Microbial

Sci Prog, 2003, 86(Pt 1-2), 115 - 37
Lethal effects of heat on bacterial physiology and structure; Russell AD; High temperatures have profound effects on the structural and physiological properties of sporulating and non-sporulating bacteria, with membranes, RNA, DNA, ribosomes, protein and enzymes all affected . Nevertheless, it is apparent that no one single event is responsible for cell death . The induction of intracellular heat-shock proteins and the activation of extracellular alarmones in vegetative cells exposed to mildly lethal temperatures are important cell responses . In bacterial spores, several factors contribute to the overall resistance to moist (wet) and dry heat; the latter, but not the former, induces mutations . Heat resistance develops during sporulation, when spore-specific heat-shock proteins are also produced . Heat sensitivity is regained during germination of spores.

Sci Prog, 2003, 86(Pt 1-2), 9 - 75
Bacterial cold shock responses; Weber MH et al.; As a measure for molecular motion, temperature is one of the most important environmental factors for life as it directly influences structural and hence functional properties of cellular components . After a sudden increase in ambient temperature, which is termed heat shock, bacteria respond by expressing a specific set of genes whose protein products are designed to mainly cope with heat-induced alterations of protein conformation . This heat shock response comprises the expression of protein chaperones and proteases, and is under central control of an alternative sigma factor (sigma 32) which acts as a master regulator that specifically directs RNA polymerase to transcribe from the heat shock promotors . In a similar manner, bacteria express a well-defined set of proteins after a rapid decrease in temperature, which is termed cold shock . This protein set, however, is different from that expressed under heat shock conditions and predominantly comprises proteins such as helicases, nucleases, and ribosome-associated components that directly or indirectly interact with the biological information molecules DNA and RNA . Interestingly, in contrast to the heat shock response, to date no cold-specific sigma factor has been identified . Rather, it appears that the cold shock response is organized as a complex stimulon in which post-transcriptional events play an important role . In this review, we present a summary of research results that have been acquired in recent years by examinations of bacterial cold shock responses . Important processes such as cold signal perception, membrane adaptation, and the modification of the translation apparatus are discussed together with many other cold-relevant aspects of bacterial physiology and first attempts are made to dissect the cold shock stimulon into less complex regulatory subunits . Special emphasis is placed on findings concerning the nucleic acid-binding cold shock proteins which play a fundamental role not only during cold shock adaptation but also under optimal growth conditions.

J Bacteriol, 2003 Jul, 185(14), 4226 - 32
A bacterial TrwC relaxase domain contains a thermally stable alpha-helical core; Arrondo JL et al.; The TrwC protein is the relaxase-helicase responsible for the initiation and termination reactions of DNA processing during plasmid R388 conjugation . The TrwC-N275 fragment comprises the 275-amino-acid N-terminal domain of the protein that contains the DNA cleavage and strand transfer activities (the relaxase domain) . It can be easily purified by keeping a cell lysate at 90 degrees C for 10 min . Infrared spectroscopy shows that this domain has a predominantly alpha/beta structure with some amount of unordered structure . Fast heating and cooling does not change the secondary structure, whereas slow heating produces two bands in the infrared spectrum characteristic of protein aggregation . The denaturation temperature is increased in the protein after the fast-heating thermal shock . Two-dimensional infrared correlation spectroscopy shows that thermal unfolding is a very cooperative two-state process without any appreciable steps prior to aggregation . After aggregation, the alpha-helix percentage is not altered and alpha-helix signal does not show in the correlation maps, meaning that the helices are not affected by heating . The results indicate that the domain has an alpha-helix core resistant to temperature and responsible for folding after fast heating and an outer layer of beta-sheet and unordered structure that aggregates under slow heating . The combination of a compact core and a flexible outer layer could be related to the structural requirements of DNA-protein binding.

Mol Genet Genomics, 2003 Aug, 269(5), 685 - 91 Epub 2003 Jun 27.
Construction and application of a bacterial artificial chromosome (BAC) library of Prunus armeniaca L . for the identification of clones linked to the self-incompatibility locus; Vilanova S et al.; To facilitate gene discovery in the Rosaceae, a bacterial artificial chromosome (BAC) library was constructed using high-molecular-weight (HMW) DNA from apricot leaves (Prunus armeniaca L.) . The library contains 101,376 clones (264, 384-well plates) with an average insert size of 64 kb, equivalent to 22-fold genome coverage . In the first application of this library, high-density filters were screened for self-incompatibility genes using apricot DNA probes . Eight positive BAC clones were detected and fingerprinted to determine clone relationships and assemble contigs . These results demonstrate the suitability of this library for gene identification and physical mapping of the apricot genome.

Nat Med, 2003 Jul, 9(7), 831 - 5
CpG motifs: the active ingredient in bacterial extracts?
Krieg AM.
The use of bacteria and bacterial extracts for immunotherapy has a checkered past . Recent developments in immunology reveal that these nonspecific immune activators actually work by triggering specific receptors that are expressed by subsets of immune cells . Identification of these receptors and the molecular signaling pathways that they activate has enabled a new era of specific targeted immunotherapy using chemically synthesized mimics of pathogen molecules.

Biomaterials, 2003 Oct, 24(22), 3877 - 84
Binding and orientation of fibronectin to silanated glass surfaces using immobilized bacterial adhesin-related peptides; Klueh U et al.; Previously, we have demonstrated the suitability of bacterial adhesin-related peptides, directly immobilized on polystyrene surfaces, to bind and orient fibronectin (FN) . For these studies a method to bind the large protein FN in a desired orientation on a solid substratum was developed which utilizes a bacterial adhesin-related peptide (designated BRP-A), which is known to bind specifically to the NH3-terminus end of FN . Glass substrata was first coated with an amine-terminated silane, followed by streptavidin (SA), which was used as an intermediate tether to bind the biotinylated bacterial adhesin-related peptide . The BRP-A peptide, used for these studies was synthesized with a terminal biotin to assure irreversible coupling of the BRP-A to the streptavidin . The biotinylated BRP-A was next immobilized on the SA-silanated glass surfaces . 125I-FN was used to quantify the amount of FN binding to the (BRP-A):SA-silanated glass surface . Monoclonal antibodies, which react with specific epitopes at either the NH3- or -COOH-termini of FN, were used to quantify the binding and orientation of FN . The results of these studies indicated: (1) FN bound to the BRP-A:SA-silanated glass surface; and (2) the bound FN was oriented such that NH2-terminal region of FN was bound towards the glass surface and the COOH-terminus was oriented away from the glass surface . These studies demonstrate that small peptides can be used to specifically bind and orient large proteins such as FN on the surfaces.

Clin Nephrol, 2003 Jun, 59(6), 447 - 54
Differences in the permeability of high-flux dialyzer membranes for bacterial pyrogens; Schindler R et al.; AIMS: The increasing use of high-flux membranes for hemodialysis has raised concerns that patients dialyzed with these membranes may be at higher risk of being exposed to cytokine-inducing bacterial substances in the dialysate than patients dialyzed with low-flux membranes . We investigated the permeability of various high-flux membranes for both purified E . coli lipopolysaccharide (LPS) as well as for LPS derived from Stenotrophomonas (Sten.) maltophilia . MATERIALS AND METHODS: An in vitro dialysis circuit with saline in the blood compartment of 3 dialyzers containing different membranes (polysulfone, helixone and Diapes) was employed . The dialysate was challenged with increasing doses of sterile filtrates derived from Sten . maltophilia cultures or with purified LPS from E . coli . Samples from the blood compartment were tested for cytokine induction (IL-1beta, IL-6 and TNF) in mononuclear cells as well as for LPS by limulus amebocyte lysate test (LAL) . RESULTS: IL-6 induction above sterile controls (< 0.02 ng/ml IL-6) was observed by samples from the blood side of DIAPES dialyzers (1.2 +/- 0.7 ng/ml IL-6) after challenging the dialysate with 4.1 +/- 3.6 U/ml E . coli LPS (9.9 +/- 4.5 ng/ml IL-6) . In contrast, at the same challenge dose no significant IL-6 induction above sterile controls was observed by blood side samples of polysulfone (0.15 +/- 0.07 ng/ml) and helixone (0.09 +/- 0.05 ng/ml) dialyzers . Increasing the amount of E . coli LPS in the dialysate further augmented IL-6 induction by blood side samples of Diapes but not of polysulfone and helixone dialyzers . Similar results were obtained for IL-1beta and TNF . After challenging the dialysate with E . coli LPS as well as with cultures of Sten . maltophilia, significantly more LAL reactivity was observed in the blood compartment of Diapes compared to polysulfone and helixone . CONCLUSIONS: There are considerable differences between high-flux membranes regarding their permeability for cytokine-inducing substances from E . coli as well as for LPS derived from E . coli and Sten . maltophilia . Dialyzers that leak CIS under aqueous conditions in vivo should not be used unless the dialysate has passed through an ultrafilter.

Acta Crystallogr D Biol Crystallogr, 2003 Jul, 59(Pt 7), 1291 - 3 Epub 2003 Jun 27.
Crystallization and preliminary X-ray data investigation of the bacterial enterocin A immunity protein at 1.65 A resolution; Dalhus B et al.; Crystals of the bacterial enterocin A immunity protein have been prepared by the hanging-drop vapour-diffusion technique at 293 K . The crystals diffract to better than 1.7 A resolution and X-ray diffraction data to 1.65 A have been collected at 110 K using synchrotron radiation . The enterocin A immunity protein crystals belong to the monoclinic crystal system, with unit-cell parameters a = 116.32, b = 42.35, c = 66.17 A, beta = 111.3 degrees . The symmetry and systematic absences in the diffraction pattern are consistent with space group C2 . The presence of two molecules in the asymmetric unit with a molecular weight of approximately 12.2 kDa gives a crystal volume per protein mass (V(M)) of approximately 3.1 A(3) Da(-1) and a solvent content of approximately 60% by volume.

Thorax, 2003 Jul, 58(7), 605 - 12
Enhancement of acute lung injury related to bacterial endotoxin by components of diesel exhaust particles; Yanagisawa R et al.; BACKGROUND: Diesel exhaust particles (DEP) synergistically aggravate acute lung injury related to lipopolysaccharide (LPS) in mice, but the components in DEP responsible for this have not been identified . A study was undertaken to examine the effects of the organic chemicals (DEP-OC) and residual carbonaceous nuclei (washed DEP) derived from DEP on LPS related lung injury . METHODS: ICR mice were divided into experimental groups and vehicle, LPS, washed DEP, DEP-OC, washed DEP+LPS, and DEP-OC+LPS were administered intratracheally . The cellular profile of the bronchoalveolar lavage (BAL) fluid, pulmonary oedema, lung histology, and expression of proinflammatory molecules and Toll-like receptors in the lung were evaluated . RESULTS: Both DEP-OC and washed DEP enhanced the infiltration of neutrophils into BAL fluid in the presence of LPS . Washed DEP combined with LPS synergistically exacerbated pulmonary oedema and induced alveolar haemorrhage, which was concomitant with the enhanced lung expression of interleukin-1beta, macrophage inflammatory protein-1alpha, macrophage chemoattractant protein-1, and keratinocyte chemoattractant, whereas DEP-OC combined with LPS did not . Gene expression of Toll-like receptors 2 and 4 was increased by combined treatment with washed DEP and LPS . The enhancement effects of washed DEP on LPS related changes were comparable to those of whole DEP . CONCLUSIONS: These results suggest that the residual carbonaceous nuclei of DEP rather than the extracted organic chemicals predominantly contribute to the aggravation of LPS related lung injury . This may be mediated through the expression of proinflammatory cytokines, chemokines, and Toll-like receptors.

FASEB J, 2003 Jul, 17(10), 1319 - 21
Bacterial DNA evokes epithelial IL-8 production by a MAPK-dependent, NF-kappaB-independent pathway; Akhtar M et al.; Recognition of bacterial products by the innate immune system is dependent on pattern-recognition receptors: toll-like receptor 9 (TLR-9) in the case of bacterial DNA . We hypothesized that bacterial DNA can directly affect enteric epithelial cells . RT-PCR revealed constitutive TLR-9 mRNA expression in three human colonic epithelial cell lines (T84, HT-29, Caco-2) and THP-1 monocytes . Epithelial cells, in six-well culture plates or on filter supports, were exposed to E . coli DNA (1-50 microg/ml), synthetic CpG-rich oligonucleotides, or calf thymus DNA for 6-48 h . Exposure to E . coli DNA resulted in an increase in IL-8 mRNA, and a time- and dose-dependent increase in IL-8 secretion . Also, CpG oligonucleotides induced epithelial IL-8 production, whereas calf thymus DNA did not . Exposure to E . coli DNA resulted in phosphorylation of ERK 1/2 MAPK and inhibitors of ERK activity (PD98059, UO126) significantly reduced the evoked IL-8 production . In contrast, inhibitors of NFkappaB activity (PDTC, SN50) did not block E . coli DNA-induced IL-8 production . Electrophoretic mobility shift assays revealed that E . coli DNA stimulated epithelial AP-1 but not NFkappaB activation . The barrier (i.e., transepithelial resistance) and ion transport parameters of epithelial monolayers (assessed in Ussing chambers) were unaltered following E . coli DNA exposure . Thus model gut epithelia express TLR-9 mRNA and, while maintaining their barrier function, can respond to E . coli DNA by increased IL-8 production.

FEMS Immunol Med Microbiol, 2003 Jul 15, 37(2-3), 129 - 34
Progress toward the development of a bacterial vaccine vector that induces high-titer long-lived broadly neutralizing antibodies against HIV-1; Fouts TR et al.; Conformationally constrained HIV-1 Env and gp120 immunogens induce broadly cross-reactive neutralizing antibodies . Thus, it is now feasible to rationally design an HIV-1 vaccine that affords protection through humoral mechanisms . This paper reviews our progress toward the development of an oral bacterial vaccine vector that is capable of delivering an HIV-1 DNA vaccine to host lymphoid tissues and inducing broadly neutralizing antibodies to HIV-1 in the mucosal and systemic immune compartments.

FEMS Microbiol Rev, 2003 Jun, 27(2-3), 215 - 37
Bacterial iron homeostasis; Andrews SC et al.; Iron is essential to virtually all organisms, but poses problems of toxicity and poor solubility . Bacteria have evolved various mechanisms to counter the problems imposed by their iron dependence, allowing them to achieve effective iron homeostasis under a range of iron regimes . Highly efficient iron acquisition systems are used to scavenge iron from the environment under iron-restricted conditions . In many cases, this involves the secretion and internalisation of extracellular ferric chelators called siderophores . Ferrous iron can also be directly imported by the G protein-like transporter, FeoB . For pathogens, host-iron complexes (transferrin, lactoferrin, haem, haemoglobin) are directly used as iron sources . Bacterial iron storage proteins (ferritin, bacterioferritin) provide intracellular iron reserves for use when external supplies are restricted, and iron detoxification proteins (Dps) are employed to protect the chromosome from iron-induced free radical damage . There is evidence that bacteria control their iron requirements in response to iron availability by down-regulating the expression of iron proteins during iron-restricted growth . And finally, the expression of the iron homeostatic machinery is subject to iron-dependent global control ensuring that iron acquisition, storage and consumption are geared to iron availability and that intracellular levels of free iron do not reach toxic levels.

Hepatogastroenterology, 2003 May-Jun, 50(51), 680 - 3
Octreotide inhibits hepatic fibrosis, bile duct proliferation and bacterial translocation in obstructive jaundice; Turkcapar N et al.; BACKGROUND/AIMS: The protective effect of octreotide on bacterial translocation, bile duct epithelial proliferation and hepatic fibrosis was studied in an experimental obstructive jaundice model . METHODOLOGY: Forty-five healthy Wistar albino rats were randomly divided into three groups . Group I (n = 15): Median laparotomy and common bile duct manipulation performed (Sham group) . Group II (n = 15): Laparotomy and common bile duct ligation performed . Group III (n = 15): After laparotomy and common bile duct ligation octreotide (Sandostatin, sandoz) was given . Simultaneously group I and II received 3 cc 0.9% NaCl and group III received 20 micrograms/kg/daily octreotide subcutaneously every 8 hours during 9 days . Two days after the procedure all rats were opened under ether anesthesia and sterile conditions . Group I had simple laparotomy but group II and III also had common bile duct ligation by 5/0 prolene . Seven days after the surgery (9th day after treatment) all rats underwent laparotomy and tests for bacterial translocation, liver biochemical tests and histopathologic analysis of liver and small bowel were carried out . RESULTS: In group II cecal population levels of bacteria were significantly higher than group I and group III (p < 0.05) . In group II there was also statistically significant bacterial translocation to the mesenteric lymph nodes . Pathological changes were found in terminal ileum samples in group II which seemed to improve in group III . Hepatocyte function was preserved with octreotide treatment which also significantly decreased bile duct proliferation and periportal fibrosis in response to biliary obstruction . CONCLUSIONS: This experimental study showed that octreotide is effective in preventing bacterial translocation, bile duct proliferation and hepatic fibrosis in obstructive jaundice.

Proc Natl Acad Sci U S A, 2003 Jul 8, 100(14), 8223 - 8 Epub 2003 Jun 25.
Quantitative modeling of sensitivity in bacterial chemotaxis: the role of coupling among different chemoreceptor species; Mello BA et al.; We propose a general theoretical framework for modeling receptor sensitivity in bacterial chemotaxis, taking into account receptor interactions, including those among different receptor species . We show that our model can quantitatively explain the recent in vivo measurements of receptor sensitivity at different ligand concentrations for both mutant and wild-type strains . For mutant strains, our model can fit the experimental data exactly . For the wild-type cell, our model is capable of achieving high gain while having modest values of Hill coefficient for the response curves . Furthermore, the high sensitivity of the wild-type cell in our model is maintained for a wide range of ambient ligand concentrations, facilitated by near-perfect adaptation and dependence of ligand binding on receptor activity . Our study reveals the importance of coupling among different chemoreceptor species, in particular strong interactions between the aspartate (Tar) and serine (Tsr) receptors, which is crucial in explaining both the mutant and wild-type data . Predictions for the sensitivity of other mutant strains and possible improvements of our model for the wild-type cell are also discussed.

Cytokine, 2003 Mar 21, 21(6), 295 - 302
Excessive increase of serum interleukin 6 jeopardizes host defense against multi-bacterial infection; Soda K et al.; Serum interleukin 6 (IL-6) is elevated among patients who have undergone surgery, trauma, and thermal injury . It is well known that the greater the increase of serum IL-6, the higher the incidence of post-injury morbidity and mortality is . However, it has not been determined whether the physiological effects of IL-6 increase the rate of morbidity and mortality or if IL-6 is just a bystander that only indicates the severity of the injury . To elucidate this, we planned to investigate the effect of IL-6 on a multi-bacterial infection, one of the most frequent post-injury complications . CDF1 male mice were administered recombinant human IL-6 (hIL-6) continuously at a dose of 0, 1, or 10 microg/day . The mice then underwent cecal ligation without puncture that induced slow multi-bacterial infection . The survival rate of mice receiving 10 microg/day of hIL-6 was significantly lower (38.5%) than the rate of those receiving 0 (83.3%) or 1 (92.3%) microg/day of hIL-6 . The result of this study showed that only excessive increases in serum IL-6, to levels that were observed among patients who underwent severe injury or extensive surgery with high incidence of post-injury infection, jeopardize the host's defense against bacterial infection.

Trends Microbiol, 2003 Jun, 11(6), 280 - 5
RNA-mediated control of virulence gene expression in bacterial pathogens; Johansson J et al.; Until recently, gene expression was thought to be controlled mainly at the level of transcription initiation by repressor or activator proteins . In some cases, transcription elongation is controlled by a so-called attenuation mechanism that involves alternative base-pairing between different regions of an mRNA transcript . Recent data reveal that other mechanisms can regulate gene expression and involve RNAs that might act as antisense RNAs, sequestering molecules, or thermosensors . This review focuses on recent studies in bacterial pathogens in which a growing list of examples show that RNA can control virulence gene expression.

Brain, 2003 Aug, 126(Pt 8), 1873 - 82 Epub 2003 Jun 23.
Lack of IL-6 augments inflammatory response but decreases vascular permeability in bacterial meningitis; Paul R et al.; Interleukin (IL)-6 is a multifunctional cytokine with diverse actions and has been implicated in the pathophysiology of many neurological and inflammatory disorders . In this study, we investigated the role of IL-6 in pneumococcal meningitis . Cerebral infection in wild-type (WT) mice caused an increase in vascular permeability and intracranial pressure (ICP), which were significantly reduced in IL-6-/- mice . In contrast, meningitis in IL-6-/- mice was associated with a significant increase in CSF white blood cell count compared with infected WT mice, indicating an enhanced inflammatory response . Analysis of mRNA expression in the brain showed an increase in tumour necrosis factor (TNF)-alpha, IL-1beta, and macrophage inflammatory protein 2 (MIP-2) levels, but decreased expression of granulocyte-macrophage colony-stimulating factor in infected IL-6-/- mice compared with infected WT controls . Similar results were obtained when rats challenged with pneumococci were systemically treated with neutralizing anti-IL-6 antibodies, resulting in an increased pleocytosis but at the same time a reduction of vascular permeability, brain oedema formation, and ICP, which was not accompanied by a downregulation of matrix metalloproteinases . Our data indicate that IL-6 plays an important anti-inflammatory role in bacterial meningitis by reducing leukocyte infiltration but contributes to the rise in intracranial pressure by increasing blood-brain barrier (BBB) permeability . These findings suggest that the migration of leukocytes across the BBB and the increase in vascular permeability are two independent processes during bacterial meningitis.

Infect Immun, 2003 Jul, 71(7), 3766 - 74
Regulation of pulmonary and systemic bacterial lipopolysaccharide responses in transgenic mice expressing human elafin; Sallenave JM et al.; The control of lung inflammation is of paramount importance in a variety of acute pathologies, such as pneumonia, the acute respiratory distress syndrome, and sepsis . It is becoming increasingly apparent that local innate immune responses in the lung are negatively influenced by systemic inflammation . This is thought to be due to a local deficit in cytokine responses by alveolar macrophages and neutrophils following systemic bacterial infection and the development of a septic response . Recently, using an adenovirus-based strategy which overexpresses the human elastase inhibitor elafin locally in the lung, we showed that elafin is able to prime lung innate immune responses . In this study, we generated a novel transgenic mouse strain expressing human elafin and studied its response to bacterial lipopolysaccharide (LPS) when the LPS was administered locally in the lungs and systemically . When LPS was delivered to the lungs, we found that mice expressing elafin had lower serum-to-bronchoalveolar lavage ratios of proinflammatory cytokines, including tumor necrosis factor alpha (TNF-alpha), macrophage inflammatory protein 2, and monocyte chemoattractant protein 1, than wild-type mice . There was a concomitant increase in inflammatory cell influx, showing that there was potential priming of innate responses in the lungs . When LPS was given systemically, the mice expressing elafin had reduced levels of serum TNF-alpha compared to the levels in wild-type mice . These results indicate that elafin may have a dual function, promoting up-regulation of local lung innate immunity while simultaneously down-regulating potentially unwanted systemic inflammatory responses in the circulation.

Arch Dis Child, 2003 Jul, 88(7), 615 - 20
Diagnosis and treatment of bacterial meningitis; El Bashir H et al.; This review focuses on recent advances of topical interest regarding the diagnosis and treatment of common causes of bacterial meningitis occurring in children beyond the neonatal period . Tuberculous meningitis is beyond the scope of this review.

Eur J Obstet Gynecol Reprod Biol, 2003 Jul 1, 109(1), 67 - 71
Efficacy of the combination of 2 g oral tinidazole and acidic buffering vaginal gel in comparison with vaginal clindamycin alone in bacterial vaginosis: a randomized, investigator-blinded, controlled trial; Milani M et al.; OBJECTIVE: To evaluate the efficacy of tinidazole (T) (Trimonase, Mipharm, Italy) and an acidic vaginal gel (Miphil) (M) in comparison with vaginal clindamycin (CL) (Cleocin Pharmacia Upjohn) in BV . DESIGN: A multicentre, randomised, investigator-blinded, controlled trial . POPULATION AND METHODS: 64 women with BV were enrolled . Thirty-two were allocated to receive oral T 2g, single dose, and 32 were assigned to CL 2% for 7 consecutive days . After week 1, T group were treated with an acidic vaginal gel, 2g every 3 days, for additional 3 weeks, whereas CL group did not received any additional treatment . Patients were evaluated at week 1 and 4 . Vaginal pH, the BV-blue test (Gryphus Diagnostics, USA) and the whiff test were performed at baseline and at week 4 . MAIN OUTCOMES MEASURES: Clinical cure rate; normalisation of vaginal pH (pH<4.5); and laboratory cure rate (defined as a clinical cure rate and a negative results of BV-blue and whiff test) . RESULTS: At baseline, vaginal pH values were (mean+/-S.D.) 5.4+/-0.7 and 5.3+/-0.5 in T and CL groups, respectively . Six patients (2 in T group and 4 in CL group) withdrew from the study due to side effects . At week 1, the clinical cure rates were 84% in both T and CL treated group (P=N.S.) . At week 4, clinical cure rates were 94% in T+M group and 77% in CL group (P=N.S.) . The laboratory cure rates were 81% in T+M group and 59% in CL group (P<0.04) . Vaginal pH normalisation (i.e . pH <4.5) was achieved in 78% and in 38% of T+M and CL groups, respectively (P<0.0007) . CONCLUSIONS: In the short term, 2g single oral dose tinidazole was at least as effective as 7-day of vaginal clindamycin . The sequential treatment of tinidazole and acidic vaginal gel was superior to vaginal clindamycin in lowering vaginal pH and achieving a higher laboratory tests normalization rate at 1-month follow-up.

Ultrason Sonochem, 2003 Jul, 10(4-5), 231 - 4
The development and evaluation of electrolysis in conjunction with power ultrasound for the disinfection of bacterial suspensions; Joyce E et al.; There is an increasing incidence in health problems related to environmental issues that originate from inadequate treatment of potable waters . This has compelled scientists and engineers to engage in innovative technologies to achieve a maximum disinfection at affordable costs . Some species of bacteria produce colonies and spores that can agglomerate in spherical clusters and thus protect organisms on the inside of the cluster against biocidal attack . Flocs of fine particles (e.g., clay) can entrap bacteria and this can also protect them against the biocides . Other bacteria have the ability to mutate, thus building up resistance to conventional biocides (e.g., chlorine).Ultrasound has been shown to be effective in improving the effectiveness of biocides such as chlorine . The aim of this present study was to investigate the effect of electrolysis and power ultrasound as a disinfection treatment and to provide a greater knowledge of the fundamentals of disinfection through the production of hypochlorite in situ from saline solution via electrolysis . The electrode materials investigated were, carbon (felt and graphite), copper and stainless steel rods . The results show that sonication appears to amplify the effect of electrolysis . A combination of both treatments is significantly better than sonication or electrolysis alone.

Am J Gastroenterol, 2003 Jun, 98(6), 1362 - 70
Small intestinal motility disturbances and bacterial overgrowth in patients with liver cirrhosis and portal hypertension; Gunnarsdottir SA et al.; OBJECTIVE: Altered small bowel motility and a high prevalence of small intestinal bacterial overgrowth (SIBO) has been observed in patients with liver cirrhosis . Our aim was to explore the relationship between motility abnormalities, portal hypertension, and SIBO . METHODS: Twenty-four patients with liver cirrhosis were included . Twelve had portal hypertension (PH) and 12 had liver cirrhosis (LC) alone . Child-Pugh score was the same in the groups . Antroduodenojejunal pressure recordings were performed, and noninvasive variceal pressure measurements were undertaken . Thirty-two healthy volunteers served as a reference group . Bacterial cultures were obtained from jejunal aspirates . RESULTS: The PH group had a higher proportion of individual pressure waves that were retrograde in the proximal duodenum during phase II (52% vs 13% vs 8% of propagated contractions; p < 0.001) as well as postprandially (49% vs 18% vs 13%; p < 0.01) compared with LC and controls, respectively . Long clusters were more common in PH than in controls (9.1 +/- 2.1 vs 4.9 +/- 0.8; p < 0.05), and a higher motility index in phase III in the proximal and distal duodenum was seen in the PH as compared with the other groups . The mean variceal pressure was 21 +/- 1 mm Hg . Motor abnormalities were not correlated to the level of variceal pressure . Thirty-three percent of the patients in the PH group but none in the LC group had SIBO . CONCLUSIONS: Abnormal small bowel motility and SIBO is common in patients with liver cirrhosis with concomitant portal hypertension . Portal hypertension per se might be significantly related to small bowel abnormalities observed in patients with liver cirrhosis.

Chin J Physiol, 2002 Jun 30, 45(2), 69 - 74
Enhancement of CREBSerine-133 phosphorylation through nitric oxide-mediated signaling induced by bacterial lipopolysaccharide in vascular smooth muscle cells from rats; Yang SN et al.; Nitric oxide (NO) induced by bacterial lipopolysaccharide (LPS) plays a critical role in various patho-physiological implications, such as atherosclerosis, vasculitis and septic shock . In addition, cAMP-responsive element binding protein (CREB), an important transcription factor for cell differentiation, has been shown to be involved in atherosclerogenesis in VSMCs . Here we investigated the possibility whether LPS-induced NO signaling led to phosphorylation of cAMP-responsive element binding protein on Serine-133 (CREBSer-133) in cultured vascular smooth muscle cells (VSMCs) from rats . Addition of LPS (1-10 microg/ml) for 48 hours increased not only the production NO, but also the phosphorylation of CREBSer-133 . The use of NOS inhibitor (100-500 microM L-NAME) blocked the magnitudes of both LPS-induced NO production and CREBSer-133 phosphorylation . In addition, either a guanylyl cyclase (GC) inhibitor (30 microM ODQ) or a cGMP-dependent protein kinase (PKG) inhibitor (20 microM (Rp)-8-pCPT-cGMPs) significantly attenuated the magnitudes of LPS-induced CREBSer-133 phosphorylation, suggesting the involvement of NO-GC-PKG signaling . Thus, the present study suggests that NO-mediated signaling activated by bacterial LPS, at least in part, enhance CREBSer-133 phosphorylation in cultured VSMCs . The findings here may provide not only signaling pathway involved in VSMC differentiation during inflammatory response, but also new insight into possible therapeutic intervention.

Philos Transact A Math Phys Eng Sci, 2003 Jun 15, 361(1807), 1283 - 312
Bacterial self-organization: co-enhancement of complexification and adaptability in a dynamic environment; Ben-Jacob E; During colonial development, bacteria generate a wealth of patterns, some of which are reminiscent of those occurring in abiotic systems . They can exhibit rich behaviour, reflecting informative communication capabilities that include exchange of genetic materials and the fact that the colony's building blocks are biotic . Each has internal degrees of freedom, informatic capabilities and freedom to respond by altering itself and others via emission of signals in a self-regulated manner . To unravel the special secrets of bacterial self-organization, we conducted an integrative (experimental and theoretical) study of abiotic and biotic systems . Guided by the notion of general biotic motives and principles, I propose that the informative communication between individuals makes possible the enhancement of the individuals' regulated freedom, while increasing their cooperation . This process is accomplished via cooperative complexification of the colony through self-organization of hierarchical spatio-temporal patterning . The colonial higher complexity provides the degree of plasticity and flexibility required for better colonial adaptability and endurability in a dynamic environment . The biotic system can modify the environment and obtain environmental information for further self-improvement . I reflect on the potential applications of the new understanding on 'engineered self-organization of systems too complex to design' and other issues.

Biotechniques, 2003 Jun, 34(6), 1220 - 2, 1224, 1226 passim
Bacterial IMPDH gene used for the selection of mammalian cell transfectants; Baccam M et al.; Stable cell transfection is used for the expression of exogenous genes or cDNAs in eukaryotic cells . Selection of these transfectants requires a dominant selectable marker . A variety of such markers has been identified and is currently in use . However, many of these are not suitable for all cell types or require unique conditions . Here we describe a simple and versatile dominant selectable marker that involves bacterial IMP dehydrogenase (IMPDH), an enzyme essential for the replication of mammalian and bacterial cells . Although IMPDH is evolutionarily conserved, the bacterial enzyme is orders of magnitude more resistant to the toxic effect of the drug mycophenolic acid, which is an IMPDH inhibitor . We have demonstrated that transfection of human, monkey or Chinese hamster cell lines with an expression vector containing bacterial IMPDH and mycophenolic acid treatment resulted in the selection of colonies with a strikingly increased resistance to mycophenolic acid toxicity . Analysis of cells derived from these colonies indicated that the acquisition of this resistance was associated with bacterial IMPDH protein expression . As a proof of principle, we showed that mammalian cell transfection with a bicistronic IMPDH/GFP expression vector and mycophenolic acid treatment can be used to successfully select transfectants that express the fluorescent protein . These results indicate that bacterial IMPDH is a practical dominant selectable marker that can be used for the selection of transfectants that express exogenous genes or cDNAs in mammalian cells.

J Bacteriol, 2003 Jul, 185(13), 3918 - 25
Archaeal homolog of bacterial type IV prepilin signal peptidases with broad substrate specificity; Albers SV et al.; A large number of secretory proteins in the thermoacidophile Sulfolobus solfataricus are synthesized as a precursor with an unusual leader peptide that resembles bacterial type IV prepilin signal sequences . This set of proteins includes the flagellin subunit but also various solute binding proteins . Here we describe the identification of the S . solfataricus homolog of bacterial type IV prepilin peptidases, termed PibD . PibD is an integral membrane protein that is phylogenetically related to the bacterial enzymes . When heterologously expressed in Escherichia coli, PibD is capable of processing both the flagellin and glucose-binding protein (GlcS) precursors . Site-directed mutagenesis of the GlcS signal peptide shows that the substrate specificity of PibD is consistent with the variations found in proteins with type IV prepilin-like signal sequences of S . solfataricus . We conclude that PibD is responsible for the processing of these secretory proteins in S . solfataricus.

Zhonghua Shao Shang Za Zhi, 2003 Apr, 19(2), 89 - 93
{An experimental study on the prevention and treatment of postburn intestinal injury and bacterial translocation by Sijunzi decoction in scalded rats}; Guo L et al.; OBJECTIVE: To explore the effects of traditional Chinese herbal medicine Sijunzi decoction on amelioration of postburn intestinal injury in scalded rats . METHODS: One hundred and eighty Wistar rats were randomly divided into 3 groups, i.e . scald and treatment (T), scald control (S) and normal control (C) groups . The rats in T group were gavaged with the decoction consisting of tangshen, tuckahoe, largehead atractylodes rhizome, glycyrrhizia and rhubarb in a dose of 2 ml twice daily, while the rats in C group were just gavaged with the same amount of distilled water . The rats were sacrificed according to the scheduled postburn observation timepoints . The contents of TNF, NO, MDA and ATPase activity in rat plasma and the intestinal mucosa and the S-IgA content in the intestinal mucus were determined respectively . The changes in histopathology of intestinal mucosa were observed . The samples from internal organ tissue and blood were obtained for bacterial culture . RESULTS: The contents of TNF, NO and MDA in the intestinal mucosa tissue and the rat plasma in scalded rats were lowered significantly by Sijunzi decoction . Furthermore, S-IgA secretion from intestinal mucous cells was maintained by Sijunzi decoction . T cell count was recovered and intestinal mucous barrier injury were lessened, and the bacterial positive rate in the internal organs was decreased . CONCLUSION: Traditional Chinese herbal medicine Sijunzi decoction might be helpful in alleviation of postburn intestinal injury and in the prevention of intestinal bacterial translocation.

