the effects of pretreatment with a monoclonal antibody to TNF alpha on PMN function, hemodynamic derangements, and alveolar capillary membrane damage in a septic porcine model . Anti-TNF alpha profoundly improved hemodynamic consequences in this model . Reduction in PMN CD11/18 receptor expression, lung myeloperoxidase activity, and attenuation of peripheral neutropenia (all P < 0.05) indicate that pretreatment significantly reduced lung sequestration of PMNs seen in septic controls . In contrast, PMN oxygen radical (O2-) generation was not significantly different from unprotected septic animals . Despite the presence of circulating PMNs primed for O2- burst, alveolar capillary membrane damage, assessed by bronchoalveolar lavage protein content and arterial PO2 was markedly attenuated in the treatment group (P < 0.05) . We conclude that anti-TNF alpha suppresses systemic hemodynamic actions of TNF alpha . Further, it prevents upregulation of PMN adhesion receptors inhibiting PMN/endothelial cell interaction . This prevents formation of a "microenvironment," protected from circulating oxidant scavengers, into which sepsis-activated PMNs release their toxic products . Pretreatment with anti-TNF alpha monoclonal antibody thus affords global protection in porcine Gram-negative sepsis.

J Infect Dis, 1993 Apr, 167(4), 890 - 8
Degradation of glycosaminoglycans and fibronectin on endotoxin-stimulated endothelium by adherent neutrophils: relationship to CD11b/CD18 and L-selectin expression; Klein NJ et al.; Vascular endothelial injury observed in overwhelming sepsis may be caused by neutrophil-derived enzymes . Adherence to the endothelium, a prerequisite for this process, is mediated sequentially by the neutrophil adhesion molecules L-selectin and the beta 2 integrins including CD11b/CD18 . The relationship between expression of these molecules, neutrophil adherence, endothelial activation, and consequent endothelial injury was assessed by changes in heparan sulfate and fibronectin matrices . Endothelial prestimulation with lipopolysaccharide caused both an increase in adherence and a generalized reduction in heparan sulfate; disruption of the fibronectin matrix occurred only on the further addition of FMLP . Although maximal disruption of these matrices was associated with elevation of neutrophil CD11b/CD18 and reduction in L-selectin expression, these changes did not determine either the nature or extent of endothelial damage . This model may provide further insights into the interrelationship between neutrophil activation and endothelial damage in gram-negative sepsis.

Proc Natl Acad Sci U S A, 1993 Mar 15, 90(6), 2179 - 83
Cytochromes c biogenesis in a photosynthetic bacterium requires a periplasmic thioredoxin-like protein; Beckman DL et al.; Rhodobacter capsulatus is a Gram-negative photosynthetic bacterium that requires c-type cytochromes for photosynthetic electron transport . Our studies demonstrate that the gene helX is required for the biogenesis of c-type cytochromes in R . capsulatus . A helX chromosomal deletion mutant cannot grow photosynthetically, due to a deficiency of all c-type cytochromes . The predicted amino acid sequence of the helX gene product (176 residues) is related to that of thioredoxin and shares active-site homology with protein disulfide isomerase . Cytochrome c2-alkaline phosphatase gene fusions are used to show that HelX is not required for the transcription, translation, or secretion of apocytochrome c2 . HelX-alkaline phosphatase and HelX-beta-galactosidase gene fusions are used to demonstrate that HelX is a periplasmic protein, which is consistent with the presence of a typical signal sequence in HelX . Based on these results, we propose HelX functions as a periplasmic disulfide oxidoreductase that is essential for cytochromes c biogenesis . This role is in accordance with the observation that both heme and the cysteines of apocytochromes c (Cys-Xaa-Yaa-Cys-His) must be in the reduced state for covalent linkage between the two moieties to occur.

Tidsskr Nor Laegeforen, 1993 Mar 10, 113(7), 859 - 61
{Chlamydia pneumoniae--pathogenesis and perspectives}; Berdal BP et al.; Chlamydia pneumoniae, a Gram-negative bacterium, formerly named TWAR but identified as a distinct species since 1988, is now considered to be the most common agent of chlamydial infection in Scandinavia . C pneumoniae has a different tissue trophism from that of Chlamydia trachomatis, since C pneumoniae may infect bronchi and lungs, macrophages, monocytes, and endothelial cells . C pneumoniae, like other chlamydiae, has a slow, intracellular life cycle . An absence of reaction from the host cells, combined with scant tissual reaction owing to the low endotoxic activity of chlamydial lipopolysaccharide, may help to explain the usually discreet clinical picture . Atherosclerosis and coronary heart disease may follow chronic lung infection, and acute pneumonic episodes can trigger myocardial infarct . Asymptomatic infection with C pneumoniae is widespread . Intriguing diagnostic questions are the possible existence of a non-pathogenic carrier state, and the conceivable sensitization of the host with respect to a heterotypic, secondary chlamydial infection by, for example, C trachomatis, giving rise to an aggravated clinical picture . Early antibiotics are indicated to avoid the development of chronic disease.

Immunol Invest, 1993 Mar, 22(2), 127 - 49
Potassium cyanide protects Escherichia coli from complement killing by the inhibition of C3 convertase activity; Bloch EF et al.; The exact mechanism by which deposited C5b-9 complexes kill Gram-negative bacteria is unclear . It has been proposed that during complement activation the membrane attack complex triggers an energy dependent process in Gram-negative bacteria that mediates destruction of the inner membrane . This observation in part resulted from the survival of Gram-negative bacteria that were incubated with an uncoupler (DNP) or an inhibitor (KCN) of oxidative phosphorylation during complement activation . In a reexamination of this issue we employed potassium cyanide (KCN) to block energy dependent pathways and observed a dose dependent inhibition of C9 uptake on E . coli J5 during serum incubation, suggesting that cyanide was interfering with complement activation . To verify the effect on complement activation we chose specifically to study the effects of KCN on the C3 convertase of the classical pathway . Sensitized sheep erythrocytes were employed as our model system . This system allowed us to construct a series of stable intermediates that were used to test the effect of cyanide on the formation and activity of precursors of the classical pathway C3 convertase . The data illustrate that the concentrations of potassium cyanide that inhibit complement killing of J5 also inhibit C3 convertase activity on sensitized sheep erythrocytes . The results of this study refute the principal observation made by other investigators, that potassium cyanide protects bacteria from complement killing by inhibiting bacterial energy dependent pathways that spark inner membrane destruction . A better scenario is that the organisms survive because cyanide inhibits complement activation.

Am J Ind Med, 1993 Mar, 23(3), 483 - 90
Investigation of an outbreak of "humidifier fever" in a print shop; Mamolen M et al.; An outbreak of "humidifier fever" affected 16 (57%) of 28 workers in a print shop . The most common symptoms were myalgia, chills or subjective fever, and cough . Illness began 5-13 hours after entering the workplace, and lasted 2-24 hours . A humidifier in use the day of the outbreak was found to be contaminated with fungi, amebae, and Gram-negative bacteria . The risk of illness was highest for those who had been on the job 3 months before the outbreak, a time when the humidifier was in constant use . Serologic studies of print shop workers showed positive reactions to extracts of organisms isolated from the humidifier, but could neither distinguish ill from well workers, nor identify causative organisms . The presence of endotoxin-producing bacteria and the clinical syndrome are consistent with an organic dust toxic syndrome . Previous exposure appeared to be the major risk factor for illness.

J Vet Intern Med, 1993 Mar-Apr, 7(2), 91 - 4
Cerebrospinal fluid composition of cattle with endotoxin-induced mastitis treated with isotonic (0.9%) or hypertonic (7.5%) sodium chloride; Tyler JW et al.; This study examined the safety of intravenous hypertonic saline in cattle with experimental gram-negative endotoxemia . Cerebrospinal fluid (CSF) composition was examined in five control cows and eight treated cows 24 hours after the intramammary infusion of 1 mg of endotoxin . Four of the endotoxin challenged cows were treated intravenously with isotonic (0.9%) sodium chloride and four cows were treated intravenously with hypertonic (7.5%) sodium chloride . Decreased CSF osmolality, and sodium and alpha globulin concentrations and increased CSF concentrations of beta globulin were observed in both endotoxin-challenged saline-treated groups . No CSF compositional differences were observed between endotoxin-challenged cows receiving isotonic or hypertonic saline . Although no cytologic or biochemical evidence of salt poisoning was observed in cows receiving hypertonic saline, significant changes were observed in the CSF composition of both endotoxin-infused saline-treated groups.

J Gen Microbiol, 1993 Mar, 139 ( Pt 3), 651 - 60
Cloning and sequencing of two type 4 (N-methylphenylalanine) pilin genes from Eikenella corrodens; Rao VK et al.; Eikenella corrodens is a Gram-negative microaerophilic rod which is gaining recognition as an important human pathogen . We have previously reported the cloning and expression in Escherichia coli of a 3.6 kb Eik . corrodens genomic DNA fragment which encodes a 31.5 kDa haemagglutinin . Maxicell analysis revealed that this fragment also encodes two proteins of approximately 14 kDa . Nucleotide sequencing of the 2.2 kb fragment upstream of the haemagglutinin gene revealed two open reading frames with strong homology to genes encoding pilin subunit proteins of the type 4 or N-methylphenylalanine class . The two pilin genes, ecpA and ecpB, are complete and are expressed in E . coli . Southern analysis of ten additional Eik . corrodens strains revealed that all possess fragments homologous to ecpA . These data represent the first molecular evidence for pili in E . corrodens.

Pharmacotherapy, 1993 Mar-Apr, 13(2), 128 - 34
Monoclonal antibody therapy for gram-negative sepsis: principles, applications, and controversies; Mehra IV et al.; Gram-negative sepsis is a common event in hospitalized patients and is a leading cause of death in the United States . Endotoxin (lipopolysaccharide, LPS), a component of the cell wall of gram-negative microorganisms, is responsible for the cascade of events leading to the sepsis syndrome consisting of fever, tachycardia, tachypnea, and evidence of organ hypoperfusion . The lipid A region of endotoxin produces most of these biologic and toxic effects . Monoclonal IgM antibodies directed against the lipid A portion of endotoxin (anti-LPS MoAb) have been developed for the treatment of gram-negative sepsis . Results of two large-scale clinical trials suggest that these antibodies offer clinically and statistically significant reductions in mortality by a factor of about one-third . However, in both trials, this apparent beneficial effect was limited to particular subsets of patients, and no overall benefit was seen . These considerations, in addition to the likely high cost of the agents, pose questions about their ultimate use in the treatment of patients with gram-negative sepsis . Nevertheless, the logic of the approach, the demonstration of efficacy in disease models, and the advances in modern techniques of molecular biology all suggest that these or other closely related products will play a significant role in the treatment of this disorder.

J Pediatr Surg, 1993 Mar, 28(3), 350 - 5; discussion 355-7
Prospective analysis of urokinase in the treatment of catheter sepsis in pediatric hematology-oncology patients; Jones GR et al.; Use of right atrial catheters (RACs) in children with cancer improves the comfort and efficacy of therapy . However, catheter-related infections are responsible for significant morbidity leading to the removal of approximately 20% of implanted RACs . Sepsis has been linked to thrombus and fibrin sheath formation within the RAC . Gram-negative and fungal infections appear to be particularly resistant to antibiotic therapy alone and most of these infections have required catheter removal . Urokinase has been effectively used for reopening thrombus occluded RACs . Theoretically, thrombolytic agents could improve the treatment of catheter-related infections by removing luminal sites of bacterial/fungal colonization . We prospectively monitored the use of urokinase and antibiotics for catheter-related sepsis in our pediatric hematology/oncology population from 1985 to 1991 . Sepsis episodes were treated with 2 doses of urokinase and antibiotics (10 to 42 days) infused through the RAC . One to 2 mL of urokinase (5,000 U/mL) was instilled in the RAC for 1 hour, then removed and repeated 24 hours later . During the study, 224 RACs were placed in 177 children . RACs were in place for a total of 71,134 days (median, 274 days) . There were 67 blood culture-positive sepsis episodes occurring in 50 RACs . Fifty-nine sepsis episodes were treated with urokinase and antibiotics and all responded by clearance of organisms from the blood . Three patients (5.1% of urokinase treated) had recurrent sepsis with the same organism within 2 months, were considered treatment failures and had RACs removed . Only 1 of 16 episodes of multiple organism/Candida sepsis led to RAC removal due to inability to cure the infection.(ABSTRACT TRUNCATED AT 250 WORDS)

Int J Clin Pharmacol Ther Toxicol, 1993 Mar, 31(3), 153 - 6
Comparison of the efficacy and safety of amikacin once or twice-a-day in the treatment of severe gram-negative infections in the elderly; Vanhaeverbeek M et al.; The aim of this study was to compare the efficacy and safety of amikacin given either as single injection or as two injections within 12-h interval in the treatment of severe gram-negative infections in elderly patients . Thirty-nine non-selected consecutive patients of a general internal medicine facility were randomized to receive the same total daily dose of amikacin either as a single dose (19 patients) or divided into two doses injected at 12-h interval (20 patients) . Amikacin was used alone or in combination with metronidazole, clindamycin, fosfomycin or a beta-lactam . Clinical and bacteriological responses were satisfactory and comparable in the two groups . There was no difference between the once/day and the twice-a-day groups with respect to drug dosage, duration of therapy and concomitant treatment . Only one patient (BID group) showed a rise of serum creatinine during the observation period . Amikacin alone or in combination can be regarded as an efficacious and safe antibiotic in the treatment of severe gram-negative infections in elderly patients, whether the daily dose is administered in a single infusion or in a BID interval.

J Clin Microbiol, 1993 Mar, 31(3), 692 - 4
Endocarditis caused by Rochalimaea quintana in a patient infected with human immunodeficiency virus; Spach DH et al.; Rochalimaea quintana and Rochalimaea henselae are closely related, fastidious, gram-negative rickettsiae . Thus far, the spectrum of human Rochalimaea sp . infections has not included endocarditis . We describe a 50-year-old human immunodeficiency virus-positive man who developed endocarditis caused by R . quintana . DNA relatedness studies, which compared our patient's blood culture isolate with known Rochalimaea species, identified the organism as R . quintana . Our report expands the spectrum of Rochalimaea sp . infections and identifies a new infectious cause of endocarditis.

J Clin Microbiol, 1993 Mar, 31(3), 528 - 32
Polymerase chain reaction-amplified nonradioactive probes for identification of Fusobacterium nucleatum; Bolstad AI et al.; A polymerase chain reaction probe with 100% sequence identity to 120 deoxyribonucleotides of Fusobacterium nucleatum Fev1, coding for a part of the 40-kDa major outer membrane protein, was labeled with the steroid hapten digoxigenin . The probe was compared with various degenerate oligonucleotide probes and found to tolerate much more stringent washing conditions . It was therefore superior in distinguishing, by means of Southern blots and slot blots, F . nucleatum from other oral gram-negative bacteria in the periodontal pocket and from other fusobacterial species and in distinguishing among different strains of F . nucleatum . F . periodonticum was found to be more similar to F . nucleatum than the other fusobacterial species tested.

Cancer, 1993 Mar 1, 71(5), 1898 - 903
Ifosfamide/etoposide combination in the treatment of recurrent malignant solid tumors of childhood . A Pediatric Oncology Group Phase II study; Kung FH et al.; BACKGROUND . The prognosis for children with recurrent or resistant malignant solid tumors remains dismal . More effective rescue therapy is needed for these children . METHODS . Between August 1987 and November 1990, 311 children with recurrent or resistant malignant solid tumors were treated by investigators in the Pediatric Oncology Group with intravenous infusions of 2.0 g/m2 of ifosfamide and 100 mg/m2 of etoposide (VP-16) plus mesna as uroprotection three times daily, with courses being repeated every 14-21 days for as long as the patients responded to therapy . RESULTS . Seventy-four percent of the 294 assessable patients entered in the study had metastatic disease and previously had been treated heavily . The complete response/partial response rate was 30%, and the overall response rate was 39.5% . Toxic effects included nephrotoxicity, mild liver dysfunction, neurotoxicity, and myelosuppression . Sixty-eight percent had an absolute neutrophil count (ANC) of less than 500/microliters . In 1606 courses of therapy administered, only 3.6% of patients developed a bacterial infection . Only two patients died of gram-negative sepsis . Four percent of the patients had gross hematuria (> 50 erythrocytes/high-power field), and 18.5% had microscopic hematuria (< 20 erythrocytes/high-power field) . Fanconi syndrome developed in eight children . CONCLUSIONS . Ifosfamide/VP-16 is an active combination in children with recurrent malignant solid tumors . Although it was myelosuppressive, the incidence of infection was quite low (3.6%) . Mesna was very effective in preventing the development of hematuria.

Transpl Int, 1993 Mar, 6(2), 77 - 84
Infections in human liver recipients: different patterns early and late after transplantation; Barkholt L et al.; The first 49 consecutive patients who underwent orthotopic liver transplantation between 1984 and 1989 in our department were studied with regard to symptomatic and asymptomatic post-transplantation infections . The major infections carrying a risk of fatal outcome are presented . During the first 4 weeks, fungal and bacterial infections predominated, the percentages of patients affected being 27% and 35%, respectively . Eight patients (17%) suffered from bacterial septicemia, which in six cases was due to gram-negative micro-organisms . The bacterial septicemia was often associated with severe ischemic damage to the graft, rejection, or cholangitis . In addition, a concomitant invasive fungal infection supervened in seven out of eight septic patients, further aggravating the patients' condition . Seventeen of the 49 patients (35%) died after transplantation within 3.3 years . Infection was the cause of death in nine patients (18%), with bacterial septicemia and/or fungemia in eight of these . Cytomegalovirus (CMV) disease was the dominant cause of illness after the 1st month . While only 5 of the 49 patients developed CMV disease during the 1st month (10%), as many as 16 of the 40 recipients who survived beyond that time suffered from symptomatic CMV viremia (40%) . CMV mismatching, i.e., the donation of a CMV-positive organ to a CMV-seronegative recipient, entailed the highest risk for CMV disease . Pneumocystis carinii pneumonia occurred within 4 months in 10% of the patients . The four liver recipients affected were among the 20 patients not receiving trimethoprim-sulfamethoxazole prophylaxis . None of the 28 patients who received this prophylaxis over a 12-month period developed this complication (P < 0.005).(ABSTRACT TRUNCATED AT 250 WORDS)

Plasmid, 1993 Mar, 29(2), 135 - 41
Characterization of an insertion sequence (IS53) located within IS51 on the iaa-containing plasmid of Pseudomonas syringae pv . savastanoi; Soby S et al.; A 2568-base pair (bp) insertion sequence (IS53) was cloned from the iaa-containing plasmid pIAA2 of Pseudomonas syringae subsp . savastanoi . IS53 is located downstream from the iaa operon and is inserted into IS51, a previously described element on pIAA2 . IS53 causes an 8-bp target duplication and has nearly perfect terminal inverted repeats of 27 bp . Several degenerate copies of the inverted repeats are located internal, and in direct orientation to the terminal inverted repeats . A putative transposase gene conforms to the transposase ORF/opposite strand overlapping ORF motif described in many gram-negative IS elements . The region 5' to the presumptive transposase gene contains sequences that are homologous to the consensus E . coli heat-shock (sigma 32) promoter.

Cleve Clin J Med, 1993 Mar-Apr, 60(2), 139 - 44
Selective bowel decontamination with quinolones and nystatin reduces gram-negative and fungal infections in orthotopic liver transplant recipients; Gorensek MJ et al.; Gram-negative and fungal infections are the most important cause of morbidity and mortality after liver transplantation, especially in the first postoperative month . From February 1989 to February 1990, all liver transplant recipients at The Cleveland Clinic Foundation, Cleveland, Ohio, were placed on a selective bowel decontamination regimen employing oral quinolones and nystatin beginning at the time they were put on the active waiting list for transplantation and continuing until the fourth postoperative week . The incidence of gram-negative and fungal infections for these patients was compared against a historical control group . Selective bowel decontamination was well tolerated and highly effective in reducing early serious gram-negative and fungal infections . This regimen may also reduce mortality.

Microb Pathog, 1993 Mar, 14(3), 177 - 85
Construction of a Brucella abortus RecA mutant and its survival in mice; Tatum FM et al.; To determine if RecA plays a role in the virulence of Brucella abortus, a B . abortus RecA mutant was constructed and its survival was examined in mice . The recA gene was cloned from a B . abortus genomic DNA library by complementation of an Escherichia coli recA mutant in the presence of methyl methanesulfonate (MMS) . The nucleotide sequence of recA was determined and the deduced protein sequence possesses extensive conservation with other RecA proteins of Gram-negative bacteria . A deletion plasmid was constructed in a suicide vector by deleting a segment of recA and inserting a kanamycin resistance gene . The deletion plasmid was introduced into B . abortus strain 2308, a virulent strain, by electroporation . Replacement of recA with the kanamycin resistance fragment was confirmed by Southern blot analysis . The RecA mutant was more sensitive than the parental strain to killing by MMS . When administered intraperitoneally to BALB/c mice, numbers of bacteria per spleen were consistently lower in animals infected with the RecA mutant than with the parental strain . However, both the RecA mutant and parental strain persisted in mice through 100 days post-infection . These results indicate that RecA is not crucial for persistence of B . abortus in mice.

Microbiol Rev, 1993 Mar, 57(1), 50 - 108
The complete general secretory pathway in gram-negative bacteria; Pugsley AP; The unifying feature of all proteins that are transported out of the cytoplasm of gram-negative bacteria by the general secretory pathway (GSP) is the presence of a long stretch of predominantly hydrophobic amino acids, the signal sequence . The interaction between signal sequence-bearing proteins and the cytoplasmic membrane may be a spontaneous event driven by the electrochemical energy potential across the cytoplasmic membrane, leading to membrane integration . The translocation of large, hydrophilic polypeptide segments to the periplasmic side of this membrane almost always requires at least six different proteins encoded by the sec genes and is dependent on both ATP hydrolysis and the electrochemical energy potential . Signal peptidases process precursors with a single, amino-terminal signal sequence, allowing them to be released into the periplasm, where they may remain or whence they may be inserted into the outer membrane . Selected proteins may also be transported across this membrane for assembly into cell surface appendages or for release into the extracellular medium . Many bacteria secrete a variety of structurally different proteins by a common pathway, referred to here as the main terminal branch of the GSP . This recently discovered branch pathway comprises at least 14 gene products . Other, simpler terminal branches of the GSP are also used by gram-negative bacteria to secrete a more limited range of extracellular proteins.

J Antimicrob Chemother, 1993 Mar, 31 Suppl C, 1 - 9
Chemistry and mode of action of macrolides; Mazzei T et al.; After the discovery of erythromycin and other natural compounds, including oleandomycin, spiramycin, josamycin and midecamycin, much research has been devoted to synthesizing derivatives or analogues with improved chemical, biological and pharmacokinetic properties . These new macrolides are semisynthetic molecules that differ from the original compounds in their substitution pattern of the lactone ring system . The chemical structure of macrolides is characterized by a large lactone ring containing from 12 to 16 atoms to which are attached, via glycosidic bonds, one or more sugars . The lactone ring is substituted by hydroxyl or alkyl groups, one ketone at C7 in 12-membered macrolides and at C9 in 14-membered macrolides, and one aldehyde group in 16-membered macrolides . The only compound with a 15-membered ring contains a tertiary amino group . Although the 12-membered macrolides have never become important in clinical practice, in recent years numerous new 14-membered macrolide derivatives of erythromycin A have shown improved pharmacokinetics due to chemical modifications of a hydroxyl group at C6, a proton at C8, or a ketone at C9 . Derivatives, such as dirithromycin, roxithromycin, clarithromycin and flurithromycin, have all been synthesized with the aim of inhibiting their decomposition under acidic conditions to inactive anhydrohemiketal derivatives . A new 15-membered macrolide, azithromycin, with a methylated nitrogen inserted into the lactone ring shows good activity against Gram-negative bacteria . The efforts expended in chemical and biochemical modifications of 16-membered macrolides have been less successful, with only a few new molecules, such as rokitamycin and miocamycin, showing improved bioavailability and activity against some resistant micro-organisms.(ABSTRACT TRUNCATED AT 250 WORDS)

Drugs, 1993 Mar, 45(3), 338 - 52
Critical care pharmacotherapy . A review; Tryba M et al.; During recent years, research in critical care medicine has focused on the role of the gastrointestinal tract in the pathogenesis of multiple organ failure and nosocomial infection, and on preventive measures . Gram-negative bacterial overgrowth of the oropharynx and stomach has been proved to be a cause of nosocomial pneumonia . Topical application of antibiotics into the oropharynx and stomach, and preservation of gastric acidity have been shown to be effective prophylaxis in ventilated patients . Recent studies have demonstrated that gastric alkalinisation is no longer necessary for the prevention of stress ulcer bleeding in critically ill patients . Tissue hypoxaemia, not gastric acidity, is the underlying pathomechanism of stress ulcer bleeding . In experimental investigations, pirenzepine and sucralfate improved gastric mucosal oxygen supply . Both compounds effectively prevent bleeding without increasing gastric pH . In mechanically ventilated patients, significantly lower rates of pneumonia occur with both of these drugs compared with antacids or histamine H2-receptor antagonists . Topical antibiotics (selective digestive decontamination) are most effective in patients with alkaline gastric juice, but of only marginal clinical relevance in those with acidic gastric contents . Isoflurane, propofol and clonidine have been recently investigated for sedation of ventilated patients . Isoflurane may lead to fluoride accumulation after more than 1 day . Propofol dosage has to be increased more often after 4 to 7 days, leading to fat overload and significantly increased costs . Clonidine was highly effective in patients with 'sympathetic overshoot', e.g . those experiencing alcohol or opioid withdrawal . Wound infections are an important problem in burn patients.(ABSTRACT TRUNCATED AT 250 WORDS)

Circ Res, 1993 Mar, 72(3), 539 - 51
Selective inhibition of endothelium-dependent vasodilator capacity by Escherichia coli endotoxemia; Parker JL et al.; Increased release of endothelium-derived relaxing factor/nitric oxide has been proposed as the final common pathway for vasodilator responses to gram-negative lipopolysaccharide (endotoxin) . To test this hypothesis, we examined endothelium-dependent and endothelium-independent vasodilator agents in vascular smooth muscle isolated from guinea pigs 16 hours after injection of saline (control group) or induction of Escherichia coli endotoxemia; aortic rings (approximately 1 mm in diameter) were studied with standard isometric tension techniques . Endotoxemia resulted in a significant loss of vasodilator responses to the endothelium-dependent receptor agonists acetylcholine (10(-10)-10(-5) M) and ADP (10(-8)-10(-5) M) . In contrast, endotoxemia did not affect vasodilator responses to either the endothelium-dependent receptor agonist substance P (10(-11)-10(-7) M), the endothelium-dependent and receptor-independent agonist A23187 (10(-9)-10(-6) M), or the endothelium-independent agonist nitroprusside (10(-10)-10(-4) M) . The nitric oxide synthase inhibitor NG-nitro-L-arginine methyl ester (L-NAME) inhibited the vasodilator response to acetylcholine more in vessels from lipopolysaccharide-injected than control guinea pigs . Unexpectedly, L-NAME converted the endothelium-dependent vasodilator action of ADP to an endothelium-dependent vasoconstrictor response that was blocked individually by the cyclooxygenase inhibitor indomethacin, the thromboxane synthase inhibitor dazoxiben, and the thromboxane A2 receptor antagonist SQ29548 . We conclude that in vivo endotoxemia inhibits the constitutive isoform of nitric oxide synthase in endothelial cells by selectively disrupting receptor-coupled activation mechanisms shared by acetylcholine and ADP . Furthermore, since L-NAME unmasks a thromboxane A2-mediated vasoconstrictor action of the endogenous purinoceptor agonist ADP, drugs that inhibit nitric oxide synthase could exacerbate sepsis-induced vasoconstriction and ischemia by synergizing with lipopolysaccharide-induced inhibition of endothelial nitric oxide synthase.

Harefuah, 1993 Feb 15, 124(4), 200 - 1, 247
{Treatment of gram-negative folliculitis with isotretinoin}; Amichai B et al.; Gram-negative folliculitis is one of the complications of antibiotic treatment of acne . Isotretinoin is recognized as extremely effective for this condition . We describe 2 patients with gram-negative folliculitis following antibiotic treatment for acne who improved after 3-months of treatment with isotretinoin.

FEMS Microbiol Lett, 1993 Feb 15, 107(1), 5 - 9
Production and characterisation of a Legionella pneumophila specific monoclonal antibody; Lever MS; To better define the antigenic structure of the outer cell membranes of Legionellae, a panel of 6 monoclonal antibodies was raised against partially purified outer membranes of Legionella pneumophila serogroup 1, Corby strain . This study describes the purification and characterization of one of these monoclonal antibodies reacting with a 135-kDa protein, which was shown to be common to all 14 serogroups of Legionella pneumophila . It shows no cross-reactivity with 20 other Legionella species, or 9 other Gram-negative species tested by SDS-PAGE and Western blotting procedures . The epitope would appear to be predominantly surface exposed and, from preliminary detergent extraction studies, not peptidoglycan-associated.

J Biol Chem, 1993 Feb 15, 268(5), 3662 - 9
DNA sequence requirements for interaction of the RK2 replication initiation protein with plasmid origin repeats; Perri S et al.; Replication of plasmid RK2 in a variety of Gram-negative bacteria requires its origin of replication and the plasmid-encoded TrfA proteins (TrfA-33 and TrfA-44) . The initiation of replication requires that the TrfA proteins bind to a series of 17-base pair (bp) direct repeats located within the RK2 origin . The conserved 17-bp repeats are arranged in tandem and are separated by less conserved spacer sequences of 4-6 bp in length . A series of plasmids containing one or two iterons, with or without the less conserved spacer sequences, were constructed to analyze the DNA sequence requirements for binding of TrfA-33 to the iterons . In addition to the analysis of TrfA binding in vitro, the plasmid constructs were examined for their ability to exert incompatibility toward an RK2 replicon in Escherichia coli . These analyses revealed that the conserved 17-bp iteron sequence itself is not sufficient for TrfA binding; the adjacent less conserved spacer sequences are also required . Site-specific mutagenesis was carried out to determine the importance of specific bases within the spacer sequence for binding activity and a consensus sequence for a TrfA-specific binding unit was determined . DNase I and methylation interference footprinting procedures were also carried out to characterize the TrfA-binding unit complex . Finally, it was shown that the binding of the TrfA-33 protein to two adjacent TrfA binding units on a DNA fragment is not substantially affected by the relative orientation or spacing between the two units.

Ned Tijdschr Geneeskd, 1993 Feb 13, 137(7), 355 - 60
{Immunotherapy using the anti-endotoxin antibody HA-1A (Centoxin) in patients with sepsis syndrome; fair results following protocol selection of patients}; Costongs LG et al.; OBJECTIVE . Evaluation of HA-1A treatment in patients with the sepsis syndrome . DESIGN . Descriptive . SETTING . Department of intensive care, Academic Medical Centre, Amsterdam . PATIENTS AND METHODS . Intensive-care patients with the sepsis syndrome and shock or organ failure with a presumptive diagnosis of Gram-negative infection were eligible for treatment with HA-1A . We analysed and compared the results with those of the double-blind, randomized HA-1A study by Ziegler et al . RESULTS . Between May 1991 and March 1992, 27 patients were treated with HA-1A . The mortality rate was 59% (16/27) . Among the 11 patients with a Gram-negative bacteraemia mortality was 7/11, much higher than in the Ziegler study (30%) . In comparison with the HA-1A study we selected sicker patients: the mean APACHE II score was higher, 93% of our patients were in shock and 85% had organ failure . More patients presented with an intra-abdominal sepsis and mortality in this group was very high (11/14) . In patients with a Gram-negative bacteraemia the delay between the onset of the sepsis syndrome and the administration of HA-1A was longer (median 22 h versus 14.3 h in the Ziegler study, mean 30 versus 20 h) . CONCLUSION . HA-1A does not appear to be beneficial in critically ill patients with a longstanding sepsis syndrome, especially not if an intra-abdominal sepsis is apparent . Therefore, we decided not to use H-1A until additional data become available . Additional objective inclusion criteria are needed to improve the identification of the patient group that may benefit from treatment with HA-1A.

Ned Tijdschr Geneeskd, 1993 Feb 13, 137(7), 350 - 4
{Immunotherapy using the anti-endotoxin antibody HA-1A in patients with sepsis syndrome; good results in relation to treatment with placebo}; Wortel CH et al.; Passive immunization with a human anti-endotoxin monoclonal IgM antibody (Centoxin, HA-1A) was recently studied in patients with suspected Gram-negative sepsis . Comparison of the results obtained in the Amsterdam subpopulation with those in a larger international study population of which the Amsterdam patient group was a part, showed that it had been possible to select a patient population in which HA-1A has an 'intention-to-treat' effect based upon clinical criteria (a decrease in mortality compared with placebo by 42% (p = 0.04) and in the larger study by 9% (p = 0.24) . Until a clinically useful test becomes available, identification of patients who have a high likelihood of Gram-negative sepsis and who would benefit from anti-endotoxin immunotherapy with HA-1A should be based upon the history and evaluation of underlying disease, infection status, severity and progression of the disease . The severely ill patients thus selected should receive treatment as early as possible.

MMWR Morb Mortal Wkly Rep, 1993 Feb 5, 42(4), 72 - 3
Capnocytophaga canimorsus sepsis misdiagnosed as plague--New Mexico, 1992.
{Structure of lipopolysaccharides from gram-negative bacteria . II . Core structure (Review)}
Knirel' IuA, Kochetkov NK.

The review summarizes the data on the core structure of bacterial lipopolysaccharides (LPS) which represents an oligosaccharide binding the lipid moiety of the macromolecule to the O-antigenic polysaccharide chain . Both S-strains with complete LPS and R-mutants having various defects of the core biosynthesis are considered . The role of the core in the outer membrane functioning and manifestation of antigenic specificity of LPS is discussed.

Biokhimiia, 1993 Feb, 58(2), 166 - 81
{Structure of lipopolysaccharides from gram-negative bacteria . I . Common characteristics of lipopolysaccharides and lipid A structure (Review)}; Knirel' IuA et al.; The data on the structure and properties of outer membrane lipopolysaccharides of gram-negative bacteria are reviewed with special reference to the general organization of the lipopolysaccharide molecule and the structure of the lipid moiety (lipid A) . The correlation between the structure of the lipopolysaccharides and lipid A and their biological properties and functions are discussed.

Environ Res, 1993 Feb, 60(2), 187 - 92
Airborne endotoxin concentrations in various work areas within a cotton mill in Central America; Christiani DC et al.; Gram-negative bacteria and their endotoxins have been shown to cause profound changes in the structure and function of mammalian lungs . Airborne exposures in humans have resulted in bronchoconstriction and symptoms of chest tightness and dyspnea . Cotton dust is often heavily contaminated and endotoxin has been postulated to be the agent in cotton dust which is responsible for the byssinosis syndrome . Previous studies of cotton dust have revealed variable amounts of contamination by bacterial endotoxin, with the variability determined in large part by botanical and climatic characteristics . We report here the results of an environmental study conducted in a cotton mill located in the tropics of Central America using locally grown cotton . Results indicate that airborne endotoxin was present in all yarn preparation and weaving areas and ranged from a low of 18 EU/m3 in weaving to a high of 3138 EU/m3 in opening areas . Airborne levels and the amount of endotoxin per nanogram of dust were not higher than values obtained in temperate climates.

Curr Opin Immunol, 1993 Feb, 5(1), 103 - 7
Bactericidal/permeability increasing protein and host defense against gram-negative bacteria and endotoxin; Elsbach P et al.; The bactericidal/permeability increasing protein is a major element in the host defense against Gram-negative bacteria and endotoxin, acting intracellularly in the polymorphonuclear leukocyte . As an isolated protein, bactericidal/permeability increasing protein also acts as an extracellular bactericidal and endotoxin-neutralizing agent and, when injected, protects animals against lethal effects of Gram-negative bacteria and endotoxin.

Am J Surg, 1993 Feb, 165(2A Suppl), 34S - 42S
Hospital-acquired gram-negative pneumonia in critically ill, injured patients; Rodriguez JL; Hospital-acquired gram-negative pneumonia is a major problem in critically ill, injured patients . The currently available therapeutic interventions to prevent the disease process are of limited usefulness . This most likely reflects an incomplete understanding of the complex pathophysiologic mechanism and thus invites examination of alternative mechanisms . We have hypothesized that the lung's response to traumatic injury may be driving the local organ injury by generating an early, local pulmonary cytokine production independent of the systemic cytokine response or the intensive care unit environment . Understanding the local pulmonary cytokine response to traumatic injury and its effect on the pulmonary airspace's immunologic contents may yield targeted and clinically relevant therapeutic interventions . Currently, the successful treatment of hospital-acquired gram-negative pneumonia depends on a clear and consistent definition of the disease process, knowledge that therapy with a single antibiotic is effective, and use of a concise treatment protocol that provides for reassessment of the patient when antibiotic therapy appears to be ineffective.

Am J Surg, 1993 Feb, 165(2A Suppl), 20S - 25S
Role of immunomodulator therapy in sepsis; Caplan ES; Endotoxin is released from the cell walls of gram-negative bacteria and causes severe systemic effects due to the release of cytokines . Monoclonal antibodies directed at endotoxin may be promising adjuncts to the standard therapeutic interventions of antibiotics and supportive measures used to treat patients with gram-negative sepsis . Monoclonal antibodies interfere with the bacteria's ability to trigger an unfavorable response . In recent clinical trials, two immunoglobulin M monoclonal antibodies have improved survival in certain small patient subgroups, although neither drug improved overall mortality in all septic patients treated . E5 murine monoclonal antibody reduced mortality in patients with gram-negative sepsis who were not in refractory shock . HA-1A human monoclonal antibody reduced mortality in patients with gram-negative infections who were bacteremic or in shock . The statistical significance and clinical importance of these benefits is not yet known . Results of these clinical trials are reviewed.

Am J Surg, 1993 Feb, 165(2A Suppl), 2S - 7S
Factors influencing the risk of infection after trauma; Polk HC Jr; Infection after injury is characterized by the degree of contamination, the duration of attendant hypotension, and the delay in definitive surgical treatment . Factors further influencing this outcome include the age of the patient and, in most U.S . urban centers, the general health of the patient . Current antibiotic treatment of such patients may soon be assisted by selective and sequential immunostimulants, which are increasingly more specific for defined defects in host defenses after injury . An ever more significant contribution to morbidity and mortality on trauma services is pneumonia, which is often caused by gram-negative aerobes . The design of clinically significant trials with respect to infection in the injury patient will be more difficult and complex.

Inflammation, 1993 Feb, 17(1), 57 - 75
Direct and indirect effects of E . coli lipopolysaccharide on isolated human polymorphonuclear granulocytes and mixed leukocytes; Opdahl H; Polymorphonuclear neutrophil granulocytes (PMN) may contribute to the lung injury induced by nonpulmonary infections with gram-negative bacteria . The direct effect of E . coli lipopolysaccharide (LPS) on isolated human PMN or mixed leukocytes (ML), as well as the priming effect of preincubating cells with LPS, was examined in assays measuring the maximal rate of oxygen consumption (OC), cell chemiluminescence (CHML), and aggregation (AGG) . LPS, 1-10 micrograms/ml, caused no acute response in PMN or ML suspended in Fisher's-HEPES medium with BSA (FHA), but increased both CHML and AGG of cells suspended in autologous plasma . Preincubation in FHA with LPS, 1 microgram/ml, for more than 15 min increased the OC of PMN activated with zymosan-activated plasma (ZAP) or n-formyl-methionyl-leu-cyl-phenylalanine (FMLP) by more than 100% . A similar increase in the CHML of such cells was seen after FMLP, but not after ZAP . ZAP, however, primed the CHML response of the cells to subsequent activation with FMLP more than did preincubation with LPS . Previous exposure to both agents had an additive effect . Preincubation of PMN with LPS decreased the time interval from addition of phorbol myristate acetate (PMA) to peak OC response, but less so than previous activation with FMLP . Neither agent affected the maximal rate of OC after addition of PMA . LPS also increased the PMN aggregation induced by ZAP and FMLP, but not by PMA . Cells preincubated with LPS, 0.01 microgram/ml, increased their CHML in response to FMLP if suspended in Krebs-Ringer balanced salt solution, but not if suspended in FHA . Such preincubation had no effect on OC of similarly activated cells in any of the media.

Crit Care Med, 1993 Feb, 21(2 Suppl), S32 - 9
Adoptive immunotherapy of gram-negative sepsis: use of monoclonal antibodies to lipopolysaccharide; Fink MP; OBJECTIVE: To provide a succinct overview of the scientific rationale for adoptive immunotherapy of Gram-negative sepsis using antibodies directed at epitopes in the core region of the lipopolysaccharide molecule . DATA SOURCES: Relevant, English-language primary and secondary (i.e., review) articles dealing with the structure or toxicity of lipopolysaccharide or adoptive anti-endotoxin immunotherapy in animals or man . STUDY SELECTION AND DATA EXTRACTION: The review emphasizes the findings obtained in several recent multicenter, randomized, prospective trials of monoclonal antibodies to core determinants of lipopolysaccharide . DATA SYNTHESIS: Lipopolysaccharide, a component of the outer cell wall of Gram-negative bacteria, is a complex molecule consisting of three regions: lipid A, core polysaccharide, and O-antigenic side chains . Epitopes in the lipid A and core polysaccharide regions are highly conserved across species and strains of Gram-negative bacteria . Because of the availability of mutant strains of bacteria that synthesize lipopolysaccharides lacking O-specific side chains and, in some instances, portions of the core polysaccharide moiety, investigators have been able to develop polyclonal and monoclonal antibodies to core-region determinants . In experimental animals, many of these antibodies have been shown to protect against the deleterious effects of either purified heterologous lipopolysaccharides or viable heterologous Gram-negative bacteria . Recently, two different monoclonal "anti-core" antibodies (HA-1A and E5) were evaluated in prospective, placebo-controlled, double-blind, randomized trials . In the pivotal trial of HA-1A, a human monoclonal IgM, no treatment effect was discernible when data from all patients enrolled in the study were analyzed . However, in a subset of patients with positive blood cultures for Gram-negative organisms, there was a decrease in all-cause mortality over a 28-day observation period in patients treated with HA-1A, as compared with those patients treated with placebo (p = .014) . Interpretation of this finding, however, is confounded because multiple other subsets and end-points also were analyzed, necessitating an adjustment in the p value necessary to reject the null hypothesis . A murine monoclonal IgM called E5 was evaluated in two separate trials . In the first trial, improved survival over a 30-day observation period was observed in a subset of patients with Gram-negative sepsis without refractory shock (p = .01) . However, in a second trial of E5, which focused on this subset, a statistically significant improvement in survival was not observed in patients with documented Gram-negative sepsis without shock . CONCLUSIONS: Adoptive immunotherapy using monoclonal anticore antibodies seems to improve survival rate in selected patients with Gram-negative sepsis . Nevertheless, because of concerns about costs and the interpretation of the results from the completed clinical trials, these new agents have generated enormous controversy . The precise role of adoptive immunotherapy against lipopolysaccharide in the practice of critical care medicine in the United States remains unclear.

Am J Ind Med, 1993 Feb, 23(2), 333 - 42
Cotton dust and gram-negative bacterial endotoxin correlations in two cotton textile mills; Christiani DC et al.; Exposure to cotton dust is known to cause both acute and chronic respiratory illness . A specific pattern of symptoms called byssinosis is well described to occur among workers in the cotton processing (e.g., yarn preparation) industry . Recent studies have implicated Gram-negative bacterial endotoxin as one of the agents responsible for acute, and possibly chronic, respiratory illness . Laboratory experiments using a model cardroom have found poor correlations between airborne dust and associated endotoxin . This study reports the results of vertical elutriated dust and endotoxin levels in 11 work areas of 2 cotton textile mills in 1986 in Shanghai, China . The overall correlation between dust and endotoxin was strong, rs = 0.66 and 0.79 (p < 0.0001) for mills 1 and 2, respectively . The dust-endotoxin correlation was relatively poor in early yarn preparation in the workshops and improved in the later preparation areas . Our findings suggest that in these mill settings, dust and endotoxin levels may be well correlated in most work areas . Therefore, dust may be a useful index for monitoring populations employed in the cotton textile industry throughout the world . Additional field studies need to be performed which consider the various determinants of dust and endotoxin levels.

J Am Geriatr Soc, 1993 Feb, 41(2), 157 - 62
Percutaneous transhepatic cholecystostomy for acute complicated calculous cholecystitis in elderly patients; Van Steenbergen W et al.; OBJECTIVE: To assess the immediate and long-term outcomes of elderly patients with acute complicated cholecystitis treated by percutaneous cholecystostomy . To assess the results of bile cultures obtained in this group of patients . DESIGN: Case series . SETTING: Tertiary care center . PATIENTS: Thirty-two patients, with a mean (+/- S.D.) age of 78 +/- 8 years (range, 58-92 years), and who presented with acute cholecystitis complicated by empyema formation . Sixty-six percent had associated disorders, which rendered them at high risk for surgical intervention . INTERVENTION: Percutaneous transhepatic catheter drainage of the gallbladder, with a mean drainage time of 20 days (range 0-84 days) . In addition, endoscopic sphincterotomy with removal of common bile duct stones was performed in six patients and percutaneous aspiration of an associated liver abscess in four cases . RESULTS: Percutaneous cholecystostomy was followed by rapid regression of clinical symptoms and of radiologic abnormalities in all patients . Sixteen cases (50%) underwent elective cholecystectomy 1-12 weeks after cholecystostomy . One of them died of aspiration pneumonia, whereas 15 had no post-operative problems and were discharged 9 days (mean) after surgery . Forty-four percent (14/32) were considered inoperable: they remained completely free of biliary symptoms and died of unrelated illness (22%) after a mean follow-up of 6 months (range, 1-22 months) or are still alive (22%) with a mean follow-up of 15 months (range, 5-36 months) . Bile cultures were positive in 75% of the patients . Escherichia coli, other aerobic Gram-negative micro-organisms, and anaerobic bacterial species accounted for 35% (16/46), 28% (13/46), and 20% (9/46) of the isolated bacteria, respectively . All aerobic Gram-negative species tested in vitro were susceptible to gentamicin and to temocillin . CONCLUSIONS: Percutaneous transhepatic cholecystostomy is a safe and effective procedure in the treatment of elderly high-risk patients with acute cholecystitis complicated by empyema formation . It can be followed by elective cholecystectomy, if possible, or by expectant conservative management in patients who are inoperable because of systemic disease.