Eur J Gastroenterol Hepatol, 2003 Jul, 15(7), 739 - 43
Spontaneous bacterial peritonitis in the Czech Republic: prevalence and aetiology; Lata J et al.; The aim of the study was to determine the prevalence and detailed data concerning the incidence of spontaneous bacterial peritonitis in the Czech Republic . Ninety-nine patients with liver cirrhosis and ascites were examined . Spontaneous bacterial peritonitis was diagnosed in 35 patients (35.4%) . It was revealed more often in patients with alcoholic aetiology of cirrhosis whose anamnesis involved sub-febrile or febrile states and the deterioration of ascites . Elevated serum leucocyte counts and increased levels of C-reactive protein can contribute to the diagnosis . A low level of total protein and albumin in ascites predisposes to the increase of this infection . The reduction of the platelet count in a set of patients with spontaneous bacterial peritonitis indicates the influence of portal hypertension in the aetiology of the disease.

Biochem Biophys Res Commun, 2003 Jul 4, 306(3), 674 - 9
Down-regulation of Toll-like receptor expression in monocyte-derived Langerhans cell-like cells: implications of low-responsiveness to bacterial components in the epidermal Langerhans cells; Takeuchi J et al.; In the skin, there are unique dendritic cells called Langerhans cells, however, it remains unclear why this particular type of dendritic cell resides in the epidermis . Langerhans cell-like dendritic cells (LCs) can be generated from CD14(+) monocytes in the presence of GM-CSF, IL-4, and TGF-beta1 . We compared LCs with monocyte-derived dendritic cells (DCs) generated from CD14(+) monocytes in the presence of GM-CSF and IL-4 and examined the effect of exposure to two distinct bacterial stimuli via Toll-like receptors (TLRs), such as peptidoglycan (PGN) and lipopolysaccharide (LPS) on LCs and DCs . Although stimulation with both ligands induced a marked up-regulation of CD83 expression on DCs, PGN but not LPS elicited up-regulation of expression CD83 on LCs . Consistent with these results, TLR2 and TLR4 were expressed on DCs, whereas only TLR2 was weakly detected on LCs . These findings suggest the actual feature of epidermal Langerhans cells with low-responsiveness to skin commensals.

Hepatol Res, 2003 Jun, 26(2), 114 - 118
Unusual hyperbilirubinemia associated with bacterial pneumonia and acute myeloid leukemia; Harada M et al.; A 77-year-old man with pneumonia associated with acute myeloid leukemia was introduced to the hepatology unit at our hospital for hyperbilirubinemia . He had been suffering from a high fever because of pneumonia . He was icteric and his serum concentrations of total and direct bilirubin were 13.1 and 7.9 mg/dl, respectively . However, the other standard biochemical examinations for hepatic function, such as serum concentrations of aspartate aminotransferase, alanine aminotransferase, gamma-glutamyl transpeptidase and alkaline phosphatase were normal except for lactate dehydrogenase . Lactate dehydrogenase isoenzyme analysis revealed that the high concentration was derived from leukemia cells . Ultrasonography of the abdomen revealed no abnormality in the liver or biliary tract . Administration of antibiotics for pneumonia decreased the serum bilirubin concentration, however, he died because of respiratory failure caused by the progression of pneumonia at 33 days after the admission . It was suggested that a disturbance in the bilirubin metabolism without hepatocyte necrosis or mechanical cholestasis might be involved in the pathogenesis of hyperbilirubinemia in patients with infectious diseases.

J Biol Chem, 2003 Aug 29, 278(35), 32778 - 83 Epub 2003 Jun 13.
The mitochondrial ornithine transporter . Bacterial expression, reconstitution, functional characterization, and tissue distribution of two human isoforms; Fiermonte G et al.; Two isoforms of the human ornithine carrier, ORC1 and ORC2, have been identified by overexpression of the proteins in bacteria and by study of the transport properties of the purified proteins reconstituted into liposomes . Both transport L-isomers of ornithine, lysine, arginine, and citrulline by exchange and by unidirectional mechanisms, and they are inactivated by the same inhibitors . ORC2 has a broader specificity than ORC1, and L- and D-histidine, L-homoarginine, and D-isomers of ornithine, lysine, and ornithine are all substrates . Both proteins are expressed in a wide range of human tissues, but ORC1 is the predominant form . The highest levels of expression of both isoforms are in the liver . Five mutant forms of ORC1 associated with the human disease hyperornithinemia-hyperammonemia-homocitrullinuria were also made . The mutations abolish the transport properties of the protein . In patients with hyperornithinemia-hyperammonemia-homocitrullinuria, isoform ORC2 is unmodified, and its presence compensates partially for defective ORC1.

J Negat Results Biomed . 2003 Jun 9;2(1):3.
Bacterial adherence to mucosal epithelium in the upper airways has less significance than believed; Ebenfelt A; BACKGROUND: Bacterial adherence to the upper airway epithelium is considered to be an important phenomenon in the pathogenesis of infections . However, the evidence for the hypothesis that bacterial adherence to mucosal epithelial cells has significance for pathogenesis of mucosal infections is based on studies using indirect techniques . We could find no biopsy studies with direct ocular observations of significant numbers of bacteria adhering to upper airway mucosal epithelial cells either in health or during disease . RESULTS: We studied specimens from healthy and infected tonsillar epithelium and specimens from the soft palate epithelium obtained by surgery . The specimens were examined by TEM . In the vast majority of specimens, we found no bacteria adhering to the epithelial cells in the mucosal line regardless of whether the patient was infected or not . Bacteria adhering to shed epithelial cells were seen in higher numbers . Furthermore, as bacteria are small compared to epithelial cells, we calculated the risk of overlooking every adhered bacteria in a section if bacterial adherence was such a significant phenomenon as earlier suggested . We found this risk to be very small . CONCLUSION: We conclude that bacterial adherence to mucosal surface epithelial cells is not a significant phenomenon, either in healthy mucosa in the upper airways or during infection . This is also in line with our earlier results, where we have shown that the site for the infectious process in pharyngotonsillitis is in the secretion on the tonsillar mucosal surface.

Plant Physiol, 2003 Jun, 132(2), 949 - 57
Identification and expression analysis of a gene encoding a bacterial-type phosphoenolpyruvate carboxylase from Arabidopsis and rice; Sanchez R et al.; Phosphoenolpyruvate carboxylase (PEPC) is distributed in plants and bacteria but is not found in fungi and animal cells . Important motifs for enzyme activity and structure are conserved in plant and bacterial PEPCs, with the exception of a phosphorylation domain present at the N terminus of all plant PEPCs reported so far, which is absent in the bacterial enzymes . Here, we describe a gene from Arabidopsis, stated as Atppc4, encoding a PEPC, which shows more similarity to Escherichia coli than to plant PEPCs . Interestingly, this enzyme lacks the phosphorylation domain, hence indicating that it is a bacterial-type PEPC . Three additional PEPC genes are present in Arabidopsis, stated as Atppc1, Atppc2, and Atppc3, encoding typical plant-type enzymes . As most plant PEPC genes, Atppc1, Atppc2, and Atppc3 are formed by 10 exons interrupted by nine introns . In contrast, Atppc4 gene has an unusual structure formed by 20 exons . A bacterial-type PEPC gene was also identified in rice (Oryza sativa), stated as Osppc-b, therefore showing the presence of this type of PEPC in monocots . The phylogenetic analysis suggests that both plant-type and bacterial-type PEPCs diverged early during the evolution of plants from a common ancestor, probably the PEPC from gamma-proteobacteria . The diversity of plant-type PEPCs in C3, C4, and Crassulacean acid metabolism plants is indicative of the evolutionary success of the regulation by phosphorylation of this enzyme . Although at a low level, the bacterial-type PEPC genes are expressed in Arabidopsis and rice.

Gut, 2003 Jul, 52(7), 927 - 32
Host gastric Lewis expression determines the bacterial density of Helicobacter pylori in babA2 genopositive infection; Sheu BS et al.; BACKGROUND AND AIMS: We tested if host gastric Lewis antigens and the babA2 genotype of Helicobacter pylori correlated with clinicohistological outcome . METHODS: We enrolled 188 dyspeptic patients (45 with duodenal ulcer, 45 with gastric ulcer, and 98 with chronic gastritis) with H pylori infection, proved by culture and gastric histology, reviewed by the updated Sydney system . Gastric expression of Lewis (Le) antigens Le(a), Le(b), Le(x), and Le(y) was determined immunochemically to determine intensity (range 0-3) . The corresponding 188 H pylori isolates were screened for babA2 genotype by polymerase chain reaction . RESULTS: All H pylori isolates had a positive babA2 genotype . We identified Le(a) in 33.5%, Le(b) in 72.9%, Le(x) in 86.2%, and Le(y) in 97.4% of biopsies from these 188 patients . Patients who expressed Le(b) had a higher H pylori density than those who did not express Le(b) (p<0.001) . Among 139 patients who expressed Le(b), H pylori density increased with a higher Le(b) intensity (p<0.05) . Gastric atrophy decreased with Le(b) intensity and thus resulted in lower H pylori density in the antrum (p<0.05) . For the 49 patients without gastric Le(b) expression, H pylori density was positively related with Le(x) and Le(a) expression (p<0.05) . CONCLUSIONS: Taiwanese H pylori isolates are 100% babA2 genopositive . Gastric Le(b) as well as Le(x) intensity may be major determinants of H pylori density . While lacking gastric Le(b) expression, Le(x) and Le(a) were closely related to H pylori colonisation.

J Biochem (Tokyo), 2003 May, 133(5), 577 - 81
Production of a functional catalytic antibody ScFv-NusA fusion protein in bacterial cytoplasm; Zheng L et al.; Functional expression of catalytic antibodies in the cytoplasm of E . coli is potentially of great interest in searching for new catalysts by genetic selection . Herein, a catalytic antibody single chain Fv (ScFv) 14D9, which catalyzes a highly enantioselective protonation, was expressed as a NusA fusion protein under the T7 promoter . A functional disulfide-containing ScFv fusion protein was obtained in the oxidizing environment of bacterial cytoplasm . The 14D9 ScFv could not be overexpressed alone without NusA fusion . The highly soluble NusA protein most likely retards aggregate formation of ScFv and indirectly supports correct folding and disulfide bridge formation in the fusion construct ScFv-NusA . The ScFv-NusA fusion product shows highly enantioselective, specific, hapten inhibited catalytic activity comparable to its parent monoclonal antibody, 14D9 . The NusA fusion method might be generally helpful for functional antibody expression in vivo and for the new development of biocatalysts by genetic selection.

Biotechnol Bioeng, 2003 Aug 20, 83(4), 474 - 8
Role of water-soluble polysaccharides in bacterial cellulose production; Ishida T et al.; Acetobacter xylinum BPR2001 produces water-insoluble bacterial cellulose (BC) and a water-soluble polysaccharide called acetan in corn steep liquor-fructose medium . Acetobacter xylinum EP1, which is incapable of acetan production was derived by disrupting the aceA gene of BPR2001 . The BC production by EP1 (2.88 g/L) was lower than that by BPR2001 (4.6 g/L) in baffled-flask culture . When purified acetan or agar was added to the medium from the start of cultivation, the BC production by EP1 was enhanced and the final BC yield of EP1 was almost the same as that of BPR2001 . A similar improvement of BC production by EP1 by the addition of agar was also confirmed by cultivation in a 50-L airlift reactor . From these results, the role of acetan in BC production is associated with the increase in the viscosity of the culture medium which may hinder coagulation of BC and cells in the culture, thereby accelerating the growth of BPR2001 and BC production by BPR2001 .

Transfus Clin Biol, 2003 Jun, 10(3), 192 - 200
{Transfusion-transmitted bacterial infection: residual risk and perspectives of prevention}; Morel P et al.; Bacterial contamination of blood components represents today the highest infectious risk of blood transfusion, the risk is particularly high when it affects platelet concentrates . The residual risk of transfusion reaction due to bacterial contamination of platelets concentrates remains stable . For all severity 1 case occurs with 25,000 distributed platelets concentrates and 1 death occurs with 200,000 distributed units . In France, efforts have focused on the prevention of contamination during donation--involving measures such as rejecting the first few millilitres of donated blood and improving skin disinfection--and the prevention of bacterial proliferation in platelets concentrates--notably by removing leukocytes and ensuring high-quality storage of donated blood . Improving strategies for reducing the risks of bacterial contamination is one of the priorities of the French National Blood Transfusion Service (l'Etablissement francais du sang-EFS) . There is currently considerable debate about the relative importance of bacterial screening methods and methods for inactivating pathogens present in PC . Automated culture (Biomerieux) and the ScanSystem (Hemosystem) and BDS (Pall) method are the most advanced detection systems available, to our knowledge . In term of pathogen inactivation system for platelets, Intercept (Baxter) is nearing the commercial market . These new prevention have logistic and/or functional consequences that will require close scrutiny methods . A national study group is currently considering the consequences of each of these methods and should give its opinion at the end of the first half of 2003.

Bioorg Med Chem Lett, 2003 Jul 7, 13(13), 2231 - 4
Discovery of a potent and selective series of pyrazole bacterial methionyl-tRNA synthetase inhibitors; Finn J et al.; Starting with a micromolar lead identified from high-throughput screening, a series of pyrazoles were discovered with significantly improved potency on bacterial methionyl-tRNA synthetase and selectivity over human methionyl-tRNA synthetase.

Infect Genet Evol, 2003 May, 3(1), 47 - 55
Mycoplasma pneumoniae: a reduced-genome intracellular bacterial pathogen; Meseguer MA et al.; Mycoplasma pneumoniae has classically been considered an extracellular (or membrane-associated) organism . Nevertheless, the recently elucidated genomic structure of this pathogen strongly suggest that this organism may have been subjected to the process of reductive genetic evolution which is characteristic of intracellular bacteria . We studied the Mycoplasma pneumoniae RYC15989 strain, recovered from a pericardial biopsy sample from a patient with atypical pneumonia and acute pericarditis . The interaction of this strain with human hepatocytes Hep-G2 and mouse neuroblastoma N2-A cell lines was investigated . Confocal laser scanning microscopy and electronic microscopy evidence is presented of the intracellular location of fluorochrome-labelled Mycoplasma pneumoniae in cell lines infected with the organism in vitro . This finding provides preliminary evidence of cellular invasive capacity of Mycoplasma pneumoniae and casts some new light on the pathogenic potential of Mycoplasma pneumoniae in host infection.

J Clin Immunol, 2003 May, 23(3), 202 - 13
Innate immune responses in lupus-prone Palmerston North mice: differential responses to LPS and bacterial DNA/CpG oligonucleotides; Lenert P et al.; Inadequate immune response to infectious danger may contribute to the pathogenesis of systemic autoimmune diseases, e.g., systemic lupus erythematosus . To test this hypothesis, we studied innate responses of prediseased lupus-prone Palmerston North (PN) mice to lipopolysaccharide (LPS), bacterial DNA, and synthetic CpG oligonucleotides . LPS and bacterial DNA/CpG oligodeoxyribonucleotides (ODNs) drove PN splenocytes into the cell cycle and protected B cells against spontaneous apoptosis, as in control lupus-free DBA-1 mice . LPS induced significantly higher IL-6 production in PN than in control splenocytes . In contrast, in PN splenocytes bacterial DNA and CpG ODNs induced approximately four- to sixfold lower IL-12p40 and approximately twofold lower IL-6 secretion than controls . This reduction in cytokine secretion in PN mice was not due to delayed kinetics but was related to significantly higher constitutive and CpG-inducible IL-10 secretion . Neutralizing anti-IL-10 antibodies almost completely restored PN IL-6 and IL-12p40 secretion to DBA-1 levels, whereas exogenous IL-10 inhibited in vitro IL-6 and IL-12p40 production in DBA-1 mice . Importantly, treatment with either IL-10 or anti-IL-10 antibody did not modulate CpG-induced cell cycle entry and apoptosis protection in either strain . In conclusion, lupus-prone PN mice show abnormal innate responses through their pattern-recognition TLR9 receptors, characterized by higher inducible IL-10 and lower IL-12p40 and IL-6 secretion, thus implying that response to infectious danger in PN mice is inappropriate and may be linked to lupus pathogenesis.

Am J Clin Oncol, 2003 Jun, 26(3), e54 - 9
Does the addition of glutamine to total parenteral nutrition have beneficial effect on the healing of colon anastomosis and bacterial translocation after preoperative radiotherapy?
El-Malt M, Ceelen W, Boterberg T, Claeys G, de Hemptinne B, de Neve W, Pattyn P.
Glutamine administration stimulates mucosal growth and preserves the morphology of the intestine . Theoretically, it could improve colonic anastomotic healing after radiotherapy (RT)-induced epithelial damage and mucosal atrophy induced by total parenteral nutrition (TPN) . To investigate this issue, the rectosigmoid colon in male Wistar rats was irradiated to a total dose of 25 Gy . Five days after the end of RT, side-to-side anastomosis was constructed between the irradiated rectosigmoid and the nonirradiated caecum . Postoperatively, animals were divided in three groups: group I, normal diet orally; group II, TPN; group III, TPN enriched with 2% glutamine (Gln-TPN) . All animals decreased in weight during RT and after surgery . Weight regain postoperatively was better in the orally fed animals in comparison with the parenterally fed animals (I vs . II and III; p < 0.01) . Colonic anastomotic bursting pressure (BP) and bursting wall tension (BWT) were significantly less in group II in comparison with groups I and III (II vs . I and III; p < 0.01) . BP and BWT were comparable in groups I and III . No significant differences were found between all the groups in gut bacterial translocation to the blood or to the mesenterical lymph nodes . Conclusively, Gln-TPN can play a role in counteracting the negative effect of food deprivation on the healing of irradiated colonic anastomoses . Postoperative Gln-TPN does not influence gut bacterial translocation in this rat model.

Eukaryot Cell, 2003 Jun, 2(3), 646 - 50
Histone-like proteins of the dinoflagellate Crypthecodinium cohnii have homologies to bacterial DNA-binding proteins; Wong JT et al.; The dinoflagellates have very large genomes encoded in permanently condensed and histoneless chromosomes . Sequence alignment identified significant similarity between the dinoflagellate chromosomal histone-like proteins of Crypthecodinium cohnii (HCCs) and the bacterial DNA-binding and the eukaryotic histone H1 proteins . Phylogenetic analysis also supports the origin of the HCCs from histone-like proteins of bacteria.

Nutr Rev, 2003 Apr, 61(4), 132 - 5
Small intestinal bacterial overgrowth: a possible risk factor for metabolic bone disease; Anantharaju A et al.; Small intestinal bacterial overgrowth (SIBO) is one of the causes of malabsorption syndromes . The prevalence of metabolic bone disease in patients with SIBO is unknown, but a recent prospective case-control study indicated significant contribution of SIBO to the development of metabolic bone disease . We review this and other reports in the literature and discuss the possible mechanisms causing metabolic bone disease in patients with SIBO.

Kyobu Geka, 2003 Jun, 56(6), 501 - 4
{Showing no increase of C-reactive protein and developing non-bacterial median chest wound dehiscence after coronary artery bypass grafting}; Suzuki S et al.; A 72-year-old male underwent coronary artery bypass grafting for 3 vessels disease . After the operation, the peak C-reactive protein (CRP) level was 0.85 mg/dl and CRP levels stayed very low . Forty days after the operation the patient developed a progressive median chest wound dehiscence but the bacterial cultures were negative . In spite of conventional therapies such as debridement of necrotic tissue or irrigation, the wound granulation was underdeveloped . After 6 months an epidermis had developed to cover the wound.

Arthritis Rheum, 2003 Jun 15, 49(3), 328 - 34
Bacterial components in the synovial tissue of patients with advanced rheumatoid arthritis or osteoarthritis: analysis with gas chromatography-mass spectrometry and pan-bacterial polymerase chain reaction; Chen T et al.; OBJECTIVE: To study the presence of bacterial components in the synovial tissue (ST) of patients with advanced rheumatoid arthritis (RA) . METHODS: ST was collected during joint surgery from 41 RA patients . Tissue from 39 patients with osteoarthritis (OA), 4 patients with undifferentiated inflammatory arthritis (UA), and 3 cases of accidental deaths served as controls . The pan-bacterial polymerase chain reaction (PCR) with primers for the 23S ribosomal RNA (rRNA) and 16S rRNA genes was used to detect bacterial DNA . In addition, synovial fluid (SF) samples from patients with chlamydial reactive arthritis (ReA) were also examined by the same method . The positive controls, bacterial DNA or ST spiked with different living bacteria, were analyzed alongside clinical samples . Most of the ST samples were also analyzed by gas chromatography-mass spectrometry (GC-MS) for determining the presence of bacteria-derived muramic acid . Strict precautions were followed in the clinics and the laboratory to prevent contamination . RESULTS: In GC-MS analysis, muramic acid was observed in the ST from 4 of 35 RA patients and from 2 of 14 OA patients, but not in ST from 2 patients with UA and 3 cadavers . Bacterial DNA was not detected by either one of the PCR primers used in ST from 42 patients with RA and 39 patients with OA . However, 5 of 15 SF samples from ReA patients were PCR positive . The sensitivity of GC-MS to detect muramic acid was 2 pg/injected amount (227 pg muramic acid/mg ST), and that of the pan-bacterial PCR was 2-20 bacteria colony forming units/reaction . CONCLUSION: These results indicate that a bacterial component, muramic acid, is detectable by GC-MS in ST from a few patients with advanced RA or OA . However, no bacterial DNA was detectable by PCR.

Sex Transm Infect, 2003 Jun, 79(3), 254 - 6
National survey of doctors' actions following the diagnosis of a bacterial STD; McCree DH et al.; OBJECTIVES: Little is known about the post-STD diagnosis management practices of community based doctors . The purpose of this study was to describe the reported actions that doctors take after diagnosing gonorrhoea, chlamydia, or syphilis and to determine if these actions differ across the three STDs . METHODS: A random national sample of 7300 doctors (70% response rate) practising in five medical specialties responded to 13 questions related to STD management . Mean differences across STDs were examined using the General Linear Model function of SPSS . RESULTS: Most doctors reported instructing patients to abstain from sex during treatment, to use condoms, and to inform their sexual partners of their exposure after diagnosing gonorrhoea, chlamydia, or syphilis . For syphilis, however, doctors were less likely to treat the patients presumptively and to give them drugs for their partners; and more likely to collect partner information, to follow up with the patient to see if the partner was referred for treatment and to send patient information to the health department . CONCLUSIONS: Doctors' post-STD diagnosis actions were similar for gonorrhoea and chlamydia compared to syphilis . Study findings suggest low levels of STD case reporting and partner follow up by doctors in the sample . Interventions are needed to educate community based doctors about the importance of partner follow up and case reporting in the management of STDs.

Microbiol Mol Biol Rev, 2003 Jun, 67(2), 213 - 25, table of contents
ADP-glucose pyrophosphorylase, a regulatory enzyme for bacterial glycogen synthesis; Ballicora MA et al.; The accumulation of alpha-1,4-polyglucans is an important strategy to cope with transient starvation conditions in the environment . In bacteria and plants, the synthesis of glycogen and starch occurs by utilizing ADP-glucose as the glucosyl donor for elongation of the alpha-1,4-glucosidic chain . The main regulatory step takes place at the level of ADP-glucose synthesis, a reaction catalyzed by ADP-Glc pyrophosphorylase (PPase) . Most of the ADP-Glc PPases are allosterically regulated by intermediates of the major carbon assimilatory pathway in the organism . Based on specificity for activator and inhibitor, classification of ADP-Glc PPases has been expanded into nine distinctive classes . According to predictions of the secondary structure of the ADP-Glc PPases, they seem to have a folding pattern common to other sugar nucleotide pyrophosphorylases . All the ADP-Glc PPases as well as other sugar nucleotide pyrophosphorylases appear to have evolved from a common ancestor, and later, ADP-Glc PPases developed specific regulatory properties, probably by addition of extra domains . Studies of different domains by construction of chimeric ADP-Glc PPases support this hypothesis . In addition to previous chemical modification experiments, the latest random and site-directed mutagenesis experiments with conserved amino acids revealed residues important for catalysis and regulation.

J Bone Joint Surg Br, 2003 May, 85(4), 490 - 4
Body-exhaust suit versus occlusive clothing . A randomised, prospective trial using air and wound bacterial counts; Der Tavitian J et al.; We randomly allocated 50 total knee replacements to scrub teams wearing body-exhaust suits (BES) or Rotecno occlusive clothing . The effectiveness of the clothing was assessed using air andwound bacterial counts . Bacteria were recovered from 62% of wounds (64% BES, 60% Rotecno) . The mean air count was 0.5 CFU/ m3 with BES and 1.0 CFU/m3 with Rotecno (p = 0.014) . The mean wound counts were 14 bacteria/wound with BES and eight bacteria/wound with Rotecno (p = 0.171) . There was no correlation between the air and wound counts (r = -0.011, Spearman's) . The higher air counts suggest that Rotecno occlusive clothing is less effective than BES, but wounds were equally contaminated with both types of clothing suggesting that at very low levels of air contamination the contribution of bacteria to the wound from the air is irrelevant . Even doubling the air counts from 0.5 to 1.0 CFU/m3 had no detectable effect on the wound . This allows a reassessment to be made of other sources of contamination the effect of which would previously have been overwhelmed by contamination from air.

Carbohydr Res, 2003 Jun 16, 338(12), 1299 - 308
Synthesis and conformational analysis of the repeating units of bacterial spore peptidoglycan; Keglevic D et al.; Deprotection of the fully blocked disacharide allyl O-(2-amino-4,6-O-benzylidene-3-O-{(R)-1-carboxyethyl}-2-deoxy-beta-D-glucopyranosyl-1',2-lactam)-(1-->4)-2-acetamido-3,6-di-O-benzyl-2-deoxy-beta-D-glucopyranoside by selective de-O-allylation and parallel removal of the benzylidene and O-benzyl groups is described . The resulting beta-muramyl lactam-(1-->4)-GlcNAc disaccharide is characterised as the per-O-acetylated derivative by 1H and 13C NMR spectroscopy and X-ray structure analysis . Conformational analysis about glycosidic bond of repeating units of bacterial spore cortex is based on experimental data and molecular modelling.

Proc Natl Acad Sci U S A, 2003 Jun 24, 100(13), 8024 - 9 Epub 2003 Jun 03.
Physical interaction between RRS1-R, a protein conferring resistance to bacterial wilt, and PopP2, a type III effector targeted to the plant nucleus; Deslandes L et al.; RRS1-R confers broad-spectrum resistance to several strains of the causal agent of bacterial wilt, Ralstonia solanacearum . Although genetically defined as recessive, this R gene encodes a protein whose structure combines the TIR-NBS-LRR domains found in several R proteins and a WRKY motif characteristic of some plant transcriptional factors and behaves as a dominant gene in transgenic susceptible plants . Here we show that PopP2, a R . solanacearum type III effector, which belongs to the YopJ/AvrRxv protein family, is the avirulence protein recognized by RRS1-R . Furthermore, an interaction between PopP2 and both RRS1-R and RRS1-S, present in the resistant Nd-1 and susceptible Col-5 Arabidopsis thaliana ecotypes, respectively, was detected by using the yeast split-ubiquitin two-hybrid system . This interaction, which required the full-length R protein, was not observed between the RRS1 proteins and PopP1, another member of the YopJ/AvrRxv family present in strain GMI1000 and that confers avirulence in Petunia . We further demonstrate that both the Avr protein and the RRS1 proteins colocalize in the nucleus and that the nuclear localization of the RRS1 proteins are dependent on the presence of PopP2.

Biochim Biophys Acta, 2003 Jun 10, 1612(2), 186 - 94
An association of 27- and 40-kDa molecules with glycolipids that bind A-B bacterial enterotoxins to cultured cells; Shimizu T et al.; It is well recognized that the Shiga-like toxins (Stxs) preferentially bind to Gb3 glycolipids and the cholera toxin (CT) and heat-labile enterotoxin (LTp) bind to GM1 gangliosides . After binding to the cell surface, A-B bacterial enterotoxins have to be internalized by endocytosis . The transport of the toxin-glycolipid complex has been documented in several manners but the actual mechanisms are yet to be clarified . We applied a heterobifunctional cross-linker, sulfosuccinimidyl-2-(p-azidosalicylamido)-1,3'-dithiopropionate (SASD), to detect the membrane proteins involved in the binding and the transport of A-B bacterial enterotoxins in cultured cells . Both Stx1 and Stx2 bound to the detergent-insoluble microdomain (DIM) of Vero cells and Caco-2 cells, which were susceptible to the toxin, but neither was bound to insusceptible CHO-K1 cells . Both CT and LTp bound to the DIM of Vero cells, Caco-2 cells, and CHO-K1 cells . In a cross-linking experiment, Stx1 cross-linked only with a 27-kDa molecule, while Stx2, which was more potently toxic than Stx1, cross-linked with 27- and 40-kDa molecules of Vero cells as well as of Caco-2 cells; moreover, no molecules were cross-linked with the insusceptible CHO-K1 cells . LTp was cross-linked only to the 27-kDa molecule of these three cell types but the CT, which was more toxic than LTp, was also cross-linked with 27- and 40-kDa molecules of Vero cells, Caco-2 cells, and CHO-K1 cells . The 27- and the 40-kDa molecules might play a role in the endocytosis and retrograde transport of A-B bacterial enterotoxins.

Solid State Nucl Magn Reson, 2003 Jun, 23(4), 198 - 212
Solid-state 13C and 1H spin diffusion NMR analyses of the microfibril structure for bacterial cellulose; Masuda K et al.; To obtain further information about the cause for the rather large splitting of the C4 resonance line into the downfield (C4D) and upfield (C4U) lines in CP/MAS 13C NMR spectra for native cellulose, 13C and 1H spin diffusion measurements have been conducted by using different types of bacterial cellulose samples . In 13C spin diffusion measurements, the C4D resonance line is selectively inverted by the Dante pi pulse sequence and the 13C spin diffusion is allowed to proceed from the C4D carbons to other carbons including the C4U carbons with use of the 13C4-enriched bacterial cellulose sample . The analysis based on the simple spin diffusion theory for the process experimentally observed reveals that the C4U carbons may be located at distances less than about 1 nm from the C4D carbons . In 1H spin diffusion measurements, poly(vinyl alcohol) (PVA) films in which ribbon assemblies of bacterial cellulose are dispersed are employed and the 1H spin diffusion process is examined from the water-swollen PVA continuous phase to the dispersed ribbon assemblies by the 13C detection through the 1H-13C CP technique . As a result, it is found that the C4D and C4U carbons are almost equally subjected to the 1H spin diffusion from the PVA phase, indicating that the C4U carbons are not localized in some limited area, e.g . in the surfacial region, but are distributed in the whole area in the microfibrils . These experimental results suggest that the C4U carbons may exist as structural defects probably due to conformational irregularity associated with disordered hydrogen bonding of the CH(2)OH groups in the microfibrils.

Microbes Infect, 2003 Jun, 5(7), 621 - 7
Inducible nitric oxide synthase and control of intracellular bacterial pathogens; Chakravortty D et al.; Inducible nitric oxide synthase (iNOS) has important functions in innate immunity and regulation of immune functions . Here, the role of iNOS in the pathogenesis of various intracellular bacterial infections is discussed . These pathogens have also evolved a broad array of strategies to repair damage by reactive nitrogen intermediates, and to suppress or inhibit functions of iNOS.

Mol Microbiol, 2003 Jun, 48(5), 1349 - 55
Oligomerization and activation of the FliI ATPase central to bacterial flagellum assembly; Claret L et al.; FliI is the peripheral membrane ATPase pivotal to the type III protein export mechanism underlying the assembly of the bacterial flagellum . Gel filtration and multiangle light scattering showed that purified soluble native FliI protein was in a monomeric state but, in the presence of ATP, FliI showed a propensity to oligomerize . Electron microscopy revealed that FliI assembles to a ring structure, the yield of which was increased by the presence of a non-hydrolysable ATP analogue . Single particle analysis of the resulting electron micrograph images, to which no symmetry was applied, showed that the FliI ring structure has sixfold symmetry and an external diameter of approximately 10 nm . The oligomeric ring has a central cavity of 2.5-3.0 nm, which is comparable to the known diameter of the flagellar export channel into which export substrates feed . Enzymatic activity of the FliI ATPase showed positive co-operativity, establishing that oligomerization and enzyme activity are coupled . Escherichia coli phospholipids increased enzyme co-operativity, and in vitro cross-linking demonstrated that they promoted FliI multimerization . The data reveal central facets of the structure and action of the flagellar assembly ATPase and, by extension, the homologous ATPases of virulence-related type III export systems.

Phys Rev E Stat Nonlin Soft Matter Phys . 2003 May;67(5 Pt 1):051921 . Epub 2003 May 23.
Applicability of the Fisher equation to bacterial population dynamics; Kenkre VM et al.; The applicability of the Fisher equation, which combines diffusion with logistic nonlinearity, to population dynamics of bacterial colonies is studied with the help of explicit analytic solutions for the spatial distribution of a stationary bacterial population under a static mask . The mask protects bacteria from ultraviolet light . The solution, which is in terms of Jacobian elliptic functions, is used to provide a practical prescription to extract Fisher equation parameters from observations and to decide on the validity of the Fisher equation.

Ann Otol Rhinol Laryngol, 2003 May, 112(5), 461 - 8
Effects of bacterial toxins on air-exposed cultured human respiratory sinus epithelium; Nell MJ et al.; The present study was designed to investigate the effects of the bacterial toxins lipopolysaccharide (LPS) and lipoteichoic acid (LTA) on air-exposed cultured human respiratory sinus epithelium . The morphological changes, proliferation, and differentiation of sphenoid sinus mucosa were examined after incubation with different LPS or LTA concentrations . Air-exposed cultured sinus mucosa differentiated from pseudostratified respiratory epithelium to squamous ciliated epithelium with few goblet cells . High concentrations of bacterial toxins induced a significant increase in mucus production and a decrease in ciliated cells . Ki67 immunostaining showed an increased cell proliferation after incubation with moderate levels of LPS or LTA . High concentrations of bacterial toxins, on the other hand, induced a decreased proliferation . Involucrin expression was clearly altered by incubation with high levels of bacterial toxins, indicating an increased degree of terminal differentiation . These results indicate that the bacterial toxins LPS and LTA both induce comparable dose-dependent morphological changes in sinus epithelium.

Ned Tijdschr Tandheelkd, 2003 May, 110(5), 178 - 80
{110th year Nederlands Tijdschrift voor Tandheelkunde . 2 . Root canal treatment, intra-canal disinfectants and bacterial culture: past and present}; Moorer WR et al.; Fifty years ago the Dutch Journal of Dentistry published methods and opinions concerning root canal treatment . Qualitative bacterial culture, inclusion of aggressive disinfectants, as well as antibiotics and widening of the apical constriction were carried out . Nowadays, because of several reasons, these are not clinical practice anymore . Controversy over the clinical consequences of bacterial presence in tubules and in the peri-apical area prevailed in the past and seem to be prevalent once again.

Calcif Tissue Int . 2003 Jun 6; {Epub ahead of print}
Systemic Effects on Bone Healing of a New Hyaluronic Acid-Like Bacterial Exopolysaccharide; Zanchetta P et al.; Critical Size Defect (CSD) technique was used to evaluate the systemic activities on bone regeneration capacity of a newly discovered hyaluronic acid-like exopolysaccharide synthesized by a bacteria originating from a deep sea hydrothermal vent . Some systemic effects were previously detected on earlier experiments . A 5 mm diameter hole was made on each parietal bone of male rats . The right hole was filled with 0.5 mg of a new bacterial exopolysaccharide referenced HE 800, while the left hole remained free of any treatment . After 21 days, the holes and surrounding tissues were examined by direct examination, X-rays, and histological staining . Using HE 800, bone healing was almost complete after only 21 days in the treated hole and always complete in the control side by some systemic effect . Neovascularization was also observed along with an organized trabecular bone on both sides . No abnormal bone growth or conjunctival abnormalities were noticed . At the end of the experiment, 90.1% (+/-5.2) bone healing (n = 20) was observed on the treated side; conversely, the control side animals demonstrate an amazing healing 100% (+/-0.5) by a systemic effect.

Sex Transm Dis, 2003 Jun, 30(6), 483 - 9
Evaluation of a new rapid diagnostic kit (FemExam) for bacterial vaginosis in patients with vaginal discharge syndrome in The Gambia; West B et al.; BACKGROUND: Diagnosis of bacterial vaginosis (BV) in resource-poor primary health care settings is often overlooked; there is a need for a cheap, rapid, objective point-of-care diagnostic test . GOAL: The goal was to determine the prevalence of BV and to evaluate the performance of a new commercial diagnostic test kit in a developing country environment . STUDY DESIGN: Vaginal and cervical swabs were collected from 230 consecutive women attending a genitourinary medicine clinic with reported symptoms of vaginal discharge and/or itching . Etiological testing was carried out . BV was diagnosed on the basis of the Nugent score, the Amsel clinical criteria, and results of FemExam card tests . Card 1 is for pH and amines, and card 2 measures proline iminopeptidase (PIP) activity . RESULTS: BV prevalence was 47.9% according to the Nugent score . When compared with the Nugent score, the Amsel clinical criteria had a sensitivity of 77.9% and specificity of 58.4%, FemExam card 1 had a sensitivity of 71.4% and specificity of 72.8%, FemExam card 2 had a sensitivity of 70% and specificity of 81.0%, and FemExam cards 1 and 2 combined had a sensitivity of 91.0% and specificity of 61.5% . Cost per patient and cost per true case detected ranged from US $0.74 and US $1.54, respectively, for Gram stain diagnosis, to US $8.32 and US $18.49 for the FemExam two-card method . CONCLUSIONS: In a setting where BV was frequently associated with vaginal discharge, the FemExam test compared favorably with conventional clinical diagnosis, and it has the advantage of being rapid, less subjective, and easily performed . Cutting its cost would provide wider accessibility in developing countries.

J Microbiol Methods, 2003 Aug, 54(2), 153 - 64
Tracking of injected and resident (previously injected) bacterial cells in groundwater using ferrographic capture; Johnson WP et al.; A high-resolution bacterial tracking technique, ferrographic capture, was used to enumerate fluorescent-stained bacterial cells that were injected into groundwater during a field experiment . The goal of the experiment was to investigate whether detachment of previously injected stained resident cells attached to aquifer sediment was enhanced in the presence of the newly injected mobile cells . This injection was an improvement on past experiments in that the attached (resident) cells were stained, allowing their concentrations to be enumerated directly by ferrographic capture (upon detachment) . Contrary to expectations based on previous experiments, enhanced detachment of stained resident cells did not occur upon the arrival of injected cells . Consistent with previous experiments, however, was the observation of ephemeral increases in unstained cell concentrations coincident with the arrival of the stained injected cells . The ephemeral pulses of unstained cells were previously speculated to represent enhanced detachment of unstained indigenous cells in response to hydrodynamic collision with injected cells . The lack of enhanced detachment of stained resident cells in the present experiments indicates that increased concentrations of unstained cells may have occurred by mechanisms other than hydrodynamic collision . Visually observed variations in stain intensity indicated that increased unstained cell concentrations may have resulted from cell division at the low-concentration fringe of the injected plume.

Eur J Obstet Gynecol Reprod Biol, 2003 Jun 10, 108(2), 146 - 51
Bacterial vaginosis: prevalence and predictive value for premature delivery and neonatal infection in women with preterm labour and intact membranes; Goffinet F et al.; OBJECTIVES: Assess the predictive values of bacterial vaginosis (BV) for preterm delivery (PD) and neonatal infection and compare them with standard markers of infection among women with preterm labour (PL) . STUDY DESIGN: Prospective blinded study in a tertiary referral centre in Paris . Women hospitalised for PL with intact membranes at a term between 24 and 34 weeks were included . Vaginal fluid, collected at inclusion was Gram-stained, scored, and interpreted according to Nugent's criteria . RESULTS: Out of 354 women tested, 254 had normal flora (72.3%), 76 intermediate (21.7%) and 24 BV (6.8%) . A history of spontaneous miscarriage after 14 weeks was the only risk factor significantly associated with BV . BV was not significantly associated with PD<35 weeks or neonatal infection . Very preterm delivery (before 33 weeks) was significantly associated with the flora grade (P=0.02): women with normal, intermediate and abnormal flora, respectively had 27 (10.6%), 14 (18.4%) and 6 (25.0%) births before 33 weeks . Of the markers tested, the highest risk of very preterm delivery was associated with BV (odds ratio 2.95, 95% CI (1.1-0.8.1)) and CRP>20mg/dl (4.23 95% CI (1.8-9.7)) . Predictive value of BV for preterm birth before 33 weeks were: sensitivity 12.8%, specificity 95.0%, positive predictive value 35.3%, and negative predictive value 84.3% . CONCLUSIONS: The frequency of BV and its association with PD are probably very variable and must be interpreted differently from one population to another . While we found an association between BV results and delivery before 33 weeks, the predictive value of BV was disappointing . Although these findings reinforce the importance of a useful marker of subclinical infection, the usefulness of testing for BV in women with PL has not been demonstrated.

Arq Gastroenterol, 2002 Jul-Sep, 39(3), 158 - 62 Epub 2003 May 21.
{Prevalence and prognosis of spontaneous bacterial peritonitis . Experience in patients from a general hospital in Porto Alegre, RS, Brazil (1991-2000)}; Coral G et al.; BACKGROUND: Spontaneous bacterial peritonitis is a frequent complication that occurs in patients with cirrhosis and ascites and has a recurrence rate of 70% in 1 year . In addition, this infection determines a poor short and long-term prognosis and a shorter survival rate . AIMS: Evaluate the prevalence of spontaneous bacterial peritonitis in cirrhotic patients with ascites and the effect of its occurrence on the survival . PATIENTS/METHODS: One thousand and thirty admissions of patients with cirrhosis and ascites were reviewed and 114 episodes of spontaneous bacterial peritonitis were documented in 94 patients . The ascitic analysis was accomplished in all patients . The diagnosis of this infection was established when the ascitic fluid polymorphonuclear count was equal or above 250 cells mm3 . RESULTS: The prevalence of this infection was 11.1% and the mortality rate 21.9% . Spontaneous bacterial peritonitis was community acquired in 61.4% and hospital acquired in 37.7% . The mortality rate was 18.6% and 27.9%, respectively . The infection resolved in 91.1% of the episodes by the analysis of ascitic fluid at 48 hours on antibiotics . The use of prophylactic antibiotics was documented in 22.3% of the episodes, but there are not significant differences on the mortality or type of bacteria isolated when comparing the patients with or without this treatment . CONCLUSIONS: Spontaneous bacterial peritonitis is a common complication in patients with cirrhosis and ascites and determines a worse prognosis, mainly when related with absence of initial response to antibiotics.

J Trauma, 2003 May, 54(5), 908 - 14
Allogeneic blood transfusion increases the risk of postoperative bacterial infection: a meta-analysis; Hill GE et al.; BACKGROUND: Immunosuppression is a consequence of allogeneic (homologous) blood transfusion (ABT) in humans and is associated with an increased risk in cancer recurrence rates after potentially curative surgery as well as an increase in the frequency of postoperative bacterial infections . Although a meta-analysis has been reported demonstrating the relationship between ABT and colon cancer recurrence, no meta-analysis has been reported demonstrating the relationship of ABT to postoperative bacterial infection . METHODS: Twenty peer-reviewed articles published from 1986 to 2000 were included in a meta-analysis . Criteria for inclusion included a clearly defined control group (nontransfused) compared with a treated (transfused) group and statistical analysis of accumulated data that included stepwise multivariate logistic regression analysis . In addition, a subgroup of publications that included only the traumatically injured patient was included in a separate meta-analysis . A fixed effects analysis was conducted with odds ratios obtained by using the conditional maximum likelihood method and 95% confidence intervals on the obtained odds ratios were determined using the mid-p technique . RESULTS: The total number of subjects included in this meta-analysis was 13,152 (5,215 in the transfused group and 7,937 in the nontransfused group) . The common odds ratio for all articles included in this meta-analysis evaluating the association of ABT to the incidence of postoperative bacterial infection was 3.45 (range, 1.43-15.15), with 17 of the 20 studies demonstrating a value of p < or = 0.05 . These results provide overwhelming evidence that ABT is associated with a significantly increased risk of postoperative bacterial infection in the surgical patient . The common odds ratio of the subgroup of trauma patients was 5.263 (range, 5.03-5.43), with all studies showing a value of p < 0.05 (0.005-0.0001) . These results demonstrate that ABT is associated with a greater risk of postoperative bacterial infection in the trauma patient when compared with those patients receiving ABT during or after elective surgery . CONCLUSION: These results demonstrate that ABT is an associated and apparently significant and frequently overlooked risk factor for the development of postoperative bacterial infection in the surgical patient . Allogeneic blood transfusion is a greater risk factor in the traumatically injured patient when compared with the elective surgical patient for the development of postoperative bacterial infection.

Acta Crystallogr D Biol Crystallogr, 2003 Jun, 59(Pt 6), 1061 - 3 Epub 2003 May 23.
Crystallization of Hfq protein: a bacterial gene-expression regulator; Vassilieva IM et al.; Hfq protein from Escherichia coli (EcoHfq) has been overproduced in E . coli, purified to homogeneity and crystallized using the hanging-drop vapour-diffusion technique . Crystallization conditions for EcoHfq were found which yielded X-ray quality crystals . Crystals of EcoHfq and of Cd-, Hg- and Se-containing derivatives grew in two months, with unit-cell parameters a = b = 127.41, c = 170.36 A . The crystals belong to space group I4 and diffract to 2.1 A resolution . Two hexamers are predicted per asymmetric unit.

Plant Mol Biol, 2003 Apr, 51(6), 803 - 15
Differential expression of genes encoding calmodulin-binding proteins in response to bacterial pathogens and inducers of defense responses; Ali GS et al.; Calmodulin (CaM) plays an important role in sensing and transducing changes in cellular Ca2+ concentration in response to several biotic and abiotic stresses . Although CaM is implicated in plant-pathogen interactions, its molecular targets and their role in defense signaling pathway(s) are poorly understood . To elucidate the signaling pathways that link CaM to defense responses, we screened a cDNA library constructed from bean leaves undergoing a hypersensitive response (HR) with radiolabeled CaM isoforms . A total of 26 putative CBPs were identified . Sequencing of the cDNAs revealed that they represent 8 different genes . They are homologues of previously identified CaM-binding proteins (CBPs) in other systems . However, some CBPs are novel members of known CBP families . The proteins encoded by these clones bound CaM in a Ca2+-dependent manner . To determine if these CBPs are involved in plant defense responses, we analyzed their expression in bean leaves inoculated with compatible, incompatible and nonpathogenic bacterial strains . Expression of three CBPs including an isoform of cyclic nucleotide-gated channels (PvCNGC-A) and two hypothetical proteins (PvCBP60-C and PvCBP60-D) was induced whereas the expression of two other isoforms of CNGCs (PvCNGC-B and PvCNGC-C) was repressed in response to incompatible pathogens . The expression of the rest, a small auxin up RNA (PvSAUR1) and two hypothetical proteins (PvCBP60-A and PvCBP60-B), was not changed . The expression of most of the pathogen-regulated genes was also affected by salicylic acid, jasmonic acid, hydrogen peroxide and a fungal elicitor, which are known to induce defense responses . Our results strongly suggest that at least five bean CBPs are involved in plant defense responses.

J Bacteriol, 2003 Jun, 185(12), 3636 - 43
Electron microscopic analysis of membrane assemblies formed by the bacterial chemotaxis receptor Tsr; Weis RM et al.; The serine receptor (Tsr) from Escherichia coli is representative of a large family of transmembrane receptor proteins that mediate bacterial chemotaxis by influencing cell motility through signal transduction pathways . Tsr and other chemotaxis receptors form patches in the inner membrane that are often localized at the poles of the bacteria . In an effort to understand the structural constraints that dictate the packing of receptors in the plane of the membrane, we have used electron microscopy to examine ordered assemblies of Tsr in membrane extracts isolated from cells engineered to overproduce the receptor . Three types of assemblies were observed: ring-like "micelles" with a radial arrangement of receptor subunits, two-dimensional crystalline arrays with approximate hexagonal symmetry, and "zippers," which are receptor bilayers that result from the antiparallel interdigitation of cytoplasmic domains . The registration among Tsr molecules in the micelle and zipper assemblies was sufficient for identification of the receptor domains and for determination of their contributions to the total receptor length . The overall result of this analysis is compatible with an atomic model of the receptor dimer that was constructed primarily from the X-ray crystal structures of the periplasmic and cytoplasmic domains . Significantly, the micelle and zipper structures were also observed in fixed, cryosectioned cells expressing the Tsr receptor at high abundance, suggesting that the modes of Tsr assembly found in vitro are relevant to the situation in the cell.

Rev Neurol (Paris), 2003 Apr, 159(4), 421 - 4
{Early diagnosis of bacterial brain abscesses: interest of diffusion-weighted MRI}; Detante O et al.; Three cases of bacterial brain abscesses, in immunocompetent patients, are reported . In all these cases, the diffusion-weighted magnetic resonance (MRI) with apparent diffusion coefficient (ADC) map has permitted an early diagnosis and a rapid treatment . This emergency MRI showed in the three cases a low signal on TI-weighted images, a high signal on T2-weighted and echo-planar images, and a decrease of ADC (0.36- 0.49 x 10(-3) mm2/s) . So, this new MRI technique provides an available and rapid element in the brain abscess diagnosis which often remains a complex clinical and radiological diagnosis.

Toxicol Sci, 2003 Aug, 74(2), 457 - 69 Epub 2003 May 28.
The coagulation system contributes to synergistic liver injury from exposure to monocrotaline and bacterial lipopolysaccharide; Yee SB et al.; Coexposure to a noninjurious dose of bacterial lipopolysaccharide (LPS; 7.4 x 106 EU/kg) and a nontoxic dose of the food-borne toxin monocrotaline (MCT; 100 mg/kg) leads to synergistic hepatotoxicity in Sprague-Dawley rats . Inflammatory factors, such as Kupffer cells (KCs), tumor necrosis factor-alpha (TNF)-alpha, and neutrophils (polymorphonuclear leukocytes; PMNs), are critical to the pathogenesis . Inasmuch as activation of the coagulation system and sinusoidal endothelial cell (SEC) injury precede hepatic parenchymal cell (HPC) injury, and since fibrin deposition occurs within liver lesions, the coagulation system might be a critical component of injury . In this study, this hypothesis is tested, and the interdependence of the coagulation system and inflammatory factors is explored . Administration of the anticoagulants heparin or warfarin to MCT/LPS-cotreated animals attenuated HPC and SEC injury . Morphometric analysis revealed that anticoagulant treatment significantly reduced the area of centrilobular and midzonal lesions . Heparin treatment also reduced fibrin deposition in these regions . Furthermore, anticoagulant treatment decreased hepatic PMN accumulation but did not affect plasma TNF-alpha concentration . Neither KC inactivation nor TNF-alpha depletion prevented activation of the coagulation system . PMN depletion, however, prevented coagulation system activation, suggesting that PMNs are needed for this response . These results provide evidence that the coagulation system and its interplay with PMNs are important in the pathogenesis of MCT/LPS-induced liver injury.

Nucleic Acids Res . 2003 Jun 1;31(11):e65.
An automated microplate-based method for monitoring DNA strand breaks in plasmids and bacterial artificial chromosomes; Rock C et al.; A method is described for high-throughput monitoring of DNA backbone integrity in plasmids and artificial chromosomes in solution . The method is based on the denaturation properties of double-stranded DNA in alkaline conditions and uses PicoGreen fluorochrome to monitor denaturation . In the present method, fluorescence enhancement of PicoGreen at pH 12.4 is normalised by its value at pH 8 to give a ratio that is proportional to the average backbone integrity of the DNA molecules in the sample . A good regression fit (r2 > 0.98) was obtained when results derived from the present method and those derived from agarose gel electrophoresis were compared . Spiking experiments indicated that the method is sensitive enough to detect a proportion of 6% (v/v) molecules with an average of less than two breaks per molecule . Under manual operation, validation parameters such as inter-assay and intra-assay variation gave values of <5% coefficient of variation . Automation of the method showed equivalence to the manual procedure with high reproducibility and low variability within wells . The method described requires as little as 0.5 ng of DNA per well and a 96-well microplate can be analysed in 12 min providing an attractive option for analysis of high molecular weight vectors . A preparation of a 116 kb bacterial artificial chromosome was subjected to chemical and shear degradation and DNA integrity was tested using the method . Good correlation was obtained between time of chemical degradation and shear rate with fluorescence response . Results obtained from pulsed- field electrophoresis of sheared samples were in agreement with those obtained using the microplate-based method.

J Exp Med, 2003 Jun 2, 197(11), 1417 - 25 Epub 2003 May 27.
Effect of anatomical distribution of mast cells on their defense function against bacterial infections: demonstration using partially mast cell-deficient tg/tg mice; Jippo T et al.; Mast cells were depleted in the peritoneal cavity of WBB6F1-tg/tg mice that did not express a transcription factor, MITF . When acute bacterial peritonitis was induced in WBB6F1-+/+, WBB6F1-W/Wv, and WBB6F1-tg/tg mice, the proportion of surviving WBB6F1-+/+ mice was significantly higher than that of surviving WBB6F1-W/Wv or WBB6F1-tg/tg mice . The poor survival of WBB6F1-W/Wv and WBB6F1-tg/tg mice was attributed to the deficient influx of neutrophils into the peritoneal cavity . The injection of cultured mast cells (CMCs) derived from WBB6F1-+/+ mice normalized the neutrophil influx and reduced survival rate in WBB6F1-W/Wv mice, but not in WBB6F1-tg/tg mice . This was not attributable to a defect of neutrophils because injection of TNF-alpha increased the neutrophil influx and survival rate in both WBB6F1-W/Wv and WBB6F1-tg/tg mice . Although WBB6F1-+/+ CMCs injection normalized the number of mast cells in both the peritoneal cavity and mesentery of WBB6F1-W/Wv mice, it normalized the number of mast cells only in the peritoneal cavity of WBB6F1-tg/tg mice . Mast cells within the mesentery or mast cells in the vicinity of blood vessels appeared to play an important role against the acute bacterial peritonitis . WBB6F1-tg/tg mice may be useful for studying the effect of anatomical distribution of mast cells on their antiseptic function.

J Biotechnol, 2003 Jun 12, 103(1), 67 - 76
Effects of bacterial treatments on wood extractives; Kallioinen A et al.; Bacterial strains were isolated from spruce wood chips and their ability to reduce the content of wood extractives was studied . Strains were screened by cultivation on liquid media containing wood extractives as the major nutrient . Some bacterial species could decrease remarkably the amount of extractives in the liquid media and reduced the amount of triglycerides, steryl esters and total extractives by 100, 20 and 39%, respectively . Spruce wood chips were treated in controlled conditions with selected bacteria to test their effects on the chips . All the bacteria grew well on wood chips . The effect of bacterial metabolism on wood extractives was significant . Bacterial treatments reduced the amount of lipophilic extractives by 16-38% in 1 week of treatment and up to 67% in 2 weeks . The most efficient strain removed 90, 66 and 50% of triglycerides, steryl esters and resin acids, respectively, in 2 weeks . These results indicate that bacteria may be promising agents for the removal of extractives for improved pulping and papermaking processes.

J Insect Physiol, 1998 Feb, 44(2), 157 - 164
The influence of bacterial species and intensity of infections on nodule formation in insects; Stanley DW et al.; Nodulation is the predominant cellular immune reaction to bacterial infection in insects . Nodulation is a complex process involving an unknown number of discrete cellular actions . Currently, there is only limited information on the signal transduction mechanisms that result in nodulation . In older larvae of the tobacco hornworm, Manduca sexta, and of the tenebrionid beetle, Zophobas atratus, eicosanoids are involved in one or more steps in the overall process, and treating these insects with inhibitors of eicosanoid biosynthesis prior to bacterial infection severely impairs their ability to form nodules . In this paper we address more detailed questions on eicosanoid-mediated nodulation . The nodulation reaction to bacterial infection occurs in all larval stages we examined, specifically, second, third, and fourth instars of M . sexta . In both species, the number of nodules formed in response to bacterial infection is related in an exponential way to the number of bacterial cells in the infection . Nodulation is also not related to larval size . We also found that nodulation intensity varies according to the species of infecting bacteria.

DNA Repair (Amst), 2003 Jun 11, 2(6), 707 - 18
Comparative analysis of 8-oxoG:C, 8-oxoG:A, A:C and C:C DNA repair in extracts from wild type or 8-oxoG DNA glycosylase deficient mammalian and bacterial cells; Dantzer F et al.; We have investigated repair of DNA containing 8-oxoguanine and certain mismatches in cell-free extracts from mouse embryonic fibroblasts (MEFs) using a plasmid substrate with a single lesion at a defined position . Repair synthesis was monitored in a small restriction fragment with different size single strands in order to follow the fate of repair reactions in both strands at the same time . An important part of the study was to assess the role of OGG1 in various repair reactions and the experiments were carried out with extracts from mouse embryonic fibroblasts diploid for a mogg1 deletion (Ogg1(-/-)) as well as wild type . In wild type, DNA containing 8-oxoG:C was repaired in the expected fashion predominantly through short-patch repair . Overall repair was reduced to 20% in the Ogg1(-/-) extracts and to 40% if only long-patch repair was considered . The 8-oxoG:A pair was processed similarly in wild type and Ogg1(-/-) extracts and repair synthesis at A as well as at 8-oxoG could be demonstrated, however, to the same extent in Ogg1(-/-) and wild type for both strands . Extracts from Ogg1(-/-) behaved normally in the correction of A:C and C:C mismatches, with a strong bias for correction of A for A:C and no significant strand discrimination for C:C . Similar experiments with extracts from Escherichia coli showed a 50% reduction in the repair of 8-oxoG:C in fpg extracts and an increase to 50% above wild type in mutY . These results show that the mouse OGG1 is the major enzyme for 8-oxoG repair in the MEF cells and does not participate in mismatch repair of A:C or C:C . Furthermore, 8-oxoG opposite A appears to be repaired by a two-step repair pathway with sequential removal of A and 8-oxoG mediated by enzymes different from OGG1.

Water Res, 2003 Jul, 37(12), 2929 - 36
Bacterial immobilization and oxidation of arsenic in acid mine drainage (Carnoulès creek, France); Casiot C et al.; The acid waters (pH=2.73-3.37) originating from the Carnoules mine tailings contain high dissolved concentrations of arsenic (1-3.5 mmol l(-1)) and iron (20-40 mmol l(-1)) . At the outlet, arsenite predominates . During the first 30 m of downflow, 20-60% is removed by coprecipitation with Fe(III) . This process results from bacterially mediated As- and Fe-oxidation . The precipitation rates in the creek depend on the oxygen concentration in spring water and are lower during the dry summer period when the anoxic character of the spring water inhibits the activity of oxidizing bacteria . Ex situ experiments show that the presence of bacteria-rich precipitates increases the As- and Fe-removal rates . Three strains of bacteria promoting the oxidation of As have been isolated, and two of them have the characteristics of Thiomonas ynys1 . The third strain, which is not identified yet, also catalyzes the oxidation of Fe.

Clin Infect Dis, 2003 Jun 1, 36(11), 1492 - 5 Epub 2003 May 16.
Clearance of 14-3-3 protein from cerebrospinal fluid heralds the resolution of bacterial meningitis; Bonora S et al.; The 14-3-3 protein, a cerebrospinal fluid (CSF) marker of neuronal damage that was recently adopted for the diagnosis of Creutzfeldt-Jakob disease, is also found in the CSF of patients with a variety of neurological disorders . We prospectively studied 12 consecutive patients with purulent bacterial meningitis and found that 14-3-3 protein was detected in all patients at admission to the hospital . All patients who recovered cleared 14-3-3 protein from the CSF before discharge from the hospital (this was the first CSF marker to clear), whereas those who died never cleared the protein.

Clin Nutr, 2003 Jun, 22(3), 277 - 81
The effect of immunonutrition on bacterial translocation, and intestinal villus atrophy in experimental obstructive jaundice; Zulfikaroglu B et al.; BACKGROUND & AIMS: Spontaneous bacterial infection and septicemia due to increased bacterial translocation (BT) in patients with obstructive jaundice result in significant morbidity and mortality . The present study evaluates the effects of enteral nutrition with immune enhancing feeds on BT and intestinal villus histopathology promoted by obstructive jaundice . METHODS: Fifty male Wistar-albino rats weighing 250-300g were assigned into five equal groups of 10 . Animals in Groups I, II, and III were fed with standard chow, those in Group IV were given glutamine 1g/kg/day and the remaining 10 animals in Group V were fed with an arginine, omega-3 fatty acids, and RNA-supplemented enteral diet for (1g/kg/day amino acid and 230 kcal/kg) 7 days preoperatively . Group I underwent sham operation and the remaining animals in all other groups underwent common bile duct ligation . After operation, Group I had standard chow, Groups II and IV had glutamine, Groups III and V had an arginine omega-3 fatty acids, and RNA-supplemented enteral diet for 7 days . All animals were sacrificed on the 8th postoperative day and evaluated both biochemically and histopathologically . Samples from blood, liver, mesenteric lymph nodes and spleen were cultured under aerobic conditions . RESULTS: Significantly less BT was observed in groups fed with an arginine, omega-3 fatty acids, and RNA-supplemented enteral diet or glutamine in pre-and postoperative periods as compared to others (P<0.001) . Histologic evaluation also showed significant reduction in villus atrophy in these groups . CONCLUSIONS: Enteral immunonutrition using glutamine or arginine, omega-3 fatty acids, and RNA-supplemented enteral diet during both pre-and postoperative periods seems to reduce BT and decrease atrophy of intestinal mucosal villi in rats with obstructive jaundice.

In Silico Biol, 2003, 3(1-2), 127 - 43 Epub 2003 Mar 16.
Analysis of bacterial RM-systems through genome-scale analysis and related taxonomy issues; Vandenbogaert M et al.; Recognition sites for type II restriction and modification enzymes in genomes of several bacteria are recognized as semi-palindromic motifs and are avoided at a significant degree . The key idea of contrast word analysis with respect to RMS recognition sites, is that under-represented words are likely to be selected against . Starting from over- or underrepresented words corresponding to RMS recognition sites in specific clades, the specificity of unknown R-M systems can be highlighted . Among the known restriction enzymes, that are described in the REBASE database of restriction and modification systems, many of their recognition sites are still uncharacterized . Eventually, this motivates studies aimed at assessing horizontal transferring events of RMS in micro-organisms through the analysis of word usage biases in well-determined genomic regions . A probabilistic model is built on a first-order Markovian chain . Statistics on the k-neighborhood of a word is carried out to assess the biological significance of a genomic motif . Efficient word counting procedures have been implemented and statistics are used for the assessment of the significance of individual words in large sequences . On the basis of the set of most avoided words, and in accordance to the IUPAC coding standards, suggestions are made regarding potential recognition sequences . In certain cases, a comparison of avoided palindromic words in taxonomically related bacteria shows a pattern of relatedness of their R-M systems . For strengthening this analysis, the primary protein structure of all type II R-M systems known in REBASE have been blasted against the nr-GENBANK database . The combination of these analyses has revealed some interesting examples of possible horizontal transfer events of R-M systems.

Oper Dent, 2003 May-Jun, 28(3), 274 - 80
Physiological remineralization of artificially demineralized dentin beneath glass ionomer cements with and without bacterial contamination in vivo; Kitasako Y et al.; This study evaluated the physiological remineralization of artificially demineralized dentin beneath glass ionomer cements with and without bacterial contamination . The artificially demineralized dentin was produced on 84 monkey teeth using a decalcifying solution . Half the samples were left open to the oral cavity for one week, then, all cavities were restored with two glass-ionomer cements: Fuji IX or Fuji II LC improved (n = 7) . The nanohardness of the artificially demineralized dentin at 3, 90 and 360 days was measured using a nanoindentation tester (ENT-1100, Elionix) and compared statistically by two-way ANOVA and Fisher's PLSD test (p < 0.05) . Each mineral (Ca, Mg, P, F) within the demineralized dentin was also analyzed using Electron Probe Microanalysis . For the samples, the mean nanohardness of the three-day samples was significantly lower than the 360-day samples (p < 0.05) . Although there was no significant difference in the mean nanohardness within all the bacterially-contaminated groups through the experimental periods (p > 0.05), the mean nanohardness of the bacterial-contaminated samples were significantly lower than the non-bacteria-contaminated samples (p < 0.05) . From the EPMA results, fluoride release from both cements to the bottom of the artificially demineralized dentin was detected within three days . Although Ca density was sparse within this demineralized dentin lesion, for the Fuji IX sample, a high Mg density within this lesion was detected at 360 days.

J Antibiot (Tokyo), 2003 Mar, 56(3), 259 - 67
Studies on novel bacterial translocase I inhibitors, A-500359s . III . Deaminocaprolactam derivatives of capuramycin: A-500359 E, F, H; M-1 and M-2; Muramatsu Y et al.; Novel derivatives of capuramycin were obtained when 10 mM of 2-aminoethyl-L-cysteine (AEC), an inhibitor of aspartokinase, was added to the culture of Streptomyces griseus SANK 60196, the producer of A-500359 . They were purified from the culture filtrate and their chemical structures were elucidated as a deaminocaprolactam derivative of capuramycin designated as A-500359 F, A-500359 E, a methyl ester of A-500359 F, and A-500359 H, a 3'-demethyl derivative of A-500359 F . Two other compounds, A-500359 M-1 and A-500359 M-2, were purified from the same medium and their structures were elucidated . A-500359 E, F, H, M-1 and M-2 inhibited bacterial translocase I with an IC50 of 0.027 microM, 1.1 microM, 0.008 microM, 0.058 microM and 0.010 microM, respectively . A-500359 E, M-1 and M-2 inhibited the growth of mycobacteria as well.

J Oral Implantol, 2003, 29(2), 80 - 5
Bacterial adhesion on titanium nitride-coated and uncoated implants: an in vivo human study; Scarano A et al.; Titanium nitride (TiN) has been used in many fields as a surgical instrument coating that makes the surgical materials more resistant to wear and corrosion . The aim of the present study was an in vivo evaluation of the bacterial adhesion to TiN-coated (test) and uncoated (control) titanium implants . Six patients aged between 21 and 25 years and in excellent systemic health participated in the study . All of the participants gave their informed consent . The participants were selected on the basis of good periodontal health and no signs of mouth breathing . In each of the 6 participants, a removable acrylic device was adapted to the molar-premolar region of each quadrant of the jaws . One 4 x 13 mm titanium implant was glued to the buccal aspect of each device . The plasma spray covered 11.5 mm of the body of the implant, whereas the neck was machined titanium . Test implants were glued to the right devices and control implants were glued to the left devices . After 24 hours, the implants were removed from each device and processed for scanning electron microscopy for evaluation of the machined portion of the implant covered by bacteria . A total of 24 implants were used in this study, 12 test and 12 control . Surface characterization of the machined portion of the neck of the implant was performed on an additional 10 implants (5 test and 5 control) . On test implants the implant surface covered by bacteria was significantly lower compared with that of control implants (P = .0001) . The surface roughness was similar in both groups . TiN surfaces showed a significant reduction of the presence of bacteria, and this fact could probably be important in the decrease of the inflammation of the peri-implant soft tissues.

Infez Med, 1999, 7(2), 108 - 112
{Bacterial pneumonia in HIV-infected patients}; De Gaetano Donati K et al.; This case-control study assessed risk factors and prognostic indicators of 350 episodes of bacterial pneumonia in 285 HIV-infected patients . On univariate analysis, intravenous drug abuse (p<0.001), regular cigarette smoking (p<0.001), cirrhosis (p=0.04), and history of a previous episode of pneumonia (p=0.04), were risk factors for community-acquired episodes of bacterial pneumonia, whereas length of hospitalization (p=0.01) was a risk factor only for nosocomial bacterial pneumonia . The small amount of circulating T CD4+ cells, (<100/mmc) was a risk factor in both groups of pneumonia (p<0.05) . Stepwise logistic regression analysis revealed that IVDA in community-acquired episodes and low levels of circulating T CD4+ cells, both in community-acquired and hospital-acquired episodes, were independent risk factors for the development of bacterial pneumonia . The case-fatality rate observed in our study was 27% . On stepwise logistic regression analysis, T CD4+ cell counts >100/mmc (p<0.02), neutropenia (p=0.04), PO2 arterial level <70 mmHg (p=0.01), and Karnofsky score <50 (p=0.04) were independent indicators of mortality . According to a personally developed prognostic score, 211 episodes of pneumonia (60%) were classified as mild, 63 (18%) as moderate, and 76 (22%) as severe . Clinicians must carefully evaluate those variables that can influence the prognosis of bacterial pneumonia to make early identification of affected patients and to promptly establish the most appropriate therapeutic strategy in each case.

J Mol Biol, 2003 May 30, 329(2), 291 - 309
A spatially extended stochastic model of the bacterial chemotaxis signalling pathway; Shimizu TS et al.; We have combined two distinct but related stochastic approaches to model the Escherichia coli chemotaxis pathway . Reactions involving cytosolic components of the pathway were assumed to obey the laws of conventional stochastic chemical kinetics, while the clustered membrane receptors were represented in two-dimensional arrays similar to the Ising model . Receptors were assumed to flip between an active and an inactive state with probabilities dependent upon three energy inputs: ligand binding, methylation level due to adaptation, and the activity of neighbouring receptors . Examination of models with different lattice size and geometry showed that the sensitivity to stimuli increases with lattice size and the nearest-neighbour coupling strength up to a critical point, but this amplification was also accompanied by a proportional increase in steady-state noise . Multiple methylation of receptors resulted in diminished signal-to-noise ratio, but showed improved stability to variation in the coupling strength and increased gain . Under the best conditions the simulated output of a coupled lattice of receptors closely matched the time-course and amplitude found experimentally in living bacteria . The model also has some of the properties of a cellular automaton and shows an unexpected emergence of spatial patterns of methylation within the receptor lattice.