J Immunol, 1993 Feb 1, 150(3), 1011 - 8
Pertussis toxin-sensitive factor differentially regulates lipopolysaccharide-induced tumor necrosis factor-alpha and nitric oxide production in mouse peritoneal macrophages; Zhang X et al.; It has been established that LPS, the major constituent of the outer membrane of gram negative bacteria, stimulates macrophages to produce numerous inflammatory mediators, including TNF-alpha and nitric oxide (NO) . Both TNF-alpha and NO are important in the macrophage-mediated cytotoxicity against invading microorganisms and tumor cells . Although many LPS-dependent immune responses have been well characterized phenomenologically, the precise signal transduction pathways in LPS-induced macrophage activation are not clear . We reported that 1) pretreatment of C3HeB/FeJ mouse peritoneal macrophages with pertussis toxin (PT) markedly enhanced LPS-induced TNF-alpha production but inhibited LPS-dependent NO production under the same conditions; 2) kinetics of the PT effects on these LPS-responses were correlated with PT-mediated ADP-ribosylation of a 41-kDa protein(s); and 3) PT pretreatment did not correct the refractory states of C3H/HeJ macrophages to wild type smooth-LPS . These results suggest that LPS stimulates TNF-alpha and NO production in mouse peritoneal macrophages through different biochemical pathways, and that the signal transduction for both pathways is regulated by a PT-sensitive factor . It is possible that this factor is a guanine nucleotide-binding regulatory protein(s) . Finally our data indicate that it is unlikely that the defect of the C3H/HeJ macrophages in response to LPS is at the level of this PT-sensitive factor.

J Bacteriol, 1993 Feb, 175(3), 905 - 8
Purification and characterization of SP21, a development-specific protein of the myxobacterium Stigmatella aurantiaca; Heidelbach M et al.; Stigmatella aurantiaca is a gram-negative bacterium with a complex life cycle, including cellular aggregation resulting in the formation of a characteristic three-dimensional structure, the so-called fruiting body . During fruiting and upon chemical induction of sporulation, a major development-specific protein, SP21, is synthesized . SP21 was purified to homogeneity from the membranous fraction of chemically induced spores . Expression of SP21 was studied with an antiserum raised against the purified protein.

Infect Immun, 1993 Feb, 61(2), 486 - 90
Bordetella pertussis and Bordetella parapertussis: two immunologically distinct species; Khelef N et al.; Bordetella pertussis and Bordetella parapertussis are closely related species . Both are responsible for outbreaks of whooping cough in humans and produce similar virulence factors, with the exception of pertussis toxin, specific to B . pertussis . Current pertussis whole-cell vaccine will soon be replaced by acellular vaccines containing major adhesins (filamentous hemagglutinin and pertactin) and major toxin (pertussis toxin) . All of these factors are antigens that stimulate a protective immune response in the murine respiratory model and in clinical assays . In the present study, we examined the protective efficacies of these factors, and that of adenylate cyclase-hemolysin, another B . pertussis toxin, against B . parapertussis infection in a murine respiratory model . As expected, pertussis toxin did not protect against B . parapertussis infection, since this bacterium did not express this protein, but the surprising result was that none of the other factors were protective against B . parapertussis infection . Furthermore, B . parapertussis adenylate cyclase-hemolysin, although it protected against B . parapertussis infection, did not protect against B . pertussis infection . Despite a high degree of homology between both B . pertussis and B . parapertussis species, no cross-protection was observed . Our results outline the fact that, as in other gram-negative bacteria, Bordetella surface proteins vary immunologically.

J Infect Dis, 1993 Feb, 167(2), 378 - 84
Therapeutic trial of lipid X in a canine model of septic shock; Danner RL et al.; Three groups of dogs were given lipid X (0, 1, or 10 mg/kg) every 8 h for for seven doses, starting simultaneously with the intraperitoneal placement of Escherichia coli-containing fibrin clots . All animals developed bacteremia, hypotension, and a pattern of decreased left ventricular ejection fraction characteristic of septic shock (P = .01) . Survival rates and survival times were not significantly different between treatment groups (P > .2) . In a similar experiment, higher doses of lipid X resulted in a significantly decreased survival time compared with concurrent controls (P = .04) . Animals receiving lipid X did not differ from controls in serial determinations of temperature, hemodynamic measurements, or laboratory parameters (except serum total protein) . Although lipid X has antiendotoxin effects, no benefit could be demonstrated in this antibiotic-treated, gram-negative bacillary-infected model of septic shock . These data do not support a therapeutic role for lipid X in the treatment of gram-negative sepsis.

Mol Microbiol, 1993 Feb, 7(3), 471 - 88
Light-induced carotenogenesis in Myxococcus xanthus: genetic analysis of the carR region; Hodgson DA; Carotenogenesis is light-inducible in the non-photosynthetic, Gram-negative, bacterium Myxococcus xanthus . We report the characterization of the carR region which controls this phenomenon . Insertion of transposon Tn5 close to the carR region caused a dominant, carotenoid-constitutive mutation because of the presence of a constitutive, outward-reading promoter in the IS50L component of Tn5 . In wild-type cells, a powerful, tightly-regulated, light-inducible promoter directs the transcription of two genetic functions . One of these functions is to activate transcription of the genetically unlinked carB gene, which is involved in carotenoid synthesis . The second function (carR) regulates the light-inducible promoter . We also report the mapping of two carotenoid constitutive mutations to the previously characterized carA locus.

J Bacteriol, 1993 Feb, 175(4), 1144 - 52
Isolation and characterization of Bordetella bronchiseptica mutants deficient in siderophore activity; Armstrong SK et al.; Iron acquisition by the gram-negative pathogens Bordetella bronchiseptica and Bordetella pertussis is thought to occur by hydroxamate siderophore-mediated transport as well as an apparently siderophore-independent process by which host transferrins bind to bacterial surface receptors . We constructed B . bronchiseptica mutants deficient in siderophore activity by insertional mutagenesis with miniTn5/lacZ1 . The mutants could be placed into four distinct complementation groups, as determined from cross-feeding assays which demonstrated restored siderophore synthesis . Mutants deficient in siderophore activity were BRM1, BRM6, and BRM9, exhibiting approximately 36 to 41% of wild-type siderophore levels, and BRM3 and BRM8, which appeared to produce very little or no detectable siderophore . Mutant BRM4 was found to be a leucine auxotroph, while mutants BRM2 and BRM7 could synthesize siderophore only in low-iron medium which was supplemented with various amino acids . Evaluation of all transcriptional fusions revealed an apparent lack of iron-regulated lacZ expression . Genomic regions flanking the transposable element in the siderophore mutants were homologous with B . pertussis chromosomal DNA, while bioassays suggested siderophore cross-feeding between B . pertussis and B . bronchiseptica . These results indicate probable similarity between the siderophore biosynthetic and transport systems of the two species.

Am J Respir Cell Mol Biol, 1993 Feb, 8(2), 134 - 44
Activated pulmonary vascular neutrophils as early mediators of endotoxin-induced lung inflammation; Williams JH Jr et al.; Pulmonary vascular sequestration of leukocytes has been reported to increase in some models of lung injury, including that induced by gram-negative bacterial lipopolysaccharide (LPS) . Neutrophils recruited to the lung likely participate in LPS-induced lung inflammation and associated injury, but the functional activities of these pulmonary vascular neutrophils have not been directly assessed . In the current study, cells were recovered by pulmonary vascular lavage (PVL) of isolated rat lungs, harvested 2 h after intravenous infusion of LPS (3 mg/kg) or saline in intact rats, at which time LPS-induced neutrophil recruitment to the lung could be appreciated histologically but not by airway lavage . Relative concentrations of leukocytes recovered from the pulmonary vasculature by PVL were compared with those present in circulating blood, normalizing for lavage dilution on the basis of erythrocyte counts . Excess neutrophils, lymphocytes, monocytes, and eosinophils were recovered from the pulmonary vasculature of controls, and LPS infusion increased recovery of neutrophils (most prominently), lymphocytes, and monocytes . Compared with cells recovered from controls, PVL neutrophils from LPS-infused animals were primed for increased zymosan-stimulated superoxide generation, determined by ferricytochrome C reduction, and were more adherent to nylon wool columns . Northern blots of extracted RNA demonstrated that LPS infusion also upregulated interleukin-1 beta (IL-1 beta) mRNA expression in PVL leukocyte samples, but not BAL or circulating blood samples . Ficoll-hypaque separation demonstrated that the LPS-induced IL-1 beta signal in PVL leukocytes was derived primarily from polymorphonuclear rather than mononuclear leukocytes . In conclusion, all circulating leukocyte populations are sequestered in rat lungs, and LPS increases pulmonary vascular sequestration of leukocytes, recruiting most prominently an activated pool of neutrophils that are more adherent, primed for increased oxygen radical production, and expressing increased IL-1 beta message . These findings suggest a more prominent role than previously appreciated for sequestered neutrophils in sepsis-induced lung inflammation.

Infect Agents Dis, 1993 Feb, 2(1), 44 - 52
Antiendotoxin therapies for septic shock; Corriveau CC et al.; Gram-negative shock is thought to result primarily from the effects of endotoxin, a component of the bacterial outer membrane . Accordingly, therapies aimed at inhibiting, neutralizing, or clearing endotoxin have been extensively explored . Despite over 30 years of research, no antiendotoxin approach to the treatment of human septic shock is of proven benefit . In recent randomized clinical trials of monoclonal antibodies against endotoxin, therapeutic efficacy was not convincingly demonstrated . This result, however, does not eliminate the possibility that other antiendotoxin therapies may be effective . The antibodies used in these clinical trials do not appear to neutralize endotoxin in vitro and are not reproducibly protective in animal models of sepsis . Newer agents with well-defined mechanisms of antiendotoxin activity may help clarify the role of endotoxin in septic shock and prove useful therapy for some patients.

New Horiz, 1993 Feb, 1(1), 110 - 9
Anti-endotoxin antibodies and other inhibitors of endotoxin; Zanetti G et al.; Neutralization of endotoxin (lipopolysaccharide) would be of considerable benefit in the treatment of Gram-negative sepsis . Administration of anti-lipopolysaccharide antibodies is an old approach that has been renewed by improvements in monoclonal antibody technology . Experimental and clinical studies of antibodies directed at the conserved core region of lipopolysaccharide or at the lipid A are discussed . Some of these antibodies appear to be protective in animal models or in clinical trials, but discrepant results have been reported and the mechanism of the postulated protection was not clarified . Despite the availability of a monoclonal anti-lipid A antibody on the market in some European countries, this type of treatment should still be considered of unclear value until further experimental and clinical studies have reinvestigated the protection afforded by these antibodies . The use of detoxified lipid A analogs with lipopolysaccharide antagonist properties is another promising strategy that is discussed briefly in this review . Recently, substantial progress has been made in understanding how lipopolysaccharide triggers the immune system . A family of proteins possessing lipopolysaccharide-binding sites has been discovered . These proteins have a striking homology in DNA sequence, but they have different functions . The biological role of these proteins is now being actively investigated . New strategies to improve the outcome of Gram-negative sepsis may result from this research.

Infect Immun, 1993 Feb, 61(2), 378 - 83
Purification and characterization of murine lipopolysaccharide-binding protein; Gallay P et al.; The serum protein lipopolysaccharide (LPS)-binding protein (LBP) seems to play an important role in regulating host responses to LPS . Complexes of LPS and LBP form in serum and stimulate monocytes, macrophages, or polymorphonuclear leukocytes after binding to CD14 . Previous reports have described the structure and properties of LBP from human and rabbit sera . Since mice are used in some experimental models of endotoxemia or gram-negative bacterial infections, information is needed about the properties of murine LBP . Murine LBP was purified by ion-exchange chromatography and high-pressure liquid chromatography; its NH2-terminal sequence (TNPGLVTRIT) was very similar to those of human and rabbit LBPs (80 to 90% amino acid identity) . Murine LBP resembled LBPs from other species in that it promoted the binding of LPS to monocytes and enhanced the sensitivity of monocytes to LPS at least 100-fold . Mouse LBP, like rabbit and human LBPs, was found to be an acute-phase protein . Further in vivo studies with mice and anti-CD14 or anti-LBP reagents should help determine the role of LBP in response to LPS challenges.

Biochem Biophys Res Commun, 1993 Jan 29, 190(2), 624 - 9
Single channel behavior of matrix porin of Escherichia coli; Buehler LK et al.; Porins are trimeric proteins in the outer membranes of Gram-negative bacteria . Several of them, among them matrix porin of Escherichia coli, form symmetrically voltage-gated ion channels in planar bilayers . Trimers exhibit negative resistance at potentials larger than +/- 90mV . Here we show that, after two pores within a trimer close irreversibly, the remaining third pore shows channel properties distinct from those observed in the trimer . This residual pore exhibits an asymmetric current-voltage dependence with a pronounced polarity-dependent shift toward low potentials and rates of channel-closing and opening that are one to two orders of magnitude faster than those observed for single channels in a reversibly voltage-dependent trimer . Rectification of single channels thus resembles that of a voltage-gated channel type observed in outer membrane patches of E.coli spheroblasts, hinting at the relevance of the phenomenon in vivo.

Med Clin (Barc), 1993 Jan 23, 100(3), 84 - 9
{A cost-efficacy analysis of treatment with antiendotoxin monoclonal antibodies in gram-negative sepsis}; Badia X et al.; BACKGROUND: The aim of this study was to determine the cost-efficacy analysis of treatment with antiendotoxin monoclonal antibodies in adult patients admitted to intensive care units for completing the decision making regulating the authorization of registry of drugs in Spain . METHODS: Two treatment strategies were considered: to treat with antiendotoxin monoclonal antibodies, addition to conventional treatment, to all patients with sepsis or to all those with septic shock versus the alternative of only carrying out conventional treatment . The economic evaluation technique which quantifies costs in pesetas and efficacy in years of life gained by the treatment alternatives was considered . RESULTS: The cost per year of additional life gained from treatment to all patients with sepsis was higher (859, 288 pesetas per year of life gained) than that in patients treated for septic shock (293, 810 pesetas per year of life gained) . The result was very sensitive to changes in those expected in survival of the patients treated . CONCLUSIONS: In agreement with this cost efficacy analysis the hypothesis to authorize drug registry in Spain for the indication of septic shock is reinforced . The results may vary upon the obtaining of more clinical and epidemiologic information of the treatment.

Science, 1993 Jan 15, 259(5093), 361 - 5
Molecular mapping and detoxification of the lipid A binding site by synthetic peptides; Rustici A et al.; Endotoxin {lipopolysaccharide (LPS)}, the major antigen of the outer membrane of Gram-negative bacteria, consists of a variable-size carbohydrate chain that is covalently linked to N,O-acylated beta-1,6-D-glucosamine disaccharide 1,4'-bisphosphate (lipid A) . The toxic activity of LPS resides in the lipid A structure . The structural features of synthetic peptides that bind to lipid A with high affinity, detoxify LPS in vitro, and prevent LPS-induced cytokine release and lethality in vivo were defined . The binding thermodynamics were comparable to that of an antigen-antibody reaction . Such synthetic peptides may provide a strategy for prophylaxis and treatment of LPS-mediated diseases.

JAMA, 1993 Jan 13, 269(2), 249 - 54
Cost-effectiveness of monoclonal antibodies to gram-negative endotoxin in the treatment of gram-negative sepsis in ICU patients; Chalfin DB et al.; OBJECTIVE--To evaluate the fiscal impact and the cost-effectiveness of monoclonal antibodies against gram-negative endotoxin (MAbGNE) in the treatment of presumed gram-negative sepsis . DESIGN--A decision analysis model was developed from (1) data from two phase III trials that studied the E5 or HA-1A MAbGNE, and (2) financial data from 1405 septic patients who required intensive care at a large tertiary hospital . SETTING--Intensive care unit (ICU) patients with presumed gram-negative sepsis . PATIENTS--The E5 trial evaluated 468 patients, and the HA-1A study enrolled 543 patients with presumed gram-negative sepsis . INTERVENTIONS--The addition of MAbGNE to standard regimens or standard regimens alone . MAIN OUTCOME MEASURES--Total expected charges and the expected probability of survival were determined for each option . Cost-effectiveness and marginal cost-effectiveness ratios were also derived . Multiple sensitivity and Monte Carlo analyses were performed to test the underlying assumptions . RESULTS--MAbGNE therapy always resulted in higher expected charges; however, these differences were less than its acquisition cost by $870 . The cost-effectiveness ratio for MAbGNE, for $2000 and $4000 acquisition costs, was $71,674 and $74,900 per probability of survival, respectively . Sensitivity analysis showed that cost-effectiveness was most affected by diagnostic accuracy, patient selection, and acquisition cost . Monte Carlo analysis showed that MAbGNE was more costly for 71% of simulations, yet the most efficacious option for 79% of simulations . CONCLUSIONS--From the perspective of acute care institutions, MAbGNE is expensive and cannot be justified on a cost-saving basis . However, it may be cost-effective throughout a reasonable range of assumptions.

Br J Hosp Med, 1993 Jan 20-Feb 2, 49(2), 121 - 2
Centoxin; Davidson J et al.; Centoxin is one of a new group of immunotherapeutic agents manufactured from 'humanized' antibodies produced using monoclonal antibody technology . It is indicated in the management of patients with Gram-negative septicaemia; its introduction represents a new approach to treatment in these patients (Luce, 1987).

Ann Ital Med Int, 1993 Jan-Mar, 8(1), 13 - 7
{Spontaneous bacterial peritonitis during liver cirrhosis: the clinical findings and therapeutic considerations}; Fiaccadori F et al.; Among 1211 consecutive patients admitted to hospital for liver cirrhosis, 625 (51.6%) had ascites . Forty-four of them (7%) had ascitic infection . Thirty-four cases (5.4%) of spontaneous bacterial peritonitis (SBP) and 10 cases (1.6%) of culture-negative neutrocytic ascites (CNNA) were diagnosed . The infecting organism was most likely Gram-negative of enteric origin (80%), CNNA mortality (30%) was lower than that of SBP (47%) . High mortality suggests to treat patients affected by either SBP or CNNA with antibiotics.

Eur Surg Res, 1993 Jan-Feb, 25(1), 1 - 10
The synergistic effect of trauma and infection on interleukin-1 but not tumor necrosis factor liberation during posttraumatic gram-negative septicemia; Rokke O et al.; The release profiles of interleukin 1 (Il-1) and tumor necrosis factor (TNF) were studied during experimental Escherichia coli septicemia and peritonitis with and without a preceding (-48 h) moderate trauma (femur marrow nailing) . The trauma alone did not induce significant Il-1 or TNF liberation to plasma . During septicemia, a rapid IL-1-rise (+30 min) and subsequent normalization (+120 min) was seen . A previous trauma delayed (30 min), but accentuated (2x) and prolonged (> 4 h) the Il-1 response . During peritonitis, a delayed (30 min) but otherwise similar Il-1-response was observed . TNF levels rose rapidly (+30 min) in all animals, and remained high throughout the experimental period (6 h) . The trauma did not influence the TNF response . We conclude that a synergism exists between trauma and infection with regard to the magnitude of the Il-1 response.

Rev Prat, 1993 Jan 1, 43(1), 19 - 25
{Other mediators of inflammation and sepsis}; Lamy M et al.; During sepsis, the systemic inflammatory response is characterized by the release of numerous mediators supporting and dispersing inflammation . In Gram negative sepsis, the endotoxins play a starting role, while in other sepsis, the triggering mediators or mechanisms are unknown but lead to a similar inflammatory reaction . Coagulation and complement cascades are activated, with the release of chemoattractive substances, mediators and proteases and the activation of phagocytic cells . Macrophages/monocytes and polymorphonuclear leucocytes produce then active oxygen species and cytokines; they degranulate (releasing active enzymes such as myeloperoxidase), they express an increasing number of membrane receptors able to interact with endothelial cells and release a supplementary lot of inflammatory mediators (prostanoids, platelet activating factor, leukotrienes .. . ) . Platelets, also activated, produce the same mediators (TXA2, PAF ...) or specific ones such as serotonine, platelet factor 4, growth factors . Last, but not least, the endothelial cells are stimulated, directly (by endotoxins) or undirectly (by cytokines, C5a, PAF ...) . These cells play then a main role by their own phagocytic activity, by alteration of their antithrombotic and fibrinolytic potential, by their secretion of inflammatory mediators and by an increased expression of receptors of adhesivity for the activated phagocytes or platelets, what leads to endothelium injury with membrane permeability alterations . These cascades of activation, these extensions of the inflammatory reaction by the mediators and by the phagocytes and platelets can explain the frequency of multiple organ failure during sepsis as well as the difficulty of an adequate pharmacological therapy.

J Vet Diagn Invest, 1993 Jan, 5(1), 47 - 51
Preliminary characterization of a pleomorphic gram-negative rod associated with avian respiratory disease; Charlton BR et al.; An unidentified, pleomorphic, gram-negative rod (PGNR) bacterium has been isolated from domestic fowl with respiratory disease . The PGNR was isolated in 5% of turkey accessions and 3% of chicken accessions, primarily from the respiratory tract . Preliminary characterization of this organism included reviewing accession records, conducting cultural and biochemical tests, and analyzing cellular fatty acids . The PGNR was also compared with other bacteria capable of inhabiting the avian respiratory system . Biochemical and cellular fatty acid analysis failed to identify the organism, however all 14 isolates were similar.

J Clin Pediatr Dent, 1993 Winter, 17(2), 99 - 104
Papillon Lefevre syndrome: treatment of two cases with a clinical microbiological and histopathological investigation; Eronat N et al.; Papillon Lefevre syndrome is presented in the cases of two female patients of the ages of 7 and 9, who exhibited all typical symptoms of the disease . Microbiological and histopathological studies were done and treatment provided . Actinobacillus actinomycetemcomitans, which is suspected as a pathogenic factor in the disease was identified as well as some other gram negative microorganisms and an antibiogram was performed in which amoxycillin plus clavulanic acid was most effective . Histopathological investigation also confirmed the presence of gram negative bacteria . Granular cell infiltration was predominant in the surface epithelium . Prosthetic appliances were provided for the patients after mechanical and chemical plaque control . In addition to this, antibiotics (amoxycillin plus clavulanic acid) were prescribed every six months . No tooth loss was observed in both patients after more than two years follow-up period . At the moment only one patient is under review and because she is uncooperative, mild periodontal inflammation is still present around the teeth which erupted before the antibiotic regime, but not in the other teeth.

Mol Microbiol, 1993 Jan, 7(2), 289 - 98
The origin of greater-than-unit-length plasmids generated during bacterial conjugation; Erickson MJ et al.; In Gram-negative bacteria, the general mechanism of conjugal plasmid transfer, which is probably similar for many different groups of plasmids, involves the transfer of a single plasmid DNA strand with 5' to 3' polarity . Transfer is initiated by nicking of the duplex DNA at a particular site, i.e . the origin of transfer (oriT) . We constructed plasmids containing two directly repeated copies of oriT, derived from the broad-host-range plasmid R1162 and flanking the lac operator . The number of lacO copies in the plasmid after transfer could be determined from the colour of transconjugant colonies on medium containing X-Gal . When the oriTs were mutated to prevent initiation and termination of transfer at the same oriT, almost all of the transconjugant cells contained greater-than-unit-length plasmids with two copies of lacO and three copies of oriT . We show that these molecules were generated by an intramolecular, conjugation-dependent mechanism unlikely to depend solely on a pre-existing population of circular or linear multimers in donor cells . We propose that the greater-than-unit-length molecules were instead generated by a rolling-circle mechanism of DNA replication.