J Mol Biol, 2003 May 30, 329(2), 283 - 90
The 9.8 kDa subunit of complex I, related to bacterial Na(+)-translocating NADH dehydrogenases, is required for enzyme assembly and function in Neurospora crassa; Marques I et al.; A nuclear gene encoding a 9.8 kDa subunit of complex I, the homologue of mammalian MWFE protein, was identified in the genome of Neurospora crassa . The gene was cloned and inactivated in vivo by the generation of repeat-induced point mutations . Fungal mutant strains lacking the 9.8 kDa polypeptide were subsequently isolated . Analyses of mitochondrial proteins from mutant nuo9.8 indicate that the membrane and peripheral arms of complex I fail to assemble . Respiration of mutant mitochondria on matrix NADH is rotenone-insensitive, confirming that the 9.8 kDa protein is required for the assembly and activity of complex I . We found a similarity between the MWFE homologues and the C-terminal part of the nqrA subunit of bacterial Na(+)-translocating NADH:quinone oxidoreductases (Na(+)-NQR), suggesting a link between proton-pumping and sodium-pumping NADH dehydrogenases.

FEMS Microbiol Lett, 2003 May 16, 222(1), 17 - 23
HutC/FarR-like bacterial transcription factors of the GntR family contain a small molecule-binding domain of the chorismate lyase fold; Aravind L et al.; Numerous bacterial transcription factors contain a DNA-binding helix-turn-helix domain and a signaling domain, linked together in a single polypeptide . Typically, this signaling domain is a small-molecule-binding domain that undergoes a conformational change upon recognizing a specific ligand . The HutC/FarR-like transcription factors of the GntR family are one of the largest groups of transcription factors in the proteomes of most free-living bacteria . Using sensitive sequence profile analysis we show that the HutC/FarR-like transcription factors contain a conserved ligand-binding domain, which possesses the same fold as chorismate lyase (Escherichia coli UbiC gene product) . This relationship suggests that the C-terminal domain of the HutC/FarR-like transcription factors binds small molecules in a cleft similar to the substrate-binding site of the chorismate lyases . The sequence diversity within the predicted binding cleft of the HutC/FarR ligand-binding domains is consistent with the ability of these transcription factors to respond to diverse small molecules, such as histidine (HutC), fatty acids (FarR), sugars (TreR) and alkylphosphonate (PhnF) . UbiC-like chorismate lyases function in the ubiquinone biosynthesis pathway, and have characteristic charged, catalytic residues . Genome comparisons reveal that chorismate lyase orthologs are found in several bacteria, chloroplasts of eukaryotic algae and euryarchaea . In contrast, the GntR transcription regulators lack the conserved catalytic residues of the chorismate lyases, and have so far been detected only in bacteria . An ancestral, generic small-molecule-binding domain appears to have given rise to the enzymatic and non-catalytic ligand-binding versions of the same fold under the influence of different selective pressures.

J Am Geriatr Soc, 2003 Jun, 51(6), 768 - 73
Prevalence of small bowel bacterial overgrowth and its association with nutrition intake in nonhospitalized older adults; Parlesak A et al.; OBJECTIVES: To determine the prevalence of small bowel bacterial overgrowth (SBBO) in older adults and to assess whether SBBO is associated with abdominal complaints and nutrient intake . DESIGN: Cross-sectional survey . SETTING: Eight senior residence sites in Stuttgart, Germany . PARTICIPANTS: Older adults living independently in senior residence houses . MEASUREMENTS: The prevalence of SBBO was measured in 328 subjects, of whom 294 were aged 61 and older, by measuring hydrogen concentration (parts per million; ppm) in exhaled air after ingestion of 50 g glucose . Anthropometric data were obtained and nutritional status was recorded with a computer-aided diet history . RESULTS: The prevalence of a positive hydrogen breath test (>10 ppm increase) was 15.6% in older adults, compared with 5.9% in subjects aged 24 to 59 . The intake of inhibitors of gastric acid production contributed significantly to the high prevalence of a positive breath test in older adults, which was associated with lower body weight, lower body mass index, lower plasma albumin concentration, and higher prevalence of diarrhea . Subjects with a positive hydrogen breath test consumed significantly less fiber, folic acid, and vitamins B2 and B6 than those without . No difference was observed in the intake of energy, protein, fat, or carbohydrates . CONCLUSION: Prevalence of SBBO is associated with reduced body weight, which is paralleled by reduced intake of several micronutrients . Malabsorption resulting from diarrhea might be an aggravating factor contributing to weight loss in these subjects.

J Biol Chem, 2003 Aug 8, 278(32), 29719 - 27 Epub 2003 May 19.
Ssq1, a mitochondrial Hsp70 involved in iron-sulfur (Fe/S) center biogenesis . Similarities to and differences from its bacterial counterpart; Dutkiewicz R et al.; The results of in vivo and in organellar experiments indicate that the Hsp70 Ssq1 and the J-protein Jac1 function together to assist in the biogenesis of iron-sulfur (Fe/S) centers in the mitochondrial matrix . Here we present biochemical evidence supporting this idea . Isu, the proposed scaffold on which Fe/S centers are assembled, is a substrate for both Jac1 and Ssq1 . Jac1 and Isu1 cooperatively stimulate the ATPase activity of Ssq1 . In addition, Jac1 facilitates the interaction of Ssq1 with Isu1 in the presence of ATP . These findings are consistent with the role in Fe/S biogenesis previously proposed for the bacterial Hsp70 Hsc66 and J-protein Hsc20 that interact with the bacterial Isu homologue IscU . However, unlike the bacterial Hsp70, we found that Ssq1 has a high affinity for nucleotide, and shares a nucleotide exchange factor, Mge1, with a second mitochondrial Hsp70, Ssc1 . Thus, whereas the bacterial and mitochondrial chaperone systems share critical features, they possess significant biochemical differences as well.

Int J Med Microbiol, 2003 Apr, 293(1), 17 - 39
Intestinal M cells and their role in bacterial infection; Clark MA et al.; M cells are located in the epithelia overlying mucosa-associated lymphoid tissues such as Peyer's patches where they function as the antigen sampling cells of the mucosal immune system . Paradoxically, some pathogens exploit M cells as a route of invasion . Here we review our current knowledge of intestinal M cells with particular emphasis on the mechanisms underlying bacterial infection of these atypical epithelial cells.

Radiats Biol Radioecol, 2003 Mar-Apr, 43(2), 176 - 8
{Some effects of radiation hormesis for bacterial and yeast cells}; Petin VG et al.; A comparative study of chronic and acute action of ionizing radiation on the processes of aging and dying off of bacterial and yeast cells was carried out . It was ascertained that chronic action of ionizing radiation, 2-10,000 times exceeded the natural background, resulted in slowing down of aging and dying off of both pro- and eukaryotic cells . A single acute irradiation of yeast also resulted in the retardation of dying off of the yeast cells surviving after irradiation . The data is presented demonstrating a great increase in the survival of yeast cells under their repeated irradiation after recovery from potentially lethal radiation.

J Bacteriol, 2003 Jun, 185(11), 3458 - 68
Tertiary structure of bacterial murein: the scaffold model; Dmitriev BA et al.; Although the chemical structure and physical properties of peptidoglycan have been elucidated for some time, the precise three-dimensional organization of murein has remained elusive . Earlier published computer simulations of the bacterial murein architecture modeled peptidoglycan strands in either a regular (D . Pink, J . Moeller, B . Quinn, M . Jericho, and T . Beveridge, J . Bacteriol . 182: 5925-5930, 2000) or an irregular (A . Koch, J . Theor . Biol . 204: 533-541, 2000) parallel orientation with respect to the plasma membrane . However, after integrating published experimental data on glycan chain length distribution and the degree of peptide side chain cross-linking into this computer simulation, we now report that the proposed planar network of murein appears largely dysfunctional . In contrast, a scaffold model of murein architecture, which assumes that glycan strands extend perpendicularly to the plasma membrane, was found to accommodate published experimental evidence and yield a viable stress-bearing matrix . Moreover, this model is in accordance with the well-established principle of murein assembly in vivo, i.e., sequential attachment of strands to the preexisting structure . For the first time, the phenomenon of division plane alternation in dividing bacteria can be reconciled with a computer model of the molecular architecture of murein.

J Bacteriol, 2003 Jun, 185(11), 3392 - 9
Modeling bacterial evolution with comparative-genome-based marker systems: application to Mycobacterium tuberculosis evolution and pathogenesis; Alland D et al.; The comparative-genomic sequencing of two Mycobacterium tuberculosis strains enabled us to identify single nucleotide polymorphism (SNP) markers for studies of evolution, pathogenesis, and epidemiology in clinical M . tuberculosis . Phylogenetic analysis using these "comparative-genome markers" (CGMs) produced a highly unusual phylogeny with a complete absence of secondary branches . To investigate CGM-based phylogenies, we devised computer models to simulate sequence evolution and calculate new phylogenies based on an SNP format . We found that CGMs represent a distinct class of phylogenetic markers that depend critically on the genetic distances between compared "reference strains." Properly distanced reference strains generate CGMs that accurately depict evolutionary relationships, distorted only by branch collapse . Improperly distanced reference strains generate CGMs that distort and reroot outgroups . Applying this understanding to the CGM-based phylogeny of M . tuberculosis, we found evidence to suggest that this species is highly clonal without detectable lateral gene exchange . We noted indications of evolutionary bottlenecks, including one at the level of the PHRI "C" strain previously associated with particular virulence characteristics . Our evidence also suggests that loss of IS6110 to fewer than seven elements per genome is uncommon . Finally, we present population-based evidence that KasA, an important component of mycolic acid biosynthesis, develops G312S polymorphisms under selective pressure.

J Appl Microbiol, 2003, 94(6), 988 - 93
Comparison of rapid methods for the extraction of bacterial DNA from colonic and caecal lumen contents of the pig; Anderson KL et al.; AIMS: The increasing uses of DNA methodologies to study the micro flora of the pig gastrointestinal tract requires an efficient recovery of bacterial DNA from the intestinal sample . Thus, the objective of this study was to determine which DNA extraction methods are most effective for luminal samples from pigs . Several routinely used nucleic acid extraction procedures were compared based upon quantity and purity of extracted DNA . METHODS AND RESULTS: DNA was extracted from pig colonic and caecal lumen samples using 19 methods for bacterial DNA extraction . The quantity of total DNA recovered by each extraction method was determined and compared . Two methods using extraction with polyvinylpolypyrrolidone (PVPP) or phenol and two methods involving bead mill homogenization were found to provide the greatest quantity of extracted DNA for both colonic and caecal lumen . Extracted DNA from these four methods was further analysed for purity based upon the presence of PCR inhibitors, which was ascertained by determining the efficiency of amplification of a segment of the 16S rDNA . PCR amplification could be readily achieved with DNA extracted by each of these four methods, but efficiency of amplification tended to be higher with DNA from two of the methods (one extracted with PVPP and one with bead mill homogenization) . CONCLUSIONS: Four extraction methods proved to be significantly superior in quantity of DNA extracted from luminal samples . Of these four, no strong inhibitors of PCR amplification were detected in any of the extracted DNA . However, the efficiency of amplification tended to be lower in DNA samples from two of the methods, suggesting the presence of low levels of PCR inhibitors . SIGNIFICANCE AND IMPACT OF THE STUDY: Results of this study provide a basis for choosing which DNA extraction procedures are most effective for use with samples of pig lumen.

Scand J Infect Dis, 2003, 35(3), 196 - 7
Spontaneous bacterial peritonitis due to Brucella melitensis; Erbay A et al.; Peritonitis is an extremely rare complication of brucellosis . A case is reported of blood and ascitic culture-proven spontaneous bacterial peritonitis caused by Brucella melitensis, in a patient who had also cirrhosis.

DNA Seq, 2003 Feb, 14(1), 33 - 52
Latent periodicity of 21 bases typical for MCP II gene is widely present in various bacterial genes; Chaley MB et al.; The existence of a typical latent periodicity of 21 bases from the Tar chemoreceptor gene of Escherichia coli (E . coli) (MCP II) in the bacterial genes has been investigated in this work . Among 583 annotated bacterial genes and ORFs in the GenBank, in which the typical periodicity has been found, the chemoreceptors' genes constituted the most numerous group (18.5%) . This typical latent periodicity of 21 bases has been revealed in many different genes of regulatory proteins, DNA polymerases, reductases, kinases and others . The numbers in such gene groups varied from 1 to 4% of the total analyzed genes . The 2D-structures analysis of the amino acid residues, which have been translated from the genes' regions with 21 bases periodicity, has shown that, though the enrichment of alpha-helical structures in such sequences is kept in all cases, it is seen that the latent periodicity of 21 bases is a very sensitively tuned basis, allowing the translated residues to smoothly change from one conformation to another . Interesting results have been obtained for 16S rRNAs genes of proteobacteria . Short sequences-determinants have been revealed in the genes, which select beta and gamma proteobacteria with an accuracy of above 90%.

Appl Microbiol Biotechnol, 2003 Aug, 62(2-3), 110 - 23 Epub 2003 May 15.
Evolution of catabolic pathways for synthetic compounds: bacterial pathways for degradation of 2,4-dinitrotoluene and nitrobenzene; Johnson GR et al.; The pathways for 2,4-dinitrotoluene (2,4-DNT) and nitrobenzene offer fine illustrations of how the ability to assimilate new carbon sources evolves in bacteria . Studies of the degradation pathways provide insight about two principal strategies for overcoming the metabolic block imposed by nitro- substituents on aromatic compounds . The 2,4-DNT pathway uses novel oxygenases for oxidative denitration and subsequent ring-fission . The nitrobenzene pathway links facile reduction of the nitro- substituent, a novel mutase enzyme, and a conserved operon encoding aminophenol degradation for mineralization of nitrobenzene . Molecular genetic analysis with comparative biochemistry reveals how the pathways were assembled in response to the recent appearance of the two synthetic chemicals in the biosphere.

Genetics, 2003 May, 164(1), 311 - 21
A bacterial artificial chromosome contig spanning the major domestication locus Q in wheat and identification of a candidate gene; Faris JD et al.; The Q locus played a major role in the domestication of wheat because it confers the free-threshing character and influences many other agronomically important traits . We constructed a physical contig spanning the Q locus using a Triticum monococcum BAC library . Three chromosome walking steps were performed by complete sequencing of BACs and identification of low-copy markers through similarity searches of database sequences . The BAC contig spans a physical distance of approximately 300 kb corresponding to a genetic distance of 0.9 cM . The physical map of T . monococcum had perfect colinearity with the genetic map of wheat chromosome arm 5AL . Recombination data in conjunction with analysis of fast neutron deletions confirmed that the contig spanned the Q locus . The Q gene was narrowed to a 100-kb segment, which contains an APETALA2 (AP2)-like gene that cosegregates with Q . AP2 is known to play a major role in controlling floral homeotic gene expression and thus is an excellent candidate for Q.

Neurosci Lett, 2003 May 29, 343(1), 25 - 8
Bacterial lipopolysaccharide shifts fasted plasma ghrelin to postprandial levels in rats; Basa NR et al.; Lipopolysaccharide (LPS) injected intraperitoneally (i.p.) is known to decrease food intake . Ghrelin is a peptide hormone produced by the stomach with a potent orexigenic effect and plasma levels that are inversely correlated with the fed state . We examined changes in plasma ghrelin levels 3 h after LPS (100 microg/kg, i.p.) in fasted rats with or without a 1 h re-feeding period . LPS injection decreased the fasting levels of ghrelin by 51+/-5% compared with preinjection values while i.p . vehicle did not modify ghrelin levels in fasted rats . LPS at this dose reduced fasting-induced food intake by 60% compared with the i.p . vehicle group . Re-feeding decreased plasma ghrelin levels by 58+/-3% compared with pre-feeding fasting values in i.p . vehicle group . These data provide the first evidence that LPS shifts fasting ghrelin levels to those observed postprandially.

Am J Obstet Gynecol, 2003 May, 188(5), 1154 - 5
The role of covering gowns in reducing rates of bacterial contamination of scrub suits; Kaplan C et al.; OBJECTIVE: This study was undertaken to determine whether covering gowns reduce the rates of contamination of surgical scrubs . STUDY DESIGN: Seventy-five clinicians had pieces of fabric from clean scrubs attached to two areas of their scrub suits . Participants wore a covering garment when wearing scrub suits off of designated areas (n = 25), did not wear a covering garment (n = 25), or wore scrub suits outside the hospital (n = 25) . Subsequently, the fabric was assessed with culture in enhanced broth media and blood agar . RESULTS: Although there was a trend toward lower rates of contamination in the group that did not wear a covering garment, the difference was not significant . At no point, and at neither site of fabric attachment, did those who wore a covering garment demonstrate any advantage in regard to levels or frequency of contamination . CONCLUSION: Wearing covering garments over scrub suits does not reduce rates of contamination.

Lab Invest, 2003 May, 83(5), 721 - 30
Inhibition of cytokine-induced fractalkine production by bacterial invasion of human-dermal fibroblasts; Fahy OL et al.; Fractalkine (FKN/CX3CL1) is an atypical chemokine, for which a major biological function has not yet emerged . However, recent data suggest a role in immune responses in the skin . In this study, we analyzed fractalkine (FKN) secretion by human-dermal fibroblasts after exposure to pro-inflammatory cytokines or to invasive and noninvasive strains of Escherichia coli . Incubation of fibroblasts with TNF-alpha and IL-1beta induced a delayed expression of soluble FKN, compared with the rapid secretion of other chemokines including IL-8 (CXCL8), monocyte chemotactic protein-1 (CCL2), and RANTES (regulated upon activation, normal T cell expressed and secreted; CCL5) . TNF-alpha and IFN-gamma gamma were more potent at inducing FKN secretion than was IL-1beta . Very little FKN was detected on the cell surface . FKN was not detected after incubation with the bacteria, regardless of the strain used . In contrast, both invasive and noninvasive E . coli triggered the release of IL-8 and monocyte chemotactic protein-1 in a dose-response manner, whereas RANTES was produced only in response to the invasive strain . Finally, incubation of fibroblasts with the invasive strain of E . coli inhibited TNF-alpha- and IFN-gamma-induced production of FKN . These results demonstrate for the first time that human-dermal fibroblasts express FKN, and that the characteristics of FKN secretion are distinct from those of other chemokines produced by these cells during immune responses in the dermis . In addition, our data indicate that bacterial invasion of dermal fibroblasts actively modulates FKN expression.

Am J Dent, 2003 Feb, 16(1), 3 - 5
Removal of bacterial endotoxin from root surface with Er:YAG laser; Folwaczny M et al.; PURPOSE: To determine the potential of 2.94 microm Er:YAG laser radiation to remove bacterial endotoxin from root surfaces . MATERIALS AND METHODS: 40 extracted teeth were divided into four groups of 10 samples each . A 16 mm2 area of the root surface on each sample was inoculated with an aliquot of 7 microl of a lipopolysaccharide suspension at a concentration of 50 IU/ml . LPS was derived from a non-oral Escherichia coli strain (E . coli 055:B5) . Source of laser radiation was an Er:YAG laser emitting pulsed infrared radiation at a wavelength of 2.94 microm, with a pulse duration of 250 micros, and a pulse repetition rate of 15 pps . Three specimen groups were irradiated with 105 laser pulses at a radiation energy of 60 mJ, 100 mJ and 140 mJ . One specimen group was untreated (control) . The LPS concentration with each sample was determined using a chromogenic, quantitative Limulus-amoebocyte-lysate assay . Statistical analysis was ANOVA and Scheffe-test . RESULTS: Mean LPS yield from the untreated control samples was 50.1 (+/- 35.9) IU/ml . Following laser irradiation the average LPS on the root surfaces was 19.86 (+/- 14.4) IU/ml at 60 mJ, 12.86 (+/- 8.1) IU/ml at 100 mJ and 8.58 (+/- 4.9) IU/ml at 140 mJ.

Chem Commun (Camb), 2003 Apr 21, (8), 960 - 1
Highly enantioselective hydrolysis of alicyclic meso-epoxides with a bacterial epoxide hydrolase from Sphingomonas sp . HXN-200: simple syntheses of alicyclic vicinal trans-diols; Chang D et al.; Hydrolysis of N-benzyloxycarbonyl-3,4-epoxy-pyrrolidine and cyclohexene oxide with the epoxide hydrolase of Sphingomonas sp . HXN-200, respectively, gave the corresponding vicinal trans-diols in high ee and yield, representing the first example of enantioselective hydrolysis of a meso-epoxide with a bacterial epoxide hydrolase.

Folia Microbiol (Praha), 2003, 48(1), 5 - 16
Molecular features of the cytolytic pore-forming bacterial protein toxins; Alouf JE; The repertoire of the cytolytic pore-forming protein toxins (PFT) comprises 81 identified members . The essential feature of these cytolysins is their capacity to provoke the formation of hydrophilic pores in the cytoplasmic membranes of target eukaryotic cells . This process results from the binding of the proteins on the cell surface, followed by their oligomerization which leads to the insertion of the oligomers into the membrane and formation of protein-lined channels . It impairs the osmotic balance of the cell and causes cytolysis . In this review the molecular aspects of a number of important PFT and their respective encoding structural genes will be briefly described.

Genome Res, 2003 Jun, 13(6A), 1123 - 32 Epub 2003 May 12.
An appraisal of the potential for illegitimate recombination in bacterial genomes and its consequences: from duplications to genome reduction; Rocha EP; An exhaustive search for shortly spaced repeats in 74 bacterial chromosomes reveals that they are much more numerous than is usually acknowledged . These repeats were divided into five classes: close repeats (CRs), tandem repeats (TRs), simple sequence repeats (SSRs), spaced interspersed direct repeats, and "others." CRs are widespread and constitute the most abundant class, particularly in coding sequences . The other classes are less frequent, but each individual element shows a higher potential for recombination, when the number of repeats and their distances are taken into account . SSRs and TRs are more frequent in pathogens, as expected given their role in contingency loci, but are also widespread in the other bacteria . The analysis of CRs shows that they have an important role in the evolution of genomes, namely by generating duplications and deletions . Several cases compatible with a significant role of small CRs in the formation of large repeats were detected . Also, gene deletion in Buchnera correlates with repeat density, suggesting that CRs may lead to sequence deletion in general and genome reductive evolution of obligatory intracellular bacteria in particular . The assembly of these results indicates that shortly spaced repeats are key players in the dynamics of genome evolution.

Mol Biochem Parasitol, 2003 May, 128(2), 123 - 33
Downregulation of Trypanosoma brucei VSG expression site promoters on circular bacterial artificial chromosomes; te Vruchte D et al.; Trypanosoma brucei has about 20 telomeric variant surface glycoprotein (VSG) gene expression sites (ESs), which are downregulated in the insect form . We investigated the transcriptional behaviour of ES promoters on bacterial artificial chromosomes (BACs) containing two different ESs and their flanking regions on fragments of about 140kb . Four different BACs containing either the 221 or the VO2 ES were introduced into insect form T . brucei . The BACs replicated as circular episomes as shown using pulsed field gel (PFG) analysis of DNA exposed to increasing doses of gamma radiation, and digestion with Dam methylation-sensitive restriction enzymes . BAC copy number per cell varied from about 3 for the 221 ES BACs to about 15 for the VO2 ES BACs . Increasing drug selection pressure on the VO2 BAC T . brucei transformants resulted in amplification to about 80 BACs per cell . Although BACs were maintained in the absence of drug selection for at least 56 days, copy number fell and there was no evidence for centromere activity . ES promoters on small plasmid episomes introduced into insect form T . brucei in transient transfections are derepressed . In contrast, ES promoters on large BAC episomes are downregulated both on the original ES BACs, and on ES BACs selected for a drug marker driven by a rDNA promoter fused to the BAC vector . This indicates that downregulation of ES promoters in insect form T . brucei is influenced by genomic context, but does not necessitate proximity to a chromosome end.

Dig Dis Sci, 2003 Apr, 48(4), 775 - 82
Time course of colonic nuclear factor-kappa B expression during bacterial peptidoglycan-polysaccharide-induced colitis in rats; Fitzpatrick LR et al.; Nuclear factor-kappa B has been proposed to play a role in the pathogenesis of bacterial peptidoglycan-polysaccharide-induced colitis . However, its colonic expression has not been defined in detail . The primary aim of this study was to profile this expression in the rat colon . Peptigoglycan-polysaccharide was administered to the rat colon by direct intramural injections . Gross colonic injury was determined at various time points . Concomitantly, colonic nuclear factor-kappa B was measured by an electrophoretic mobility shift assay and by immunohistochemistry . Gross colonic injury and colonic nuclear factor-kappa B expression showed similar time courses following peptigoglycan-polysaccharide administration . Peak colonic injury and nuclear factor-kappa B expression were found on day 21 . Nuclear factor-kappa B was mainly expressed in submucosal inflammatory cells . In conclusion, the administration of bacterial peptidoglycan-polysaccharide to the rat colon caused a chronic colitis, which was characterized by up-regulated colonic nuclear factor-kappa B.

Microb Ecol, 2003 Jul, 46(1), 62 - 72 Epub 2003 May 13.
Benthic bacterial production and protozoan predation in a silty freshwater environment; Wieltschnig C et al.; The interrelation of heterotrophic bacteria with bacterivorous protists has been widely studied in pelagic environments, but data on benthic habitats, especially in freshwater systems, are still scarce . We present a seasonal study focusing on bacterivory by heterotrophic nanoflagellates (HNF) and ciliates in the silty sediment of a temperate macrophyte-dominated oxbow lake . From January 2001 to February 2002 we monitored the standing stock of bacteria and protozoa, bacterial secondary production (BSP, (3)H-thymidine, and (14)C-leucine incorporation), and grazing rates of HNF and ciliates on bacteria (FLB uptake) in the oxic sediment of the investigated system . BSP ranged from 470 to 4050 micro g C L(-1) wet sediment h(-1) . The bacterial compartment turned out to be highly dynamic, indicated by population doubling times (0.6-10.0 d), which were comparable to those in the water column of the investigated system . Yet, the control mechanisms acting upon the bacterial population led to a relative constancy of bacterial standing stock during a year . Ingestion rates of protozoan grazers were 0-20.0 bacteria HNF(-1) h(-1) and 0-97.6 bacteria ciliate(-1) h(-1) . HNF and ciliates together cropped 0-14 (mean 4)% of BSP, indicating that they did not significantly contribute to benthic bacterial mortality during any period of the year . The low impact of protozoan grazing was due to the low numbers of HNF and ciliates in relation to bacteria (1.8-3.5 x 10(4) bacteria HNF(-1), 0.9-3.1 x 10(6) bacteria ciliate(-1)) . Thus, grazing by HNF and ciliates could be ruled out as a parameter regulating bacterial standing stock or production in the sediment of the investigated system, but the factors responsible for the limitation of benthic protistan densities and the fate of benthic BSP remained unclear.

Microb Ecol, 2003 Jul, 46(1), 73 - 82 Epub 2003 May 13.
Simultaneous measurements of organic carbon mineralization and bacterial production in oxic and anoxic lake sediments; Bastviken D et al.; Based on work in marine sediments it can be hypothesized that (i) overall OM mineralization depends on the enzymatic capacity and is largely independent from the energy yield, (ii) similar oxic and anoxic rates are expected for fresh OM, while oxic rates should be faster for old OM that is partially degraded or adsorbed to particles, and (iii) that the thermodynamic energy yield does not regulate mineralization, but primarily determines the energy fraction allocated to bacterial production (BP) . We addressed these hypotheses by simultaneous measurements of mineralization rates (MR) and BP in sediments from a eutrophic lake, along with MR measurements in sediments of a dystrophic lake . Anoxic MR were 44 and 78% of oxic MR in the eutrophic and dystrophic lake, respectively, which was always higher than expected given the theoretical energy yields . The BP:MR ratio was 0.94 and 0.24 in the oxic and anoxic treatments, respectively, in accordance with the expected energy yields . Thus, the results support all three hypotheses above . We also critically discuss BP measurements in sediments and suggest that bacterial growth efficiency values from simultaneous MR and BP measurements can be used to evaluate the reliability of BP estimates.

FASEB J . 2003 May 8; {Epub ahead of print}
Bacterial DNA evokes epithelial IL-8 production by a MAPK-dependent, NFkappaB-independent pathway; Akhtar M et al.; Recognition of bacterial products by the innate immune system is dependent on pattern-recognition receptors: toll-like receptor 9 (TLR-9) in the case of bacterial DNA . We hypothesized that bacterial DNA can directly affect enteric epithelial cells . RT-PCR revealed constitutive TLR-9 mRNA expression in three human colonic epithelial cell lines (T84, HT-29, Caco-2) and THP-1 monocytes . Epithelial cells, in six-well culture plates or on filter supports, were exposed to E . coli DNA (1-50 &mgr;g/ml), synthetic CpG-rich oligonucleotides, or calf thymus DNA for 6-48 h . Exposure to E . coli DNA resulted in an increase in IL-8 mRNA, and a time- and dose-dependent increase in IL-8 secretion . Also, CpG oligonucleotides induced epithelial IL-8 production, whereas calf thymus DNA did not . Exposure to E . coli DNA resulted in phosphorylation of ERK 1/2 MAPK and inhibitors of ERK activity (PD98059, UO126) significantly reduced the evoked IL-8 production . In contrast, inhibitors of NFkappaB activity (PDTC, SN50) did not block E . coli DNA-induced IL-8 production . Electrophoretic mobility shift assays revealed that E . coli DNA stimulated epithelial AP-1 but not NFkappaB activation . The barrier (i.e., transepithelial resistance) and ion transport parameters of epithelial monolayers (assessed in Ussing chambers) were unaltered following E . coli DNA exposure . Thus model gut epithelia express TLR-9 mRNA and, while maintaining their barrier function, can respond to E . coli DNA by increased IL-8 production.

Clin Diagn Lab Immunol, 2003 May, 10(3), 352 - 6
Respiratory disease in kennelled dogs: serological responses to Bordetella bronchiseptica lipopolysaccharide do not correlate with bacterial isolation or clinical respiratory symptoms; Chalker VJ et al.; The role of Bordetella bronchiseptica in a natural outbreak of canine infectious respiratory disease was investigated both by culture and serological analysis . B . bronchiseptica was found in the lungs of a large proportion of clinically healthy dogs and in a greater proportion of dogs with respiratory disease . Using a lipopolysaccharide (LPS) antigen-based enzyme-linked immunosorbent assay, we analyzed the serological responses of a large number of dogs . Dogs with high antibody levels showed no protection from disease, and there was no correlation between the development of disease and rising antibody titer . Similarly, there was no difference in antibody levels in dogs with and without B . bronchiseptica in the lungs . Antibodies to LPS have no predictive value in determining which animals will contract respiratory disease, how severe the disease will be, or which dogs will have B . bronchiseptica colonizing the lungs.

Obstet Gynecol, 2003 May, 101(5 Pt 1), 862 - 8
Bacterial vaginosis, vaginal fluid neutrophil defensins, and preterm birth; Balu RB et al.; OBJECTIVE: To examine the association between bacterial vaginosis, vaginal fluid neutrophil defensins, and preterm birth . METHODS: Vaginal fluid specimens were obtained at 24-29 weeks' gestation from 242 cases with preterm birth and 507 noncases sampled using a case-cohort study design . We tested for bacterial vaginosis by Gram staining and Nugent scores and assayed for neutrophil defensins by enzyme-linked immunosorbent assay . Bacterial vaginosis was studied as a categoric variable (negative, intermediate, and positive), whereas defensins were studied as a continuous, categoric (based on percentiles), and dichotomous measure (presence versus absence) . Three gestational age cut points were used to define preterm birth . Modified Cox proportional hazard models were used to evaluate the associations between bacterial vaginosis, defensins, and degree (less than 32, less than 34, and less than 37 weeks) and type (premature rupture of membranes, preterm labor) of preterm birth . RESULTS: Elevated vaginal fluid neutrophil defensins were not associated with birth before 37 weeks . Compared with women who did not have measurable vaginal fluid defensins, women with higher defensin levels (0-2.8 micro g/mL, 2.8-8.2 micro g/mL, and greater than 8.2 micro g/mL) had a greater risk of delivering before 32 weeks . Hazard ratios adjusted for maternal race and vaginal bleeding during pregnancy and 95% confidence intervals for these defensin levels were 1.7 (0.4, 6.9), 2.4 (0.7, 7.9), and 3.1 (1.0, 9.8), respectively . Bacterial vaginosis status did not influence the association between defensins and preterm birth . CONCLUSION: Elevated concentrations of vaginal fluid neutrophil defensins at 24-29 weeks' gestation might predict preterm birth before 32 weeks.

Microbes Infect, 2003 Apr, 5(5), 405 - 11
Vitamin A deficiency leads to severe functional disturbance of the intestinal epithelium enzymes associated with diarrhoea and increased bacterial translocation in gnotobiotic rats; Kozakova H et al.; A disturbance of the integrity of the intestinal epithelium with an increased risk for bacterial translocation is one of the suggested factors underlying the increased incidence of infections and septicaemia during vitamin A deficiency . In the present study the effects of vitamin A deficiency on the enzymic activity of enterocytes in response to bacterial colonization with a non-pathogenic Escherichia coli strain were studied in monocolonized and conventional Wistar rats . The monocolonized, but not the conventional, vitamin A-deficient rats had markedly reduced weight compared to their pair-fed controls and presented neurological symptoms, such as hind leg weakness, tremor and slow gait . Moreover, only in the monocolonized vitamin A-deficient rats were severe diarrhoea and bacterial translocation to extraintestinal sites-mainly kidneys-detected . Measurements of enterocyte brush-border enzyme activities revealed that lactase, sucrase, gamma-glutamyltranspeptidase (GGT) and dipeptidyl peptidase IV (DPP IV) were significantly reduced in the monocolonized vitamin A-deficient rats compared to the pair-fed controls, indicating a severe functional disturbance of the enterocytes . In conventional vitamin A-deficient rats only sucrase activity was markedly lower than in the respective controls . Our observation, that the deficient vitamin A status led to a strong reduction of enterocyte enzymic activities, associated with diarrhoea and increased bacterial translocation, mainly in the gnotobiotic rats, suggests that the composition of the bacterial flora, i.e . the colonization state, has a strong influence on triggering the severity of the functional disturbances of the intestinal epithelium, and adds to the clinical manifestations of vitamin A deficiency.

Infez Med, 1999, 7(3), 192 - 194
Bacterial esophagitis in patients with HIV disease; Mastroianni A et al.; The authors carried out a retrospective study by reviewing all patients with HIV disease presenting esophageal symptoms who were evaluated by upper endoscopy . Three cases of bacterial esophagitis are reported and discussed according to literature data.