Ophthal Plast Reconstr Surg, 1993, 9(1), 38 - 41; discussion 42
Management of acute dacryocystitis in adults; Cahill KV et al.; Acute dacryocystitis frequently is extremely painful and slow to resolve even with systemic antibiotic therapy . We have identified that incision, drainage, and direct application of antibiotics inside the infected sac result in almost immediate resolution of pain and rapid control of infection . This also provides optimal culture material . Twelve consecutive patients treated in this manner had rapid control of the acute infectious process . All eight patients subsequently undergoing dacryocystorhinostomy were fully cured . A total of 58.3% of the patients were infected with gram-negative rods; 50% of the isolates were resistant to most oral antibiotics.

Trends Biochem Sci, 1993 Jan, 18(1), 7 - 12
Membrane traffic wardens and protein secretion in gram-negative bacteria; Salmond GP et al.; Recent progress in the genetic analysis of protein secretion in diverse Gram-negative bacteria has revealed three major, highly conserved but functionally independent pathways that involve accessory apparatus proteins . Protein secretion via the Type I pathway is signal sequence-independent with no free periplasmic intermediate . Secretion by the Type II pathway is signal sequence-dependent and via the periplasm . Recent results also suggest that a third (Type III) secretory pathway exists in which protein secretion is signal sequence-independent.

J Cell Biochem, 1993 Jan, 51(1), 69 - 74
Signal transduction in chemotaxis mediated by the bacterial phosphotransferase system; Titgemeyer F; Gram-negative bacteria are able to respond chemotactically to carbohydrates which are substrates of the bacterial phosphoenolpyruvate:sugar phosphotransferase system (PTS) . The mechanism of signal transduction in PTS-mediated chemotaxis is different from the well-studied mechanism involving methyl-accepting chemotaxis proteins (MCPs) . In PTS-mediated chemotaxis, carbohydrate transport is required, and phosphorylation seems to be involved in both excitation and adaptation . In this review the roles of the components of the PTS in chemotactic signal transduction are discussed.

J Cell Biochem, 1993 Jan, 51(1), 62 - 8
Regulatory interactions involving the proteins of the phosphotransferase system in enteric bacteria; Saier MH Jr; Sugar uptake and cytoplasmic inducer generation as well as cyclic AMP synthesis are regulated by the phosphoenolpyruvate:sugar phosphotransferase system (PTS) in Gram-negative enteric bacteria . In these organisms, the free form of the glucose-specific Enzyme IIA (IIAglc) of the PTS, which can be phosphorylated on a histidyl residue by PEP and the PTS energy coupling proteins, inhibits the activities of non-PTS carbohydrate permeases and catabolic enzymes . By contrast, the phosphorylated form of IIAglc appears to activate adenylate cyclase, the cyclic AMP biosynthetic enzyme . What is known of the molecular details of these regulatory interactions will be summarized, and a novel regulatory mechanism involving the fructose repressor, FruR, which controls the transcription of genes encoding enzymes which catalyze reactions in central pathways of carbon metabolism, will be presented.

Genomics, 1993 Jan, 15(1), 188 - 90
The genes for the lipopolysaccharide binding protein (LBP) and the bactericidal permeability increasing protein (BPI) are encoded in the same region of human chromosome 20; Gray PW et al.; The lipopolysaccharide binding protein is an acute-phase reactant produced during gram-negative bacterial infections . The bactericidal/permeability increasing protein is associated with human neutrophil granules and has bactericidal activity on gram-negative organisms . In addition to their functional relationship, both proteins share extensive structural similarity . This article demonstrates that the genes for both proteins are in the same region of human chromosome 20, between q11.23 and q12.

Ann Pharmacother, 1993 Jan, 27(1), 36 - 7
Ceftriaxone-induced acute pancreatitis; Zimmermann AE et al.; OBJECTIVE: To report a case probable ceftriaxone-induced acute pancreatitis . CASE SUMMARY: A patient with a history of short-bowel syndrome on home total parenteral nutrition developed fever, chills, and right flank pain . She was diagnosed with gram-negative catheter sepsis and prescribed antibiotic therapy to be administered for four weeks . After completion of the first week of therapy, the antibiotic regimen was changed to intravenous injections of ceftriaxone to be given daily at home . Prior to discharge the patient developed acute abdominal pain, leukocytosis, jaundice, and markedly elevated lipase and amylase concentrations consistent with acute pancreatitis . The patient's condition improved upon discontinuation of the ceftriaxone and the remainder of her stay was uneventful . DISCUSSION: There is only one other case report in the literature of probable ceftriaxone-induced pancreatitis . Multiple other medications have been implicated in causing acute pancreatitis . The exact mechanism of this uncommon adverse effect of ceftriaxone is unknown . CONCLUSIONS: There was a temporal relationship between the development of this patient's signs and symptoms and the administration of ceftriaxone . We could not identify any other factors that may have been responsible for the development of her acute pancreatitis . Ceftriaxone should be considered as a possible etiologic agent in patients who present with acute abdominal pain and elevated lipase and amylase concentrations.

Hum Antibodies Hybridomas, 1993 Jan, 4(1), 36 - 9
Pilot study of anti-lipopolysaccharide human monoclonal antibody MAB-T88 in patients with gram-negative sepsis; Daifuku R et al.; MAB-T88 is a human monoclonal IgM antibody directed at the lipopolysaccharide of gram-negative bacteria . A protocol was designed to identify a group of septic patients with a very high likelihood of gram-negative bacteremia . All 6 patients entered in the protocol had a gram-negative source, and 4 of 6 had gram-negative bacteremia . In this patient population, MAB-T88 was shown to be safe with an effective half-life of 19.1 hours.

Am J Physiol, 1993 Jan, 264(1 Pt 2), R90 - 6
Effect of fibronectin on permeability of normal and TNF-treated lung endothelial cell monolayers; Wheatley EM et al.; Fibronectin is found in a soluble form in plasma and lymph and in an insoluble form in the extracellular matrix . Plasma fibronectin can incorporate into the tissue pool of fibronectin where its adhesive properties may influence cell-cell interaction, cell adhesion to a collagenous matrix, and vascular integrity . Elevation of plasma fibronectin can attenuate the increase in lung vascular permeability in sheep during postoperative gram-negative bacteremia, and plasma fibronectin deficiency can magnify the increase in lung vascular permeability with postoperative sepsis . Using pulmonary endothelial monolayers, we determined if exogenous human plasma fibronectin (pFn) would influence the protein permeability of pulmonary endothelial monolayers as determined by transendothelial clearance (microliters/min) of 125I-albumin after they were exposed to human recombinant tumor necrosis factor-alpha . Treatment of endothelial monolayers with tumor necrosis factor (TNF) (200 U/ml) for 18 h resulted in a significant (P < 0.05) increase in protein permeability . Addition of intact purified human plasma fibronectin to normal confluent endothelial monolayers to yield a medium concentration of 300, 600, and 900 micrograms/ml for 18 h had no effect on baseline protein permeability . In contrast, whereas addition of lower amounts of human plasma fibronectin (300 micrograms/ml) did not attenuate the TNF-induced increase in monolayer permeability, the higher concentrations of 600 or 900 micrograms pFn/ml significantly decreased (P < 0.05) protein permeability . The ability of soluble plasma fibronectin to attenuate the TNF-induced increase in endothelial protein permeability required an incubation time of at least 2-3 h, perhaps due to a lag time required for its incorporation into the extracellular matrix.(ABSTRACT TRUNCATED AT 250 WORDS)

Int J Syst Bacteriol, 1993 Jan, 43(1), 135 - 42
Thauera selenatis gen . nov., sp . nov., a member of the beta subclass of Proteobacteria with a novel type of anaerobic respiration; Macy JM et al.; A recently isolated, selenate-respiring microorganism (strain AXT {T = type strain}) was classified by using a polyphasic approach in which both genotypic and phenotypic characteristics were determined . Strain AXT is a motile, gram-negative, rod-shaped organism with a single polar flagellum . On the basis of phenotypic characteristics, this organism can be classified as a Pseudomonas sp . However, a comparison of the 16S rRNA sequence of strain AXT with the sequences of other organisms indicated that strain AXT is most similar to members of the beta subclass (level of similarity, 86.8%) rather than to members of the gamma subclass (level of similarity, 80.2%) of the Proteobacteria . The presence of the specific polyamine 2-hydroxyputrescine and the presence of a ubiquinone with eight isoprenoid units in the side chain (ubiquinone Q-8) excluded strain AXT from the authentic genus Pseudomonas and allowed placement in the beta subclass of the Proteobacteria . Within the beta subclass, strain AXT is related to Iodobacter fluvatile . The phylogenetic distance (level of similarity, less than 90%), as well as a lack of common phenotypic characteristics between these organisms, prevents classification of strain AXT as a member of the genus Iodobacter . In addition, strain AXT possesses a unique mechanism for anaerobic respiration, which allows it to utilize selenate as an electron acceptor without interference by nitrate . Therefore, we propose that strain AXT should be the first member of a new genus and species, Thauera selenatis.

J Am Osteopath Assoc, 1993 Jan, 93(1), 87 - 91
Demonstration of Helicobacter pylori in tracheal secretions; Mitz HS et al.; A new gram-negative bacterium, Helicobacter pylori, has been found in gastric secretions . In view of an almost 9% incidence of unidentified gram-negative bacteria in aspiration pneumonia, the authors set out to prove that H pylori could be found in tracheal secretions . Eighteen sequential patients admitted to the intensive care unit who had endotracheal or nasogastric intubation for 24 hours or longer were studied . Of 20 sets of specimens from 18 patients . Helicobacter was recovered from 2(10%) of the endotracheal specimens . The authors conclude that H pylori can gain access to the endobronchial tree and therefore may contribute to the origin of pneumonia due to unidentified gram-negative bacteria.

South Med J, 1993 Jan, 86(1), 46 - 51
Aminoglycoside dosing: a randomized prospective study; Kemme DJ et al.; Although clinical benefits for aminoglycoside dosing services have been suggested, this has not been clearly documented in a prospective fashion . Therefore, we randomly assigned patients to be dosed (1) by their physician (Physician dosing method), (2) by predicting an initial dosage (Predictive dosing method), or (3) by calculating an initial dosing regimen by measuring the pharmacokinetics for the individual patients after a loading dose (Individual dosing method) . The patients' clinical response and nephrotoxicity were then evaluated . The individual dosing method resulted in erratic aminoglycoside levels, necessitating its elimination from the study . This group was not included in the final analysis . Of the 164 patients entering the study, 41 had a documented gram-negative infection, received aminoglycosides for more than 2 days, and had serum aminoglycoside levels measured . The predictive dosing method in these 41 patients produced statistically significant higher peak and lower trough levels, but there was no difference in the incidence of nephrotoxicity or clinical response . The 95% confidence intervals precluded any major clinical benefit in these patients with documented gram-negative infections . We question the previous findings of increased efficacy and decreased nephrotoxicity with the use of an aminoglycoside dosing service and suggest that larger studies be done.

Arch Surg, 1993 Jan, 128(1), 47 - 53; discussion 53-4
Insulinlike growth factor 1 (IGF-1) reduces gut atrophy and bacterial translocation after severe burn injury; Huang KF et al.; Bacterial translocation after severe burns is associated with gut mucosal atrophy and increased mucosal permeability . Insulinlike growth factor 1 (IGF-1) levels are low after trauma and do not respond to growth hormone treatment . Since IGF-1 receptors have been demonstrated in gut mucosa, we proposed that treatment with IGF-1 would reduce mucosal atrophy and bacterial translocation . Rats received 50% total body surface area full-thickness burn or sham burn . They were treated with a continuous, subcutaneous infusion of either IGF-1 (approximately 3 mg/kg per day) or placebo (0.01 mol of acetate) for up to 5 days after receiving the burn . The mesenteric lymph node and liver were cultured for gram-negative bacteria . The small intestinal mucosa was scraped, weighed, and analyzed for DNA and protein content . Treatment with IGF-1 improved body weight, spleen weight, and gut mucosal weight . It stimulated mucosal DNA and protein content and reduced the incidence of bacterial translocation to the mesenteric lymph node from 89% to 30% . Insulinlike growth factor may reduce gut barrier failure by decreasing mucosal atrophy and subsequent barrier failure . In addition to its general anabolic effects, recombinant human IGF-1 may improve gut mucosal function and reduce infectious morbidity in severely traumatized or septic patients by reducing gut atrophy and reducing bacterial translocation.

J Exp Med, 1993 Jan 1, 177(1), 89 - 97
Assessment of ability of murine and human anti-lipid A monoclonal antibodies to bind and neutralize lipopolysaccharide; Warren HS et al.; The use of monoclonal antibodies (mAbs) directed to lipid A for the therapy of gram-negative sepsis is controversial . In an attempt to understand their biologic basis of action, we used a fluid-phase radioimmunoassay to measure binding between bacterial lipopolysaccharide (LPS) and two IgM mAbs directed to lipid A that are being evaluated for the treatment of gram-negative bacterial sepsis . Both antibodies bound 3H-LPS prepared from multiple strains of gram-negative bacteria when large excesses of antibody were used, although binding was modest and only slightly greater than control preparations . We also studied the ability of each anti-lipid A antibody to neutralize some of the biological effects of LPS in vitro . Despite large molar excesses, neither antibody neutralized LPS as assessed by the limulus lysate test, by a mitogenic assay for murine splenocytes, or by the production of cytokines interleukin (IL)-1, IL-6, or tumor necrosis factor from human monocytes in culture medium or in whole blood . Our experiments do not support the hypothesis that either of these anti-lipid A mAbs function by neutralizing the toxic effects of LPS.

J Antimicrob Chemother, 1993 Jan, 31(1), 89 - 103
Activity of 13 beta-lactam agents combined with BRL 42715 against beta-lactamase producing gram-negative bacteria compared to combinations with clavulanic acid, tazobactam and sulbactam; Piddock LJ et al.; The activity of six cephalosporins, six penicillins and one monobactam combined with BRL 42715, clavulanic acid, sulbactam or tazobactam at 0.1, 0.5, 1, 2, 5 and 10 mg/L was determined for 45 beta-lactamase producing Gram-negative bacteria . The combination of BRL 42715 with any of the agents was more active than any of the other inhibitor and beta-lactam combinations . Unlike the other beta-lactamase inhibitors, BRL 42715 enhanced the activity of the beta-lactams for strains that constitutively expressed Richmond & Sykes Class I beta-lactamase and against strains expressing extended-spectrum plasmid-mediated beta-lactamases.

J Periodontal Res, 1993 Jan, 28(1), 1 - 9
Monoclonal antibodies to lipopolysaccharide of four oral bacteria associated with periodontal disease; Shelburne CE et al.; Periodontal disease is a common inflammatory disease which erodes the supporting structures of the teeth, and is initiated by a subgingival infection with selected Gram-negative bacteria . Monoclonal antibodies (mAb) to lipopolysaccharide (LPS) of four periodontal pathogens, A . actinomycetemcomitans, P . intermedia, F . nucleatum and P . gingivalis were examined for specificity and their ability to bind these pathogens in a particle concentration fluorescence immunoassay (PCFIA) . The mAb selected were specific for their homologous bacteria and when tested against a large battery of other bacteria, including 16 genera and 46 species, were found not to cross-react with heterologous species . When each of the mAb was challenged with 40 or more homologous freshly isolated bacteria, more than 90% were positive . Non-cellular antigens in the form of soluble LPS and extracellular vesicles were examined for their ability to bind to assay components and alter the apparent results of the assay . LPS was found to have potential as an interfering agent if bound to assay components prior to sample treatment, but this non-specific binding was significantly reduced when a surfactant was added to the buffers . Extracellular vesicles had no significant effect on the estimation of P . gingivalis by the assay.

Life Sci, 1993, 53(13), 1099 - 104
Reduced incidence of stress ulcer in germ-free Sprague Dawley rats; Pare WP et al.; Recent findings with respect to the role of spiral gram-negative bacteria in peptic ulcer disease have stimulated interest in discerning the role of these agents in stress ulcer disease . We tested the hypothesis that a standard restraint-cold ulcerogenic procedure would fail to produce ulcers in axenic rats . Axenic, as well as normal Sprague Dawley rats, were exposed to a cold-restraint procedure . The germ-free condition was maintained throughout the study in the axenic rats . Axenic rats had significantly fewer ulcers as compared to normal rats exposed to the standard cold-restraint procedure, as well as handling control rats . The data represent the first report suggesting a microbiologic component in the development of stress ulcer using the rat model.

Agents Actions Suppl, 1993, 42, 179 - 93
Cytokine regulation of endothelial cell extracellular proteolysis; Niedbala MJ; The vascular endothelium plays a central role in the regulation of extrinsic fibrinolysis and thus maintains vascular patency through clot dissolution . Plasminogen activation provides an important source of localized proteolytic activity not only during fibrinolysis but also during a variety of other physiological and pathological processes . Numerous studies have indicated that human endothelial cells can directly synthesize and secrete plasminogen activators (PA) and inhibitors of these activators . PAs specifically hydrolyse a single arginine-valine bond in plasminogen, an abundant and widely distributed plasma zymogen, to form the broad spectrum serine protease, plasmin . Tissue type-PA (t-PA) and urokinase type PA (u-PA) forms of PA have been described in endothelial cells, although t-PA production and secretion is elevated most frequently . The tPA form of PA functions predominantly in endothelial cell mediated fibrinolysis, while uPA is involved in tissue remodeling . During inflammatory reactions activated mononuclear phagocytes produce a variety of cytokines which may influence the phenotype of the endothelium through a process termed "endothelial cell activation" . Tumor necrosis factor alpha (TNF alpha), a mononuclear cytokine, is a distinct polypeptide of Mr 17,000 and has been implicated as a mediator of gram negative induced sepsis as well as angiogenesis . TNF alpha is known to interact with specific endothelial cell receptors and to alter endothelial coagulant and anticoagulant properties implying that cytokines may be potent modulators of hemostasis . Recent observations have indicated that TNF alpha and lymphotoxin (TNF beta) can promote the expression, synthesis and secretion of urokinase plasminogen activator (uPA) in human endothelial cells . The upregulation of uPA results in an alteration in the fibrinolytic capacity of endothelial cells and allows cells the selective ability to degrade and invade underlying subendothelial extracellular matrix (ECM) . Endothelial cells treated with TNF alpha also display, in an in vitro angiogenic assay, the ability to invade Matrigel and reorganize into tube-like structures, unlike control cultures . The effects of TNF alpha on the PA proteolytic system of endothelial cells, the biological significance of this event and potential in vivo consequences will be discussed . In addition, the influence of cytokine regulatory control systems will be described, since it is becoming increasingly clear that cytokines do not act in isolation . The vascular endothelium serves as a widely distributed anatomical interface between the blood and tissue with diverse capabilities, performing distinctive biologic functions at different sites and within specific organs.(ABSTRACT TRUNCATED AT 400 WORDS)

Agents Actions Suppl, 1993, 42, 125 - 43
Factor XII activation and inhibition in inflammation; Colman RW; Biochemical observations during clinical sepsis using functional and immunological measurements of enzymes, cofactors and inhibitors of the kallikrein-kinin system indicate that activation of these proteases occur during hypotensive gram-negative septicemia and adult respiratory distress syndrome . Using animal models of septicemia, we demonstrated that protease inhibitors or neutralizing monoclonal antibodies to proteins of the contact system inhibit or prevent the formation of kallikrein and the decrease in kininogen . In addition, the irreversible phase of hypotension can be prevented and survival prolonged . Thus, bradykinin is one of the important mediators of hypotension . In contrast, the contact system plays little role in the associated DIC . In cardiopulmonary bypass, the formation of kallikrein leads to neutrophil degranulation and release of elastase . Selective inhibitors of kallikrein not only block its activation but play a predominant role in inhibiting elastase release.

Int Rev Immunol, 1993, 10(1), 17 - 36
Genetic deficiencies of the complement system and association with disease--early components; Kolble K et al.; Genetic deficiency of one of the early components of the classical pathway of complement (C1q, C1r, C1s, C4 and C2) is often associated with clinical symptoms and immunochemical abnormalities common in idiopathic autoimmune diseases, such as lupus erythematosus, but also with an increased incidence of various, local and generalized infections . These observations are consistent with the current view of the complement system's role in handling immune complexes and combating microbial invasion . However, the absence of absolute correlations in these experiments of nature suggests that genetic defects of the classical pathway act only epistatically to other host factors and the primary etiologies of the associated diseases . In contrast, the strong association of properdin and factor D deficiency with serious infections caused by encapsulated Gram-negative bacteria suggests a more immediate involvement of the alternative pathway in a specific segment of immunity and its pathology . This concept is also supported by the primordial role of the alternative pathway in the evolution of the complement system and the apparent lethality of factor B deficiency . The gene structures of most of these early components have now been elucidated providing the basis for detailed analyses of the defective alleles, the determination of carrier status, and prenatal diagnosis.

Res Microbiol, 1993 Jan, 144(1), 35 - 46
Balneatrix alpica gen . nov., sp . nov., a bacterium associated with pneumonia and meningitis in a spa therapy center; Dauga C et al.; In 1987, an outbreak of pneumonia and meningitis caused by an unknown bacterium occurred in a spa therapy centre . Nine isolates of this pathogen constituted a tight DNA hybridization group . rRNA-DNA hybridization and 16S rRNA sequencing showed that the studied bacteria represented a new branch in superfamily II (= gamma subclass) of the Proteobacteria, close to the genus Oceanospirillum . The new bacterium was highly polymorphic and, in young cultures, had curved Gram-negative cells, motile by polar single flagella . The new bacterium differed from the genus Oceanospirillum by its lacking the NaCl requirement and by reducing nitrate into nitrite, producing indole from tryptophan and producing acid from carbohydrates . The name Balneatrix alpica gen . nov., sp . nov . is proposed for the studied organism . The type strain is strain 4-87 (= CIP 103589).

Anaesthesiol Reanim, 1993, 18(6), 164 - 70
{Selective gut decontamination in ventilated intensive therapy patients}; Grundling M et al.; Treatment of long-term artificially ventilated patients is often complicated by nosocomial infections . The infection that occurs with the highest frequency during intensive care treatment is pneumonia (22-63%) . Ninety per cent of nosocomial infections of intensive care patients are endogenous infections caused by mainly gram-negative aerobic microorganisms that have colonized in the gastrointestinal tract . Selective decontamination of the intestine provides a method that prevents nosocomial infections . In a prospective study 13 patients whose oropharynx and gastrointestinal tract had been decontaminated (SDD) were compared to 17 patients in a control group . In a third group twelve patients were decontaminated in the gastrointestinal tract (SGD) only, and in a fourth group 16 patients were decontaminated in the oropharynx (SMD) only . Trachea, oropharynx and faeces of the patients belonging to the control group (KG) were colonized to almost 100% with gram-negative bacteria . Only 10% of the patients of the SDD and SMD groups showed gram-negative bacteria located in the trachea and oropharynx after one week of decontamination . No gram-negative aerobic bacteria were present after seven days in the faeces of the patients of the SDD and SGD groups . There was no difference with regard to the trachea and oropharynx between the control group and the SGD group . The gram-negative aerobic intestinal flora was not affected by the selective mouth decontamination . The average rate of pneumonia occurrence within the 15-day observation period was 28.2% for the control group, 14% for the SGD group, and 9.6% for the SDD group, and 4.1% for the SMD group . Decontamination of the oropharynx of patients is essential in order to successfully prevent pneumonia.(ABSTRACT TRUNCATED AT 250 WORDS)

Br J Neurosurg, 1993, 7(5), 501 - 5
Antibiotic prophylaxis in unrepaired CSF fistulae; Eljamel MS; The value of antibiotic prophylaxis in patients with Cerebrospinal Fluid Leakage (CSF) is debatable . The aim of this study was to determine the value of prophylactic antibiotics in these patients . The study population comprised 253 patients with definite CSF leaks, of whom 106 received adequate antibiotic prophylaxis (Group A) and 109 were not treated with antibiotics (Group B) . Thirty-eight patients were excluded from the analysis because they received antibiotics for reasons other than the CSF leakage . The two groups were closely matched for age, sex, type of CSF fistula, site, and duration of CSF leakage and presence or absence of skull fractures, but there were more patients with facial fractures and pneumocephalus in those who were treated with antibiotics . The first week meningitis rate was 6.6 and 9.17% in the treated and untreated groups, respectively, while the annual risk of meningitis was 7.6% in the treated and 11.9% in the untreated group . However, these differences did not reach significance (P > 0.05) . The survival curves of meningitis-free survival were similar in the two groups, particularly during the first 4 weeks during which antibiotics were given (Log Rank test, p > 0.05) . Furthermore, there were more cases of Gram-negative infection and of partially-treated meningitis in the treated group . Although this was a retrospective, non-randomized study, it confirms the conclusions of previous smaller series, that prophylactic antibiotics do not significantly reduce the risk of meningitis in these patients . It is ethically justifiable to withhold antibiotic prophylaxis in patients with CSF fistulae until a prospective controlled double blind trial has settled the question.

Annu Rev Microbiol, 1993, 47, 855 - 74
The stationary phase of the bacterial life cycle; Kolter R et al.; In the natural environment bacteria seldom encounter conditions that permit periods of exponential growth . Rather, bacterial growth is characterized by long periods of nutritional deprivation punctuated by short periods that allow fast growth, a feature that is commonly referred to as the feast-or-famine lifestyle . In this chapter we review the recent advances made in our understanding of the molecular events that allow some gram-negative bacteria to survive prolonged periods of starvation . After an introductory description of the properties of starved gram-negative bacteria, the review presents three aspects of stationary phase: entry into stationary phase, responses during prolonged starvation, and reentry into the growth cycle.

Wien Klin Wochenschr, 1993, 105(18), 527 - 9
{Course and management of tetanus}; Pfausler B et al.; Within a span of 18 months 4 patients (2 men, 2 women) were admitted with generalized tetanus to our neurological intensive care unit . The "period of onset" ranged from 2 to 14 days . All patients needed artificial ventilation over 17 to 38 days . The clinical course was complicated by gram-negative pneumonia in 3 of the 4 patients . Time of hospitalization ranged between 35 and 60 days . One patient had never been immunized against tetanus . The other 3 had not received a booster dose for over 10 years . All 4 patients survived, 3 of them without sequelae.

Swed Dent J Suppl, 1993, 90, 1 - 46
Actinobacillus actinomycetemcomitans and localized juvenile periodontitis . Clinical, microbiologic and histologic studies; Christersson LA; The present studies examined Actinobacillus actinomycetemcomitans and its role in localized juvenile periodontitis (LJP) . The distribution of the bacteria was studied in healthy normals, patients with adult periodontitis, diabetics, and those with LJP . Over 95% of the LJP patients harbored A . actinomycetemcomitans, whereas only 17% of healthy subjects, 21% of adult periodontitis patients, and 5% of diabetics were positive . All members of a LJP family harboring the organism yielded isolates of the same biotype and serotype . The transmission of the bacteria was studied after transfer of the bacteria, with periodontal probes from infected to healthy gingival sites, within the oral cavity of LJP patients . Newly colonized gingival sites, 50% of those involved, became free of A . actinomycetemcomitans after only 3 weeks . A purposely forceful inoculation contributed to a more predictable colonization (89%), but only prolonged the colonization with one week . Treatment of LJP lesions with scaling and root planing resulted in minimal clinical and microbiological changes during a 16 week follow-up period . However, gingival curettage and modified Widman flap surgery suppressed A . actinomycetemcomitans in 75% and 89% of the sites, and resulted in resolution of periodontal pocket depth and gain in attachment level . Gingival tissue specimens, from 35 LJP sites, 3 control sites, and one monkey biopsy, were studied to verify the hypothesis of gingival infiltration of A . actinomycetemcomitans . Bacteria were identified immunohistologically with rabbit antisera serospecific to the three A . actinomycetemcomitans serotypes . Positive staining was observed in the tissue from all but one LJP patient . Twenty-eight (80%) lesions were positive for A . actinomycetemcomitans antigens in the gingival connective tissue, often with antigens located both between and within cells . The specimen from a culture positive control demonstrated no signs of invasion, similar to the monkey specimen . Transmission electron microscopic examination verified gram-negative bacteria that appeared as single microbes or small clusters in the connective tissue . The viability of the invading bacteria was studied by selective culture of minced tissue specimens and a series of washing solutions, after a rigorous surface disinfection . The washings successfully removed the bacteria in most cases, and after mincing 73% of the biopsies showed to be culture positive . The culture recovery of A . actinomycetemcomitans from minced tissues correlated exceedingly well with the presence of specific antigens in the gingival tissues from corresponding gingival biopsies . To further demonstrate the importance of A . actinomycetemcomitans in LJP, six culture positive LJP patients were treated by systemic tetracycline alone.(ABSTRACT TRUNCATED AT 400 WORDS)

Rev Hosp Clin Fac Med Sao Paulo, 1993 Jan-Feb, 48(1), 8 - 12
{Infection involving arterial prosthesis: clinical picture, etiology, and predisposing factors}; Aguiar ET et al.; Thirty patients were operated for arterial reconstruction with synthetic grafts . They presented one of the following complications: 1) prosthesis exposed by a cutaneous fistula; 2) prosthesis in communication with a hollow viscus; 3) prosthesis involved by pus; 4) positive culture of a fragment of the prosthesis or of the surrounding secretion . The clinical manifestations were cutaneous fistula in 21 patients (70%), external bleeding in 14 (47%), exposition of the prosthesis in five (17%), anastomotic aneurysm in five (17%), and enteric fistula in four (13%) . The infection became evident during the first postoperative year in half of the patients . The most important agents of infection were staphyilococci and Gram negative bacteria.

Intensive Care Med, 1993, 19(6), 358 - 60
Multiple organ failure after mitral valve repair with intravascular hemolysis and its recovery due to mitral valve replacement; Seyr M et al.; We present a patient with evidence of severe intravascular hemolysis after mitral valve repair, an established method for the surgical treatment of mitral valve disease to avoid prosthesis related complications . The coincidence of this uncommon complication with hemodynamic instability due to pre-existing myocardial dysfunction and Gram-negative pneumonia promoted the development of simultaneous dysfunction of liver, kidney and the cardio-respiratory system . Elimination of the source of hemolysis by re-operation with mitral valve replacement on the ninth postoperative day allowed prompt recovery from severe organ dysfunction . Free hemoglobin may have perpetuated progressive organ failure in our patient.

Brain Res Bull, 1993, 32(6), 581 - 7
Differential induction of c-Fos immunoreactivity in hypothalamus and brain stem nuclei following central and peripheral administration of endotoxin; Wan W et al.; Lipopolysaccharide (LPS), an endotoxin associated with gram-negative bacteria, is a potent activator of the immune system . We have tested the effects of ICV infusions of LPS (10 ng) or Ringer's solution on the induction of the proto-oncogene protein c-Fos in the brain as well as plasma levels of corticosterone and splenic concentrations of norepinephrine (NE) and VIP . At 3 h post-ICV infusion of LPS, numerous labeled neurons were observed in the paraventricular nucleus (PVN) of the hypothalamus and the nucleus tractus solitarius (A2) region of the brain stem . Also, corticosterone and splenic NE and VIP levels were all elevated post-ICV LPS . Analysis of the time course for the induction of c-Fos protein in the brain following IP injections of LPS indicated that, relative to control injections, increased numbers of c-Fos-positive cells were detected in the PVN 0.5 h following IP injections (100 micrograms), peaked at 2-3 h postinjection, and then returned to control levels at later intervals . Additional dose-response data for IP LPS indicated a small increase in the number of labeled cells at a dose of 4.0 micrograms, and the number and staining intensity increased up to a dose of 100 micrograms . Corticosterone levels followed a similar pattern and were elevated at the 4.0 micrograms IP dose of LPS and increased to peak levels at 40 micrograms and higher . In contrast to ICV injections, splenic NE levels were unaltered by IP injections of LPS.(ABSTRACT TRUNCATED AT 250 WORDS)

Eur J Cancer B Oral Oncol, 1993 Jan, 29B(1), 1 - 2
Irradiation mucositis: a reappraisal; Martin MV; Irradiation mucositis is a complication of anticancer therapy . It is regarded as an unavoidable consequence of treatment . Recent studies have shown that this condition is probably a consequence of abnormal Gram negative bacillary carriage in the oral cavity . If this abnormal carriage is avoided then prevention or amelioration of irradiation mucositis may be possible.

Genetica, 1993, 90(2-3), 115 - 32
Integrons: novel DNA elements which capture genes by site-specific recombination; Hall RM et al.; Integrons are unusual DNA elements which include a gene encoding a site-specific DNA recombinase, a DNA integrase, and an adjacent site at which a wide variety of antibiotic resistance and other genes are found as inserts . One or more genes can be found in the insert region, but each gene is part of an independent gene cassette . The inserted genes are expressed from a promoter in the conserved sequences located 5' to the genes, and integrons are thus natural expression vectors . A model for gene insertion in which circular gene cassettes are inserted individually via a single site-specific recombination event has been proposed and verified experimentally . The gene cassettes include a gene coding region and, at the 3' end of the gene an imperfect inverted repeat, a 59-base element . The 59-base elements are a diverse family of elements which function as sites recognized by the DNA integrase . Site-specific insertion of individual genes thus represents a further mechanism which contributes to the evolution of the genomes of Gram-negative bacteria and their plasmids and transposons . Members of the most studied class of integrons, which include the sulI gene in the conserved sequences, are believed to be mobile DNA elements on the basis that they are found in many independent locations, and a discrete boundary is found at the outer end of the 5'-conserved segment . However, the length of the 3'-conserved segment is variable in the integrons examined to date, and it is likely that this variability has arisen as the result of insertion and deletion events . Though the true extent of the 3'-conserved segment remains to be determined, it seems likely that these integrons are mobile DNA elements . The second known class of integrons comprises members of the Tn7 transposon family.

Antonie Van Leeuwenhoek, 1993-94, 64(3-4), 231 - 51
Pseudomonas classification . A new case history in the taxonomy of gram-negative bacteria; Palleroni NJ; Various criteria that have been used in the development of a system of classification of Pseudomonas species, as well as in the precise circumscription of the genus on phenotypic and molecular bases, are discussed . Pseudomonas taxonomy has transcended its own limits by suggesting a general strategy for the definition of taxonomic hierarchies at and above the genus level . A selection of studies on the biochemical and physiological properties of members of the genus is critically examined in relation to the current taxonomic scheme as a frame of reference.

Trends Biotechnol, 1993 Jan, 11(1), 6 - 10
Practical applications of engineering gram-negative bacterial cell surfaces; Georgiou G et al.; The recent development of systems for the expression of heterologous proteins on the surface of Gram-negative bacteria has stimulated considerable interest in practical applications . Areas in which surface expression is particularly important include the development of live bacterial vaccines, the display and selection of peptide and antibody libraries, the production of whole cell adsorbents, and the preparation of microbial biocatalysts.

Folia Microbiol (Praha), 1993, 38(3), 163 - 70
Characterization of two Metabacterium sp . from the gut of rodents . 1 . Morphology and histochemical examination of a new Metabacterium sp . from the gut of the European hamster (Cricetus cricetus)
Schiel R, Kunstyr I, Uhr G, Kaup FJ.
A new giant Gram-negative non-cultivatable symbiotic endospore-forming bacterium was found in the gut of the European hamster . This "Metabacterium" sp., provisionally named "Metabacterium criceti", sp . n., has a length of approximately 20 microns and thickness of 4 microns . It forms 1 to 2 cylindrical endospores, approximately 9 microns long and 1.4 microns thick . TEM-micrographs show a cell wall structure characteristic of Gram-negative bacteria . Vegetative cells are filled with granules 0.3 micron in diameter which resemble starch granules . The reproduction occurs with binary fission and by formation of two endospores . Of thirteen biochemical components sought, four, i.e . glycogen, triacylglycerols, peroxidase and alkaline phosphatase, were not found . Starch, acid mucosubstances, DNA, RNA, lipids, proteins, adenosine triphosphatase and acid phosphatase were found in different patterns, depending on the developmental stage of the bacterium . In the vegetative cell stage all these components, with the exception of starch, were found . In the endospore-bearing cell stage, only the starch-like cell component granules could be detected . In free endospores only DNA, RNA and acid phosphatase were found . Some of the components, i.e . DNA, lipids, starch-like granules, were linked to certain cell substructures, the distribution of others, viz . polysaccharides, RNA, adenosine triphosphatase and proteins was diffuse . The lipids, found only in vegetative cells, were associated with the cell wall.

Blood Purif, 1993, 11(2), 134 - 40
Regulation of the response to bacterial lipopolysaccharide by endogenous and exogenous lipopolysaccharide binding proteins; Marra MN et al.; Bactericidal/permeability-increasing protein (BPI) is a natural constituent of human neutrophils . Recombinant BPI has been shown to bind to bacterial lipopolysaccharide (LPS), and to neutralize the ability of LPS to stimulate inflammatory cells in vitro and in vivo . BPI shares sequence homology and immunocrossreactivity with another endogenous LPS binding protein, lipopolysaccharide binding protein (LBP) . Despite the homology, these proteins have opposite effects on LPS . LBP mediates cell activation by low, otherwise nonstimulatory concentrations, while BPI neutralizes LPS bioactivity . Exogenous LPS binding proteins in the form of monoclonal antibodies have been developed with the goal of generating antiendotoxin therapeutics to treat gram-negative sepsis and related syndromes . Here we show that LPS-binding and neutralizing properties of BPI compare favorably with two monoclonal antibodies tested, HA-1A and XMMEN-OE5 . BPI also competes effectively with LBP for LPS . Thus, BPI may represent an endogenous LPS-regulatory molecule suitable for use as a potent antiendotoxin therapeutic.

Biochem Biophys Res Commun, 1992 Dec 30, 189(3), 1498 - 502
Acid pH decreases OmpF and OmpC channel size in vivo; Todt JC et al.; To be effective against gram-negative organisms, beta-lactam antibiotics must be able to penetrate the outer membrane . For Escherichia coli, these compounds generally cross this barrier through non-specific channels in porins OmpF and OmpC . In vitro studies have shown that increased pH induces a switch in the structure of OmpF and OmpC from a small channel conformation to a set of larger-sized channel conformations . In this study, the permeability of two cephalosporins into cells producing either OmpC or OmpF was examined at various pHs . The results suggest that the pH-induced switch in channel size observed in vitro also occurs in vivo.

Proc Natl Acad Sci U S A, 1992 Dec 15, 89(24), 12058 - 62
Translocation of a folded protein across the outer membrane in Escherichia coli; Pugsley AP; A mutation in the Escherichia coli dsbA gene (coding for a periplasmic disulfide oxidoreductase) reduces the rate of disulfide bond formation in the enzyme pullulanase and also reduces the rate at which the enzyme is secreted to the cell surface, as measured by protease accessibility . The enzyme did not become protease accessible when disulfide bond formation was completely prevented in the mutant strain by carboxymethylation . These results indicate that a disulfide bond may be required for, and certainly does not impede, the translocation of pullulanase across the outer membrane . Since it is unlikely that a disulfide bond could be formed and then reduced again in the periplasm, these results would appear to strengthen the argument that pullulanase polypeptides fold into or close to their final conformation before they are transported across the outer membrane . It is suggested that this might be a feature common to all proteins that are secreted by other Gram-negative bacteria by a pullulanase-like pathway.

Biochemistry, 1992 Dec 8, 31(48), 12132 - 40
Sequence of the gene encoding flavocytochrome c from Shewanella putrefaciens: a tetraheme flavoenzyme that is a soluble fumarate reductase related to the membrane-bound enzymes from other bacteria; Pealing SL et al.; Flavocytochrome c from the Gram-negative, food-spoiling bacterium Shewanella putrefaciens is a soluble, periplasmic fumarate reductase . We have isolated the gene encoding flavocytochrome c and determined the complete DNA sequence . The predicted amino acid sequence indicates that flavocytochrome c is synthesized with an N-terminal secretory signal sequence of 25 amino acid residues . The mature protein contains 571 amino acid residues and consists of an N-terminal cytochrome domain, of about 117 residues, with four heme attachment sites typical of c-type cytochromes and a C-terminal flavoprotein domain of about 454 residues that is clearly related to the flavoprotein subunits of fumarate reductases and succinate dehydrogenases from bacterial and other sources . A second reading frame that may be cotranscribed with the flavocytochrome c gene exhibits some similarity with the 13-kDa membrane anchor subunit of Escherichia coli fumarate reductase . The sequence of the flavoprotein domain demonstrates an even closer relationship with the product of the yeast OSM1 gene, mutations in which result in sensitivity to high osmolarity . These findings are discussed in relation to the function of flavocytochrome c.

N Engl J Med, 1992 Dec 3, 327(23), 1625 - 31
Isolation of Rochalimaea species from cutaneous and osseous lesions of bacillary angiomatosis; Koehler JE et al.; BACKGROUND . Bacillary angiomatosis is characterized by vascular lesions, which occur usually in patients infected with the human immunodeficiency virus (HIV) . A newly described gram-negative organism, Rochalimaea henselae, has been associated with cutaneous bacillary angiomatosis, but no organism has been isolated and cultivated directly from cutaneous tissue . METHODS . We used two methods to isolate the infecting bacterium from four HIV-infected patients with cutaneous lesions suggestive of bacillary angiomatosis: cultivation with eukaryotic tissue-culture monolayers and direct plating of homogenized tissue onto agar . The patients' blood was cultured with the lysis-centrifugation method . Isolates recovered from skin and blood were identified by sequencing all or part of the 16S ribosomal RNA gene amplified with the polymerase chain reaction . RESULTS . R . quintana, historically known as the agent of trench fever, was isolated from cutaneous lesions in three patients, after tissue homogenates were cultivated with endothelial-cell monolayers; R . henselae was isolated from a cutaneous lesion in one patient . In two patients, R . quintana was isolated from both cutaneous tissue and blood; in one patient it was also isolated from bone . CONCLUSIONS . In bacillary angiomatosis, either of two species of rochalimaea--R . quintana or R . henselae--can be isolated from cutaneous lesions or blood, providing an additional method of diagnosis.

Laryngorhinootologie, 1992 Dec, 71(12), 644 - 8
{Chronic otitis externa from the dermatologic viewpoint}; Krahl D; Chronic external otitis may be divided into several diagnostic categories . Disposition for psoriasis, seborrhoeic and atopic eczema are main endogenous reasons . Exogenous pathogens for external otitis are microbes and allergens . There are numerous interrelations by coincidence of dispositional diseases, e.g . psoriasis and atopic eczema and by combination of exogenous and endogenous pathogens . This holds good for the yeast Pityrosporum ovale vs . orbiculare in seborrhoeic eczema and for the susceptibility to contact (type IV) and respiratory (type I) allergy in atopic individuals as well . Mycotic and bacterial, especially gram negative external otitis are linked to predisposing factors like eczema, long-term microbicidal therapy, hot and humid environment . Contact allergic external otitis may occur during long lasting local therapy with various substances including vehicles, the most common allergen being neomycin . Mucosal allergic reactions (Type I) in the upper respiratory tract may impair ventilation of the Eustachian tube and middle ear and therefore epithelial migration, as a drainage mechanism of the auditory canal . Examination should include functional assessment of the Eustachian tube and middle ear and allergy testing (patch, prick test) . Preparations for local therapy should contain a limited number of constituents and avoid common allergens . Surgical procedures to reestablish ventilation of the middle ear are also a therapy for chronic external otitis.

Environ Res, 1992 Dec, 59(2), 416 - 26
Dose-dependent pulmonary effects of inhaled endotoxin in guinea pigs; Gordon T; As a cell wall component of gram-negative bacteria, endotoxin is thought to play a significant role in the respiratory effects of inhaled organic dusts which are microbially contaminated . Assessment of occupational survey data and clinical studies suggests that few measureable, acute functional changes occur below 30-50 ng/m3 endotoxin (as sampled in airborne dust with a vertical elutriator) . Little information is available on the inflammatory effects of inhaled endotoxin at these low concentrations . The present study examined the dose-response relationship between inhaled endotoxin and functional, biochemical, and histological endpoints in the lungs of guinea pigs . Animals were exposed to 0.03 to 50.5 micrograms/m3 aerosolized endotoxin or the vehicle water for 4 hr . At 2 hr into exposure, significant decreases in specific airway conductance were observed only in animals exposed to 9.6 and 50.5 micrograms/m3 endotoxin (17.3 +/- 1.2 and 35.5 +/- 0.5% decreases from baseline values, respectively (mean +/- SE)) . Total cell count and lactate dehydrogenase levels in bronchoalveolar lavage fluid were significantly elevated at 24 hr after exposure in all endotoxin-exposed groups except the lowest dose, 0.03 micrograms/m3 (P < 0.05) . Polymorphonuclear leukocyte influx into the alveolar region was also dependent on the concentration of inhaled endotoxin . Thus, LDH activity, a biochemical marker of cell injury, and total cell counts and polymorphonuclear leukocytes, markers of inflammation, were more sensitive indices of adverse pulmonary effects from inhaled endotoxin than a functional measurement . These results suggest that subtle inflammatory changes may occur at airborne endotoxin concentrations which may produce no acute respiratory symptoms.

Arch Ophthalmol, 1992 Dec, 110(12), 1763 - 4
Rapid detection of gram-negative endotoxin contamination of contact lens saline solutions; Alfonso EC et al.; The use of the limulus amoebocyte lysate for the early and rapid detection of gram-negative endotoxin contamination of contact lenses and their solutions could reduce the risk of a keratitis developing that is associated with these devices . Using multiple aliquots from 17 unopened brands of commercially available contact lens saline solutions (15 for soft, two for hard), plus multiple aliquots from these solutions mixed with bacterial endotoxin, we evaluated the ability of two limulus amoebocyte lysate (0.125 endotoxin units/mL) products to detect the presence or absence of gram-negative endotoxin contamination . Sensitivity ranged from 65% (11/17) to 82% (14/17) when the solutions were tested undiluted . When diluted 1:8, the sensitivity increased to 100% . Specificity was 100% for undiluted and diluted specimens . The solutions used for hard contact lenses had the highest false-negative results . The limulus amoebocyte lysate can be used to detect the presence of gram-negative endotoxin in contact lens solutions.