J Biol Chem, 2003 Jul 11, 278(28), 25867 - 71 Epub 2003 May 07.
Binding of the chemotaxis response regulator CheY to the isolated, intact switch complex of the bacterial flagellar motor: lack of cooperativity; Sagi Y et al.; In bacteria, the chemotactic signal is greatly amplified between the chemotaxis receptors and the flagellar motor . In Escherichia coli, part of this amplification occurs at the flagellar switch . However, it is not known whether the amplification results from cooperativity of CheY binding to the switch or from a post-binding step . To address this question, we purified the intact switch complex (constituting the switch proteins FliG, FliM, and FliN and the scaffolding protein FliF) in quantities sufficient for biochemical work and used it to investigate whether the binding of CheY to the switch complex is cooperative . As a negative control, we used complexes of switchless basal bodies, formed from the proteins FliF and FliG and similarly isolated . Using double-labeling centrifugation assays for binding, we found that CheY binds to the isolated, intact switch complex in a phosphorylation-dependent manner . We observed no significant phosphorylation-dependent binding to the negative control of the switchless basal body . The dissociation constant for the binding between the switch complex and phosphorylated CheY (CheY approximately P) was 4.0 +/- 1.1 microm, well in line with the published range of CheY approximately P concentrations to which the flagellar motor is responsive . Furthermore, the binding was not cooperative (Hill coefficient approximately 1) . This lack of CheY approximately P-switch complex binding cooperativity, taken together with earlier in vivo studies suggesting that the dependence of the rotational state of the motor on the fraction of occupied sites at the switch is sigmoidal and very steep (Bren, A., and Eisenbach, M . (2001) J . Mol . Biol . 312, 699-709), indicates that the chemotactic signal is amplified within the switch, subsequent to the CheY approximately P binding.

Biol Proced Online, 1998 May 14, 1, 17 - 26
Methods for the Measurement of a Bacterial Enzyme Activity in Cell Lysates and Extracts; Burns B et al.; The kinetic characteristics and regulation of aspartate carbamoyltransferase activity were studied in lysates and cell extracts of Helicobacter pylori by three diffirent methods . Nuclear magnetic resonance spectroscopy, radioactive tracer analysis, and spectrophotometry were employed in conjunction to identify the properties of the enzyme activity and to validate the results obtained with each assay . NMR spectroscopy was the most direct method to provide proof of ACTase activity; radioactive tracer analysis was the most sensitive technique and a microtitre-based colorimetric assay was the most cost-and time-efficient for large scale analyses . Freeze-thawing was adopted as the preferred method for cell lysis in studying enzyme activity in situ . This study showed the benefits of employing several different complementary methods to investigate bacterial enzyme activity.

J Immunol, 2003 May 15, 170(10), 5165 - 75
The Toll-like receptor 5 stimulus bacterial flagellin induces maturation and chemokine production in human dendritic cells; Means TK et al.; Toll-like receptors (TLRs) are pattern recognition receptors that serve an important function in detecting pathogens and initiating inflammatory responses . Upon encounter with foreign Ag, dendritic cells (DCs) go through a maturation process characterized by an increase in surface expression of MHC class II and costimulatory molecules, which leads to initiation of an effective immune response in naive T cells . The innate immune response to bacterial flagellin is mediated by TLR5, which is expressed on human DCs . Therefore, we sought to investigate whether flagellin could induce DC maturation . Immature DCs were cultured in the absence or presence of flagellin and monitored for expression of cell surface maturation markers . Stimulation with flagellin induced increased surface expression of CD83, CD80, CD86, MHC class II, and the lymph node-homing chemokine receptor CCR7 . Flagellin stimulated the expression of chemokines active on neutrophils (IL-8/CXC chemokine ligand (CXCL)8, GRO-alpha/CXCL1, GRO-beta/CXCL2, GRO-gamma/CXCL3), monocytes (monocyte chemoattractant protein-1/CC chemokine ligand (CCL)2), and immature DCs (macrophage-inflammatory protein-1 alpha/CCL3, macrophage-inflammatory protein-1 beta/CCL4), but not chemokines active on effector T cells (IFN-inducible protein-10 kDa/CXCL10, monokine induced by IFN-gamma/CXCL9, IFN-inducible T cell alpha chemoattractant/CXCL11) . However, stimulating DCs with both flagellin and IFN-inducible protein-10 kDa, monokine induced by IFN-gamma, and IFN-inducible T cell alpha chemoattractant expression, whereas stimulation with IFN-beta or flagellin alone failed to induce these chemokines . In functional assays, flagellin-matured DCs displayed enhanced T cell stimulatory activity with a concomitant decrease in endocytic activity . Finally, DCs isolated from mouse spleens or bone marrows were shown to not express TLR5 and were not responsive to flagellin stimulation . These results demonstrate that flagellin can directly stimulate human but not murine DC maturation, providing an additional mechanism by which motile bacteria can initiate an acquired immune response.

J Microbiol Methods, 2003 Jul, 54(1), 75 - 9
Monitoring promoter activity in a single bacterial cell by using green and red fluorescent proteins; Hakkila K et al.; We investigated the possibility of monitoring promoter activity with flow cytometry by using green fluorescent protein (GFPmut2) and red fluorescent protein (drFP583) in a single bacterial cell . The drFP583 was used as an intrinsic marker of the bacterial cells, because it was expressed constantly in Escherichia coli MC1061 strain . The GFPmut2 expressed under the control of the Hg(2+) ion inducible mer promoter/operator, was used to study promoter activity . Over 75% of the cells were positive for red and green fluorescence in flow cytometric analysis . The average green fluorescence of the whole population increased from 6.7 to 1700 when the mercury concentration was increased from 0 to 1 x 10(-4) M, while the red fluorescence was unaffected by the mercury concentration . These results show that gfpmut2 and drFP583 could be expressed under different promoters in one bacterial cell and measured independently with a flow cytometer.

Curr Opin Microbiol, 2003 Apr, 6(2), 163 - 72
Regulation by proteolysis in bacterial cells; Jenal U et al.; Regulation by proteolysis plays a major role in bacterial stress responses, the cell cycle and development . Key regulators of these processes are subject to conditional proteolysis that depends on complex cellular information processing . This information includes temporal and spatial cues, and recent research has revealed a striking potential for multiple signal integration.

Curr Opin Microbiol, 2003 Apr, 6(2), 140 - 5
The bacterial universal stress protein: function and regulation; Kvint K et al.; The universal stress protein A (UspA) superfamily encompasses an ancient and conserved group of proteins that are found in bacteria, Archea, fungi, flies and plants . The Escherichia coli UspA is produced in response to a large number of different environmental onslaughts and UspA is one of the most abundant proteins in growth-arrested cells . Although insights into the regulation of the E . coli uspA gene have been gained, the exact roles of the Usp proteins and Usp domains remain enigmatic; they appear, in some cases, to be linked to resistance to DNA-damaging agents and to respiratory uncouplers.

Curr Opin Microbiol, 2003 Apr, 6(2), 135 - 9
Function of the universally conserved bacterial GTPases; Caldon CE et al.; The GTPase superfamily of cellular regulators is well represented in bacteria . A small number are universally conserved over the entire range of bacterial species . Such a pervasive taxonomic distribution suggests that these enzymes play important roles in bacterial cellular systems . Recent advances have demonstrated that bacterial GTPases are important regulators of ribosome function, and important for the distribution of DNA to daughter cells following cell division . In addition, the atomic structure of a unique GTPase, EngA, has recently been established . Unlike any other GTPase, EngA contains tandem GTP-binding domains . This structural study suggests that the GTPase cycles of the domains are regulated differentially in a manner that remains to be elucidated.

Curr Opin Microbiol, 2003 Apr, 6(2), 109 - 13
Eyeing bacterial genomes; Ochman H et al.; The density of information in a bacterial genome allows its history, organization and encoded functions to be distilled into a single graphical representation . These features have made it possible to discern the forces acting in and on bacterial genomes at levels not attainable in eukaryotes.

BMC Genomics . 2003 May 05;4(1):17.
Wavelet to predict bacterial ori and ter: a tendency towards a physical balance; Song J et al.; BACKGROUND: Chromosomal DNA replication in bacteria starts at the origin (ori) and the two replicores propagate in opposite directions up to the terminus (ter) region . We hypothesize that the two replicores need to reach ter at the same time to maintain a physical balance; DNA insertion would disrupt such a balance, requiring chromosomal rearrangements to restore the balance . To test this hypothesis, we needed to demonstrate that ori and ter are in a physical balance in bacterial chromosomes . Using wavelet analysis, we documented GC skew, AT skew, purine excess and keto excess on the published bacterial genomic sequences to locate the turning (minimum and maximum) points on the curves . Previously, the minimum point had been supposed to correlate with ori and the maximum to correlate with ter . RESULTS: We observed a strong tendency of the bacterial chromosomes towards a physical balance, with the minima and maxima corresponding to the known or putative ori and ter and being about half chromosome separated in most of the bacteria studied . A nonparametric method based on wavelet transformation was employed to perform significance tests for the predicted loci . CONCLUSIONS: The wavelet approach can reliably predict the ori and ter regions and the bacterial chromosomes have a strong tendency towards a physical balance between ori and ter.

Glia, 2003 Jun, 42(4), 433 - 46
Oncostatin M enhances the expression of prostaglandin E2 and cyclooxygenase-2 in astrocytes: synergy with interleukin-1beta, tumor necrosis factor-alpha, and bacterial lipopolysaccharide; Repovic P et al.; Oncostatin M (OSM), a cytokine of the interleukin-6 family, is expressed in rheumatoid arthritis, multiple sclerosis, multiple myeloma, and other inflammatory and neoplastic conditions . Prostaglandin E(2) (PGE(2)), an eicosanoid also associated with inflammation and cancer, has recently been shown to induce OSM expression . We report here that OSM in turn induces PGE(2) production by astrocytes and astroglioma cells . More importantly, in combination with the inflammatory mediators IL-1beta, tumor necrosis factor-alpha, and lipopolysaccharide, OSM exhibits a striking synergy, resulting in up to 50-fold higher PGE(2) production by astrocytes, astroglioma, and neuroblastoma cell lines . Enhanced PGE(2) production by OSM and IL-1beta treatment is explained by their effect on cyclooxygenase-2 (COX-2), an enzyme that catalyzes the committed step in PGE(2) synthesis . Of the enzymes involved in PGE(2) biosynthesis, only COX-2 mRNA and protein levels are synergistically amplified by OSM and IL-1beta . Nuclear run-on assays demonstrate that OSM and IL-1beta synergistically upregulate transcription of the COX-2 gene, and the mRNA stability assay indicates that COX-2 mRNA is posttranscriptionally stabilized by OSM and IL-1beta . To effect synergy on the PGE(2) level, OSM signals in part through its gp130/OSMRbeta receptor, since neutralizing antibodies against gp130 and OSMRbeta, but not LIFRbeta, decrease PGE(2) production in response to OSM plus IL-1beta . SB202190 and U0126, inhibitors of p38 MAPK and ERK1/2 activation, respectively, inhibit IL-1beta and OSM upregulation of COX-2 and PGE(2), indicating that these MAPK cascades are utilized by both stimuli . This mechanism of PGE(2) amplification may be active in brain pathologies where both OSM and IL-1beta are present, such as glioblastomas and multiple sclerosis .

Infez Med, 1998, 6(4), 225 - 227
{Neurologic signs in bacterial endocarditis: an unusual case report}; Tarquini P et al.; We describe the case of a 35-year-old woman, hospitalised due to a fever lasting two days, with signs and symptoms of cerebro-spinal meningitis . The subsequent occurrence of a thromboembolic event with stroke suggests that the neurological symptoms were secondary to a centre of infective endocarditis (instrumentally proved) in the absence of further outstanding symptoms of endocarditis . This event is very unusual during the course of endocardic disease.

Toxicol Sci, 2003 Jul, 74(1), 203 - 14 Epub 2003 May 02.
Endothelial cell injury and coagulation system activation during synergistic hepatotoxicity from monocrotaline and bacterial lipopolysaccharide coexposure; Yee SB et al.; A small, noninjurious dose of bacterial lipopolysaccharide (LPS; 7.4 x 106 EU/kg) administered 4 h after a small, nontoxic dose of monocrotaline (MCT; 100 mg/kg) produces synergistic hepatotoxicity in rats within 6 to 12 h after MCT exposure . The resulting centrilobular (CL) and midzonal (MZ) liver lesions are characterized by hepatic parenchymal cell (HPC) necrosis . Pronounced hemorrhage, disruption of sinusoidal architecture, and loss of central vein intima suggest that an additional component to injury may be the liver vasculature . In the present investigation, the hypothesis that sinusoidal endothelial cell (SEC) injury and coagulation system activation occur in this model was tested . Plasma hyaluronic acid (HA) concentration, a biomarker for SEC injury, was significantly increased in cotreated animals before the onset of HPC injury and remained elevated through the time of maximal HPC injury (i.e., 18 h) . SEC injury was confirmed by immunohistochemistry and electron microscopy . Pyrrolic metabolites were produced from MCT by SECs in vitro, which suggests that MCT may injure SECs directly through the formation of its toxic metabolite, monocrotaline pyrrole . Inasmuch as SEC activation and injury can promote hemostasis, activation of the coagulation system was evaluated . Coagulation system activation, as marked by a decrease in plasma fibrinogen, occurred before the onset of HPC injury . Furthermore, extensive fibrin deposition was observed immunohistochemically within CL and MZ regions after MCT/LPS cotreatment . Taken together, these results suggest that SEC injury and coagulation system activation are components of the synergistic liver injury resulting from MCT and LPS coexposure.

J Mol Biol, 2003 May 9, 328(4), 927 - 37
Structural and functional defects caused by point mutations in the alpha-crystallin domain of a bacterial alpha-heat shock protein; Lentze N et al.; The diverse family of alpha-crystallin-type small heat shock proteins (alpha-Hsps or sHsps) is characterised by a central, moderately conserved alpha-crystallin domain . Oligomerisation followed by dissociation of subparticles is thought to be a prerequisite for chaperone function . We demonstrate that HspH, a bacterial alpha-Hsp from the soybean-symbiont Bradyrhizobium japonicum, assembles into dynamic complexes freely exchanging subunits with homologous and heterologous complexes . The importance of the alpha-crystallin domain for oligomerisation and chaperone activity was tested by site-directed mutagenesis of 12 different residues . In contrast to mammalian alpha-Hsps, the majority of these mutations elicited severe structural and functional defects in HspH . The individual exchange of five amino acid residues throughout the alpha-crystallin domain was found to compromise oligomerisation to various degrees . Assembly defects resulting in complexes of reduced size correlated with greatly decreased or abolished chaperone activity, reinforcing that complete oligomerisation is required for functionality . Mutation of a highly conserved glycine (G114) at the C-terminal end of the alpha-crystallin domain specifically impaired chaperone activity without interfering with oligomerisation properties, indicating that this residue is critical for substrate interaction . The structural and functional importance of this and other residues is discussed in the context of a modeled three-dimensional structure of HspH.

Arch Gynecol Obstet, 2003 Oct, 268(4), 251 - 5 Epub 2003 May 01.
Bacterial endocarditis complicating pregnancy: case report and systematic review of the literature; Campuzano K et al.; INTRODUCTION . Infectious endocarditis is a rare life-threatening complication of pregnancy . We report a pregnancy complicated by a 3.5-cm infected vegetation of the tricuspid valve initially presenting as unilateral hip pain as well as systematic review of this entity . SOURCES . A MEDLINE review of the English language literature from 1965 to present using the search terms 'endocarditis', 'pregnancy' and 'infection' as well as review of references was performed . RESULTS . Sixty-eight cases of infectious endocarditis complicating pregnancy were identified . The calculated maternal and fetal mortality rates were 22.1% and 14.7% respectively . CONCLUSIONS . With persistent symptomatic lesions, delivery should be considered without regard to measures of fetal lung maturity because of high fetal mortality rates.

Nat Rev Mol Cell Biol, 2003 May, 4(5), 385 - 96
Phagocyte sabotage: disruption of macrophage signalling by bacterial pathogens; Rosenberger CM et al.; Macrophages function at the front line of immune defences against incoming pathogens . But the ability of macrophages to internalize bacteria, migrate, recruit other immune cells to the site of infection and influence the nature of the immune response can provide unintended benefits for bacterial pathogens that are able to subvert or co-opt these normally effective defences . This review highlights recent advances in our understanding of the many interference strategies that are used by bacterial pathogens to undermine macrophage signalling.

Infez Med, 1999, 7(1), 62 - 63
4th Congress Bacterial Infections and Therapeutic Perspectives (IBAT) 1999 in Naples; Esposito S; Not available

Biochem Biophys Res Commun, 2003 May 16, 304(4), 795 - 800
Induction of neurite outgrowth in PC12 cells by the bacterial nucleoside N6-methyldeoxyadenosine is mediated through adenosine A2a receptors and via cAMP and MAPK signaling pathways; Charles MP et al.; We have previously shown that N(6)-methyldeoxyadenosine (MDA) is an inducer of differentiation in several tumor cells . Here we show that in addition to its ability to induce neurite-outgrowth in PC12 cells, MDA also significantly enhances the nerve-growth factor-mediated neurite outgrowth of these cells . Thus, MDA acts synergistically with NGF to repress cdc2 and cdk2 synthesis and to enhance tyrosine hydroxylase synthesis . To further elucidate the mechanisms of action of MDA, we investigated the effect of this drug on various signaling pathways . The neuritogenesis observed in PC12 following MDA treatment is mediated through activation of adenylyl cyclase in a PKA independent process and through the recruitment of the p44/p42 MAPK pathway . Furthermore, the adenosine A(2a) receptor antagonist ZM 241385 prevents the MDA-induced neuritogenesis, suggesting that MDA mediates its effect via this adenylyl cyclase-coupled A(2a) receptor . Collectively, these findings suggest that, in PC12 cells, the MDA-induced neuritogenesis requires the recruitment of adenosine A(2a) receptor, the stimulation of adenylate cyclase, and the activation of the p44/42MAP kinase cascade.

Plasmid, 2003 Mar, 49(2), 184 - 7
Development of a novel bacterial artificial chromosome cloning system for functional studies; Al-Hasani K et al.; Bacterial artificial chromosome (BAC) cloning systems currently in use generate high quality genomic libraries for gene mapping, identification, and sequencing . However, the most commonly used BAC cloning systems do not facilitate functional studies in eukaryotic cells . To overcome this limitation, we have developed pEBAC190G, a new BAC vector that combines the features of the first generation PAC/BAC vectors with eukaryotic elements that facilitate the transfection, episomal maintenance, and functional analysis of large genomic fragments in eukaryotic cells . A number of different cloning strategies may be used to retrofit genomic fragments from existing libraries into the new vector . The system was tested by the retrofitting of a 170kb NotI genomic fragment from the RPCI-11 BAC library into the NotI site of pEBAC190G . Clones from any eukaryotic genomic library harboured in this vector can be transferred from bacteria directly to eukaryotic cells for functional analysis.

Virology, 2003 Apr 25, 309(1), 108 - 13
Only one pRNA hexamer but multiple copies of the DNA-packaging protein gp16 are needed for the motor to package bacterial virus phi29 genomic DNA; Shu D et al.; A common feature in the maturation of linear dsDNA viruses is that the lengthy viral genome is translocated with remarkable velocity into a limited space within a preformed protein shell using ATP as motor energy . Most biomotors, such as myosin, kinesin, DNA-helicase, and RNA polymerase, contain one ATP-binding component that acts processively . An examination of the well-studied dsDNA viruses reveals that DNA packaging motors involve two nonstructural components . Which component of the motor is the integrated processive factor to turn the motor has not been identified . In bacterial virus phi 29, these two components consist of a gp16 protein and an RNA molecule called pRNA . We have previously predicted and recently confirmed that gp16 binds ATP . It is generally believed that gp16 serves as an ATP-binding and processive component to drive the motor . In this article, phi 29 DNA-packaging intermediates were purified in quantity and examined to differentiate the role between gp16 and pRNA . It was found that the pRNA hexamer is an integral motor component, while gp16 is not stably bound . Only one pRNA hexamer, but multiple copies of gp16, were needed to accomplish DNA packaging . pRNA functions continuously during the entire DNA translocation process, suggesting that pRNA is a vital part of the DNA packaging motor.

Biochem J, 2003 Aug 1, 373(Pt 3), 949 - 55
On the substrate specificity of bacterial DD-peptidases: evidence from two series of peptidoglycan-mimetic peptides; Anderson JW et al.; The reactions between bacterial DD-peptidases and beta-lactam antibiotics have been studied for many years . Less well understood are the interactions between these enzymes and their natural substrates, presumably the peptide moieties of peptidoglycan . In general, remarkably little activity has previously been demonstrated in vitro against potential peptide substrates, although in many cases the peptides employed were non-specific and not homologous with the relevant peptidoglycan . In this paper, the specificity of a panel of DD-peptidases against elements of species-specific D-alanyl-D-alanine peptides has been assessed . In two cases, those of soluble, low-molecular-mass DD-peptidases, high activity against the relevant peptides has been demonstrated . In these cases, the high specificity is towards the free N-terminus of the peptidoglycan fragment . With a number of other enzymes, particularly high-molecular-mass DD-peptidases, little or no activity against these peptides was observed . In separate experiments, the reactivity of the enzymes against the central, largely invariant, peptide stem was examined . None of the enzymes surveyed showed high activity against this structural element although weak specificity in the expected direction towards the one structural variable (D-gammaGln versus D-gammaGlu) was observed . The current state of understanding of the activity of these enzymes in vitro is discussed.

Biofizika, 2003 Mar-Apr, 48(2), 273 - 80
{Kinetic analysis of behavior of chemotactic bacterial population at the interface of two media}; Zaval'skii LIu; The behavior of a population of bacteria at the boundary with a chemotactically active water medium containing the sites of specific binding to cell receptors was analyzed . A kinetic model of chemotaxis was used for the analysis . Differences in the behavior of strains metabolizing and not metabolizing the substrate were revealed . Six phases of interface taxis were distinguished and characterized . The results of the analysis were confirmed by the densitometric data.

Genetika, 2003 Mar, 39(3), 376 - 82
{Expression of a thermostable bacterial cellulase in transgenic tobacco plants}; Abdeev RM et al.; The bacterial gene of the thermostable endo-beta-1,4-glucanase (cellulase) was shown to retain its activity and substrate specificity when expressed in transgenic tobacco plants . The leader peptide of the carrot extensin was efficient in transferring the bacterial enzyme into the apoplast . The expression of the bacterial cellulase gene leads to changes in the plant tissue morphology . In the transgenic plant lines, regeneration of primary shoots from callus occurred at the three to five times higher cytokinin (6-BAP) concentration than in control plants . The transgenic plants that expressed the bacterial gene exhibited increased business and altered leaf shape . The transgenic plants developed can be used as models for studying the cellulases role and function in plants.

Proc Natl Acad Sci U S A, 2003 May 13, 100(10), 6115 - 20 Epub 2003 Apr 29.
Folding quality control in the export of proteins by the bacterial twin-arginine translocation pathway; DeLisa MP et al.; To examine the relationship between folding and export competence by the twin-arginine translocation (Tat) pathway we analyzed the subcellular localization of fusions between a set of eight putative Tat leader peptides and alkaline phosphatase in isogenic Escherichia coli strains that either allow or disfavor the formation of protein disulfide bonds in the cytoplasm . We show that export by the Tat translocator is observed only in strains that enable oxidative protein folding in the cytoplasm . Further, we show that other disulfide-containing proteins, namely single-chain Fv and heterodimeric F(AB) antibody fragments, are export-competent only in strains having an oxidizing cytoplasm . Functional, heterodimeric F(AB) protein was exported from the cytoplasm by means of a Tat leader peptide fused to the heavy chain alone, indicating that the formation of a disulfide-bonded dimer preceeds export . These results demonstrate that in vivo only proteins that have attained the native conformation are exported by the Tat translocator, indicating that a folding quality-control mechanism is intrinsic to the export process . The ability to export proteins with disulfide bonds and the folding proofing feature of the Tat pathway are of interest for biotechnology applications.

J Ind Microbiol Biotechnol, 2003 Apr, 30(4), 239 - 44 Epub 2003 Apr 26.
Detection of bacterial contamination in starch and resin-based papermaking chemicals using fluorescence techniques; Nohynek L et al.; Rapid fluorescence techniques were evaluated for the detection of bacterial contaminants in papermaking chemicals including starch and the resin-based sizes and starch slurries used in the paper industry . Viable and non-viable bacterial cells were visualised by fluorescent probes and detected by epifluorescence microscopy and flow cytometry . The best discrimination ability was obtained with the fluorescent probes LIVE/DEAD and SYBR Green, based on the staining of cellular nucleic acid, and ChemChrome V3, which demonstrated cellular enzymatic activity . The process samples had to be diluted and filtered before fluorescence staining and analysis because they were viscous and contained solid particles . Fluorescence microscopic counts of bacteria in highly contaminated process samples were similar to plate counts, but flow cytometric enumeration of bacterial cells in process samples yielded 2- to 10-fold lower counts compared with plate counts, depending on the consistency of the sample . The detection limits in flow cytometric analysis and in epifluorescence microscopy were 10(3)-10(6) cells ml(-1) and 10(5)-10(6) cells ml(-1), respectively . Intrinsic bacterial contamination was detectable with fluorescence techniques and highly contaminated process samples could be analysed with fluorescence methods.

Biophys J, 2003 May, 84(5), 2943 - 56
Perfect and near-perfect adaptation in a model of bacterial chemotaxis; Mello BA et al.; The signaling apparatus mediating bacterial chemotaxis can adapt to a wide range of persistent external stimuli . In many cases, the bacterial activity returns to its prestimulus level exactly, and this perfect adaptability is robust against variations in various chemotaxis protein concentrations . We model the bacterial chemotaxis signaling pathway, from ligand binding to CheY phosphorylation . By solving the steady-state equations of the model analytically, we derive a full set of conditions for the system to achieve perfect adaptation . The conditions related to the phosphorylation part of the pathway are discovered for the first time, while other conditions are generalizations of the ones found in previous works . Sensitivity of the perfect adaptation is evaluated by perturbing these conditions . We find that, even in the absence of some of the perfect adaptation conditions, adaptation can be achieved with near-perfect precision as a result of the separation of scales in both chemotaxis protein concentrations and reaction rates, or specific properties of the receptor distribution in different methylation states . Since near-perfect adaptation can be found in much larger regions of the parameter space than that defined by the perfect adaptation conditions, their existence is essential to understand robustness in bacterial chemotaxis.

Genesis, 2003 Apr, 35(4), 220 - 6
Expression of Nkx2-5-GFP bacterial artificial chromosome transgenic mice closely resembles endogenous Nkx2-5 gene activity; Chi X et al.; Mouse Nkx2-5 gene is essential for early heart development and it is regulated by a complex array of regulatory modules . In order to establish an efficient in vivo system for mapping the Nkx2-5 genomic locus for regulatory regions, we developed improved homologous recombination technology for use in Escherichia coli and then knocked an IRES-hrGFP reporter gene into Nkx2-5 gene in a 120 kb Nkx2-5 bacterial artificial chromosome (BAC) clone . We employed the recombination genes redalpha and redbeta under the pBAD promoter, which was specifically induced by the addition of L-arabinose . Recombination was selected for by our universal targeting cassette which conferred kanamycin resistance in bacterial cells and neomycin resistance in mammalian cells . Transgenic mouse lines generated from this modified BAC clone closely resembled the endogenous Nkx2-5 expression in the heart, pylorus sphincter, and spleen, but expression was not detected in the tongue . Nkx2-5 BAC-GFP expression was copy number-dependent and locus site-independent . BAC transgenics using the GFP reporter offers an efficient model system to study gene expression and regulation .

Hepatology, 2003 May, 37(5), 1043 - 55
Toll-like receptor 4 mediates inflammatory signaling by bacterial lipopolysaccharide in human hepatic stellate cells; Paik YH et al.; Bacterial lipopolysaccharide (LPS) stimulates Kupffer cells and participates in the pathogenesis of alcohol-induced liver injury . However, it is unknown whether LPS directly affects hepatic stellate cells (HSCs), the main fibrogenic cell type in the injured liver . This study characterizes LPS-induced signal transduction and proinflammatory gene expression in activated human HSCs . Culture-activated HSCs and HSCs isolated from patients with hepatitis C virus-induced cirrhosis express LPS-associated signaling molecules, including CD14, toll-like receptor (TLR) 4, and MD2 . Stimulation of culture-activated HSCs with LPS results in a rapid and marked activation of NF-kappaB, as assessed by in vitro kinase assays for IkappaB kinase (IKK), IkappaBalpha steady-state levels, p65 nuclear translocation, NF-kappaB-dependent luciferase reporter gene assays, and electrophoretic mobility shift assays . Lipid A induces NF-kappaB activation in a similar manner . Both LPS- and lipid A-induced NF-kappaB activation is blocked by preincubation with either anti-TLR4 blocking antibody (HTA125) or Polymyxin B . Lipid A induces NF-kappaB activation in HSCs from TLR4-sufficient (C3H/OuJ) mice but not from TLR4-deficient (C3H/HeJ) mice . LPS also activates c-Jun N-terminal kinase (JNK), as assessed by in vitro kinase assays . LPS up-regulates IL-8 and MCP-1 gene expression and secretion . LPS-induced IL-8 secretion is completely inhibited by the IkappaB super repressor (Ad5IkappaB) and partially inhibited by a specific JNK inhibitor, SP600125 . LPS also up-regulates cell surface expression of ICAM-1 and VCAM-1 . In conclusion, human activated HSCs utilize components of TLR4 signal transduction cascade to stimulate NF-kappaB and JNK and up-regulate chemokines and adhesion molecules . Thus, HSCs are a potential mediator of LPS-induced liver injury.

Prof Nurse, 2003 Apr, 18(8), 437 - 8
Bacterial STIs; Jones M; The three most common bacterial sexually transmitted infections--chlamydia, gonorrhoea and syphilis--and are now more common than viral STIs in the UK.

Wiad Lek, 2002, 55(11-12), 699 - 705
{Bacterial flora as a potential etiopathogenic factor for nasal polyps}; Rostkowska-Nadolska B et al.; Nasal polyps develop in patients with disturbed local homeostasis of nasal mucosa . Research of Bernstein and Stoop showed the influence of inflammation of nasal mucosa on growth of nasal polyps . Bacterial and viral infections play significant role in development and intensification of inflammatory and immunological process . Many bacteria produce factors which damage cilia cells, cause dyskinesis of cilia and destroy respiratory epithelium . Disturbed muco-ciliar cleaning permits bacteria to penetrate through mucous layer and facilitate their adhesion and colonization on the cells of respiratory epithelium . Adhesion and successive colonization with bacteria multiplication can lead to inflammatory process . Nose swabs results were different in each examined group . The aim of this work was to define bacterial flora in patients with nasal polyps and in patients after polypectomia and to compare it to bacterial flora in healthy patients . Nose swabs were performed in 51 patients with nasal polyps before polypectomia and additional in 15 patients after polypectomia . 25 healthy medical students were a control group . The patients were divided into two groups: I . patients with nasal eosinophilic polyps, II . patients with nasal neutrophilic polyps . In the group of patients with both neutrophilic and eosinophilic polyps dominating bacterial flora included bacteria which usually are the cause of inflammation . In the group of patients after polypectomia bacterial flora was not different from that which was found in patients with polyps.

Mol Genet Genomics, 2003 Apr, 269(1), 126 - 36 Epub 2003 Mar 01.
A bacterial artificial chromosome (BAC) library of sugar beet and a physical map of the region encompassing the bolting gene B; Hohmann U et al.; In sugar beet (Beta vulgaris L.), early bolting is caused by a single dominant gene, designated B . Twenty AFLP markers selected from a 7.8-cM segment of the B region on chromosome 2 were used to screen a YAC library, and a first-generation physical map including the B gene, made up of 11 YACs, was established . Because the genome coverage of the YAC library was low, a BAC library was constructed in the vector pBeloBAC11 . This library consists of 57,600 clones with an average insert size of 116 kb, corresponding to 8.8 genome equivalents . Screening of the BAC library with chloroplast and mitochondrial DNA probes indicated that less than 0.1% of the clones contained organelle-derived DNA . To fill the gaps in the physical map around the B gene, the BAC library was screened with four AFLP markers and 10 YAC-derived probes . In total, 54 different BACs were identified . Overlaps between BACs were detected by using BAC termini amplified by PCR as probes, and by RFLP fingerprinting . In this way, a minimal tiling path of the central 4.6-cM region was constructed, which consists of 14 BACs . The B locus was localized to a 360-kb contig, a size which makes positional cloning of the gene feasible.

J Biol Chem, 2003 Jul 4, 278(27), 24586 - 93 Epub 2003 Apr 24.
Translocation of phospholipids is facilitated by a subset of membrane-spanning proteins of the bacterial cytoplasmic membrane; Kol MA et al.; The mechanism by which phospholipids are transported across biogenic membranes, such as the bacterial cytoplasmic membrane, is unknown . We hypothesized that this process is mediated by the presence of the membrane-spanning segments of inner membrane proteins, rather than by dedicated flippases . In support of the hypothesis, it was demonstrated that transmembrane alpha-helical peptides, mimicking the membrane-spanning segments, mediate flop of 2-6-(7-nitro-2,1,3-benzoxadiazol-4-yl) aminocaproyl (C6-NBD)-phospholipids (Kol, M . A., de Kroon, A . I., Rijkers, D . T., Killian, J . A., and de Kruijff, B . (2001) Biochemistry 40, 10500-10506) . Here the dithionite reduction assay was used to measure transbilayer equilibration of C6-NBD-phospholipids in proteoliposomes, composed of Escherichia coli phospholipids and a subset of bacterial membrane proteins . It is shown that two well characterized integral proteins of the bacterial cytoplasmic membrane, leader peptidase and the potassium channel KcsA, induce phospholipid translocation, most likely by their transmembrane domains . In contrast, the ATP-binding cassette transporter from the E . coli inner membrane MsbA, a putative lipid flippase, did not mediate phospholipid translocation, irrespective of the presence of ATP . OmpT, an outer membrane protein from E . coli, did not facilitate flop either, demonstrating specificity of protein-mediated phospholipid translocation . The results are discussed in the light of phospholipid transport across the E . coli inner membrane.

Blood, 2003 Sep 1, 102(5), 1740 - 2 Epub 2003 Apr 24.
Bacterial lipopolysaccharide directly induces angiogenesis through TRAF6-mediated activation of NF-kappaB and c-Jun N-terminal kinase; Pollet I et al.; The intracellular pathways by which inflammatory mediators transmit their angiogenic signals is not well studied . The effects of a potent inflammatory mediator, bacterial lipopolysaccharide (LPS), are transmitted through Toll-like receptors (TLRs) . A major, although not exclusive, LPS/TLR intracellular signaling pathway is routed through TNF (tumor necrosis factor) receptor associated factor 6 (TRAF6) . In this report we demonstrate that LPS directly stimulates endothelial sprouting in vitro . By blocking TRAF6 activity using retroviral expression of a dominant-negative TRAF6 in endothelial cells, we show that TRAF6 is absolutely required for the LPS-initiated angiogenic response in vitro and in vivo . Inhibition of either c-Jun N-terminal kinase (JNK) activity or nuclear factor kappaB (NF-kappaB) activity, downstream of TRAF6, is sufficient to inhibit LPS-induced endothelial sprouting . In contrast, only inhibition of NF-kappaB, but not JNK, activity blocks basic fibroblast growth factor (bFGF)-induced angiogenesis . Our findings thus demonstrate a direct endothelial-stimulatory role of LPS in initiating angiogenesis through activation of TRAF6-dependent signaling pathways.