J Bacteriol, 1992 Dec, 174(24), 8119 - 32
Definition of a minimal plasmid stabilization system from the broad-host-range plasmid RK2; Roberts RC et al.; The stable inheritance of the broad-host-range plasmid RK2 is due at least in part to functions within a region located at coordinates 32.8 to 35.9 kb, termed the RK2 par locus . This locus encodes four previously identified genes in two operons (parCBA and parD; M . Gerlitz, O . Hrabak, and H . Schwab, J . Bacteriol . 172:6194-6203, 1990, and R . C . Roberts, R . Burioni, and D . R . Helinski, J . Bacteriol . 172:6204-6216, 1990) . The parCBA operon is functional in resolving plasmid multimers to monomers . Analysis of the plasmid stabilization capacity of deletions within this region, however, indicates that this multimer resolution operon is required for stabilization only in certain Escherichia coli strains and under specific growth conditions . The deletion analysis further allowed a redefinition of the minimal functional region as 790 bp in length, consisting of the parD gene (243 bp) and its promoter as well as sequences downstream of parD . This minimal region stabilizes an RK2-derived minireplicon in several different gram-negative bacteria and, at least in E . coli, in a vector-independent manner . By insertional mutagenesis, both the parD gene and downstream (3') regions were found to be required for plasmid stabilization . The downstream DNA sequence contained an open reading frame which was subsequently shown by transcriptional and translational fusions to encode a protein with a predicted size of 11,698 Da, designated ParE . Since the parDE operon requires the presence of the parCBA operon for efficient stabilization under certain growth conditions, the potential role of multimer resolution in plasmid stabilization was tested by substituting the ColE1 cer site for the parCBA operon . While the cer site did function to resolve plasmid multimers, it was not sufficient to restore stabilization activity to the parDE operon under growth conditions that require the parCBA operon for plasmid stability . This suggests that plasmid stabilization by the RK2 par locus relies on a complex mechanism, representing a multifaceted stabilization system of which multimer resolution is a conditionally dispensable component, and that the function(s) encoded by the parDE operon is essential.

J Clin Microbiol, 1992 Dec, 30(12), 3225 - 9
Cellular fatty acids in Fusobacterium species as a tool for identification; Tuner K et al.; Identification of fusobacteria from clinical specimens currently requires analysis of metabolic end products by gas-liquid chromatography in addition to certain biochemical and enzymatic tests because of the relative biochemical inactivity of these bacteria . Even the finding of pointed, thin gram-negative cells on Gram-stained slides can no longer be relied on for identification of Fusobacterium nucleatum, since at least four other species of fusobacteria have been seen to exhibit similar morphology . We examined 46 clinical isolates and six American Type Culture Collection type strains of fusobacteria by conventional methods and by the Microbial ID Systems MIDI software package for analyzing cellular fatty acid patterns measured by capillary column gas-liquid chromatography . Distinctive patterns of major fatty acids could be used to reliably identify most clinical isolates to the species level . The MIDI system identified 89% of the isolates correctly and provides an alternative to conventional methods.

Infect Immun, 1992 Dec, 60(12), 4995 - 5003
Purification and immunological characterization of a major low-molecular-weight lipoprotein from Borrelia burgdorferi; Katona LI et al.; Borrelia burgdorferi resembles gram-negative bacteria in having both cellular and outer membranes . We previously showed that a lipopolysaccharide (LPS)-like material could be extracted from B . burgdorferi with phenol-chloroform-petroleum ether (PCP) . The PCP extract of B . burgdorferi exhibited biological activity in several in vitro assays (e.g., mitogenicity, pyrogenicity, and cytokine release) . These activities suggested the presence of endotoxin . The PCP extract of B . burgdorferi, however, also contained a small amount of protein . Preliminary studies showed that monoclonal antibody prepared against this protein inhibited the mitogenic activity of the PCP extract toward murine spleen cells . The current study was therefore undertaken to characterize this protein and to establish methods for its separation from the LPS . The PCP-extracted protein consisted of a single, low-molecular-weight lipoprotein (apparent M(r), 10,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis) (SDS-PAGE) . By protein analysis, it accounted for 2% of the dry weight of defatted cells, thus making it a major constituent of the spirochete . It was purified from the LPS by initial extraction into 10% Triton X-100 followed by immunoaffinity chromatography in the presence of detergent . On removal of the LPS, the purified lipoprotein formed aggregates stable to SDS-PAGE which were detectable on Western blots (immunoblots) probed with either the monoclonal antibody or polyclonal antiserum . From a plot of the aggregate molecular weight versus aggregate size, a monomer molecular weight of 7,500 was obtained . Indirect immunofluorescence with the monoclonal antibody showed that the lipoprotein was exposed at the surface of the spirochete in only a small percentage of cells . The lipoprotein was present in several strains of B . burgdorferi but absent in other Borrelia spp., treponemes, and gram-negative human pathogens, indicating species specificity.

Eur J Immunol, 1992 Dec, 22(12), 3097 - 101
Interferon-alpha prevents endotoxin-induced mortality in mice; Tzung SP et al.; Endotoxins, the lipopolysaccharide (LPS) moieties on the bacterial cell wall, cause many of the pathological features of Gram-negative septicemia . Tumor necrosis factor (TNF), primarily a product of monocyte/macrophages, has been shown to mediate many of the pathophysiological effects of endotoxin . Kupffer cells, the largest macrophage population in the body, release TNF when stimulated by LPS in vitro . A recombinant human hybrid interferon-alpha A/D (rIFN-alpha) markedly inhibited this LPS-elicited TNF production by Kupffer cells . The effects of rIFN-alpha were further tested in C57BL/6 mice receiving a lethal dose (400 micrograms/mouse) of LPS . All LPS-treated mice died within 2 days . Pretreatment with rIFN-alpha 1 h before LPS challenge improved the survival at 3 days to 22% (5/23, p < 0.04) . In contrast, rIFN-alpha was more effective when administered 20 min after LPS injection, increasing the survival rate to 81% (13/16, p < 0.0001) . TNF mRNA expression in the liver and spleen 50 min after LPS challenge, and plasma TNF 1.5 h after LPS were also reduced by either pretreatment or post-treatment with rIFN-alpha . Subsequently, experiments were carried out to test the efficacy of delayed rIFN-alpha treatment . A significant protective effect was still apparent when rIFN-alpha was administered 6, 10 and even 14 h (81%, 62% and 28% survival, respectively) after LPS challenge when serum TNF levels had already returned to near baseline . These experimental results suggest that rIFN-alpha might have a therapeutic potential for the prevention and treatment of the deleterious effects associated with endotoxemia besides mechanisms initially blocking TNF production.

J Infect Dis, 1992 Dec, 166(6), 1367 - 74
Effectiveness of a human monoclonal anti-endotoxin antibody (HA-1A) in gram-negative sepsis: relationship to endotoxin and cytokine levels; Wortel CH et al.; Gram-negative sepsis is caused by endotoxin-induced release of tumor necrosis factor (TNF) and other cytokines . HA-1A is a human monoclonal antibody that binds specifically to endotoxin . HA-1A should prevent death in endotoxemic patients and reduce serum levels of TNF and interleukin-6 (IL-6) . This hypothesis was tested in 82 septic patients who were randomly allocated to receive a single intravenous 100-mg dose of HA-1A or placebo . Pretreatment endotoxemia was detected in 27 patients (33%) . Death occurred within 28 days of treatment in 8 (73%) of 11 placebo recipients and in 5 (31%) of 16 HA-1A recipients (P = .02) . The median decrease in serum TNF level 24 h after treatment was 12 ng/L in patients given HA-1A and 0 ng/L in placebo recipients (n = 65; P = .04) . For IL-6, this was 204 ng/L in patients given HA-1A and 44 ng/L in placebo recipients (n = 67; P = .4) . Thus, HA-1A reduces mortality in septic patients with endotoxemia and lowers serum TNF levels.

EMBO J, 1992 Dec, 11(13), 4747 - 56
Interactive surface in the PapD chaperone cleft is conserved in pilus chaperone superfamily and essential in subunit recognition and assembly; Slonim LN et al.; The assembly of adhesive pili in Gram-negative bacteria is modulated by specialized periplasmic chaperone systems . PapD is the prototype member of the superfamily of periplasmic pilus chaperones . Previously, the alignment of chaperone sequences superimposed on the three dimensional structure of PapD revealed the presence of invariant, conserved and variable amino acids . Representative residues that protruded into the PapD cleft were targeted for site directed mutagenesis to investigate the pilus protein binding site of the chaperone . The ability of PapD to bind to fiber-forming pilus subunit proteins to prevent their participation in misassembly interactions depended on the invariant, solvent-exposed arginine-8 (R8) cleft residue . This residue was also essential for the interaction between PapD and a minor pilus adaptor protein . A mutation in the conserved methionine-172 (M172) cleft residue abolished PapD function when this mutant protein was expressed below a critical threshold level . In contrast, radical changes in the variable residue glutamic acid-167 (E167) had little or no effect on PapD function . These studies provide the first molecular details of how a periplasmic pilus chaperone binds to nascently translocated pilus subunits to guide their assembly into adhesive pili.

Am J Physiol, 1992 Dec, 263(6 Pt 2), R1324 - 32
Sepsis- and endotoxin-induced increase in organ glucose uptake in leukocyte-depleted rats; Lang CH et al.; Both gram-negative infection and bacterial endotoxin (lipopolysaccharide, LPS) produce a marked neutropenia and increase glucose disposal by peripheral tissues . The purpose of the present study was to determine whether leukocyte depletion before these insults would diminish the commonly observed increases in tissue glucose uptake . Rats were depleted of circulating and marginated leukocytes with cyclophosphamide (CPA) . Under basal postabsorptive conditions the subcutaneous injection of live Escherichia coli into control animals enhanced whole body glucose disposal that resulted in part from a stimulation of glucose uptake by the liver, spleen, intestine, and lung . These increases in tissue glucose uptake were not associated with an increase in neutrophil number, as assessed by myeloperoxidase (MPO) activity . CPA-induced leukopenia did not alter the sepsis-induced increase in glucose uptake by these tissues and whole body glucose use remained elevated . In contrast, skin and muscle proximal to the site of infection showed an increase in both glucose uptake and MPO activity . Furthermore, leukocyte depletion attenuated the elevated glucose uptake by skin and muscle near the inflammatory focus . The intravenous injection of LPS also increased whole body glucose disposal and enhanced glucose uptake by the lung, liver, spleen, intestine, and skin in saline-treated rats . Of these tissues the lung, liver, and spleen had a corresponding increase in neutrophil number . The LPS-induced increases in tissue glucose uptake in leukopenic rats were comparable, with the exception of liver and lung . In these tissues the incremental increase in glucose uptake after LPS was reduced 40-50% in leukopenic animals.(ABSTRACT TRUNCATED AT 250 WORDS)

J Periodontol, 1992 Dec, 63(12 Suppl), 1093 - 101
Bacterial concentration fluorescence immunoassay (BCFIA) for the detection of periodontopathogens in plaque; Wolff LF et al.; A bacterial concentration fluorescence immunoassay (BCFIA) was developed to rapidly detect periodontopathic bacteria in human plaque samples . The BCFIA utilized fluorescent-tagged monoclonal antibodies (MAbs) directed against the lipopolysaccharide of selected Gram-negative bacteria . Microorganisms identified in plaque using the BCFIA included Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Prevotella intermedia, and Fusobacterium nucleatum . The immunoassay procedure involved combining a patient's plaque sample with a species-specific fluorescein isothiocyanate-labeled MAb and then incubating the mixture in a specialized microtiter plate allowing the MAb to bind to its homologous bacteria . Bound and unbound fluorescent-tagged MAbs were separated by filtration and total bound bacterial fluorescence was determined with a fluorimeter . The relative number of a bacterial species in a given plaque sample was estimated by reference to a standard curve carried through the BCFIA . The BCFIA had a lower detection limit of near 10(4) specific bacterial cells in a mixed bacterial preparation or plaque sample . When compared to cultivable flora procedures in detecting the 4 periodontopathogens, the BCFIA had high levels of statistical sensitivity, 97% to 100%, while statistical specificity ranged between 57% and 92% . There was a 71% to 82% agreement between BCFIA and DNA probe methodology in detecting periodontopathogens in plaque . The BCFIA, when compared to cultivable flora, offers the advantage of evaluating both live and dead bacterial cells in plaque . This may in part, if not fully, explain the lower specificity values of the BCFIA when compared to cultivable flora . Screening plaque samples for periodontopathic bacteria is considerably faster and results in a greater frequency of detection with BCFIA than cultivable flora based methods.

Kidney Int, 1992 Dec, 42(6), 1309 - 18
Porins and lipopolysaccharide stimulate platelet activating factor synthesis by human mesangial cells; Camussi G et al.; Porins, a family of hydrophobic proteins located in the outer membrane of the cell wall of gram-negative bacteria and lipopolysaccharide (LPS), were shown to stimulate the synthesis of platelet activating factor (PAF), a phospholipid mediator of inflammation and endotoxic shock, by cultured human glomerular mesangial cells (MC) . The synthesis of PAF induced by porins was rapid (peak at 20 min) and independent either from contamination by LPS or from generation of an endotoxin-induced cytokine such as tumor necrosis factor (TNF) since it was not prevented by cycloheximide, an inhibitor of protein synthesis or anti-TNF blocking antibodies . LPS also stimulated PAF synthesis by MC . However, the kinetic of PAF synthesis induced by LPS was biphasic with an early and transient peak at 10 minutes and a second and sustained peak at three to six hours . This second peak required an intact protein synthesis and was prevented by anti-TNF antibodies, suggesting the dependency on LPS-induced synthesis of TNF . Experiments with labeled precursors demonstrated that in MC, either after stimulation with porins or LPS, PAF was synthesized via the remodeling pathway that involves acetylation of 1-0-alkyl-sn-glyceryl-3-phosphorylcholine (2-lyso-PAF) generated from 1-0-alkyl-2-acyl-sn-glyceryl-3-phosphorylcholine by phospholipase A2 (PLA2) activity . Porins and LPS, indeed, induced PLA2-dependent mobilization of {14C}-arachidonic acid that was inhibited by p-bromodiphenacylbromide (PBDB) . PBDB, an inhibitor of PLA2, also blocked PAF synthesis by preventing the mobilization of 2-lyso-PAF, the substrate for PAF-specific acetyltransferase.(ABSTRACT TRUNCATED AT 250 WORDS)

Biochem J, 1992 Dec 1, 288 ( Pt 2), 337 - 40
Evidence for copper and 3,4,6-trihydroxyphenylalanine quinone cofactors in an amine oxidase from the gram-negative bacterium Escherichia coli K-12; Cooper RA et al.; The cofactors present in a amine oxidase induced in Escherichia coli K-12 by growth on 2-phenylethylamine have been studied by spectroscopic methods . E.s.r . spectroscopy establishes the presence of cupric copper while resonance Raman spectroscopy on the phenylhydrazine derivative of the enzyme provides strong evidence for the oxidized form of 3,4,6-trihydroxyphenylalanine (TOPA) quinone . The amine oxidase should accordingly be classified as EC 1.4.3.6 . This is the first report of such an amine oxidase in a Gram-negative bacterium.

Med J Malaysia, 1992 Dec, 47(4), 311 - 5
A case of empyema thoracis caused by actinomycosis; Hooi LN et al.; A female patient who presented with left empyema thoracis caused by Actinomyces odontolyticus is reported . She responded to treatment with penicillin and metronidazole but after 3 weeks developed leucopenia complicated by gram-negative septicaemia . Leucopenia improved rapidly on withdrawal of metronidazole . Treatment was continued with a prolonged course of penicillin and she made an uneventful recovery.

Unfallchirurg, 1992 Dec, 95(12), 641 - 3
{Candida infection in the severely burned patient--a successful treatment concept with liposomal amphotericin B}; Pallua N et al.; Candida sepsis is a very serious complication in severely burned patients . This mainly affects patients whose immune system is weakened by illness and/or by drugs . Often diagnosis is difficult because candida sepsis occurs after an initial infection, but therapy is always difficult . Good fungicidal drugs are available, but their side effects limit their effectivity . Two severely burned patients who were suffering from a gram-negative sepsis confirmed by clinical and laboratory data developed candida sepsis . Conventional therapy failed, and both patients suffered from renal failure with constantly high candida-latex-antigen titre . By means of the liposomal encapsulated amphotericin B, which has the same fungicidal effect as amphotericin B, but without its limiting side effects, both, patients could be saved . The kidneys functioned as normal again, the laboratory findings were normal when the patients were discharged.

Am J Pathol, 1992 Nov, 141(5), 1197 - 207
Efficacy of monoclonal antibody against human recombinant tumor necrosis factor in E . coli-challenged swine; Jesmok G et al.; Monoclonal antibody against human tumor necrosis factor alpha (TNF MAb) prevents death induced by intravenous gram-negative bacteria or lipopolysaccharide (LPS) in primates . Although these studies have demonstrated that TNF plays a prominent role in the development of lethal septic shock, exploration of dose-response relationships and possible mechanisms of protection have been limited . We addressed these questions in a series of experiments conducted in E . coli-challenged pigs . First, we determined that TNF MAb neutralized the cytotoxic activity found in septic pig plasma and in culture media from pig monocytes incubated with LPS . Second, we demonstrated that pretreatment with TNF MAb promotes survival, in a dose-dependent fashion, in an otherwise lethal E . coli bacteremic pig model . The results of the survival study highly correlate (r = 0.96, P < 0.01) the presence of TNF in the circulation with mortality . In an additional series of physiologic monitoring experiments designed to delineate possible mechanisms of protection, the authors demonstrate that TNF MAb pretreatment abrogates the prolonged leukopenia, thrombocytopenia, and microvascular leakiness resulting from intravenous bacterial challenge and maintains arterial blood pressure while diminishing pulmonary edema . These findings may provide a mechanism whereby neutralization of TNF systemically affords protection against the lethal sequelae of bacteremia.

Biochemistry, 1992 Nov 3, 31(43), 10479 - 82
Involvement of histidine-21 in the pH-induced switch in porin channel size; Todt JC et al.; Porin is a channel-forming protein in the outer membrane of Gram-negative bacteria . In the previous paper (Todt et al., 1992), we showed that the pH induced a switch in the channel size in vitro for the porins OmpF, OmpC, and PhoE . In the results presented here, His21 of OmpC and OmpF from Escherichia coli was chemically modified with diethyl pyrocarbonate . Functional analysis of these modified porins at different pHs suggested that this histidine is involved in the pH-induced switch in channel size . Secondary structure analysis of porins at various pHs using Fourier transform infrared spectroscopy indicated that there was no global change in structure accompanying the pH-induced switch in channel size.

Biochemistry, 1992 Nov 3, 31(43), 10471 - 8
Effects of pH on bacterial porin function; Todt JC et al.; Porin is a trimeric channel-forming protein in the outer membrane of Gram-negative bacteria . Functions of the porins OmpF, OmpC, and PhoE from Escherichia coli K12 were analyzed at various pHs . Preliminary results from bilayer lipid membrane and liposome swelling assays indicated that in vitro porin has at least two open-channel configurations with a small and a large size . The small channels were stabilized at low pH while the larger channels were detected under basic conditions . The size switch occurred over a very narrow range near neutral pH, and the two major open-channel configurations responded differently to variations in voltage . The presence of two or more pH-dependent substates of porin could explain the variability in pore diameter measured by others and suggests a more dynamic role for porin in the cell.

Immunology, 1992 Nov, 77(3), 473 - 6
Lipopolysaccharide synergizes with tumour necrosis factor-alpha in cytotoxicity assays; Pfister H et al.; Lipopolysaccharide (LPS) from Escherichia coli was found to synergize with human recombinant tumour necrosis factor-alpha (TNF-alpha) in the lysis of L929 and WEHI 164 (clone 13) murine fibroblasts, two cell lines classically used in TNF-alpha bioassays . The effect was noted with TNF-alpha at low (sublytic or lightly lytic) concentrations and was significant for LPS concentrations in the ng range . The LPS effect could be inhibited by polymyxin B, and was not observed when the TNF-alpha assay was performed in the absence of actinomycin D . Enhancement of TNF-alpha lysis by LPS occurred in several assays for determining TNF-alpha, including MTT cleavage, crystal violet staining and lactate dehydrogenase release . Synergism was obtained only when LPS and TNF-alpha were added to cells simultaneously, but not when applied in sequence . The reported synergism may be relevant for TNF-alpha determinations by bioassay, and for the understanding of pathophysiology of Gram-negative sepsis.

J Periodontol, 1992 Nov, 63(11), 897 - 901
Endotoxin levels in periodontally healthy and diseased sites: correlation with levels of gram-negative bacteria; Fine DH et al.; This study investigated the correlation between endotoxin levels and the percentage of Gram-negative bacteria in healthy sites and in periodontitis sites . Twelve healthy adults participated . Each subject provided 3 periodontitis sites with 5 to 8 mm probing depths that bled on gentle probing and 3 healthy sites with sulcus depths of 1 to 3 mm that did not bleed . Clinical examinations and sterile paper point sampling of all study sites were conducted on days 0, 7, and 14, and site-specific endotoxin levels and percentage of Gram-negative bacteria were determined . There were significant differences in both endotoxin levels and percentage Gram-negative bacteria between healthy and periodontitis sites across all 3 sampling periods, but no difference across sampling periods in the healthy sites and the periodontitis sites, respectively . Correlation coefficients revealed a high degree of correlation between site-specific endotoxin levels and percentage of Gram-negative organisms . Using a sample dilution of 1 x 10(4), endotoxin levels differentiated healthy from periodontitis sites with a specificity of approximately 91% and a sensitivity of approximately 90%.

Surg Endosc, 1992 Nov-Dec, 6(6), 302 - 4
Diagnostic laparoscopy in critically ill intensive-care-unit patients; Bender JS et al.; The diagnosis of intraabdominal sepsis in critically ill intensive-care-unit patients remains a challenge . Diagnostic laparoscopy has been performed in seven such patients following admission for coronary artery bypass surgery, gram-negative sepsis, major burns, pneumonia, myocardial infarction, and post-pneumonectomy . Laparoscopy revealed acalculous cholecystitis in two patients (one removed laparoscopically), gangrenous colon in two, cirrhosis with liver infarction in one, and, in two patients, no pathology . Although five patients died postoperatively, none was related to the laparoscopy . There were no intraoperative complications and no known pathology was missed . Because of its ease and accuracy, diagnostic laparoscopy should be considered in all critically ill patients suspected of harboring intraabdominal pathology . Further studies are needed to fully establish its efficacy and safety.