Biosens Bioelectron, 2003 Jul, 18(7), 917 - 23
Palladium-bacterial cellulose membranes for fuel cells; Evans BR et al.; Bacterial cellulose is a versatile renewable biomaterial that can be used as a hydrophilic matrix for the incorporation of metals into thin, flexible, thermally stable membranes . In contrast to plant cellulose, we found it catalyzed the deposition of metals within its structure to generate a finely divided homogeneous catalyst layer . Experimental data suggested that bacterial cellulose possessed reducing groups capable of initiating the precipitation of palladium, gold, and silver from aqueous solution . Since the bacterial cellulose contained water equivalent to at least 200 times the dry weight of the cellulose, it was dried to a thin membranous structure suitable for the construction of membrane electrode assemblies (MEAs) . Results of our study with palladium-cellulose showed that it was capable of catalyzing the generation of hydrogen when incubated with sodium dithionite and generated an electrical current from hydrogen in an MEA containing native cellulose as the polyelectrolyte membrane (PEM) . Advantages of using native and metallized bacterial cellulose membranes in an MEA over other PEMs such as Nafion 117 include its higher thermal stability to 130 degrees C and lower gas crossover.

J Clin Pharm Ther, 2003 Apr, 28(2), 131 - 6
A randomized controlled trial of a new ovule formulation of ornidazole for the treatment of bacterial vaginosis; Baloglu E et al.; OBJECTIVE: To compare the efficacy of 500 mg ornidazole vaginal ovules (VO) and vaginal tablets (VT) in the treatment of bacterial vaginosis . METHOD: Patients were allocated at random to one group of 50 subjects to be treated with a VO (500 mg) prepared in our laboratory and to a second group of 50 subjects to be treated with a VT of ornidazole (500 mg) . Therapeutic efficacy was assessed by Nugent's scoring system and clinical criteria (Amsel's criteria) before and 1 week after treatment . RESULTS: At the first follow-up visit, complete disappearance of the signs and symptoms or highly significant reduction in intensity of symptoms was observed in both treatment groups . No significant difference was evident between the two ornidazole formulations.

Environ Microbiol, 2003 May, 5(5), 383 - 94
The gene cassette metagenome is a basic resource for bacterial genome evolution; Holmes AJ et al.; Lateral gene transfer has been proposed as a fundamental process underlying bacterial diversity . Transposons, plasmids and phage are widespread and have been shown to significantly contribute to lateral gene transfer . However, the processes by which disparate genes are assembled and integrated into the host regulatory network to yield new phenotypes are poorly known . Recent discoveries about the integron/gene cassette system indicate it has the potential to play a role in this process . Gene cassettes are small mobile elements typically consisting of a promoterless orf and a recombination site . Integrons are capable of acquisition and re-arrangement of gene cassettes and of the expression of their associated genes . The potential of the integron/gene cassette system is thus largely determined by the diversity contained within the cassette pool and the rate at which integrons sample this pool . We show here using a polymerase chain reaction (PCR) approach by which the environmental gene cassette (EGC) metagenome can be directly sampled that this metagenome contains both protein-coding and non-protein coding genes . Environmental gene cassette-associated recombination sites showed greater diversity than previously seen in integron arrays . Class 1 integrons were shown to be capable of accessing this gene pool through tests of recombinational activity with a representative range of EGCs . We propose that gene cassettes represent a vast, prepackaged genetic resource that could be thought of as a metagenomic template for bacterial evolution.

Pol Merkuriusz Lek, 2003 Jan, 14(79), 50 - 4
{Role of bacterial infection in the pathophysiology of benzylpenicillin allergy}; Zdziarski P; In this paper current data on the pathogenesis of benzylpenicillin allergy is presented . Individual allergic reaction and susceptibility for the drug perhaps is not related to chemical properties of benzylpenicillin . An association between bacterial infection and allergy to benzylpenicillin is reviewed.

J Virol Methods, 2003 May, 109(2), 235 - 44
Use of PEG to acquire highly soluble DNA-packaging enzyme gp16 of bacterial virus phi29 for stoichiometry quantification; Huang LP et al.; All linear dsDNA viruses package their genome into a preformed procapsid via a ATP-driving motor involving two nonstructural enzymes or ATPase . This essential viral replication step has been investigated in the quest for new antiviral drugs . These DNA-packaging motors could be potential parts in nanotechnology . But both the low solubility and self-aggregation of all nonstructural enzymes have seriously hampered studies on these motors . Bacterial virus phi29 DNA-packaging motor has been well characterized . But the role of the nonstructural ATPase gp16 has not been well defined due to its hydrophobicity, low solubility, and self-aggregation . Here we report a novel approach to obtain affinity-purified, soluble, and highly active native gp16 with the aid of polyethylene glycol (PEG) or acetone . With several thousand-fold increase in specific activity in comparison to the traditional method, this unique approach has made the quantification of gp16 feasible . The basic functional unit of gp16 in solution was found to be a monomer, as determined by sedimentation and size exclusion chromatography . This result leads to a subsequent finding that the stoichiometry of gp16 for phi29 DNA-packaging was about 11+/-2 . These findings will facilitate the study on this novel motor that involves three pRNA dimers and a 12-subunit connector.

Rheumatology (Oxford), 2003 May, 42(5), 617 - 21
Serious bacterial infections in patients with rheumatoid arthritis under anti-TNF-alpha therapy; Kroesen S et al.; OBJECTIVE: With rising numbers of anti-tumour necrosis factor alpha (TNF-alpha) treatments for rheumatoid arthritis (RA), Crohn's disease and other conditions, physicians unaware of potential pitfalls are increasingly likely to encounter associated severe infections . Our purpose was to assess the incidence and nature of severe infections in our RA patients under anti-TNF-alpha therapy . METHODS: We reviewed patient charts and records of the Infectious Disease Unit for serious infections in patients with RA in the 2 yr preceding anti-TNF-alpha therapy and during therapy . RESULTS: Serious infections affected 18.3% of patients treated with infliximab or etanercept . The incidence was 0.181 per anti-TNF-alpha treatment year vs 0.008 in the 2 yr preceding anti-TNF-alpha therapy . In several cases, only a few signs or symptoms indicated the severity of developing infections, including sepsis . CONCLUSIONS: A high level of suspicion of infection is necessary in patients under anti-TNF-alpha therapy . We suggest additional strategies for the prevention, rapid identification and pre-emptive therapy of such infections.

Mol Biochem Parasitol, 2003 Apr 25, 128(1), 11 - 9
Bacterial-like energy metabolism in the amitochondriate protozoon Hexamita inflata; Biagini GA et al.; Hexamita inflata is an amitochondriate flagellated protozoon which inhabits O(2)-limited environments . With the aid of 1H NMR spectroscopy, analysis of the metabolic fluxes in H . inflata grown in complex media under limited O(2) was performed . Almost complete carbon recovery from maltose (the principle carbohydrate source in the medium) catabolism was calculated from the measured increase in concentration of ethanol, alanine, acetate and lactate (and estimated CO(2) production) . Difference spectra and amino acid analysis also identified changes in concentration of metabolites belonging to the arginine dihydrolase (ADH) pathway . The enzymes of the ADH pathway were detected in extracts with the following activities (in nmoles min(-1) x (mg of protein) x (-1)): arginine deiminase, 3.30; catabolic ornithine carbamyltransferase (OCT), 1.3; anabolic OCT, 93.0; and carbamate kinase, 1829 . The organism metabolized the ornithine produced from catabolic OCT activity to putrescine via ornithine decarboxylase (ODC) . The polyamines, spermidine and spermine, were formed by the sequential addition of the aminopropyl group of decarboxylated S-adenosyl-L-methionine (SAM) by the respective polyamine synthases . In addition, asparaginase activity was confirmed in H . inflata, catalysing the deamination of asparagine generating aspartate and ammonia . This study also indicates that, as with other amitochondriate protozoa and some bacteria, the ADH pathway significantly contributes to the energy yield of the cell, particularly under O(2)-limited conditions.

Vaccine, 2003 May 16, 21(17-18), 1862 - 6
Evaluation of endotoxin content of diphtheria-tetanus-acellular pertussis combined (DTaP) vaccines that interfere with the bacterial endotoxin test; Ochiai M et al.; Applicability of the endotoxin test to diphtheria-tetanus-acellular pertussis combined (DTaP) vaccines was examined . We found some DTaP vaccines that strongly interfered with Limulus amoebocyte lysate (LAL) activity of endotoxin without affecting lethal activity of endotoxin in D-galactosamine-treated mice . LAL activity that was interfered in such vaccines increased apparently after the treatment with phosphate buffer at 4 degrees C for a week . The DTaP vaccines that interfered with the endotoxin test showed no significant effect on endotoxin activity in inducing tumor necrosis factor-alpha (TNF-alpha) in rabbit peripheral blood . The in vitro TNF-alpha induction assay was, therefore, suggested to be an appropriate assay method for the quantitative detection of the endotoxin activity in DTaP vaccines.

Biosens Bioelectron, 2003 May, 18(5-6), 683 - 7
SNP detection in transforming growth factor-beta1 gene using bacterial magnetic particles; Ota H et al.; A single nucleotide polymorphism (SNP) within the transforming growth factor-beta1 (TGF-beta1) gene was detected by hybridization-based method using bacterial magnetic particles (BMPs) . TGF-beta1 is commonly associated with a single base change resulting in a Leu(10)-->Pro (T(869)-->C) polymorphism and is a genetic marker for susceptibility to osteoporosis . Short (9 bases) and specific probes were designed to detect SNP in TGF-beta1 . Detection probes were immobilized on BMPs using cross-linking reagents . TGF-beta1 PCR products (139 bp) were labeled with the fluorescent dye coumarin and hybridized with detection probes on BMPs . Complementary hybridized targets gave over four times higher fluorescent intensities, compared with one base mismatched hybridizations . The SNP genotype was successfully discriminated using this technique.

Biosens Bioelectron, 2003 May, 18(5-6), 661 - 6
Single nucleotide polymorphism genotyping of aldehyde dehydrogenase 2 gene using a single bacterial magnetic particle; Yoshino T et al.; A single nucleotide polymorphism (SNP) genotyping for aldehyde dehydrogenase 2 gene (ALDH2) has been developed by using a nano-sized magnetic particle, which was synthesized intracellularly by magnetic bacteria . Streptavidin-immobilized on bacterial magnetic particles (BMPs) were prepared using biotin labeled cross-linkers reacting with the amine group on BMPs . ALDH2 fragments from genomic DNA were amplified using a TRITC labeled primer and biotin labeled primer pair, and conjugated onto BMP surface by biotin-streptavidin interaction . PCR product-BMP complex was observed at a single particle level by fluorescence microscopy . These complexes were treated with restriction enzyme, specifically digesting the wild-type sequence of ALDH2 (normal allele of ALDH2) . The homozygous (ALDH2*1/*1), heterozygous (ALDH2*1/*2), and mutant (ALDH2*2/*2) genotypes were discriminated by three fluorescence patterns of each particle . SNP genotyping of ALDH2 has been successfully achieved at a single particle level using BMP.

No Shinkei Geka, 2003 Apr, 31(4), 443 - 8
{A case of a bacterial brain abscess presenting as symptoms of 'sudden stroke-like' onset}; Mori K et al.; We report a case of a bacterial brain abscess presenting symptoms of 'sudden stroke-like' onset, associated with infective endocarditis . A 59-year-old woman experienced a sudden stroke-like onset of left hemiplegia . Computed tomography (CT) and magnetic resonance imaging (MRI) were performed on the day of ictus . No lesion responsible for the symptom was seen on either CT or a T2 weighted image (T2WI), but a diffusion-weighted image (DWI) revealed focal increased signal intensity in the right frontal lobe . An initial diagnosis of acute embolic infarction associated with infective endocarditis was made . Although the patient's neurological state had been stable, motor paresis of her left extremities became worse starting one month after her admission . MRI with gadolinium-diethylenetriaminepenta-acid (Gd-DTPA) at 37 days after admission showed an irregular-shaped ring-enhancement lesion located at the same place as the initial infarction, and in the left frontal lobe . Surgical drainage of the lesion in the right frontal lobe was performed, and diagnosed as a bacterial abscess . The exact mechanism of a bacterial brain abscess presenting with 'sudden stroke-like' onset is unknown, but various hypotheses have been proposed . One is that paroxysmal septic emboli lead to abscess formation within or near the area of embolic infarction . Our case showed that the creation of a brain abscess followed embolic strokes, and that this hypothesis was demonstrated by MRI carried out on the day of ictus.

Microb Ecol, 2003 May, 45(4), 353 - 61 Epub 2003 Apr 28.
Ultraviolet radiation alters maize phyllosphere bacterial diversity; Kadivar H et al.; Epiphytic bacteria are subjected to very stressful environments, including UV radiation . Bacterial assemblages on Zea mays (maize) leaves exposure were examined with and without UV-B radiation . Culture-independent molecular techniques were utilized for bacterial identification, diversity analysis and selection of putative UV exposure marker sequences . Few sequences corresponded to previously characterized phyllosphere bacteria . There was a strong tendency toward increased 16S rDNA sequence diversity in UV samples . Overall community structure was assessed using denaturing gel gradient electrophoresis; significant alterations in community structure were found in comparisons of phyllosphere bacterial samples from control and solar UV-B exposed plants.

Microb Ecol, 2003 May, 45(4), 384 - 98 Epub 2003 Apr 22.
Interaction of nutrient limitation and protozoan grazing determines the phenotypic structure of a bacterial community; Matz C et al.; We examined the impact of nutrient conditions (carbon and phosphorus limitation) and grazing by protozoans on the phenotypic community structure of freshwater bacteria in continuous culture systems . Lakewater bacteria were grown on mineral medium, which was supplemented with glucose and amino acids and adjusted by different phosphorus concentrations to achieve either carbon or phosphorus limitation . Each nutrient treatment was inoculated with the same bacterial community and consisted of a nongrazing and a grazing treatment, to which the heterotrophic nanoflagellates Spumella sp . and Ochromonas sp . were added . We found that nutrient conditions alone resulted in differences in the phenotypic structure of the bacterial community: small and motile bacteria dominated under C limitation while large, elongated, and capsulated bacteria were characteristic for P limitation . The genotypic community composition as measured by T-RFLP (terminal restriction fragment length polymorphism) was not severely influenced by the two nutrient treatments . In the presence of flagellate predators, grazing-resistant bacteria developed under both nutrient conditions, but with different survival mechanisms: highly motile bacteria prevailed under C limitation, whereas the P-limited grazing treatment was dominated by filamentous forms . T-RFLP analysis revealed only moderate changes in bacterial community composition due to grazing, which were most pronounced under P limitation . Analysis by video microscopy revealed that high swimming speed is an efficient nonmorphological survival mechanism for bacteria to reduce the capture success of the flagellate predator . The rejection of optimal-sized, nonmotile bacteria under P limitation suggests the importance of other nonmorphological, surface-located cell properties . Our results illustrate that the realized mechanisms of grazing resistance are linked to the actual limitation conditions, and that the combined effects of nutrient limitation and grazing are major determinants of bacterial community structure.

Proc Natl Acad Sci U S A, 2003 Apr 29, 100(9), 5449 - 54 Epub 2003 Apr 18.
A sensitive, versatile microfluidic assay for bacterial chemotaxis; Mao H et al.; We have developed a microfluidic assay for bacterial chemotaxis in which a gradient of chemoeffectors is established inside a microchannel via diffusion between parallel streams of liquid in laminar flow . The random motility and chemotactic responses to L-aspartate, L-serine, L-leucine, and Ni(2+) of WT and chemotactic-mutant strains of Escherichia coli were measured . Migration of the cells was quantified by counting the cells accumulating in each of 22 outlet ports . The sensitivity of the assay is attested to by the significant response of WT cells to 3.2 nM L-aspartate, a concentration three orders of magnitude lower than the detection limit in the standard capillary assay . The response to repellents was as robust and easily recorded as the attractant response . A surprising discovery was that L-leucine is sensed by Tar as an attractant at low concentrations and by Tsr as a repellent at higher concentrations . This assay offers superior performance and convenience relative to the existing assays to measure bacterial tactic responses, and it is flexible enough to be used in a wide range of different applications.

Infect Immun, 2003 May, 71(5), 2902 - 6
Influence of Lewis antigen expression by Helicobacter pylori on bacterial internalization by gastric epithelial cells; Lozniewski A et al.; The role of Helicobacter pylori lipopolysaccharide (LPS) Lewis antigens in infection is still not well known . We investigated the influence of Lewis antigen expression by H . pylori on its internalization by AGS cells and the epithelium of human gastric xenografts in nude mice using isogenic mutants in LPS biosynthetic genes . In vivo, colonization rates were unaffected by the change in H . pylori Lewis antigen expression, whereas the number of viable intracellular bacteria was significantly higher with wild-type H . pylori strains expressing Lewis antigens when compared to the isogenic mutants in both models . H . pylori strains expressing more Lewis X antigens (Le(x)) were internalized at a higher rate than those expressing less Le(x), type II Lewis antigens (Le(a) or Le(b)) alone, or no Lewis antigens . Thus, Lewis antigens appear to be involved in the internalization of H . pylori by the gastric epithelium.

Infect Immun, 2003 May, 71(5), 2766 - 74
Inhibition of gamma interferon decreases bacterial load in peritonitis by accelerating peritoneal fibrin deposition and tissue repair; Qiu G et al.; Bowel perforation can lead to significant bacterial spillage, which may then cause septic peritonitis, characterized by a systemic inflammatory response and organ dysfunction . There are several reports that have shown that the development of peritoneal adhesions is dependent on inflammatory cytokine levels and that these adhesions can reduce bacterial spread, possibly by sealing off the cecum in the cecal ligation and puncture (CLP) model of septic peritonitis . There have not, however, been any studies that have utilized a strategy to accelerate tissue repair in order to seal off the injured cecum and reduce bacterial spread as well as ameliorate systemic inflammation . In the present study, we demonstrate that the administration of anti-gamma interferon (IFN-gamma) antibody (1.2 mg/kg of body weight, intravenously) accelerated tissue repair via increased fibrin deposition 12 and 24 h after CLP in rats . This increase in fibrin deposition was associated with peritoneal adhesion 24 h after CLP and a reduction in bacterial load compared to the bacterial load of rats given irrelevant antibody . Plasma fibrin levels, however, were not altered after IFN-gamma antibody administration, suggesting that the inhibition of IFN-gamma activity specifically increased fibrin deposition to the site of injury . Furthermore, plasma interleukin-6, used as a marker of systemic inflammatory response, was reduced in CLP rats given IFN-gamma antibody compared to that found in those given irrelevant antibody . These results suggest that the early inhibition of IFN-gamma activity in the CLP model is beneficial by accelerating fibrin deposition in cecal tissue to prevent bacterial spread and reduce the systemic inflammatory response . Importantly, increased fibrin deposition in the ceca was not associated with increased plasma fibrin whereas the latter may have detrimental effects associated with coagulation disorders.

Arch Histol Cytol, 2003 Mar, 66(1), 53 - 62
Bacterial lipopolysaccharide-induced expression of the IkappaB protein MAIL in B-lymphocytes and macrophages; Kitamura H et al.; Molecule possessing ankyrin-repeats induced by lipopolysaccharide (MAIL), a recently cloned nuclear IkappaB protein induced by lipopolysaccharide (LPS) stimulation in lymphoid organs, is involved in the regulation of inflammatory responses . The present in situ hybridization and immunohistochemical analyses revealed the distinct expression of the MAIL mRNA and protein in B-lymphocytes of the white pulp of the spleen and cortical lymphoid follicles of lymph nodes in LPS-injected mice . MAIL signals were also localized in F4/80-positive macrophages in these organs . LPS clearly induced MAIL expression in cultured B-lymphocytes and monocytes/macrophages, but only faintly so in T-lymphocytes, fibroblasts, and endothelial cells . MAIL was also induced by inflammatory cytokines such as interleukin-1 and -6, and tumor necrosis factor in cultured cells . Northern blot, Western blot, and in situ hybridization analyses showed that the major expression product of the Mail gene was a long splicing variant (MAIL-L) rather than a short one, both in lymphoid organs and cultured cells . These results collectively indicate that LPS induces MAIL-L predominantly in B-lymphocytes and macrophages.

J Bacteriol, 2003 May, 185(9), 2723 - 30
Chaperone-subunit-usher interactions required for donor strand exchange during bacterial pilus assembly; Barnhart MM et al.; The assembly of type 1 pili on the surface of uropathogenic Escherichia coli proceeds via the chaperone-usher pathway . Chaperone-subunit complexes interact with one another via a process termed donor strand complementation whereby the G1beta strand of the chaperone completes the immunoglobulin (Ig) fold of the pilus subunit . Chaperone-subunit complexes are targeted to the usher, which forms a channel across the outer membrane through which pilus subunits are translocated and assembled into pili via a mechanism known as donor strand exchange . This is a mechanism whereby chaperone uncapping from a subunit is coupled with the simultaneous assembly of the subunit into the pilus fiber . Thus, in the pilus fiber, the N-terminal extension of every subunit completes the Ig fold of its neighboring subunit by occupying the same site previously occupied by the chaperone . Here, we investigated details of the donor strand exchange assembly mechanism . We discovered that the information necessary for targeting the FimC-FimH complex to the usher resides mainly in the FimH protein . This interaction is an initiating event in pilus biogenesis . We discovered that the ability of an incoming subunit (in a chaperone-subunit complex) to participate in donor strand exchange with the growing pilus depended on a previously unrecognized function of the chaperone . Furthermore, the donor strand exchange assembly mechanism between subunits was found to be necessary for subunit translocation across the outer membrane usher.

Bioorg Med Chem Lett, 2003 May 5, 13(9), 1557 - 60
Macrocyclic inhibitors of the bacterial cell wall biosynthesis enzyme MurD; Horton JR et al.; Computer-based molecular design has been used to produce a series of new macrocyclic systems targeted against the bacterial cell wall biosynthetic enzyme MurD . Following their preparation, which involved a novel metathesis-based cyclisation as the key step, these systems were found to show good inhibition when assayed against the MurD enzyme.

Mikrobiologiia, 2003 Jan-Feb, 72(1), 5 - 13
{Bacterial cold shock response at the level of DNA transcription, translation and chromosome dynamics}; Golovlev EL; The review is an attempt to analyze the data available in the literature concerning the response of mesophilic bacteria to cold shock at the level of DNA transcription, translation, and chromosome dynamics, i.e., in terms of cell biology . The review considers relevant molecular mechanisms and particular regulatory systems within the framework of a general cell response to cold shock . It is suggested that a short-term response to cold shock is necessary for bacteria to transit to a viable but nonculturable state and/or for their physiological and genetic adaptation to psychrotrophic life . It is emphasized that cell responses to cold and heat shocks are different and that DNA dynamics (i.e., its supercoiling, multiple bending, and condensation) and the rearrangement of the protein-synthesizing apparatus of cells (including the induction of alternative translational mechanisms) may play a central role in cell response to cold shock . The role of molecular chaperones in cold shock response is presumably of less importance than it is in the case of heat shock.

Appl Microbiol Biotechnol, 2003 May, 61(3), 234 - 9 Epub 2003 Feb 26.
High-level expression of a lacZ gene from a bacterial artificial chromosome in Escherichia coli; Chang TS et al.; The GlnAP2 element has been proved to be an effective and inducible-by exogenous acetate-promoter in Escherichia coli with glnL/pta double mutations . Based on this feature, a single-copy expression vector was constructed via coupling of the glnAP2 promoter-regulated T7 RNA polymerase gene and the T7-promoter-controlled lacZ gene on a bacterial artificial chromosome . After induction with 20 mM potassium acetate, the glnL/pta double mutant E . coli harboring the single-copy plasmid produced 47,500 Miller units of beta-galactosidase activity . This high level expression, corresponding to 27% of total cell protein, was comparable to that determined with the commercial multi-copy expression vector, pET-14b, in strain E . coli Tuner (DE3) (64,300 Miller units, 41% of total cell protein) . Moreover, this single-copy expression vector could be maintained for at least 150 generations even in the presence of inducers . In contrast, the multi-copy expression vector was extensively lost after induction . The results indicate that the single-copy expression system has the potential for high-level heterologous protein production for industrial applications.

Kobe J Med Sci, 2003, 49(1-2), 17 - 24
Intraperitoneal administration of oxygenated perfluorochemical inhibits bacterial translocation associated with severe acute pancreatitis; Shinzeki M et al.; Bacterial translocation from gut has been assumed to be an infectious source in severe acute pancreatitis . The purpose of this study was to test the effect of intraperitoneal administration of oxygenated perfluorochemical on bacterial translocation associated with rat experimental acute necrotizing pancreatitis . Severe necrotizing pancreatitis was induced by retrograde injection of 3% sodium deoxycholate into the biliopancreatic ducts of male Wistar rats . Although mortality rate was not improved by the treatment, intraperitoneal administration of oxygenated perfluorochemical, perfulorodecalin reduced incidence of bacterial translocation to the mesenteric lymph nodes from 60% to 37% 12 hours after development of pancreatitis, and significantly reduced number of bacterial colonies detected after 24 hours . The treatment did not alter the villous height and crypt depth of the ileum . In this model for pancreatitis, however, accelerated apoptosis of the intestinal epithelium was detected histochemically by TUNEL staining and biochemically by DNA fragmentation ELISA, and the apoptotic changes were significantly suppressed by the treatment . These results indicate that intraperitoneal administration of oxygenated perfluorochemical inhibits apoptosis of intestinal epithelium and bacterial translocation induced in severe acute pancreatitis.

Water Res, 2003 Apr, 37(8), 1711 - 22
Bacterial contamination of Mediterranean coastal seawater as affected by riverine inputs: simulation approach applied to a shellfish breeding area (Thau lagoon, France); Fiandrino A et al.; Consequences of short-term changes in thermotolerant coliform loads on their spatio-temporal distribution in a Mediterranean lagoon with large-scale mollusk farming (Thau lagoon, France) were explored using a simulation approach . Simulations were based on bacterial transport and survival coupled models forced by the input of bacterial loads from the two main rivers (Vene and Pallas) that flow into the lagoon . Different flow types (reference, sudden and constant), bringing the same bacterial load, were considered and subsequent spatial and temporal bacterial contamination of lagoon surface water and shellfish was estimated . Simulation results showed that as long as loads were high, hydrodynamical processes governed the distribution of bacterial abundance in receiving areas . As soon as loads decreased or when time supply increased, biological die-off processes became dominant . Bacterial contamination of shellfish induced by the different flow types appeared to depend on the receiving area . In the case of Pallas River area, a sudden input of bacteria led to a high bacterial contamination of shellfish but only during a short period ( approximately 1 day) . A constant input of the same amount of bacteria induced a lower but significant contamination during all the simulation period (10 days) . On the contrary, bacterial inputs from the Vene River led to shellfish contamination only when bacteria were delivered through a flood event . Exposure time of bacteria to adverse environmental conditions appeared to be the main explanation to the above-mentioned differences . Consequences of our results in terms of environmental management strategy were discussed.

Genome Res, 2003 May, 13(5), 991 - 8 Epub 2003 Apr 14.
Comparing bacterial genomes through conservation profiles; Martin MJ et al.; We constructed two-dimensional representations of profiles of gene conservation across different genomes using the genome of Escherichia coli as a model . These profiles permit both the visualization at the genome level of different traits in the organism studied and, at the same time, reveal features related to the genomes analyzed (such as defective genomes or genomes that lack a particular system) . Conserved genes are not uniformly distributed along the E . coli genome but tend to cluster together . The study of gene distribution patterns across genomes is important for the understanding of how sets of genes seem to be dependent on each other, probably having some functional link . This provides additional evidence that can be used for the elucidation of the function of unannotated genes . Clustering these patterns produces families of genes which can be arranged in a hierarchy of closeness . In this way, functions can be defined at different levels of generality depending on the level of the hierarchy that is studied . The combined study of conservation and phenotypic traits opens up the possibility of defining phenotype/genotype associations, and ultimately inferring the gene or genes responsible for a particular trait.

Yi Chuan Xue Bao, 2002 Dec, 29(12), 1126 - 31
{Research on the preparation of bacterial artificial chromosome (BAC) vector DNA}; Jiang T et al.; Bacterial artificial chromosome (BAC) library plays a pivotal role in genomics studies . A crucial step in BAC library construction is preparation of BAC vector DNA . Preparation of highly purified vector DNA is affected by a series of factors including digestion of restriction enzyme and dephosphorylation of linearized vector DNA . In our study, the BAC vector pECBAC1 was digested by the restriction enzyme of BamHI and dephosphorylated by HK phosphatase respectively . In order to improve the ligation capability of vector DNA, gel purification of linearized vector DNA was also conducted in our study . At the same time, we did a series of experiments to get high quality of vector DNA for construction of BAC library . They included the optimal concentration of restriction enzyme, optimal digestion time, the type of phosphatase and gel purification of linearized vector DNA.

Nihon Kokyuki Gakkai Zasshi, 2002 Dec, 40(12), 929 - 35
{Evaluation of community-acquired pneumonia guidelines of Japanese Respiratory Society: differentiation of atypical pneumonia and bacterial pneumonia}; Ishida T et al.; To evaluate the usefulness of differentiation of atypical pneumonia and bacterial pneumonia in the community-acquired pneumonia guidelines of the Japanese Respiratory Society, we investigated 124 cases of three atypical pneumonias (Mycoplasma pneumonia, 62 cases; Chlamydia pneumoniae pneumonia, 46 cases; Chlamydia psittaci pneumonia, 13 cases) and 403 cases of bacterial pneumonia at our hospital over seven years . Overall, the sensitivity and specificity of the criteria in the guideline were 70.4% and 91.8%, respectively . High accordance was recognized in patients under 60 years old with atypical pneumonia . Items in the criteria that included subjective factors were considered inassessable . We found that the differentiation of pneumonias in the guideline is useful for the diagnosis of atypical pneumonia among younger patients, but it should be concise and objective . We therefore propose that the criteria would be more effective if they consisted of only 4 items: age under 60 years, no underlying disorders, presence of stubborn dry cough, and normal peripheral white blood cell count.

J Virol, 2003 May, 77(9), 5073 - 83
Cloning of the full-length rhesus cytomegalovirus genome as an infectious and self-excisable bacterial artificial chromosome for analysis of viral pathogenesis; Chang WL et al.; Rigorous investigation of many functions encoded by cytomegaloviruses (CMVs) requires analysis in the context of virus-host interactions . To facilitate the construction of rhesus CMV (RhCMV) mutants for in vivo studies, a bacterial artificial chromosome (BAC) containing an enhanced green fluorescent protein (EGFP) cassette was engineered into the intergenic region between unique short 1 (US1) and US2 of the full-length viral genome by Cre/lox-mediated recombination . Infectious virions were recovered from rhesus fibroblasts transfected with pRhCMV/BAC-EGFP . However, peak virus yields of cells infected with reconstituted progeny were 10-fold lower than wild-type RhCMV, suggesting that inclusion of the 9-kb BAC sequence impeded viral replication . Accordingly, pRhCMV/BAC-EGFP was further modified to enable efficient excision of the BAC vector from the viral genome after transfection into mammalian cells . Allelic exchange was performed in bacteria to substitute the cre recombinase gene for egfp . Transfection of rhesus fibroblasts with pRhCMV/BAC-Cre resulted in a pure progeny population lacking the vector backbone without the need of further manipulation . The genomic structure of the BAC-reconstituted virus, RhCMV-loxP(r), was identical to that of wild-type RhCMV except for the residual loxP site . The presence of the loxP sequence did not alter the expression profiles of neighboring open reading frames . In addition, RhCMV-loxP(r) replicated with wild-type kinetics both in tissue culture and seronegative immunocompetent macaques . Restriction analysis of the viral genome present within individual BAC clones and virions revealed that (i) RhCMV exhibits a simple genome structure and that (ii) there is a variable number of a 750-bp iterative sequence present at the S terminus.

J Microbiol Methods, 2003 Jun, 53(3), 355 - 63
Locating transposable element polymorphisms in bacterial genomes; Yesilkaya H et al.; Although whole-genome sequencing is greatly extending our knowledge of the genetic capacity of those bacterial species, it is only directly informative for the particular strain sequenced . Many bacterial species exhibit more or less genetic polymorphism within their populations and characterising this variety is an extremely important way of elucidating the biology of these species . Often genomic polymorphisms are associated with multicopy elements, particularly transposable elements . We describe a novel method that efficiently characterises the sequences of such polymorphisms . We have optimised heminested inverse PCR (hINVPCR) to assess the diversity of insertional polymorphisms of a transposable element (IS6110) in clinical isolates of Mycobacterium tuberculosis . To increase the yield of information, genomic DNA was digested with different endonucleases (Bsp1286I, HaeII or PvuI), and primers based on both the 5' and 3' ends of IS6110 were used to amplify and determine the genomic sequence upstream (or downstream) of the transposable element . We found that both the choice of restriction enzyme and the use of primers at both ends of the transposable element significantly increased the diversity of the insertion sites identified . Band stabbing was incorporated into the method as an alternative to cloning in order to screen large number of isolates at a sequence level in a rapid and labour-efficient fashion . We describe some of the purposes to which such data can be put.

Shock, 2003 Apr, 19(4), 378 - 82
Supplementation and inhibition of nitric oxide synthesis influences bacterial transit time during bacterial translocation in rats; Samel S et al.; In the obstructed gut, nitric oxide (NO) may influence intestinal barrier function and translocation of bacteria . By using a novel experimental approach, we investigated the effect of supplementation and inhibition of NO synthesis on the time interval necessary for translocation of green fluorescent protein-transfected Escherichia coli (GFP-uv E . coli) in a rat model of small bowel obstruction . In anesthetized Wistar rats, 4 x 10(8) GFP-uv E . coli were administered into a reservoir of terminal ileum formed by ligature . Animals were randomized to receive either i.v . arginine (10 mg/kg), aminoguanidine (300 mg/kg), L-NAME (25 mg/kg), or saline (control) . Translocation of GFP-uv E . coli was assessed using intravital video microscopy . Minimal transit time of translocation was measured as time from injection of GFP-uv E . coli into the gut lumen until bacteria were observed in the lamina submucosa and as time from injection of bacteria into the gut lumen until bacteria were observed in the lamina muscularis propria . Minimal transit times were expressed as mean +/- SD . Bacterial translocation into the submucosa and muscularis propria took 36 +/- 7 min and 81 +/- 9 min, respectively in control animals receiving saline . Aminoguanidine and L-NAME caused a marked delay of minimal transit time into the submucosa (63 +/- 5 min and 61 +/- 7 min, respectively; P < 0.05) . Arginine significantly accelerated bacterial translocation into the muscularis propria (61 +/- 9 min, P < 0.05) . GFP-uv E . coli were detected on frozen sections of small bowel, mesentery, liver, and spleen 2 h after GFP-uv E . coli administration in all animals . A marked upregulation of inducible NO synthase (NOS) in the obstructed bowel segment was demonstrated on immunohistochemistry . The assessment of a newly defined parameter, minimal bacterial transit time, may serve as an additional functional aspect of intestinal barrier function for pathophysiological and pharmacological studies . Aminoguanidine, L-NAME, and arginine were effective in influencing minimal transit time of E . coli during small bowel obstruction.

Bioresour Technol, 2003 Feb, 86(3), 215 - 9
Increased production of bacterial cellulose by Acetobacter sp . V6 in synthetic media under shaking culture conditions; Son HJ et al.; Acetobacter strains are bacteria that can synthesize cellulose when grown in a complex medium containing glucose . The effect of the components of a synthetic medium on bacterial cellulose (BC) production by a newly isolated Acetobacter sp . V6 in shaking cultures was investigated . BC production was dependent on the presence of MgSO4 x 7H2O and cosubstrates such as ethanol and lactic acid in the medium . The optimal synthetic medium contained 1.5% glucose, 0.2% (NH4)2SO4, 0.3% KH2PO4, 0.3% Na2HPO4 x 12H2O, 0.08% MgSO4 x 7H2O, 0.0005% FeSO4 x 7H2O, 0.0003% H3BO3, 0.00005% nicotinamide, and 0.6% ethanol . A maximum BC concentration of 4.16 g/l was achieved after 8 days of cultivation at 200 rpm . The production of BC by Acetobacter sp . V6 was higher in synthetic medium than complex medium (Hestrin and Schramm medium) traditionally used for Acetobacter strains.

J Clin Microbiol, 2003 Apr, 41(4), 1785 - 7
BIBI, a bioinformatics bacterial identification tool; Devulder G et al.; BIBI was designed to automate DNA sequence analysis for bacterial identification in the clinical field . BIBI relies on the use of BLAST and CLUSTAL W programs applied to different subsets of sequences extracted from GenBank . These sequences are filtered and stored in a new database, which is adapted to bacterial identification.

FEBS Lett, 2003 Apr 10, 540(1-3), 234 - 40
Photo-accumulation of the P+QB- radical pair state in purple bacterial reaction centres that lack the QA ubiquinone; Wakeham MC et al.; Photo-excitation of membrane-bound Rhodobacter sphaeroides reaction centres containing the mutation Ala M260 to Trp (AM260W) resulted in the accumulation of a radical pair state involving the photo-oxidised primary electron donor (P) . This state had a lifetime of hundreds of milliseconds and its formation was inhibited by stigmatellin . The absence of the Q(A) ubiquinone in the AM260W reaction centre suggests that this long-lived radical pair state is P(+)Q(B)(-), although the exact reduction/protonation state of the Q(B) quinone remains to be confirmed . The blockage of active branch (A-branch) electron transfer by the AM260W mutation implies that this P(+)Q(B)(-) state is formed by electron transfer along the so-called inactive branch (B-branch) of reaction centre cofactors . We discuss how further mutations may affect the yield of the P(+)Q(B)(-) state, including a double alanine mutation (EL212A/DL213A) that probably has a direct effect on the efficiency of the low yield electron transfer step from the anion of the B-branch bacteriopheophytin (H(B)(-)) to the Q(B) ubiquinone .

FEBS Lett, 2003 Apr 10, 540(1-3), 26 - 34
Coupling of nuclear wavepacket motion and charge separation in bacterial reaction centers; Shuvalov VA et al.; The mechanism of the charge separation and stabilization of separated charges was studied using the femtosecond absorption spectroscopy . It was found that nuclear wavepacket motions on potential energy surface of the excited state of the primary electron donor P* leads to a coherent formation of the charge separated states P(+)B(A)(-), P(+)H(A)(-) and P(+)H(B)(-) (where B(A), H(B) and H(A) are the primary and secondary electron acceptors, respectively) in native, pheophytin-modified and mutant reaction centers (RCs) of Rhodobacter sphaeroides R-26 and in Chloroflexus aurantiacus RCs . The processes were studied by measurements of coherent oscillations in kinetics at 890 and 935 nm (the stimulated emission bands of P*), at 800 nm (the absorption band of B(A)) and at 1020 nm (the absorption band of B(A)(-)) as well as at 760 nm (the absorption band of H(A)) and at 750 nm (the absorption band of H(B)) . It was found that wavepacket motion on the 130-150 cm(-1) potential surface of P* is accompanied by approaches to the intercrossing region between P* and P(+)B(A)(-) surfaces at 120 and 380 fs delays emitting light at 935 nm (P*) and absorbing light at 1020 nm (P(+)B(A)(-)) . In the presence of Tyr M210 (Rb . sphaeroides) or M195 (C . aurantiacus) the stabilization of P(+)B(A)(-) is observed within a few picosseconds in contrast to YM210W . At even earlier delay (approximately 40 fs) the emission at 895 nm and bleaching at 748 nm are observed in C . aurantiacus RCs showing the wavepacket approach to the intercrossing between the P* and P(+)H(B)(-) surfaces at that time . The 32 cm(-1) rotation mode of HOH was found to modulate the electron transfer rate probably due to including of this molecule in polar chain connecting P(B) and B(A) and participating in the charge separation . The mechanism of the charge separation and stabilization of separated charges is discussed in terms of the role of nuclear motions, of polar groups connecting P and acceptors and of proton of OH group of TyrM210 .

Biochemistry, 2003 Apr 15, 42(14), 4064 - 74
Primary quinone (QA) binding site of bacterial photosynthetic reaction centers: mutations at residue M265 probed by FTIR spectroscopy; Wells TA et al.; In the primary quinone (Q(A)) binding site of Rb . sphaeroides reaction centers (RCs), isoleucine M265 is in extensive van der Waals contact with the ubiquinone headgroup . Substitution of threonine or serine for this residue (mutants M265IT and M265IS), but not valine (mutant M265IV), lowers the redox midpoint potential of Q(A) by about 100 mV (Takahashi et al . (2001) Biochemistry 40, 1020-1028) . The unexpectedly large effect of the polar substitutions is not due to reorientation of the methoxy groups as similar redox potential changes are seen for these mutants with either ubiquinone or anthraquinone as Q(A) . Using FTIR spectroscopy to compare Q(A)(-)/Q(A) IR difference spectra for wild type and the M265 mutant RCs, we found changes in the polar mutants (M265IT and M265IS) in the quinone C{double bond}O and C{double bond}C stretching region (1600-1660 cm(-1)) and in the semiquinone anion band (1440-1490 cm(-1)), as well as in protein modes . Modeling the mutations into the X-ray structure of the wild-type RC indicates that the hydroxyl group of the mutant polar residues, Thr and Ser, is hydrogen bonded to the peptide C{double bond}O of Thr(M261) . It is suggested that the mutational effect is exerted through the extended backbone region that includes Ala(M260), the hydrogen bonding partner to the C1 carbonyl of the quinone headgroup . The resulting structural perturbations are likely to include lengthening of the hydrogen bond between the quinone C1{double bond}O and the peptide NH of Ala(M260) . Possible origins of the IR spectroscopic and redox potential effects are discussed.

Russ J Immunol, 2001 Jul, 6(2), 131 - 146
Peripheral T Cell Apoptosis and Its Role in Generalized Bacterial Infections: A Minireview; Chernykh HR et al.; In the present review we have attempted to analyze recent findings concerning apoptosis of mature peripheral T cells . The great attention is made to the factors underlying resistance or sensitivity of mature T lymphocytes to activation-induced cell death . The role of preactivation and altered costimulation is discussed in this regard . Besides, the possible role of cytokines in the modulation of T cell apoptosis is emphasized . Particular attention is paid to the studies of apoptosis disorders in the pathogenesis of generalized bacterial infections . In this connection some own results are summarized as well . To characterize T cell death and its role in the pathogenesis of bacterial infections an anti-CD3-mAb or Con A-induced apoptosis in patients with severe and generalized forms of surgical infections have been investigated . We have found a significant increase of activation-induced lymphocyte apoptosis and a high level of apoptosis in freshly isolated lymphocytes in patients with surgical infections . Alternatively, peripheral blood mononuclear cells from surgical patients without infectious complications did not exhibit a marked enhancement of activation-induced cell death . Activation-induced T cell death in surgical infections appeared to be Fas-dependent, involved reactive oxygen intermediates and was partly prevented by pro-inflammatory cytokines, among which IL-2 exhibited the most pronounced anti-apoptotic activity . Likewise, APACHE II score, as a marker of the infection severity, directly correlated with a rate of activation-induced T cell apoptosis . Accelerated T cell apoptosis at the early stage of infection was revealed in survivors and non-survivors, that appears to designate a common pathway for the restriction of systemic inflammation . At the late stage of infection altered T cell apoptosis could account for different outcomes, since the patients with lethal outcome showed 2-fold increase in activation-induced cell death compared to the opposite group . Immunotherapy with rIL-2 declined anti-CD3-induced apoptosis and promoted a reduction in the mortality rate from 29% to 13%.

Russ J Immunol, 1998 Jul, 3(2), 141 - 146
Competitive Specificity Analysis of Natural Antibodies against Epitope of Bacterial Cell Wall Peptidoglycoan: Glucosaminylmuramyl Dipeptide Carrying Adjuvant Activity; Pinegin BV et al.; The natural antibodies against glucosaminylmuramyl dipeptide (GMDP), the epitope of peptidoglycan of bacterial cell wall, isolated from human serum by thermal extraction possess a capability to cross-react with determinant of glycan chain - tetrasaccharide consisting of N-acetylglucosamine and N-acetylmuramic acid . The intensity of interaction of natural anti-GMDP-antibodies with specific ligand is significantly higher than with tetrasaccharide . The natural antibodies against tetrasaccharide carry properties of heteroclitic antibodies, i.e . the intensity of their interaction with heterologous ligand, GMDP, is significantly higher than with homologous one, tetrasaccharide GMGM . GMDP is supposed to be specific antigenic peptidoglycan determinant against which the antibodies reacting with various intensity to homologous (GMDP) and relative (tetrasaccharide) hapten are formed in the process of natural immunization.

Russ J Immunol, 1997 Dec, 2(3-4), 177 - 182
Affinity of Serum Natural Antibodies against Peptidoglycan Bacterial Cell Component with Adjuvant Activity; Kulakov AV et al.; We studied affinity of natural antibodies of human serum against glucosaminylmuramyl dipeptide (GMDP), the epitope peptidoglycan bacterial cell wall component which carries adjuvant activity . Antibodies against GMDP were isolated from the blood sera of healthy donors using thermal extraction of antibodies from specific ligand on plastic . Determination of the dissociation constant (K(d)) showed equal K(d) in the serum and affinity-purified anti-GMDP-antibodies, i.e . extraction by this method led to the isolation of all subpopulations of antibodies in spite of their affinity . K(d) of serum, affinity-purified and monoclonal anti-GMDP-antibodies proved of low value - 10(-6) M, and according to this index anti-GMDP-antibodies may be classified between anti-protein and anti-carbohydrate antibodies.

Eur Biophys J, 2003 Sep, 32(6), 537 - 43 Epub 2003 Mar 18.
Electron transport dynamics at the quinone acceptor site of bacterial photosynthetic reaction centers as probed using fast temperature changes; Chamorovsky SK et al.; Methods of laser-induced temperature jumps and fast freezing were used for testing the rates of thermoinduced conformational transitions of reaction center (RC) complexes in chromatophores and isolated RC preparations of various photosynthesizing purple bacteria . An electron transfer reaction from primary to secondary quinone acceptors was used as a probe of electron transport efficiency . The thermoinduced transition of the acceptor complex to the conformational state facilitating electron transfer to the secondary quinone acceptor was studied . To investigate the dynamics of spontaneous decay of the RC state induced by the thermal pulse, the thermal pulse was applied either before or during photoinduced activation of electron transport reactions in the RC acceptor complex . The maximum effect was observed if the thermal pulse was applied against the background of steady-state photoactivation of the RC . It was shown that neither the characteristic time of the thermoinduced transition within the temperature range 233-253 K nor the characteristic time of spontaneous decay of this state at 253 K exceeded several tens of milliseconds . Independent support of the estimates was obtained from experiments with varied cooling rates of the samples tested.

Eur Biophys J, 2003 May, 32(2), 159 - 62 Epub 2003 Mar 22.
Observing bacterial activity interferometrically; Jardim DF et al.; It is shown that bacterial activity, even of slowly growing species, can be detected by precise interferometric measurements of refractive index changes of the culture medium . The bacteria-containing sample is kept in an isothermal block together with a reference liquid without bacteria . The biological activity is obtained from the difference of the index changes of these samples . Experiments were performed with Bacilo Calmette-Guerin . The order of magnitude of the observed total refractive index change was compatible with theoretical estimates based on the amount of available oxygen . An unexpected positive index change during the lag phase was observed, which might permit fast diagnostics in medical applications . This technique may provide cheap and quick tests of bacterial susceptibility with respect to antibiotics.

Cell, 2003 Apr 4, 113(1), 1 - 2
A stress sensor for the bacterial periplasm; Young JC et al.; DegS, the periplasmic stress sensor, becomes activated when its PDZ domain recognizes the improperly exposed C-terminal sequences of outer membrane porins . This interaction relieves the inhibition of the neighboring protease domain of DegS, triggering a proteolysis cascade that leads to the sigma(E)-driven expression of periplasmic chaperones.

Curr Pharm Biotechnol, 2003 Apr, 4(2), 99 - 107
Enhanced efficacy of DNA vaccines against an intracellular bacterial pathogen by genetic adjuvants; Leclercq S et al.; After 200 years of practice, vaccinology has proved to be very effective in preventing infectious diseases . However, several human and animal pathogens exist for which vaccines need to be improved or simply have not yet been discovered . The era of molecular genetic has given a new breath for vaccine development with the achievement of the "Third Generation of Vaccines": the DNA vaccine . In this article, we reviewed strategies that have been used to improve and modulate the immune response induced by DNA vaccines, using as a model the intracellular bacterial pathogen Brucella abortus . First, we described different approaches used to isolate and to identify genes that encode potential immunogens . Secondly, we reported the use of cytokine genes and genetic adjuvants that could improve the immunogenicity of target genes . And finally, we discussed the "Expression Library Immunization"-(ELI) strategy and the recent results obtained against Brucella abortus infection.

Cell Mol Life Sci, 2003 Feb, 60(2), 390 - 400
Biochemical characterization and bacterial expression of an odorant-binding protein from Locusta migratoria; Ban L et al.; Analysis of soluble proteins from different body parts of Locusta migratoria revealed a fast-migrating component in native electrophoresis, unique to antennae of both sexes . N-terminal sequence analysis and cloning identified this protein as a member of the insect odorant-binding proteins, carrying a well-conserved six-cysteine motif . Mass spectrometry analysis confirmed the occurrence of two distinct polypeptide species determined by nucleotide sequencing and demonstrated that the cysteine residues are paired in an interlocked fashion . The protein was expressed in a bacterial system with yields of about 10 mg/l of culture, mostly present as inclusion bodies . However, this recombinant product was solubilized after disulfide reduction . Air oxidation yielded a species with all disulfides spontaneously formed as in the native counterpart . Both native and recombinant proteins migrated as a dimer in gel filtration chromatography . Ligand binding was measured, using N-phenyl-1-naphthylamine as the fluorescent probe; the affinity of other ligands was measured in competitive binding assays . The protein exhibited great resistance to thermal denaturation even following prolonged treatment at 100 degrees C . A structural model for this dimeric species was generated on the basis of its sequence homology with Bombyx mori pheromone-binding protein, whose three-dimensional structure has been resolved as an unbound species and in complex with its physiological ligand . This is the first report of an odorant-binding protein identified and characterized from Orthoptera.

Appl Environ Microbiol, 2003 Apr, 69(4), 2058 - 64
Coupling of bacterial endosymbiont and host mitochondrial genomes in the hydrothermal vent clam Calyptogena magnifica; Hurtado LA et al.; The hydrothermal vent clam Calyptogena magnifica (Bivalvia: Vesicomyidae) depends for its nutrition on sulfur-oxidizing symbiotic bacteria housed in its gill tissues . This symbiont is transmitted vertically between generations via the clam's eggs; however, it remains uncertain whether occasionally symbionts are horizontally transmitted or acquired from the environment . If symbionts are transmitted strictly vertically through the egg cytoplasm, inheritance of symbiont lineages should behave as if coupled to the host's maternally inherited mitochondrial DNA . This coupling would be obscured, however, with low rates of horizontal or environmental transfers, the equivalent of recombination between host lineages . Population genetic analyses of C . magnifica clams and associated symbionts from eastern Pacific hydrothermal vents clearly supported the hypothesis of strictly maternal cotransmission . Host mitochondrial and symbiont DNA sequences were coupled in a clam population that was polymorphic for both genetic markers . These markers were not similarly coupled with sequence variation at a nuclear gene locus, as expected for a randomly mating sexual population . Phylogenetic analysis of the two cytoplasmic genes also revealed no evidence for recombination . The tight association between vesicomyid clams and their vertically transmitted bacterial endosymbionts is phylogenetically very young (<50 million years) and may serve as a model for the origin and evolution of eukaryotic organelles.

Cell Microbiol, 2003 Apr, 5(4), 233 - 43
Redox-active metals in commercial preparations of lipopolysaccharide: implications for studies of cellular responses to bacterial products; Brubacher JL et al.; The mechanisms by which lipopolysaccharide (LPS) activates cells have been the subject of intense investigation for many years . Whereas much information on this process has been collected for mammalian species, little is known about the signalling path-ways operative in other animals . One general mode of cellular activation that has been recently pro-posed for pathways independent of the primary mammalian LPS receptor, CD14, involves reactive oxygen species (ROS) as intermediates in LPS-induced signalling pathways . Therefore, we used 2',7'-dichlorodihydrofluorescein, a fluorogenic probe of redox activity, to examine LPS-induced oxidative responses of a macrophage-like cell line from the rainbow trout, RTS11 . Lipopolysaccharide dose-dependently increased oxidation of this probe by RTS11 cells, and a variety of other cell lines . This process was inhibited by catalase, superoxide dismutase and NG-methylarginine citrate, an inhibitor of nitric oxide synthases, suggesting the involvement of a diverse assortment of cellular ROS . More careful dissection of this phenomenon led us to conclude that the increase in oxidation was, in fact, due almost entirely to metals, particularly copper, in some LPS preparations, which is something to consider when experimenting with LPS.

Theor Appl Genet, 2003 Apr, 106(6), 1102 - 6 Epub 2002 Dec 04.
Construction and characterization of a bacterial artificial chromosome library of banana (Musa acuminata Colla); Vilarinhos AD et al.; A bacterial artificial chromosome (BAC) library for banana was constructed from leaves of the wild diploid 'Calcutta 4' clone (Musa acuminata subsp . Burmannicoides 2n = 2 x = 22) . 'Calcutta 4' is widely used in breeding programs for its resistance to the current major disease of banana and is being used to build a genetic reference map of banana . As banana leaves are particularly rich in polyphenols and polysaccharides a protocol was adapted to isolate intact nuclei and high-molecular-weight (HMW) DNA . A total of 55,152 clones with an average insert size of 100 kb were picked . The frequency of BAC clones carrying inserts derived from chloroplast and mitochondrial DNA was estimated to be 1.5% . The coverage of the library is equivalent to 9.0-times the haploid genome . The BAC library was screened with 13 RFLP probes belonging to the 8 linkage groups of the consensus molecular map of banana . A total of 135 clones were identified giving an average of 10.38 clones for each locus . This BAC library will be a valuable starting tool for many of the goals of the recently emerged International Musa Genomic Consortium . One of our initial objectives will be to develop a banana physical map by BAC-FISH (fluorescent in situ hybridization) viewing the characterization of translocation break points.

Biophys J, 2003 Apr, 84(4), 2634 - 7
Single particle tracking of correlated bacterial dynamics; Soni GV et al.; Pattern formation in 3D random media has been a topic of interest in soft matter and biological systems . However, the onset of long-range microscopic ordering has not been explored in randomly moving self-propelled particles due to a lack of model systems as well as local probe techniques . In this article, we report on a novel experiment, using motile Escherichia coli bacteria as a model system, to study the onset of dynamic correlation and collective movement in three-dimension . We use fluctuation of an optically trapped micron-size bead as a detector of correlated bacterial motion, and further study this behavior by analyzing the motility of fluorescent bacteria in a confocal volume . We find evidence of dynamic correlation at very low volume fractions (0.01) . We show that the magnitude of this correlation strongly depends on the interbacterial distances and their coupling modes . This opens up possibilities to probe long-range pattern formation in actively propelled cells or organisms coupled through hydrodynamics and/or chemical signaling.

Transfus Clin Biol, 2003 Feb, 10(1), 6 - 9
Bacterial contamination and transfusion safety: experience in the United States; Dodd RY; In the United States, septic reactions from bacterial contamination of blood components are considered to be the most frequent and serious infectious outcomes of transfusion, reflecting 77 of 694 transfusion-related deaths reported to the FDA, during the period 1985-1999 . A number of recent surveillance programs have emphasized this, with nationally reported rates of about 10 per million platelet units transfused and about 1.4 per 10,000 transfusions in one hospital: significant fatality rates were noted in each setting . Although there is currently no regulatory requirement to undertake additional measures to reduce this adverse outcome, a number of approaches are under consideration . These include increased attention to skin preparation, diversion of the initial volume at phlebotomy, and the use of automated bacterial culture . A number of research studies are directed towards accessible methods for the detection of bacterial contamination . These include a modified culture approach for use in the blood center and tests that may be applied to platelet units shortly before transfusion . In addition, there is clear evidence that platelets prepared by apheresis offer lower overall risk to recipients as a result of the lower number of discrete products received by each patient . It is anticipated that pathogen reduction procedures may also impact bacterial contamination, but such procedures are not yet available in the US.

Arch Orthop Trauma Surg, 2003 Apr, 123(2-3), 121 - 4 Epub 2003 Mar 28.
Posterior dislocation of a cruciate-retaining total knee arthroplasty following an acute bacterial infection; Chu CM et al.; BACKGROUND: We report a rare complication of posterior dislocation of a cruciate-retaining total knee arthroplasty following an acute bacterial infection . The mechanism of dislocation proved to be septic loosening of the femoral component and a tear of the posterior cruciate ligament near to the femoral insertion site . The tear arose during the treatment of acute septic arthritis following total knee arthroplasty when the patient attempted full weight-bearing with the affected limb in a semiflexion position and twisted the knee . METHODS AND RESULTS: Successful treatment was provided with subsequent surgical debridement, removal of the loosened prosthesis, the application of systemic antibiotics, and a revision total knee arthroplasty utilizing a posteriorly stabilized prosthesis after adequate control of the infection . CONCLUSION: Soft-tissue protection from full weight-bearing of the knee during the treatment of an acute infection following total knee arthroplasty and timely removal of the loosened total knee prosthesis are recommended in order to prevent such a complication.

Curr Allergy Asthma Rep, 2003 May, 3(3), 232 - 7
Relationships between atopy and bacterial infections; Mucha SM et al.; Atopy in its most common forms (asthma, allergic rhinitis, and atopic dermatitis) has a significant impact on society in terms of health care costs and quality of life . Aside from having significant morbidity from these diseases, patients with atopy have also been noted to have a high incidence of comorbidities, including bacterial infections such as otitis media and sinusitis . In this paper, current evidence is reviewed that supports the close associations among allergic rhinitis and the two commonly diagnosed bacterial diseases, otitis media and sinusitis.

J Hum Nutr Diet, 2003 Apr, 16(2), 119 - 22
Small bowel bacterial overgrowth and rice malabsorption in healthy and physically disabled older adults; Mitsui T et al.; AIM: To determine whether there are differences in small bowel bacterial overgrowth (SBBO) and rice digestion between healthy and disabled older adults and to estimate the influence of physical activity on these nutritional statuses . METHOD: Fifteen disabled adults who commute to a day-care centre and 11 healthy older adults participated in this study . SBBO and rice absorption were judged using a breath hydrogen test . Physical activity was estimated using a pedometer . RESULTS: The average number of steps taken per day by the disabled was 1056 +/- 243, which was statistically lower than that of the healthy, 6904 +/- 782 (P < 0.001) . No SBBO-positive subject was seen in the healthy group, whereas five (33.3%) of 15 disabled older adults were SBBO-positive . After ingesting glucose solution, the triangle up H2 of disabled subjects was higher than that of the healthy subjects (7.6 +/- 2.7 versus 0.5 +/- 0.3 p.p.m., P < 0.05) . Rice malabsorption was seen in one (9.1%) of 11 in the healthy and two (14.3%) of 14 in the disabled groups, which was not statistically significant . CONCLUSIONS: Disabled older people who have a physically inactive lifestyle are at risk of SBBO, probably because of a reduction in their intestinal motility . SBBO has no influence on absorption of rice, and some older adults, independent of physical condition, can not absorb rice adequately.

Biotechniques, 2003 Mar, 34(3), 626 - 8, 630-2
PCR-assisted contig extension: stepwise strategy for bacterial genome closure; Carraro DM et al.; Finishing is rate limiting for genome projects, and improvements in the efficiency of complete genome-sequence compilation will require improved protocols for gap closure . Here we report a novel approach for extending shotgun contigs and closing gaps that we termed PCR-assisted contig extension (PACE) . PACE depends on the capture of rare mismatched interactions that occur between arbitrary primers and template DNA of unknown sequence, even under highly stringent conditions, by means of elevated PCR-cycle repetition and the use of specific anchoring primers corresponding to adjacent regions of known sequence . Using PACE, we have generated extensions with an average of 1 kb from all contigs generated from the shotgun sequencing of a 5-Mb genome, which closed the majority of gaps with a single round of experimentation . This included the generation of multiple extensions for contigs that terminated in one of the eight copies of the rRNA operon . We calculate that the switch from shotgun sequencing to PACE should occur between 5- and 8-fold genome coverage for maximum benefit and minimum overall cost . PACE is a robust and straightforward strategy that should simplify the finishing phase of bacterial genome projects.

Prostate, 2003 May 1, 55(2), 105 - 10
Negative bacterial polymerase chain reaction (PCR) findings in prostate tissue from patients with symptoms of chronic pelvic pain syndrome (CPPS) and localized prostate cancer; Leskinen MJ et al.; BACKGROUND: The etiology of chronic pelvic pain syndrome (CPPS) remains obscure . Although, bacterial etiology has frequently been suggested, evidence of both bacterial involvement in CPPS and the presence of normal bacterial flora in the prostate remain uncertain . MATERIALS AND METHODS: We investigated the presence of bacterial DNA using polymerase chain reaction (PCR) techniques on prostatic tissue samples obtained in radical prostatectomy from 10 patients with moderate to severe symptoms of CPPS and 10 nonsymptomatic patients with localized prostate cancer . For symptom evaluation we used the National Institutes of Health-Chronic Prostatitis Symptom Index (NIH-CPSI) . RESULTS: All but one sample were negative for bacterial DNA . The PCR from a symptomatic patient was reproducibly positive in 16S rDNA PCR but negative in 23S rDNA PCR . Bacterial DNA was found in only one out of two sample aliquots and cloning yielded different sequences in two PCR products . CONCLUSIONS: A bacterial etiology for CPPS symptoms could not be demonstrated in patients with prostate cancer . The results also suggest that the prostate is unlikely to harbor bacterial normal flora .

Fish Shellfish Immunol, 2003 Apr, 14(4), 347 - 62
Bacterial kidney disease as a model for studies of cell mediated immunity in rainbow trout (Oncorhynchus mykiss); Jansson E et al.; A cell mediated immune (CMI) response was measured in vitro to heat-killed and to paraformaldehyde fixed Renibacterium salmoninarum (Rs) in rainbow trout (Oncorhynchus mykiss) experimentally challenged with live Rs . The mitogenic response to the T lymphocyte mitogen Concanavalin A (Con A) was reduced during samplings 4 to 6 weeks after immersion, but no effect of the response to the B lymphocyte mitogen lipopolysaccharide (LPS) was detected . The subpopulation of lymphocytes, detected by the monoclonal antibody 1C2, was decreased from the 4th week to the 5th week of infection, and remained at the decreased level up to 10 weeks post immersion . The proportion of Immunoglobulin (Ig) bearing lymphocytes was not affected during the Rs infection period . The humoral antibody level to heat-stable Rs-antigens was increased up to 10 weeks after immersion but after 27 weeks was reduced to a level similar to that of the non-challenged fish . An anamnestic response was demonstrated in challenged fish, as intraperitoneal injection of heat-treated Rs bacteria into Rs challenged fish elicited a stronger humoral antibody response compared with injection into non-challenged fish.

Int J Gynecol Cancer, 2003 Mar-Apr, 13(2), 159 - 63
Is bacterial vaginosis associated with cervical intraepithelial neoplasia?
Boyle DC, Barton SE, Uthayakumar S, Hay PE, Pollock JW, Steer PJ, Smith JR.
Previous research has produced conflicting results regarding the association of bacterial vaginosis (BV) and cervical intraepithelial neoplasia (CIN) . These studies have been weakened in their conclusions mainly by failure to adequately control for the presence of sexually transmitted infections (STIs) . One proposed mechanism suggesting that carcinogenic nitrosamines acting either independently or via human papilloma virus (HPV) has not been fully tested previously . We undertook a prospective, case-controlled, cross-sectional study where the presence of STIs, in particular human papillomavirus (HPV) which is known to be associated with the development of CIN, was controlled for . Women with BV were not found to have CIN more frequently than women with normal vaginal flora and the quantities of nitrosamines produced by women with BV did not differ significantly from women without BV . We thus found that BV is not associated with CIN.

J Gen Virol, 2003 Apr, 84(Pt 4), 1013 - 20
In vitro cell-free conversion of bacterial recombinant PrP to PrPres as a model for conversion; Kirby L et al.; Prion diseases are associated with the conversion of the normal cellular prion protein, PrP(C), to the abnormal disease-associated protein, PrP(Sc) . This conversion can be mimicked in vitro using PrP(Sc) isolated from the brains of scrapie-infected animals to induce conversion of recombinant PrP(C) into a proteinase K-resistant isoform, PrP(res) . Traditionally, the 'cell-free' conversion assay has used, as substrate, recombinant PrP(C) purified from mammalian tissue culture cells or, more recently, from baculovirus-infected insect cells . The cell-free conversion assay has been modified by replacing the tissue culture-derived PrP(C) with recombinant PrP purified from bacteria . Bacterial expression and chromatographic purification give high yields of recombinant radiolabelled untagged protein, eliminates artefacts that may be due to cellular factors or antibody fragments normally present in labelled PrP preparations and allows accurate and rapid variation of protein sequence using standard molecular biological techniques . In addition, these cell-free conversion assays were carried out under more physiological conditions, giving more relevance to the assay as a model for conversion . To validate its use in this assay, this bacterial recombinant PrP has been shown to have the conversion properties of mammalian PrP(C): (i) it converts to a proteinase K-resistant isoform in the presence of PrP(Sc); (ii) the efficiency of this conversion by PrP(Sc) of different strains and species parallels that found in vivo; and (iii) its cell-free conversion is inhibited by Congo Red analogues in a structure-dependent manner similar to that seen in in vivo and in vitro cell assays.

Vector Borne Zoonotic Dis, 2001 Fall, 1(3), 197 - 210
Acarologic risk of exposure to emerging tick-borne bacterial pathogens in a semirural community in northern California; Lane RS et al.; An acarologic study was conducted in a semirural community in northern California to determine the relative abundance of, and the prevalence of infection with, three emerging bacterial pathogens in the western black-legged tick (Ixodes pacificus) . These included the agents causing Lyme disease (Borrelia burgdorferi), human granulocytic ehrlichiosis {Ehrlichia phagocytophila (formerly Ehrlichia equi)}, and human monocytic ehrlichiosis (Ehrlichia chaffeensis) . The study area in Sonoma County consisted of two properties each with four residents and an uninhabited adjacent comparison area . Six of the eight residents had been either physician-diagnosed or serodiagnosed previously with Lyme disease, and, of these, one also had been serodiagnosed with human monocytic ehrlichiosis . Direct immunofluorescent/culture assays and bacterial species-specific polymerase chain reaction assays were used to test whole ticks individually for presence of B . burgdorferi and Ehrlichia spp., respectively . Overall, 6.5% of the nymphal (n = 589) and 1.6% of the adult ticks (n = 318) from the same generational cohort were found to contain B . burgdorferi . In contrast, none of 465 nymphs and 9.9% of 202 adults were infected with E . phagocytophila . Excised tissues from another 95 adult ticks yielded a comparable E . phagocytophila infection prevalence of 13.7% . E . chaffeensis was not detected in either nymphal or adult ticks . Using a combination of culture and polymerase chain reaction assays, coinfection of I . pacificus adults with B . burgdorferi and E . phagocytophila was demonstrated for the first time . The marked disparity in the infection prevalence of these pathogens in nymphal and adult ticks suggests that their maintenance cycles are inherently different.

Curr Opin Cardiol, 2003 Mar, 18(2), 106 - 10
Diagnosis and management of bacterial endocarditis in 2003; Murtagh B et al.; The diagnosis of infective endocarditis has been notoriously difficult . Over the last decade, the modified Duke criteria have assumed an increasingly important role in the early detection of this often occult disease . Echocardiography has assumed increasing importance . Transesophageal echocardiography is recognized as more sensitive and specific than transthoracic echocardiography at detecting vegetations less than 10 mm in diameter . Vegetations greater than 10 mm in diameter are thought to be at increased risk of embolizing . Combined medical and surgical medical management result in the lowest mortality for those patients with hemodynamic compromise.

Microbes Infect, 2003 Feb, 5(2), 143 - 50
Helicobacter pylori CagA--a potential bacterial oncoprotein that functionally mimics the mammalian Gab family of adaptor proteins; Hatakeyama M; Helicobacter pylori virulence factor CagA is injected into gastric epithelial cells and undergoes tyrosine phosphorylation . Similar to mammalian Gab protein, tyrosine-phosphorylated CagA recruits and activates SHP-2 phosphatase at the plasma membrane, thereby inducing a growth factor-like effect . CagA-SHP-2 interaction may play an important role in bacterial pathogenesis, leading to gastric carcinoma.

Transgenic Res, 2003 Feb, 12(1), 123 - 6
Coexpression of the soybean vegetative storage protein beta subunit (S-VSPbeta) either with the bacterial feedback-insensitive dihydrodipicolinate synthase or with S-VSPalpha stabilizes the S-VSPbeta transgene protein and enhances lysine production in transgenic tobacco plants; Guenoune D et al.; Soybean vegetative storage proteins (S-VSPs) are lysine-rich leaf proteins, originally found to accumulate to high levels in depodded soybean plants . In the present study, we overexpressed S-VSPbeta, the ruminant stable subunit of the S-VSP genes, in transgenic tobacco plants . The S-VSPbeta protein accumulated in all organs studied, but its level declined drastically with leaf age . This instability of S-VSPbeta could be overcome either by elevating free lysine levels or by coexpressing S-VSPbeta with S-VSPbeta . High levels of rumen-stable, lysine-rich proteins is expected to improve absorption of lysine by ruminants . Furthermore, the expression of S-VSPs in heterologous plants led to a significant increase in total soluble lysine, suggesting that these proteins may also permit better assimilation of lysine by humans and monogastric animals.

Mol Plant Microbe Interact, 2003 Mar, 16(3), 217 - 25
Expression of the bacterial catalase genes during Sinorhizobium meliloti-Medicago sativa symbiosis and their crucial role during the infection process; Jamet A et al.; Sinorhizobium meliloti possesses three distinct catalases to cope with oxidative stress: two monofunctional catalases (KatA and KatC) and one bifunctional catalase-peroxydase (KatB) . The katB gene is constitutively expressed during growth in batch culture and is not induced under oxidative stress conditions . In contrast, the expression of katA and katC genes is mainly regulated at the transcription level in these conditions . A differential expression of kat genes was observed during the development of the nodule . A high expression of katA gene was detected in bacteroids, suggesting that the nitrogen-fixation process induces a strong oxidative stress . In contrast, bacteria express katB and katC genes and not the H2O2-inducible katA gene in infection threads despite the detection of H2O2 around the bacteria . A katB katC double mutant nodulated poorly and displayed abnormal infection . After nonefficient release into plant cells, bacteria failed to differentiate into bacteroids and rapidly underwent senescence . Our results indicate that these two catalases are essential for the establishment of the symbiosis . They also suggest that the bacteria are in a nonexponential growth phase in infection threads and corroborate previous studies on the growth rate of bacteria inside the plant.