Clin Infect Dis, 1992 Nov, 15(5), 866 - 73
Factors influencing prognosis in bacteremia due to gram-negative organisms: evaluation of 448 episodes in a Turkish university hospital; Uzun O et al.; A total of 448 episodes of bacteremia due to gram-negative organisms observed during the 7-year period between 1983 and 1989 at Hacettepe University Hospitals were studied for evaluating the factors influencing the prognosis . The overall mortality rate was 45.0% . The mortality rates were not significantly different in "rapidly fatal" and "ultimately fatal" disease groups (48.3% and 45.5%, respectively), whereas it was significantly less (34.8%) in the "nonfatal" disease group compared with the "rapidly fatal" category . There were great differences in the mortality rates among different diseases within the same disease category . Shock, multi-organ failure, source of infection, hospital service, appropriateness of antibiotic therapy, and place of acquisition of infection were found to affect prognosis significantly in multivariate analysis . In conclusion, the identification of prognostic factors is a further step for making necessary interventions in reducing the mortality rate associated with bacteremia due to gram-negative organisms . Underlying disease is still an important prognostic factor; however, a new approach is needed for classification of underlying diseases.

Clin Infect Dis, 1992 Nov, 15(5), 840 - 54
Antibiotic-induced release of endotoxin: a reappraisal; Hurley JC; A three- to 20-fold increase in the total concentration of endotoxin occurs as a consequence of antibiotic action on gram-negative bacteria both in vitro and in vivo . There is considerable overlap between the effect of beta-lactam antibiotics and non-beta-lactam antibiotics . Moreover, there is an unexplained delay between the lethal activity of antibiotics and the release of endotoxin . Hence, the mechanism whereby antibiotic action leads to the release of endotoxin is unclear, and mechanisms other than bacterial lysis warrant consideration . The evidence that the release of endotoxin has clinical importance is conflicting, and the issue is unresolved . However, nonlytic release may have implications for the therapeutic efficacy of antiendotoxin immunotherapy . Although frequently cited in the context of the antibiotic-induced release of endotoxin, a number of important differences pertain to conditions, such as the Jarisch-Herxheimer reaction and the tumor lysis syndrome, for which there is clear evidence of an initial deterioration with effective therapy.

Arch Ophthalmol, 1992 Nov, 110(11), 1625 - 9
Toxicity of intravitreous ceftazidime in primate retina; Campochiaro PA et al.; Squirrel monkeys were anesthetized and given intravitreous injections of 0.1 mL of balanced salt solution containing 0 mg (two eyes), 1 mg (three eyes), 2.25 mg (three eyes), or 10 mg (three eyes) of ceftazidime, a third-generation cephalosporin that provides excellent coverage for gram-negative infections . Ophthalmoscopic examinations were performed 48 hours after the injections and results were completely normal in all eyes except for those that were injected with 10 mg of ceftazidime, all three of which showed the appearance of cystic change in the macula . The monkeys were killed and the eyes were removed and examined by light and electron microscopy . All eyes were normal by light microscopy except for those injected with 10 mg of ceftazidime, which showed disruption of photoreceptors (primarily outer segments) in the foveas with cystic changes in all three and macular holes in two of three . Electron microscopy showed mild swelling of mitochondria and perinuclear halos around photoreceptor nuclei in both control eyes and in eyes injected with 1 and 2.25 mg . An eye injected with 10 mg showed severe damage to central photoreceptor outer segments consisting of disruption of plasma membranes and accumulation of intracytoplasmic granular material . Inner segments showed mild changes and there was loss of apical microvilli of the retinal pigmented epithelium . The inner retina was normal . These data suggest that high doses of intravitreous ceftazidime show toxicity primarily to photoreceptor cells, but a dose of 2.25 mg is safe as studied in this model and can be used instead of intravitreous aminoglycosides that have a narrow therapeutic window.

J Lab Clin Med, 1992 Nov, 120(5), 740 - 5
Respiratory epithelial carbohydrate levels of rats with gram-negative bacillary colonization; Mason CM et al.; One mechanism by which severe illness or stress might facilitate adherence and colonization of GNB to respiratory epithelium is by altering epithelial cell surface carbohydrates . To investigate this possibility we used radiolabeled lectins to quantitate carbohydrate levels on intact buccal and tracheal epithelium . A rat model of GNB colonization, in which renal infarction was performed to produce colonization, was used . Buccal and tracheal epithelial surface carbohydrate levels from normal rats and rats 48 hours after renal infarction were compared . Buccal and tracheal epithelium from the renal infarction animals had decreased amounts of sialic acid and fucose, and decreased levels of these sugars occurred at the same time that heavy oropharyngeal GNB colonization developed . Tracheas obtained from the infarcted animals bound three times more Type 1 piliated GNB than normal tracheas . Sialic acid and fucose levels are decreased early after stress, and we speculate that altered epithelial carbohydrates may predispose to GNB colonization by exposing binding sites for GNB.

Cleve Clin J Med, 1992 Nov-Dec, 59(6), 608 - 15
New strategies in nonantibiotic treatment of gram-negative sepsis; Cohn J et al.; Gram-negative bacterial infections are difficult to control and often lead to septic shock or septic syndrome . Many physiologic changes in sepsis are due to bacterial triggering of host responses . Improved understanding of these mechanisms has led to new treatment modalities that aim to block the runaway inflammatory process of sepsis . New therapeutic agents are currently being evaluated in animal and human studies . By combining these advances with adequate antibiotic therapy, it may be possible to improve overall survival in patients with gram-negative sepsis.

J Infect Dis, 1992 Nov, 166(5), 1045 - 50
Toxicity in neuronal cells caused by cerebrospinal fluid from pneumococcal and gram-negative meningitis; Tauber MG et al.; To identify neurotoxic factors in meningitis, a neuronal cell line (HN33.1) was exposed to cerebrospinal fluid (CSF) obtained from rabbits with pneumococcal meningitis or Escherichia coli meningitis or 2 h and 6 h after meningitis was induced by proinflammatory bacterial products (pneumococcal cell walls, endotoxin) . CSF from all types of meningitis induced similar degrees of cytotoxicity . When a soluble tumor necrosis factor (TNF) receptor that completely blocked TNF-mediated toxicity at 10(-7) M was used, all toxicity in meningitis caused by E . coli, endotoxin, or pneumococcal cell wall administration (2 h afterwards) was mediated by TNF . In contrast, CSF from animals with meningitis caused by live pneumococci or pneumococcal cell wall injection (6 h afterwards) retained cytotoxicity in the presence of the TNF receptor . Thus, in established pneumococcal meningitis, but not in the other forms of meningitis, TNF is not the only component toxic in this neuronal cell line.

J Bacteriol, 1992 Nov, 174(21), 6857 - 61
Purification and characterization of phosphoenolpyruvate phosphomutase from Pseudomonas gladioli B-1; Nakashita H et al.; Phosphoenolpyruvate phosphomutase (PEPPM) catalyzes C-P bond formation by intramolecular rearrangement of phosphoenolpyruvate to phosphonopyruvate (PnPy) . We purified PEPPM from a gram-negative bacterium, Pseudomonas gladioli B-1 isolated as a C-P compound producer . The equilibrium of this reaction favors the formation of the phosphate ester by cleaving the C-P bond of PnPy, but the C-P bond-forming reaction is physiologically significant . The C-P bond-forming activity of PEPPM was confirmed with a purified protein . The molecular mass of the native enzyme was estimated to be 263 and 220 kDa by gel filtration and polyacrylamide gel electrophoresis, respectively . A subunit molecular mass of 61 kDa was determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, indicating that the native protein was a tetramer . The optimum pH and temperature were 7.5 to 8.0 and 40 degrees C, respectively . The Km value for PnPy was 19 +/- 3.5 microM, and the maximum initial velocity of the conversion of PnPy to phosphoenolpyruvate was 200 microM/s/mg . PEPPM was activated by the presence of the divalent metal ion, and the Km values were 3.5 +/- 1.4 microM for Mg2+, 16 +/- 5 nM for Mn2+, 3.0 +/- 1.5 microM for Zn2+, and 1.2 +/- 0.2 microM for Co2+.

Infect Immun, 1992 Nov, 60(11), 4819 - 25
Opsonic antibody activity against Actinobacillus actinomycetemcomitans in patients with rapidly progressive periodontitis; Sjostrom K et al.; Actinobacillus actinomycetemcomitans has been closely associated with early-onset, severe periodontitis, and such patients often have serum immunoglobulin G (IgG) antibodies reactive with antigens of this gram-negative pathogen . We examined the functionality and potential importance of these antibodies . The opsonic activity against A . actinomycetemcomitans of sera from 30 patients with rapidly progressive periodontitis (RPP) and from 28 periodontally normal subjects was tested by using polymorphonuclear leukocyte (PMN) chemiluminescence and bactericidal assays . Peak chemiluminescence values correlated strongly with killing observed in the PMN-dependent bactericidal assay (r = 0.88; P < 0.001) . Neither the mean IgG titer nor the mean peak chemiluminescence differed significantly between the two groups . However, when the relationship between chemiluminescence and titer was examined, regression analysis showed that antibodies present in low-titer normal sera were significantly more effective at opsonizing A . actinomycetemcomitans than antibodies present in low-titer RPP patient sera (P = 0.04) . Thus, periodontally normal individuals may be better able than RPP patients to clear A . actinomycetemcomitans in early stages of colonization, and anti-A . actinomycetemcomitans antibodies in RPP patients may be relatively ineffective in preventing infection by this organism.

J Hosp Infect, 1992 Nov, 22 Suppl A, 69 - 74
Comparative clinical and microbiological study of amoxycillin-clavulanic acid and ciprofloxacin in acute purulent exacerbations of chronic bronchitis; Legnani D et al.; In a retrospective study, the clinical and microbiological efficacy of amoxycillin-clavulanic acid and ciprofloxacin were evaluated in outpatients observed within the previous year who were affected by acute purulent exacerbations of chronic bronchitis . Of the 95 patients included in the trial, 50 received amoxycillin 875 mg-clavulanic acid 125 mg 8-hourly for 10 days and 45 received ciprofloxacin 500 mg 12-hourly before meals for 10 days . Of the amoxycillin-clavulanic acid-treated patients, 90% showed clear clinical improvement and in 10% treatment failed . In the ciprofloxacin group, 75.5% of patients showed improvement and in 24.5% treatment failed . All pathogens isolated prior to therapy were susceptible to the antibiotic used for therapy . At the end of treatment, in the amoxycillin-clavulanic acid-treated group, 84% of strains were eradicated and 8% persisted; others were superinfections . In the ciprofloxacin group, 57.7% of strains were eradicated, 26.6% persisted and 15.5% were superinfections . No clinically significant side effects were observed in either group . Overall, amoxycillin-clavulanic acid demonstrated superior clinical and microbiological efficacy to ciprofloxacin, although this might be attributable to the higher proportion of aerobic Gram-negative pathogens in the ciprofloxacin group.

Presse Med, 1992 Oct 17, 21(34), 1625 - 30
{Bacillary angiomatosis}; Robert C et al.; Bacillary angiomatosis (BA) is a recently described infection usually found in patients with human immunodeficiency virus disease . BA is caused by a Gram-negative coccobacillus . This organism is primarily responsible for skin lesions of the pseudo-botryomycoma type or inflammatory nodules, but it also produces fever, degradation of the general condition and visceral lesions involving the lymph nodes, the liver, the spleen and the bones . Histology shows vascular proliferation with turgid endothelial cells and mostly neutrophilic inflammatory infiltrates . BA is susceptible to many antibiotics . The authors describe the history of the disease and its clinical and histological features, discuss its differential diagnosis and principally deal with the relationship between BA and cat-scratch disease and between BA and verruca peruana . They also present the molecular biology technique which enables a genotypic diagnosis of the disease to be made, replacing a deficient phenotype.

Science, 1992 Oct 16, 258(5081), 471 - 5
Formation of a gated channel by a ligand-specific transport protein in the bacterial outer membrane; Rutz JM et al.; The ferric enterobactin receptor (FepA) is a high-affinity ligand-specific transport protein in the outer membrane of Gram-negative bacteria . Deletion of the cell-surface ligand-binding peptides of FepA generated mutant proteins that were incapable of high-affinity uptake but that instead formed nonspecific, passive channels in the outer membrane . Unlike native FepA, these pores acted independently of the accessory protein TonB, which suggests that FepA is a gated porin and that TonB acts as its gatekeeper by facilitating the entry of ligands into the FepA channel . The sequence homology among TonB-dependent proteins suggests that all ligand-specific outer membrane receptors may function by this gated-porin mechanism.

Oral Microbiol Immunol, 1992 Oct, 7(5), 291 - 8
Characterization of shared antigens of Fusobacterium nucleatum and Fusobacterium necrophorum; Kaur M et al.; Fusobacterium nucleatum and Fusobacterium necrophorum are gram-negative, non-spore-forming anaerobic rods, frequently isolated from the normal flora and diseased lesions of the human oral cavity, gastrointestinal and genitourinary tracts . F . necrophorum is also known to be an animal pathogen . Studies were undertaken with rabbit anti-F . nucleatum sera and with human adult periodontitis (AP) sera that demonstrated the sharing of antigens between the two species . Immunodiffusion and immunoelectrophoresis studies of Fusobacterium species with rabbit anti-F . nucleatum sera demonstrated the presence of shared antigen(s) between F . nucleatum and F . necrophorum . Adsorption studies of AP sera in an enzyme-linked immunosorbent assay demonstrated the presence of antibodies reacting with the shared antigens of the two species . Immunoblot (IB) analysis of a soluble protein preparation of the two species of Fusobacterium, when allowed to react with rabbit anti-F . nucleatum 10197 serum, demonstrated 53 kDa and 30 kDa bands present in members of the two species . Further, IB analysis of protein preparations of the two species with AP sera indicated the presence of antibodies reacting with the shared 53 kDa band and in some cases the 30 kDa band . During serological testing with antisera or host immune studies with human sera to these species, the presence of shared antigens must be considered.

J Surg Res, 1992 Oct, 53(4), 384 - 90
Optimal method for culturing vascular prosthetic grafts; Padberg FT Jr et al.; Vascular prosthetic infection may be underrecognized when identified by standard culture techniques . Improved microbiologic methodology may enhance detection of bacteria in prosthetic graft specimens, and thus may alter clinical decisions . Quantitative culture techniques were employed to compare three methods of enhancing bacterial recovery from Dacron graft cylinders seeded with commonly encountered bacterial pathogens . Methods included: (1) ultrasonic bath treatment, (2) direct ultrasonic disruption, and (3) agitation on a Vortex mixer . Ultrasonic bath treatment released bacteria with colony counts that were consistently greater by 1 log than direct ultrasonic disruption and Vortex agitation . Direct ultrasonic disruption at high energy levels selectively killed gram-negative bacteria by as much as a 4 log decrease in viable organisms . Agitation (Vortex mixing) of the specimen produced the lowest counts among the three methods tested . These data would indicate that a 5-min ultrasonic bath treatment was the optimal method of preparation of vascular prostheses for bacterial culture.

Pharmacoeconomics, 1992 Nov, 2(5), 408 - 13
The economic impact of HA-1A (Centoxin) against endotoxin; Barriere SL; Monoclonal antibodies have been shown to reduce morbidity and mortality in selected subsets of patients with Gram-negative sepsis and/or septic shock . However, the acquisition costs of the antibody products are expected to be in the range of $US3500 to $US4000 per course of therapy and precise identification of patients who will benefit may be difficult . Therefore, the economic impact of these antibodies will be significant . We have performed a model cost-effectiveness and cost-benefit analysis specific to our institution based on previously reported mortality figures . Our data suggest that the cost-effectiveness of HA-1A (Centoxin) will be comparable with that of a variety of commonly used medical interventions, but will produce an incremental increase in costs of at least $US7000 per patient because of the acquisition cost of the drug, as well as an increase in numbers of survivors whose hospitalisation will be prolonged.

J Infect Dis, 1992 Oct, 166(4), 847 - 53
Cytokine release from microglia: differential inhibition by pentoxifylline and dexamethasone; Chao CC et al.; Cytokines have been implicated in the pathogenesis of gram-negative bacterial meningitis . The effects of pentoxifylline and dexamethasone on the release of tumor necrosis factor (TNF), interleukin (IL)-1, and IL-6 from primary murine microglial cell cultures were explored using bioassays . When added concomitantly with lipopolysaccharide, pentoxifylline blocked the release of TNF and IL-1 but not IL-6, while dexamethasone inhibited the release of TNF and IL-6 . After a 2-h exposure of microglia to lipopolysaccharide, pentoxifylline but not dexamethasone still inhibited the release of TNF . Release of TNF was enhanced 20-fold by priming of the microglia with interferon-gamma; only pentoxifylline blocked the priming effect of interferon-gamma on TNF release . These results demonstrate that pentoxifylline and dexamethasone differentially regulate the release of cytokines in microglial cell cultures and provide potential insight into their role in the treatment of gram-negative bacterial meningitis.

Can J Vet Res, 1992 Oct, 56(4), 318 - 25
Ultrastructure and molecular characterization of Fusobacterium necrophorum biovars; Garcia MM et al.; The ultrastructural features and molecular components of 18 strains of Fusobacterium necrophorum biovars A, AB and B, isolated from animal and human infections, were examined by electron microscopy, multilocus enzyme electrophoresis (MEE) and by sodium dodecyl sulfate-gradient polyacrylamide gel electrophoresis (SDS-PAGE) . High resolution scanning electron microscopy revealed that the strains possessed a convoluted surface pattern . Transmission electron microscopy showed that all strains possessed a cell wall structure typical of gram-negative bacteria . Bleb formation was not uncommon . Numerous extracellular materials, resembling lipopolysaccharide (LPS) fragments, surrounded cells of both human strains and biovar B animal strains . Biovar A field strains revealed capsules as stained by ruthenium red whereas a stock culture strain showed the capsule only when immunostabilized with hyperimmune serum . Starch gel electrophoresis showed all strains to possess adenyl kinase, glutamate dehydrogenases and lactate dehydrogenase; each enzyme migrated uniformly (monomorphic) among the strains and represented an electrotype . However, SDS-PAGE indicated differences in the protein profiles between all of the strains; the most distinctly different was a human isolate (FN 606) . Silver staining to detect LPS showed extensive "ladder" patterns among the majority of biovar A strains but not in the animal biovar B strains . Immunoblotting of LPS with a rabbit antiserum prepared against phenol extracted LPS from a biovar A animal isolate (LA 19) suggested marked variability in the LPS antigens among the isolates studied.

Lab Anim, 1992 Oct, 26(4), 288 - 94
Spiral bacterium associated with gastric, ileal and caecal mucosa of mice; Queiroz DM et al.; A spiral shaped bacterium was seen in smears and histological sections (stained by carbolfuchsin) of gastric, ileal and caecal mucosa as well as in stool smears from mice . A significant correlation between the presence of the spiral bacterium and the occurrence of gastritis was observed but the ileal and caecal mucosa seemed unaffected . The bacterium was Gram negative and grew on BHM and Skirrow's medium, under microaerophilic conditions, at 37 degrees C . Its major biochemical characteristics included positive catalase and oxidase reactions and a rapidly positive urease test . There were 2 or 3 spiral turns per cell and a tuft of up to 12 sheathed flagella on each pointed end . Entwined, braided periplasmic fibrils covered the surface of the cell . This spiral bacterium seemed to be part of the normal intestinal flora but was associated with gastritis.

Antimicrob Agents Chemother, 1992 Oct, 36(10), 2349 - 51
Phase I study of antilipopolysaccharide human monoclonal antibody MAB-T88; Daifuku R et al.; Monoclonal antibody MAB-T88 is a human monoclonal immunoglobulin M antibody directed at the lipopolysaccharide of gram-negative bacteria . In this study, nine patients who were expected to become neutropenic from antineoplastic chemotherapy received an infusion of MAB-T88, three patients at each of three doses: 1, 4, and 8 mg/kg of body weight . MAB-T88 was shown to be safe, with an effective half-life in plasma of 25.4 h, and no patient developed immunoglobulin G antibody to MAB-T88.

Antimicrob Agents Chemother, 1992 Oct, 36(10), 2139 - 46
Endotoxin concentration in neutropenic patients with suspected gram-negative sepsis: correlation with clinical outcome and determination of anti-endotoxin core antibodies during therapy with polyclonal immunoglobulin M-enriched immunoglobulins; Behre G et al.; We carried out a study in patients with severe neutropenia from hematologic malignancy and suspected gram-negative sepsis to evaluate the clinical significance of endotoxin concentrations in plasma before and during a therapeutic intervention with a human polyclonal immunoglobulin M (IgM)-enriched immunoglobulin preparation (Pentaglobin; Biotest, Dreieich, Germany) . Twenty-one patients with acute leukemia or non-Hodgkin's lymphoma entered the study upon the development of clinical signs of gram-negative sepsis and received the IgM-enriched immunoglobulin preparation every 6 h for 3 days (total dose, 1.3 liter with 7.8 g of IgM, 7.8 g of IgA, and 49.4 g of IgG), in addition to standardized antibiotic treatment . Concentrations of endotoxin and IgM and IgG antibodies against lipid A and Re lipopolysaccharide (LPS) in plasma were determined by a modified chromogenic Limulus amebocyte lysate test and semiquantitative enzyme linked immunosorbent assay, respectively, before each immunoglobulin infusion and during the following 25 days . Seventeen patients were endotoxin positive; in five of these patients, gram-negative infection was confirmed by microbiologic findings . Prior to therapy, endotoxemia correlated significantly with the occurrence of fever, and a quantitative correlation between the endotoxin concentration and body temperature was found during the individual course of infection in 8 of the 17 patients . Overall mortality from endotoxin-positive sepsis was 41% (7 of 17) and 64% (7 of 11) in patients with symptoms of septic shock . Nonsurvivors had significantly higher maximum concentration of endotoxin in plasma compared with those of survivors at the first study day (median of 126 versus 34 pg/ml; P < 0.05) and during the whole septic episode (median of 126 versus 61 pg/ml; P < 0.05) . In survivors, immunoglobulin therapy resulted in a significant decrease in endotoxin levels in plasma within the initial 18-h treatment period, from a pretreatment median value of 28 pg/ml to a value of 8 pg/ml (P< 0.05) . In the seven patients who died from uncontrollable infection, no effect of therapy on endotoxin levels in plasma was observed . IgM and IgG antibodies against lipid A and Re LPS increased significantly under immunoglobulin treatment, with significant correlations between antibodies against lipid A and Re LPS . These data strongly suggest a prognostic significance of the endotoxin levels in plasma and a potential effect of treatment with a polyclonal IgM-enriched immunoglobulin preparation . Further studies are needed to substantiate these findings and to assess the impact on the clinical course by way of a prospective placebo-controlled clinical trial.

Am Rev Respir Dis, 1992 Oct, 146(4), 1059 - 66
Nosocomial bronchopneumonia in the critically ill . Histologic and bacteriologic aspects; Rouby JJ et al.; To provide a comprehensive description of the histologic and bacteriologic characteristics of human nosocomial bronchopneumonia (BPN), the lungs of 83 critically ill patients decreased after a period of mechanical ventilation were examined in the immediate postmortem period . In addition, the accuracy of the protected minibronchoalveolar lavage (BAL) technique in the diagnosis of nosocomial BPN was evaluated . In each patient, a surgical pneumonectomy was performed at the bedside within 30 min following death . Each pulmonary lobe was sampled and bacteriologically analyzed using semiquantitative cultures in 50 patients and quantitative cultures in 33 patients . The entire lung was histologically analyzed using 5 to 10 slices per lung segment . In 69 patients, the bacteriologic result of a protected mini-BAL performed within 48 h preceding death was compared with histologic and bacteriologic results of study of the lung tissue itself . Histologic lesions of BPN were found in 43 of the 83 lungs examined . These lesions were (1) severe in the majority of patients (confluent BPN, n = 23; lung abscess, n = 6), (2) preferentially found in dependent lung segments, (3) often associated with nonspecific alveolar damage, (4) associated with positive lung cultures in 65% of patients (53% with gram-negative bacteria), (5) polymicrobial in 28% of patients, (6) characterized by a lobar bacterial burden greater than 10(3) cfu/g in 32% of cases . Using semiquantitative bacteriologic analysis, the sensitivity and the specificity of the protected mini-BAL in the diagnosis of nosocomial BPN were found to be 70 and 69%, respectively . Protected mini-BAL identified 77% of causative microorganisms of BPN.(ABSTRACT TRUNCATED AT 250 WORDS)

Oral Surg Oral Med Oral Pathol, 1992 Oct, 74(4), 431 - 6
Effect of extraction of partly erupted third molars on subgingival microorganisms; Rajasuo A et al.; This study was made to investigate the effect of extraction of third molars on subgingival microbes in 39 generally and gingivally healthy men with an average age of 20.2 years (SD 0.9) . Microbial samples were taken from the pericoronal space of symptom-free partly erupted lower third molars and from the adjacent gingival pockets of the second molars . The samples were cultivated anaerobically . All partly erupted third molars were extracted from 20 subjects . A control group of 19 subjects was left untreated . Microbe sampling was repeated 2 and 5 months postoperatively with highly significant results . It was shown that at baseline the number of black-pigmented gram-negative bacteria and Fusobacterium species was more frequent in third molar than in second molar sites . The total bacterial count decreased significantly at the second molar sites after extraction of the third molars when compared with the control group . Before the extractions, black-pigmented gram-negative bacteria were detected in 45% of the test subjects and Actinobacillus actinomycetemcomitans in 20% . The respective postoperative figures were 30% for black-pigmented gram-negative bacteria and 10% for Actinobacillus actinomycetemcomitans . Capnocytophaga species were not affected by the extractions . The findings suggest that erupting third molars may harbor harmful bacteria that can be reduced by eradicating the foci.

J Bacteriol, 1992 Oct, 174(20), 6617 - 23
Electrotransformation of Thiobacillus ferrooxidans with plasmids containing a mer determinant; Kusano T et al.; The mer operon from a strain of Thiobacillus ferrooxidans (C . Inoue, K . Sugawara, and T . Kusano, Mol . Microbiol . 5:2707-2718, 1991) consists of the regulatory gene merR and an operator-promoter region followed by merC and merA structural genes and differs from other known gram-negative mer operons . We have constructed four potential shuttle plasmids composed of a T . ferrooxidans-borne cryptic plasmid, a pUC18 plasmid, and the above-mentioned mer determinant as a selectable marker . Mercury ion-sensitive T . ferrooxidans strains were electroporated with constructed plasmids, and one strain, Y4-3 (of 30 independent strains tested), was found to have a transformation efficiency of 120 to 200 mercury-resistant colonies per microgram of plasmid DNA . This recipient strain was confirmed to be T . ferrooxidans by physiological, morphological, and chemotaxonomical data . The transformants carried a plasmid with no physical rearrangements through 25 passages under no selective pressure . Cell extracts showed mercury ion-dependent NADPH oxidation activity.

J Bacteriol, 1992 Oct, 174(19), 6018 - 24
Identification of a Bdellovibrio bacteriovorus genetic locus, hit, associated with the host-independent phenotype; Cotter TW et al.; Bdellovibrios invade and grow within the periplasmic space of suitable gram-negative bacteria . Wild-type bdellovibrios are obligately dependent on host cells for growth, but spontaneous host-independent (H-I) mutants that grow axenically on standard rich culture media can be isolated . Such mutants generally retain the ability to grow intraperiplasmically, although the plaques that they produce on lawns of host cells are smaller and more turbid than those produced by wild-type bdellovibrios . Here, we identify the first genetic locus associated with the H-I phenotype: hit (host interaction) . We show that three individual H-I mutants suffered mutations at the hit locus and that recombination of wild-type hit sequences into the genomes of the H-I mutants greatly enhanced their plaquing ability . DNA sequence analysis localized the hit mutation in each of the H-I mutants to a 135-bp region of the genome . Mutations at hit may not fully account for the H-I phenotype, however, as recombination of wild-type hit sequences into the genomes of the H-I mutants had little effect on the axenic-growth phenotype of the mutants . Possible explanations for this result and potential roles for the hit locus are discussed.

J Bacteriol, 1992 Oct, 174(19), 6011 - 7
A conjugation procedure for Bdellovibrio bacteriovorus and its use to identify DNA sequences that enhance the plaque-forming ability of a spontaneous host-independent mutant; Cotter TW et al.; Wild-type bdellovibrios are obligate intraperiplasmic parasites of other gram-negative bacteria . However, spontaneous mutants that can be cultured in the absence of host cells occur at a frequency of 10(-6) to 10(-7) . Such host-independent (H-I) mutants generally display diminished intraperiplasmic-growth capabilities and form plaques that are smaller and more turbid than those formed by wild-type strains on lawns of host cells . An analysis of the gene(s) responsible for the H-I phenotype should provide significant insight into the nature of Bdellovibrio host dependence . Toward this end, a conjugation procedure to transfer both IncQ and IncP vectors from Escherichia coli to Bdellovibrio bacteriovorus was developed . It was found that IncQ-type plasmids were capable of autonomous replication in B . bacteriovorus, while IncP derivatives were not . However, IncP plasmids could be maintained in B . bacteriovorus via homologous recombination through cloned B . bacteriovorus DNA sequences . It was also found that genomic libraries of wild-type B . bacteriovorus 109J DNA constructed in the IncP cosmid pVK100 were stably maintained in E . coli; those constructed in the IncQ cosmid pBM33 were unstable . Finally, we used the conjugation procedure and the B . bacteriovorus libraries to identify a 5.6-kb BamHI fragment of wild-type B . bacteriovorus DNA that significantly enhanced the plaque-forming ability of an H-I mutant, B . bacteriovorus BB5.

Infect Immun, 1992 Oct, 60(10), 4133 - 9
Differential effects of monoclonal antibodies to tumor necrosis factor alpha and gamma interferon on induction of hepatic nitric oxide synthase in experimental gram-negative sepsis; Evans T et al.; To investigate the stimuli required for the induction of nitric oxide synthase (NOS) in sepsis, we have analyzed the levels of this enzyme in the livers of mice infected with a 90% lethal dose of Escherichia coli in a model of gram-negative sepsis . Hepatic NOS levels are markedly induced in this model, with peak values occurring 12 to 22 h following infection . Treatment with TN3-19.12, a neutralizing monoclonal antibody to tumor necrosis factor alpha (TNF-alpha), resulted in complete protection from death in this model of sepsis but had no significant effect on the level of induction of hepatic NOS . Treatment with H22, a monoclonal antibody to gamma interferon (IFN-gamma), also gave significant protection against death and, in addition, did lead to a decrease in the level of induction of the hepatic NOS . Treatment of mice with pure TNF-alpha (0.2 microgram), IFN-gamma (2,000 U), or a combination of the two did not induce the hepatic NOS, but treatment with the combination led to significant mortality (probability of survival at 22 h, 0.32) . Thus, the level of induction of NOS within the liver either in sepsis or by the coadministration of TNF-alpha and IFN-gamma does not correlate with death.

Am J Respir Cell Mol Biol, 1992 Oct, 7(4), 399 - 405
Chinese hamster ovarian cell glycoproteins that mediate type 1 piliated gram-negative bacterial adherence; Dal Nogare AR et al.; We used Chinese hamster ovary (CHO) cell lines to define the structures of glycoproteins responsible for Type 1 piliated bacterial adherence . CSH 50 Escherichia coli, a Type 1 piliated bacteria, adhered significantly better than an isogenic nonpiliated E . coli to all CHO lines tested . CSH 50 E . coli adhered least well to CHO cells expressing intact complex type oligosaccharides on cell surface glycoproteins . CSH 50 adherence increased when shorter oligosaccharides were present and was maximal when mannose groups were present in terminal, nonreducing positions . Five high mannose type glycoproteins, with molecular weights of 79, 75, 55, 50, and 37 kD, were identified as high affinity ligands for Type 1 piliated bacteria . Our results suggest that alterations in cell surface carbohydrates may increase adherence of Type 1 piliated gram-negative bacteria to cells.

Rev Med Chil, 1992 Oct, 120(10), 1140 - 3
{Gonococcal urethritis in men: clinical experience in 1978-1988}; Benavides MI et al.; Nine hundred thirty four samples of urethral secretion proceeding from male subjects without previous treatment were analyzed with Gram stain . These subjects consulted in Santiago Western area, and belong to medium and low socioeconomical status . In 82.5% of samples, intracellular gram-negative diplococci were found, that made the presumptive diagnosis of gonococcal urethritis . The importance of direct examination of urethral secretion for the diagnosis and immediate treatment of gonococcal urethritis is emphasized.

J Bacteriol, 1992 Oct, 174(20), 6377 - 85
Roles of the Tn21 merT, merP, and merC gene products in mercury resistance and mercury binding; Hamlett NV et al.; The mercury resistance (mer) operon of the gram-negative transposon Tn21 encodes not only a mercuric reductase and regulatory genes but also two inner membrane proteins (MerT and MerC) and a periplasmic protein (MerP) . Although the merT, merP, and merC genes have been implicated in Hg(II) transport, the individual roles of these genes have not been established . We created in vitro precise deletion and frameshift mutations that eliminated each of the genes singly and in combination . Our results show that both merT and merP are required for Hg(II) binding but that merC is not . Both merT and merP are required for full expression of Hg(II) resistance, but loss of merP is less deleterious than loss of merT . Furthermore, mutations eliminating both merT and merP decrease resistance more than the single mutations do . In contrast, mutating merC had no effect on Hg(II) resistance . Both the merT and merP mutations increase the threshold Hg(II) concentration for induction of merA-lacZ transcriptional fusions and cause an increase in the maximal expression level . In contrast, the merC mutation had little effect on the threshold inducing concentration of Hg(II) but decreased the level of expression . Our results show that merT and merP alone are sufficient to specify a mercury transport system . The role of merC remains obscure.

J Bacteriol, 1992 Oct, 174(19), 6294 - 7
Decreased function of the class B tetracycline efflux protein Tet with mutations at aspartate 15, a putative intramembrane residue; McMurry LM et al.; The aspartate 15 residue within the first predicted intramembrane helix of the tetracycline efflux protein Tet has been conserved in four tetracycline resistance determinants from gram-negative bacteria . Its replacement in class B Tet by tyrosine, histidine, or asparagine resulted in a 60 to 85% loss of tetracycline resistance and a similar loss of tetracycline-proton antiport . The tyrosine and histidine substitutions lowered the Vmax of the efflux system by some 90% but did not alter the Km . The asparagine substitution raised the Km over 13-fold, while the Vmax was equal to or greater than that of the wild type . Therefore, although the nature of its role is unclear, aspartate 15 is important for normal Tet function.

J Endod, 1992 Oct, 18(10), 501 - 4
Endotoxin and gram-negative bacteria in the rat periapical lesions; Yamasaki M et al.; The purpose of this study was to measure the amount of endotoxin as well as to identify Gram-negative bacteria in experimental periapical lesions in rats . Molar pulps were exposed and infected and the amount of endotoxin in the periapical tissue of the right mandibular first molar was measured by Endospecy, while the colony number of Gram-negative bacteria was determined in the same region of the left mandibular first molar . In the control animals, the amount of endotoxin in the periapical tissues did not change at all during the experimental period, and no Gram-negative bacteria were isolated . In the experimental animals, the amount of endotoxin in the periapical tissues increased gradually from 1 to 70 days, and its level was significantly greater than that of control animals after 7 days . Gram-negative bacteria were isolated from the periapical tissues and their number gradually increased from 1 to 14 days (26 to 82%), but decreased at 21 days . It was approximately 60% from 28 to 70 days . The results of this study showed that the amount of endotoxin in the periapical tissues gradually increased with increasing time and that Gram-negative bacteria were isolated from the same region but did not increase in number concurrently with the increase in the amount of endotoxin.

Antimicrob Agents Chemother, 1992 Oct, 36(10), 2280 - 5
Salicylate-inducible antibiotic resistance in Pseudomonas cepacia associated with absence of a pore-forming outer membrane protein; Burns JL et al.; The most common mechanism of antibiotic resistance in multiply resistant Pseudomonas cepacia is decreased porin-mediated outer membrane permeability . In some gram-negative organisms this form of antibiotic resistance can be induced by growth in the presence of weak acids, such as salicylates, which suppress porin synthesis . To determine the effects of salicylates on outer membrane permeability of P . cepacia, a susceptible laboratory strain, 249-2, was grown in 10 mM sodium salicylate . Antibiotic susceptibility and uptake, as well as outer membrane protein patterns, were compared between strain 249-2 grown with and without salicylates . The MICs of chloramphenicol, trimethoprim, ciprofloxacin, and ceftazidime were compared between organisms grown in standard and salicylate-containing medium and are as follows: chloramphenicol, 12.5 versus 100 micrograms/ml; trimethoprim, 0.78 versus 3.125 micrograms/ml; ciprofloxacin, 0.4 versus 1.56 micrograms/ml; ceftazidime, 3.125 versus 3.125 micrograms/ml . The permeability of beta-lactam antibiotics was calculated from the rate of hydrolysis of the chromogenic cephalosporin, PADAC . There was no significant difference between strains grown in the presence and absence of salicylate . By using high-pressure liquid chromatography quantitation of loss from culture medium, the effect of 10 mM salicylate on the cellular permeability of chloramphenicol was measured in strain 249-2 by introduction of a plasmid which encodes production of chloramphenicol acetyltransferase . After 1 h of incubation, 18.5% +/- 1.54% versus 70.1% +/- 3.52%, and after 2 h, 4.20% +/- 1.65% versus 41.90% +/- 2.16% remained in supernatants from organisms grown in the absence and presence of 10 mM salicylate, respectively . Outer membrane protein pattern analysis demonstrated the absence of a protein of apparent molecular weight of 40,000 when strain 249-2 was grown in the presence of 10 mM salicylate . To determine whether this protein functioned as a porin, reconstituted membrane vesicles were constructed to assess antibiotic permeability . Vesicles constructed with this salicylate-suppressible outer membrane protein (OpcS) were permeable to chloramphenicol but not to penicillin G . These findings suggest that OpcS is a selective, antibiotic-permeable porin which can be suppressed by growth in the presence of salicylate . Further investigation will be required to determine the biochemical effects of salicylate on porin synthesis.

J Biol Chem, 1992 Sep 15, 267(26), 18896 - 901
Purification and characterization of a 50-kDa cysteine proteinase (gingipain) from Porphyromonas gingivalis; Chen Z et al.; Porphyromonas gingivalis, a Gram-negative anaerobic rod, has been closely associated with the initiation and progression of periodontal disease . This organism has been shown to produce a large number of proteolytic enzymes which can degrade a variety of tissue proteins, and these are considered to be major virulence factors . One of the proteinases produced by this organism, referred to as gingipain-1, has been purified to homogeneity from P . gingivalis culture medium by a combination of gel filtration and ion-exchange chromatography . The enzyme was found to have a molecular mass near 50 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, a pH optimum in the neutral to alkaline range, and a requirement for cysteine for activation and Ca2+ for stabilization . Amino-terminal sequence analysis indicated that gingipain belongs to a new, so far unknown, subfamily of cysteine proteinases . Three unusual features of this proteinase are: (a) the stimulation of amidolytic activity by glycine-containing dipeptides; (b) a narrow specificity which is limited to peptide bonds containing arginine residues; and (c) resistance to inhibition by proteinase inhibitors in human plasma.

J Biol Chem, 1992 Sep 15, 267(26), 18702 - 7
A novel 3-deoxy-D-manno-octulosonic acid transferase from Chlamydia trachomatis required for expression of the genus-specific epitope; Belunis CJ et al.; DNA cloned from Chlamydia trachomatis is able to direct the formation of the genus-specific lipopolysaccharide epitope of chlamydiae in enteric Gram-negative bacteria . We now demonstrate that a single C . trachomatis gene (gseA) is sufficient to impart this trait to Escherichia coli . The deduced amino acid sequence of gseA shows 23% identity (66% similarity) to kdtA, an E . coli gene that codes for a bifunctional enzyme catalyzing the addition of two 3-deoxy-D-manno-octulosonic acid (Kdo) residues to lipid A precursors (Clementz, T., and Raetz, C . R . H . (1991) J . Biol . Chem . 266, 9687-9696) . Extracts of E . coli expressing gseA transfer at least one additional Kdo unit from CMP-Kdo to precursors already bearing the two Kdo residues attached by the kdtA gene product . Introduction of gseA into an E . coli mutant with a thermolabile kdtA gene product endows cell extracts with the ability to transfer not only the third but also the first two Kdos to lipid A precursors, demonstrating that the C . trachomatis enzyme is at least trifunctional . Given the similarities of these two Kdo transferases and the essentiality of Kdo in Gram-negative bacteria, lipopolysaccharide biosynthesis may be a target for development of novel drugs effective against chlamydiae.

Biochemistry, 1992 Sep 8, 31(35), 8344 - 8
Aspartic acid-66 is the only essential negatively charged residue in the putative hydrophilic loop region of the metal-tetracycline/H+ antiporter encoded by transposon Tn10 of Escherichia coli; Yamaguchi A et al.; Of the 16 acidic amino acid residues located in the hydrophilic region of the metal-tetracycline/H+ antiporter of transposon Tn10, five glutamic acids and three aspartic acids are conserved among the tetracycline/H+ antiporters of Gram-negative bacteria . When these conserved acidic residues were each replaced by a neutral polar residue, glutamine or asparagine, only the Asp66 substitution mutants completely lost their transport activity . The substitution of Glu274, Asp120, Glu181, or Asp38 caused significant reduction of the transport activity, whereas the substitution of the other three residues had no detectable effect on the activity . These findings led to the conclusion that only Asp66 is essential for the transport function.

Med J Aust, 1992 Sep 7, 157(5), 308 - 11
Prospective audit of an aminoglycoside consultative service in a general hospital; Li SC et al.; OBJECTIVE: To investigate the impact of the introduction of a consultative service on the use, efficiency of dosing and clinical toxicology of the aminoglycoside antibiotics, gentamicin and tobramycin, in a general hospital . METHODS: Two audits were conducted six months and 18 months after the introduction of the consultative service . The audits reviewed the use of drug assay services, the adequacy of drug administration (as measured by serum antibiotic concentrations), indications for prescription, adverse outcomes (by noting markers of nephrotoxicity) and the antibiotic sensitivity of Gram-negative pathogens . The results were compared with the results of an audit conducted before the consultative service was instituted . RESULTS: There was a significant (P < 0.001 by chi 2 test) increase in the use of assays, with drug assays performed in 67% (first audit) and 77% (second audit) of aminoglycoside courses compared with 48.2% in the pre-intervention audit . Sample timing was greatly improved, with more than 70% of the samples collected at the appropriate times . Assay wastage in terms of uninterpretable assay results decreased significantly (P < 0.001) from 42.9% of total assays to 6.3% at the first audit and 3.8% at the second audit . The percentage of assay results in the desirable range increased significantly (P < 0.001) from 39.1% to 71.9% (first audit) and 75.4% (second audit) . Pharmacokinetic recommendations were made in 39.1% and 64% of all aminoglycoside courses during the first and second audits respectively, with clinician acceptance of dosage recommendations at 83.1% and 82.8% respectively . For aminoglycoside courses prescribed for therapeutic reasons, 97.9% (first audit, n = 325) and 98.6% (second audit, n = 280) of indications for use were judged as clinically appropriate . The incidence of suspected aminoglycoside-induced nephrotoxicity was reduced from 8.9% of patients to 1.6% (first audit, P < 0.001) and 2.4% (second audit) . Bacterial sensitivity audits showed that the great majority of clinical isolates of target organisms (n = 3523, Year 1 and n = 3385, Year 2) were sensitive to gentamicin (92.2% and 91.5% respectively) and tobramycin (98.1% and 98.8% respectively); these aminoglycosides exceeded all alternative agents in effectiveness, including first and third generation cephalosporins . CONCLUSIONS: The overall results indicate that introduction of the consultative service had a positive impact on the effective use of aminoglycosides, with a marked decrease in clinical toxicity . These influences were shown to persist for at least 18 months . The availability of reliable predictive techniques to reduce toxicity allows active promotion of aminoglycosides as the agents of choice on grounds of efficacy and economy.

J Lab Clin Med, 1992 Sep, 120(3), 465 - 70
Diltiazem treatment of endotoxic shock in suckling rats; Goto M et al.; Gram-negative sepsis septic shock continues to produce significant mortality and therefore remains a major medical problem . Vasodilators have been studied in the treatment of circulatory shock . However, the effectiveness of calcium channel blockers in the treatment of newborn endotoxic shock has not been well documented . In the present study, diltiazem, a calcium channel blocker, and nitroprusside, a vasodilator, were used for the treatment of endotoxic shock in 10-day-old rats . Mortality rate, hemodynamics, and glucose metabolism were monitored . Diltiazem at a dose of 0.3 mg/kg attenuated the hypotension, bradycardia, hypoglycemia, and lactacidemia in newborn endotoxic shock . Diltiazem treatment resulted in reduced 24-hour mortality . However, 0.6 mg/kg diltiazem enhanced the hypotension, bradycardia, and lactacidemia in endotoxic shock . Nitroprusside blunted the hypoglycemia and decreased the mortality rate among rats with endotoxic shock . Afterload reduction may be responsible for the beneficial effects of 0.3 mg/kg diltiazem and nitroprusside . Diltiazem at a dose of 0.3 mg/kg reduced the lactacidemia of endotoxic shock more than nitroprusside . Therefore the effects of diltiazem may be due not only to afterload reduction but also to inhibition of cellular calcium influx . We conclude that 0.3 mg/kg diltiazem and 1.0 mg/kg nitroprusside are beneficial for the treatment of endotoxic shock in newborn rats.

Cleft Palate Craniofac J, 1992 Sep, 29(5), 463 - 9
Microbiota associated with residual clefts and neighboring teeth in patients with cleft lip, alveolus, and palate; Mombelli A et al.; Twenty patients with residual clefts or pronounced soft tissue grooves, treated for uni- or bilateral cleft lip, alveolus, and palate were included in this study . Ten patients were recalled for dental prophylaxis at regular intervals, 10 patients were not . One microbiologic sample was obtained from the cleft area and two samples from a tooth adjacent to the cleft (sites adjacent and distant to the cleft) . Between the recall and the nonrecall group there were notable differences in the presence of anaerobic Gram-negative organisms . Fusobacterium spp., Prevotella melaninogenica, and P . intermedia were more often found in nonrecall patients . While rarely seen in recall patients, spirochetes and motile rods were a common feature of nonrecall patients . The putative periodontal pathogens Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis were not detected in either group . The differences between the recall and the nonrecall groups were more pronounced when the respective samples from teeth were related to each other than when the samples obtained from the clefts were compared . The cleft flora was less complex irrespective of how good maintenance was and resembled the flora of teeth of well-maintained patients . Samples from clefts were never Wolinella positive, and harbored significantly less Capnocytophaga and Actinomyces viscosus than samples from dental sites.

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