J Biol Chem, 2003 Jun 13, 278(24), 21336 - 43 Epub 2003 Mar 20.
Detection of an intermediate during unfolding of bacterial cell division protein FtsZ: loss of functional properties precedes the global unfolding of FtsZ; Santra MK et al.; Using environment-sensitive fluorescence of 1-anilinonaphthalene-8-sulfonic acid, polarization of fluorescein 5'-isothiocyanate-labeled FtsZ, and far-UV circular dichroism spectroscopy, the chemical unfolding of FtsZ was found to proceed through two steps . The first step of the urea-induced unfolding produced an intermediate, which then unfolded at higher concentrations of urea . The intermediate state contains native-like secondary structure and much less tertiary structure compared with the native state . It is distinct from the native state as well as from the unfolded state . Similar to urea-induced unfolding of FtsZ, thermal unfolding of FtsZ also occurs in two steps . The midpoints for the first and second thermal unfolding transitions were found to be 38 +/- 4 and 77 +/- 5 degrees C, respectively . Further, the functional properties of FtsZ are extremely sensitive to urea, guanidium chloride, and sodium dodecyl sulfate . For example, 50% inhibition of the FtsZ assembly and GTP hydrolysis occurred at 0.1 and 0.2 m of urea, respectively . FtsZ lost its functional properties before any significant perturbation in the secondary or tertiary structure was detected by using several fluorescence techniques and far UV-CD indicating preferential local unfolding of the functional region(s) . In addition, the unfolded FtsZ regains its ability to polymerize fully upon removal of urea . The data taken together suggest that FtsZ unfolds reversibly through a multistep process, and local responses that inhibit functional properties precede the global transition of FtsZ to the unfolded state.

Infect Dis Obstet Gynecol, 2002, 10(4), 203 - 7
Comparison of Gram stain and Pap smear procedures in the diagnosis of bacterial vaginosis; Vardar E et al.; OBJECTIVE: The purpose of this study was to examine the characteristics of Gram stain versus Pap smear in diagnosis of bacterial vaginosis (BV) . METHODS: One-thousand and sixty women were enrolled in this study . All cases with symptoms of BV were determined by Amsel's criteria, which were accepted as the gold standard for diagnosis of BV . Pap smear and Gram stain evaluations were compared according to Amsel's criteria, without viewing the clinical results of the patients . Gram stain and Pap smear results were determined as negative or positive according to Amsel's criteria . Sensitivity, specifity and positive predictive values were calculated . RESULTS: After accepting the cases that were diagnosed as BV according to Amsel's criteria as reference cases, the sensitivity of the Gram stain method was calculated as 97% and the sensitivity of the Pap smear method as 93% . Similar specificity rates were obtained with both methods in diagnosis of BV related to the clinical results . There were no statistically significant differences in diagnosis of BV between these two groups . CONCLUSION: If Amsel's criteria are accepted as the gold standard for diagnosis of BV, Gram stain and Pap smear methods will give similar results in diagnosis.

Cell Mol Biol (Noisy-le-grand), 2002, 48 Online Pub, OL279 - 88
Inhibition of DNA replication by berenil of bacterial plasmids containing poly(dA)-poly(dT) sequences . 2D gel analysis of replicative intermediates; Snowden TE et al.; Berenil, an aromatic compound used in veterinary medicine to treat trypanosome infections in livestock, has been shown to interfere with kinetoplast DNA replication . The drug is thought to bind to the minor groove of DNA and form hydrogen bonds between opposite A/T pairs . Studies utilizing Trypanosoma cruzi, revealed that minicircle DNA, which is 60% A-T rich, and also the major component of kinetoplast DNA networks, is one of the targets for berenil . In order to better understand the mode of action of berenil and its effect on DNA replication, we have studied the effect of the drug on pBR322 derived plasmids containing poly(dA)-poly(dT) sequences . The resulting plasmids were pVL26, which contained 240 bp of poly(dA)-poly(dT) inserted at the EcoRV site of pBR322 and pKH47, which contained 100 bp of poly(dA)-poly(dT) inserted at the PvuII site of pBR322 . When cultures containing all of these plasmids were exposed to berenil, plasmids pVL26 and pKH47 were found to have significantly lower yields than pBR322, with pKH47 being the most sensitive to berenil . In the present study we show that the poly(dA)-poly(dT) sequences in plasmids pVL26 and pKH47 are not very stably maintained . However, the resulting deletion mutants containing a fraction of the poly(dA)-poly(dT) sequences were still sensitive to berenil . We also analyzed by 2D agarose gel electrophoresis the progression of the replication fork through the homopolymer region in plasmid pVL26d but failed to detect a replication barrier in this region in the presence of berenil.

Proc Natl Acad Sci U S A, 2003 Apr 1, 100(7), 4132 - 7 Epub 2003 Mar 17.
Impaired adrenocorticotropic hormone response to bacterial endotoxin in mice deficient in prostaglandin E receptor EP1 and EP3 subtypes; Matsuoka Y et al.; Sickness evokes various neural responses, one of which is activation of the hypothalamo-pituitary-adrenal (HPA) axis . This response can be induced experimentally by injection of bacterial lipopolysaccharide (LPS) or inflammatory cytokines such as IL-1 . Although prostaglandins (PGs) long have been implicated in LPS-induced HPA axis activation, the mechanism downstream of PGs remained unsettled . By using mice lacking each of the four PGE receptors (EP1-EP4) and an EP1-selective antagonist, ONO-8713, we showed that both EP1 and EP3 are required for adrenocorticotropic hormone release in response to LPS . Analysis of c-Fos expression as a marker for neuronal activity indicated that both EP1 and EP3 contribute to activation of neurons in the paraventricular nucleus of the hypothalamus (PVN) . This analysis also revealed that EP1, but not EP3, is involved in LPS-induced activation of the central nucleus of the amygdala . EP1 immunostaining in the PVN revealed its localization at synapses on corticotropin-releasing hormone-containing neurons . These findings suggest that EP1- and EP3-mediated neuronal pathways converge at corticotropin-releasing hormone-containing neurons in the PVN to induce HPA axis activation upon sickness.

Proc Natl Acad Sci U S A, 2003 Apr 1, 100(7), 4221 - 6 Epub 2003 Mar 17.
Ring-like pore structures of SecA: implication for bacterial protein-conducting channels; Wang HW et al.; SecA, an essential component of the general protein secretion pathway of bacteria, is present in Escherichia coli as soluble and membrane-integral forms . Here we show by electron microscopy that SecA assumes two characteristic forms in the presence of phospholipid monolayers: dumbbell-shaped elongated structures and ring-like pore structures . The ring-like pore structures with diameters of 8 nm and holes of 2 nm are found only in the presence of anionic phospholipids . These ring-like pore structures with larger 3- to 6-nm holes (without staining) were also observed by atomic force microscopic examination . They do not form in solution or in the presence of uncharged phosphatidylcholine . These ring-like phospholipid-induced pore-structures may form the core of bacterial protein-conducting channels through bacterial membranes.

Am Surg, 2003 Feb, 69(2), 131 - 4; discussion 134-5
Bacterial lipopolysaccharide-stimulated release of tumor necrosis factor-alpha from the isolated rat heart: the effect of aprotinin and forskolin; McKinney W et al.; Aprotinin has been reported to reduce plasma levels of inflammatory cytokines associated with cardiopulmonary bypass (CPB) . Because CPB is also associated with elevated levels of bacterial lipopolysaccharide (LPS) and LPS stimulates release of inflammatory cytokines from the heart we tested the hypothesis that aprotinin would inhibit cardiac release of tumor necrosis factor-alpha (TNF) provoked by LPS . Isolated rat hearts were perfused Langendorf style . After 30 minutes of equilibration LPS (100 ng/mL) was infused for 60 minutes . Timed samples of coronary effluent were collected at 0, 30, 60, 90, 120, and 150 minutes after the initiation of LPS for the measurement of coronary flow and the determination of TNF and cyclic AMP . Other hearts were perfused with buffer containing aprotinin {137 kallikrein-inhibiting units (KIU)/mL or 250 KIU/mL} and then infused with LPS . An additional group received forskolin (10 microM) and LPS . In hearts perfused as controls with buffer alone no TNF was detected in the coronary effluent . In hearts perfused with LPS TNF was reliably detected in the coronary effluent at 60 minutes (606 +/- 450 pg/min) and increased with time to a level of 1792 +/- 650 pg/min at 150 minutes . The addition of aprotinin had no significant effect on LPS-stimulated TNF release . For instance in hearts perfused with 137 KIU/mL aprotinin LPS-stimulated release at 150 minutes was 2141 +/- 732 pg/min and in hearts perfused with 250 KIU/mL LPS-stimulated TNF release was 2049 +/- 789 pg/min . Forskolin administration was associated with release of cyclic AMP from the heart and completely inhibited LPS-stimulated TNF release . We conclude that LPS stimulated release of TNF from the heart . Adding aprotinin to the perfusion buffer in either high or low concentrations did not attenuate LPS-stimulated cytokine release . Elevating myocardial cyclic AMP with forskolin completely attenuated LPS-stimulated TNF release.

J Gastroenterol, 2003, 38(2), 149 - 52
Interleukin 6 concentration in ascitic fluid of cirrhotic patients: relationship with previous episodes of spontaneous bacterial peritonitis; Souza MH et al.; BACKGROUND: We aimed to determine whether inter-leukin (IL)-6 concentrations in plasma and ascitic fluid from cirrhotic patients with previous episodes of spontaneous bacterial peritonitis (SBP) differred from those with no previous episodes of SBP . METHODS: Cirrhotic patients with ascites were divided into two groups: G1, without previous episodes of SBP (n = 6), and G2, with previous episodes of SBP (n = 6) . Patients with chronic heart failure (n = 5) with ascites, without hepatic diseases, were included as controls (G3) . IL-6 concentration was determined in ascitic fluid and plasma by enzyme-linked immunosorbent assay (ELISA) . A group of healthy subjects (n = 8) was used as a control for the blood IL-6 assay . RESULTS: IL-6 concentration in ascitic fluid was significantly higher in G1 (P < 0.05) than in G2 or G3 . No significant difference in plasma IL-6 concentrations was observed between the groups or between each group and the healthy subjects . A significant positive linear correlation (P < 0.002) was observed between IL-6 concentration and total protein in ascitic fluid . CONCLUSIONS: A low concentration of IL-6 in ascitic fluid could predispose cirrhotic patients to SBP.

Vaccine, 2003 Apr 2, 21(15), 1665 - 71
Immunization onto shaved skin with a bacterial enterotoxin adjuvant protects mice against respiratory syncytial virus (RSV); Godefroy S et al.; This study evaluated the potential of the skin as a non invasive route for RSV vaccination using two G protein-derived molecules, G2Na and G5 in mice . G2Na contains T and B-cell epitopes whether G5 is a pure B-cell epitope . In contrast to G5, G2Na coadministered with CT three times at 1 month interval onto 1cm of square area shaved skin, elicited a consistent serum anti-G2Na and anti-CT IgG response . The anti-G2Na IgG response was dominated by IgG1 isotype, an indirect marker of a Th2 type of response . Dramatic reduction and decrease of RSV titers in lung tissues and in the nasal tract, respectively, following intranasal virus challenge revealed biological relevance of the transcutaneous immunization in the context of RSV vaccine . These results suggest that the transcutaneous route may offer a promising potential for novel RSV vaccine strategy, simple, painless and economical.

J Biol Chem, 2003 May 16, 278(20), 18588 - 96 Epub 2003 Mar 10.
Crystal structure of Mycobacterium tuberculosis diaminopimelate decarboxylase, an essential enzyme in bacterial lysine biosynthesis; Gokulan K et al.; The Mycobacterium tuberculosis lysA gene encodes the enzyme meso-diaminopimelate decarboxylase (DAPDC), a pyridoxal-5'-phosphate (PLP)-dependent enzyme . The enzyme catalyzes the final step in the lysine biosynthetic pathway converting meso-diaminopimelic acid (DAP) to l-lysine . The lysA gene of M . tuberculosis H37Rv has been established as essential for bacterial survival in immunocompromised mice, demonstrating that de novo biosynthesis of lysine is essential for in vivo viability . Drugs targeted against DAPDC could be efficient anti-tuberculosis drugs, and the three-dimensional structure of DAPDC from M . tuberculosis complexed with reaction product lysine and the ternary complex with PLP and lysine in the active site has been determined . The first structure of a DAPDC confirms its classification as a fold type III PLP-dependent enzyme . The structure shows a stable 2-fold dimer in head-to-tail arrangement of a triose-phosphate isomerase (TIM) barrel-like alpha/beta domain and a C-terminal beta sheet domain, similar to the ornithine decarboxylase (ODC) fold family . PLP is covalently bound via an internal aldimine, and residues from both domains and both subunits contribute to the binding pocket . Comparison of the structure with eukaryotic ODCs, in particular with a di-fluoromethyl ornithine (DMFO)-bound ODC from Trypanosoma bruceii, indicates that corresponding DAP-analogues might be potential inhibitors for mycobacterial DAPDCs.

Environ Sci Technol, 2003 Feb 15, 37(4), 781 - 5
Analysis of bacterial random motility in a porous medium using magnetic resonance imaging and immunomagnetic labeling; Sherwood JL et al.; In this study, we demonstrate the application of immunomagnetic labeling and magnetic resonance imaging (MRI) for the noninvasive visualization of changes in bacterial density distributions as a function of time in a water-saturated porous medium . Magnetite particles (50-60 nm diameter) were attached via a monoclonal antibody to the surface' of Escherichia coli K12 NR50 cells . The cells maintained their motility after labeling, and the presence of the magnetite did not significantly alter cell swimming speed . Diffusive migration for both motile and nonmotile E . coli through a porous medium with a particle-diameter distribution of 250-300 microm was compared . The movement of the nonmotile cells was described by an effective random motility coefficient consistent with Brownian diffusion of a nonmotile colloid . An effective coefficient determined a priori from bacterial motility in an aqueous medium and properties of the porous medium adequately described the movement of the motile cells . The ability to noninvasively visualize bacterial concentrations within an opaque porous medium in real time provides researchers with a powerful tool for studying bacterial transport in porous media . This is important for understanding the impact of bacterial transport on remediation strategies for environmental cleanup of polluted groundwater.

Microbiology, 2003 Mar, 149(Pt 3), 547 - 56
Moving folded proteins across the bacterial cell membrane; Palmer T et al.; The Tat protein export system is located in the bacterial cytoplasmic membrane and operates in parallel to the well-known Sec pathway . While the Sec system only transports unstructured substrates, the function of the Tat pathway is to translocate folded proteins . The Tat translocase thus faces the formidable challenge of moving structured macromolecular substrates across the bacterial cytoplasmic membrane without rendering the membrane freely permeable to protons and other ions . The substrates of the Tat pathway are often proteins that bind cofactor molecules in the cytoplasm, and are thus folded, prior to export . Such periplasmic cofactor-containing proteins are essential for most types of bacterial respiratory and photosynthetic energy metabolism . In addition, the Tat pathway is involved in outer membrane biosynthesis and in bacterial pathogenesis . Substrates are targeted to the Tat pathway by amino-terminal signal sequences harbouring consecutive, essentially invariant, arginine residues, and movement of proteins through the Tat system is energized by the transmembrane proton electrochemical gradient . The TatA protein probably forms the transport channel while the TatBC proteins act as a receptor complex that recognizes the signal peptide of the substrate protein.

Trends Biochem Sci, 2003 Mar, 28(3), 121 - 4
The NIT domain: a predicted nitrate-responsive module in bacterial sensory receptors; Shu CJ et al.; A nitrate- and nitrite-sensing (NIT) domain as found in the NasR protein, has been detected in various receptor components of signal transduction pathways in different bacterial lineages . Cellular functions controlled by receptors that contain this novel domain include regulation of gene expression (transcription anti-terminators and histidine kinases), cell motility (chemotaxis receptors) and enzyme activity (diguanylate cyclases and phosphodiesterases).

Proc Natl Acad Sci U S A, 2003 Mar 18, 100(6), 3027 - 30 Epub 2003 Mar 11.
Type III secretion systems and bacterial flagella: insights into their function from structural similarities; Blocker A et al.; Type III secretion systems and bacterial flagella are broadly compared at the level of their genetic structure, morphology, regulation, and function, integrating structural information, to provide an overview of how they might function at a molecular level.

Eur J Biochem, 2003 Mar, 270(6), 1222 - 30
Disease-related mutations in cytochrome c oxidase studied in yeast and bacterial models; Bratton M et al.; Mitochondrial cytochrome c oxidase is a key protonmotive component of the respiratory chain . Mutations in the mitochondrially-encoded subunits of the complex have been reported in association with a range of diseases . In this work we used yeast and bacterial mutants to assess the effect of human mutations in subunit 1 (L196I) and subunit 3 (G78S, A200T, Delta F94-F98, F251L and W249Stop) . While the stop mutation at the C-terminus of subunit 3 and the short deletion were highly deleterious and abolished the assembly of the mitochondrial enzyme, the four missense mutations caused little or no effect on the respiratory function . Detailed analysis of G78S, A200T and Delta F94-F98 in Rhodobacter sphaeroides confirmed and extended these observations . We show in this study that the combination of yeast and bacterial models is a useful tool to elucidate the effect of mutations in the catalytic core of cytochrome oxidase . The yeast enzyme is highly similar to the human enzyme and provides a good model to assess the deleterious effect of reported mutations . The bacterial system allows detailed biochemical analysis of the effect of the mutations on the function and assembly of the catalytic core of the enzyme.

J Exp Med, 2003 Mar 17, 197(6), 735 - 42 Epub 2003 Mar 10.
Role of adhesin release for mucosal colonization by a bacterial pathogen; Coutte L et al.; Pathogen attachment is a crucial early step in mucosal infections . This step is mediated by important virulence factors called adhesins . To exert these functions, adhesins are typically surface-exposed, although, surprisingly, some are also released into the extracellular milieu, the relevance of which has previously not been studied . To address the role of adhesin release in pathogenesis, we used Bordetella pertussis as a model, since its major adhesin, filamentous hemagglutinin (FHA), partitions between the bacterial surface and the extracellular milieu . FHA release depends on its maturation by the specific B . pertussis protease SphB1 . We constructed SphB1-deficient mutants and found that they were strongly affected in their ability to colonize the mouse respiratory tract, although they adhered even better to host cells in vitro than their wild-type parent strain . The defect in colonization could be overcome by prior nasal instillation of purified FHA or by coinfection with FHA-releasing B . pertussis strains, but not with SphB1-producing FHA-deficient strains, ruling out a nonspecific effect of SphB1 . These results indicate that the release of FHA is important for colonization, as it may facilitate the dispersal of bacteria from microcolonies and the binding to new sites in the respiratory tract.

J Mol Biol, 2003 Mar 21, 327(2), 309 - 15
Locking TolC entrance helices to prevent protein translocation by the bacterial type I export apparatus; Eswaran J et al.; The periplasmic entrance of the TolC channel tunnel is sealed by close-packing of inner and outer coiled-coils, and it has been proposed that opening of the entrance is achieved by an iris-like realignment of the inner coiled-coils . This is supported by experimental disruption of the key links connecting them, which effects transition to the open state in TolC inserted into planar lipid bilayers . Here we provide in vivo evidence for this "twist to open" mechanism by constraining the coiled coils with disulphide bonds, either self-locking or bridged by a chemical cross-linker, and reconstituting the resulting TolC variants into the type I protein export system in Escherichia coli . Introducing an intermonomer disulphide bridge between Ala159 and Ser350 caused a fivefold reduction in export, and when the coiled coils were cross-linked at the entrance constriction, between Asp374 of adjacent monomers or between Asn156 and Ala375, TolC-dependent export was abolished . In vivo cross-linking showed that the locked non-exporting TolC variants were still recruited to assemble the type I export apparatus . The data show that untwisting the entrance helices is essential for the export function of TolC in E.coli, specifically to allow access and passage of substrates engaged at the inner membrane translocase.

Biochemistry, 2003 Mar 18, 42(10), 3105 - 12
Demonstration of two independently folding domains in the alpha subunit of bacterial luciferase by preferential ligand binding-induced stabilization; Noland BW et al.; The alpha subunit of bacterial luciferase unfolds and refolds reversibly by a three-state mechanism in urea-containing buffer . It has been proposed that the three-state unfolding of the alpha subunit arises from a stepwise unfolding of a C-terminal folding domain at lower concentrations of urea, followed by unfolding of the N-terminal domain at higher concentrations of urea (Noland, B . W., Dangott, L . J., and Baldwin, T . O . (1999) Biochemistry 38, 16136-16145) . The location of an anion binding site in the proposed N-terminal folding domain allowed the folding mechanism to be probed in the context of the intact polypeptide . Anions preferentially stabilized the N-terminal domain in a concentration-dependent manner . The polyvalent anions sulfate and phosphate were found to be more stabilizing than monovalent chloride ion . Cations did not show a similar stabilizing effect, demonstrating that the stabilization was due to the anions alone . The purified N-terminal domain prepared by limited proteolysis and anion exchange chromatography was found to refold cooperatively with a midpoint approximately that of the second unfolding transition of the alpha subunit . Phosphate ion stabilized this fragment to roughly the same extent as it did the alpha subunit . The results presented are consistent with the proposed two-domain folding model and demonstrate that anion binding to the N-terminal folding domain stabilizes the alpha subunit of bacterial luciferase.

Nat Biotechnol, 2003 Apr, 21(4), 443 - 7 Epub 2003 Mar 10.
Engineering the mouse genome with bacterial artificial chromosomes to create multipurpose alleles; Testa G et al.; The mouse is the leading vertebrate model because its genome can be altered by both random transgenesis and homologous recombination with targeting constructs . Both approaches have been hindered by the size and site limitations implicit in conventional Escherichia coli DNA-engineering methods . Homologous recombination in E . coli, or 'recombineering', has overcome these limitations for bacterial artificial chromosome (BAC) transgenesis . Here we applied Red/ET recombineering (using the lambda Redalpha/Redbeta recombinase pair) to generate a 64 kilobase targeting construct that carried two selectable cassettes permitting the simultaneous mutation of the target gene, Mll, at sites 43 kb apart in one round of mouse embryonic stem (ES) cell targeting . The targeting frequency after dual selection was 6% . Because the two selectable cassettes were flanked by FRT or loxP sites, three more alleles can be generated by site-specific recombination . Our approach represents a simple way to introduce changes at two or more sites in a genetic locus, and thereafter generate allele combinations . The size of BAC templates offers new freedom for the design of targeting constructs . Combined with the use of two selectable cassettes placed far apart, BAC-based targeting constructs may be applicable to tasks such as regional exchanges, deletions, and insertions.

Nat Biotechnol, 2003 Apr, 21(4), 447 - 51 Epub 2003 Mar 10.
Site-specific gene targeting in mouse embryonic stem cells with intact bacterial artificial chromosomes; Yang Y et al.; Homologous recombination in Escherichia coli simplifies the generation of gene targeting constructs for transduction into mouse embryonic stem (ES) cells . Taking advantage of the extensive homology provided by intact bacterial artificial chromosomes (BACs), we have developed an efficient method for preparing targeted gene disruptions in ES cells . Correctly integrated clones were identified by a simple screening procedure based on chromosomal fluorescence in situ hybridization (FISH) . To date, five mutant lines have been generated and bred to homozygosity by this approach.

Crit Care Med, 2003 Mar, 31(3), 729 - 37
Bacterial colonization of the respiratory tract following tracheal intubation-effect of gravity: an experimental study; Panigada M et al.; OBJECTIVE: To explore the role of the horizontal orientation of endotracheal tube and neck on bacterial colonization of the respiratory tract in anesthetized sheep on mechanical ventilation, without use of antibiotics . DESIGN: Prospective animal study . SETTING: National Institutes of Health research laboratory . SUBJECTS: Anesthetized, paralyzed, and ventilated sheep . INTERVENTIONS: Sheep were randomized into five groups and managed as follows: Group IS contained sheep that were not intubated and were immediately killed . Group HU4 contained six sheep that were mechanically ventilated for 4 hrs, with head and endotracheal tube elevated 30 degrees from horizontal . Group HU72 contained seven sheep that were prone, mechanically ventilated for 72 hrs, and managed the same as group HU4 . Groups G and Gf each contained seven sheep that were prone on a lateral body rotation device, mechanically ventilated for 72 hrs, with neck and endotracheal tube horizontal . Group Gf received nasogastric enteral feeding . MEASUREMENTS AND MAIN RESULTS: At the end of the study, sheep were examined postmortem, and a total of 11 tissue samples were taken from the trachea, the five lobar bronchi, and the five lobar parenchyma, for qualitative and quantitative culture . Group HU72 had significant decrease in Pao2/Fio2 and heavy bacterial colonization in all sheep . Groups G and Gf retained excellent lung function; lung bacterial colonization was no different from the IS group . CONCLUSIONS: The horizontal orientation of the endotracheal tube and neck, through lateral body rotation, showed no altered airway colonization and maintained excellent gas exchange and lung function in our animal model.

Nucleic Acids Res, 2003 Mar 15, 31(6), 1780 - 9
ZCURVE: a new system for recognizing protein-coding genes in bacterial and archaeal genomes; Guo FB et al.; A new system, ZCURVE 1.0, for finding protein- coding genes in bacterial and archaeal genomes has been proposed . The current algorithm, which is based on the Z curve representation of the DNA sequences, lays stress on the global statistical features of protein-coding genes by taking the frequencies of bases at three codon positions into account . In ZCURVE 1.0, since only 33 parameters are used to characterize the coding sequences, it gives better consideration to both typical and atypical cases, whereas in Markov-model-based methods, e.g . Glimmer 2.02, thousands of parameters are trained, which may result in less adaptability . To compare the performance of the new system with that of Glimmer 2.02, both systems were run, respectively, for 18 genomes not annotated by the Glimmer system . Comparisons were also performed for predicting some function-known genes by both systems . Consequently, the average accuracy of both systems is well matched; however, ZCURVE 1.0 has more accurate gene start prediction, lower additional prediction rate and higher accuracy for the prediction of horizontally transferred genes . It is shown that the joint applications of both systems greatly improve gene-finding results . For a typical genome, e.g . Escherichia coli, the system ZCURVE 1.0 takes approximately 2 min on a Pentium III 866 PC without any human intervention . The system ZCURVE 1.0 is freely available at: tju.edu.cn/Zcurve_B/.

Nucleic Acids Res, 2003 Mar 15, 31(6), 1656 - 64
Targeted and random bacterial gene disruption using a group II intron (targetron) vector containing a retrotransposition-activated selectable marker; Zhong J et al.; Mobile group II introns have been used to develop a novel class of gene targeting vectors, targetrons, which employ base pairing for DNA target recognition and can thus be programmed to insert into any desired target DNA . Here, we have developed a targetron containing a retrotransposition-activated selectable marker (RAM), which enables one-step bacterial gene disruption at near 100% efficiency after selection . The targetron can be generated via PCR without cloning, and after intron integration, the marker gene can be excised by recombination between flanking Flp recombinase sites, enabling multiple sequential disruptions . We also show that a RAM-targetron with randomized target site recognition sequences yields single insertions throughout the Escherichia coli genome, creating a gene knockout library . Analysis of the randomly selected insertion sites provides further insight into group II intron target site recognition rules . It also suggests that a subset of retrohoming events may occur by using a primer generated during DNA replication, and reveals a previously unsuspected bias for group II intron insertion near the chromosome replication origin . This insertional bias likely reflects at least in part the higher copy number of origin proximal genes, but interaction with the replication machinery or other features of DNA structure or packaging may also contribute.

Clin Diagn Lab Immunol, 2003 Mar, 10(2), 286 - 92
Constitutive expression of a bacterial pattern recognition receptor, CD14, in human salivary glands and secretion as a soluble form in saliva; Uehara A et al.; Saliva contains a number of proteins and glycoproteins that protect oral tissues, but little is known about the role of human saliva in innate immunity . Here we showed that human major salivary gland cells constitutively expressed a bacterial pattern recognition receptor, CD14, by immunohistochemistry . Human salivary gland cells in culture express CD14 mRNA and a 55-kDa CD14 protein in, but not on the cells, and secrete a soluble form with the same molecular mass . Human whole saliva contains a 55-kDa CD14, and the concentration of parotid saliva was 10-fold higher than whole saliva, which is comparable to that of serum CD14 . Levels of CD14 in unstimulated whole and parotid saliva were unchanged before and after a meal and between unstimulated and stimulated saliva, indicating that saliva CD14 is constitutively secreted into the oral cavity . In contrast, lipopolysaccharide (LPS)-binding protein was below the detectable level . The saliva CD14 is functionally active in that it mediated the activation of CD14-lacking intestinal epithelial cells by LPS in a Toll-like receptor 4-dependent manner . These results suggested that saliva CD14 is important for the maintenance of oral health and possibly intestinal homeostasis.

Microbiology, 2003 Feb, 149(Pt 2), 363 - 8
Studies on distant regulation of bacterial growth and light emission; Trushin MV; Reciprocal interactions of two Escherichia coli MC1061 cultures separated by a glass window were investigated . The growth parameters and light emission from these cultures were analysed . A link between light emission and the growth parameters was observed.

J Clin Microbiol, 2003 Mar, 41(3), 960 - 6
Interaction between porcine reproductive-respiratory syndrome virus and bacterial endotoxin in the lungs of pigs: potentiation of cytokine production and respiratory disease; van Gucht S et al.; Porcine reproductive-respiratory syndrome virus (PRRSV) is a key agent in multifactorial respiratory disease of swine . Intratracheal administration of bacterial lipopolysaccharides (LPSs) to PRRSV-infected pigs results in markedly enhanced respiratory disease, whereas the inoculation of each component alone results in largely subclinical disease . This study examines whether PRRSV-LPS-induced respiratory disease is associated with the excessive production of proinflammatory cytokines in the lungs . Gnotobiotic pigs were inoculated intratracheally with PRRSV and then with LPS at 3, 5, 7, 10, or 14 days of infection and euthanatized 6 h after LPS inoculation . Controls were inoculated with PRRSV or LPS only or with phosphate-buffered saline . Virus titers, (histo)pathological changes in the lungs, numbers of inflammatory cells, and bioactive tumor necrosis factor alpha (TNF-alpha), interleukin-1 (IL-1), and IL-6 levels in bronchoalveolar lavage fluids were examined . All pigs inoculated with PRRSV-LPS developed severe respiratory disease, whereas the controls that were inoculated with PRRSV or LPS alone did not . PRRSV infection significantly enhanced cytokine production in response to LPS . Peak TNF-alpha, IL-1, and IL-6 titers were 10 to 100 times higher in the PRRSV-LPS-inoculated pigs than in the pigs inoculated with PRRSV or LPS alone; and the titers correlated with the respiratory signs . The levels of neutrophil infiltration and the pathological changes detected in the lungs of PRRSV-LPS-inoculated pigs resembled those detected when the effects of PRRSV and LPS inoculated alone are combined, but with no synergistic effects between PRRSV and LPS . These data demonstrate a synergism between PRRSV and LPS in the induction of proinflammatory cytokines and an association between induction of these cytokines and disease.

J Biol Chem, 2003 May 9, 278(19), 16834 - 43 Epub 2003 Mar 05.
Essential steps in the ppGpp-dependent regulation of bacterial ribosomal RNA promoters can be explained by substrate competition; Jores L et al.; Transcription of stable RNA genes is known to be dramatically reduced in the presence of guanosine tetraphosphate (ppGpp), the mediator of the stringent response . Using in vitro transcription systems with ribosomal RNA P1 promoters, we have analyzed which step of the initiation cycle is inhibited by the effector ppGpp . We show that formation of the ternary transcription initiation complex consisting of RNA polymerase holoenzyme, the promoter DNA, and the first initiating nucleotide triphosphate is the major step at which ppGpp exerts its regulation . Neither primary binding of RNA polymerase to the promoter nor isomerization to the open binary complexes or the subsequent promoter clearance steps contributes notably to the observed inhibition . The effect of ppGpp-dependent inhibition in the formation of the ternary transcription initiation complex could be mimicked by nucleotide derivatives known to bind to the RNA polymerase active center . Using these model compounds, almost identical inhibition characteristics were observed as seen with ppGpp . The results support the previously published model, which suggests that ppGpp-dependent inhibition is based on competition between the inhibitor molecules and NTP substrates for access to the active center of RNA polymerase.

Appl Environ Microbiol, 2003 Mar, 69(3), 1800 - 9
Soil type is the primary determinant of the composition of the total and active bacterial communities in arable soils; Girvan MS et al.; Degradation of agricultural land and the resulting loss of soil biodiversity and productivity are of great concern . Land-use management practices can be used to ameliorate such degradation . The soil bacterial communities at three separate arable farms in eastern England, with different farm management practices, were investigated by using a polyphasic approach combining traditional soil analyses, physiological analysis, and nucleic acid profiling . Organic farming did not necessarily result in elevated organic matter levels; instead, a strong association with increased nitrate availability was apparent . Ordination of the physiological (BIOLOG) data separated the soil bacterial communities into two clusters, determined by soil type . Denaturing gradient gel electrophoresis and terminal restriction fragment length polymorphism analyses of 16S ribosomal DNA identified three bacterial communities largely on the basis of soil type but with discrimination for pea cropping . Five fields from geographically distinct soils, with different cropping regimens, produced highly similar profiles . The active communities (16S rRNA) were further discriminated by farm location and, to some degree, by land-use practices . The results of this investigation indicated that soil type was the key factor determining bacterial community composition in these arable soils . Leguminous crops on particular soil types had a positive effect upon organic matter levels and resulted in small changes in the active bacterial population . The active population was therefore more indicative of short-term management changes.

Environ Monit Assess, 2003 Jan-Feb, 81(1-3), 289 - 300
Characterization and statistical modeling of bacterial (Escherichia coli) outflows from watersheds that discharge into southern Lake Michigan; Olyphant GA et al.; Two watersheds in northwestern Indiana were selected for detailed monitoring of bacterially contaminated discharges (Escherichia coli) into Lake Michigan . A large watershed that drains an urbanized area with treatment plants that release raw sewage during storms discharges into Lake Michigan at the outlet of Burns Ditch . A small watershed drains part of the Great Marsh, a wetland complex that has been disrupted by ditching and limited residential development, at the outlet of Derby Ditch . Monitoring at the outlet of Burns Ditch in 1999 and 2000 indicated that E . coli concentrations vary over two orders of magnitude during storms . During one storm, sewage overflows caused concentrations to increase to more than 10,000 cfu/100 mL for several hours . Monitoring at Derby Ditch from 1997 to 2000 also indicated that E . coli concentrations increase during storms with the highest concentrations generally occurring during rising streamflow . Multiple regression analysis indicated that 60% of the variability in measured outflows of E . coli from Derby Ditch (n = 88) could be accounted for by a model that utilizes continuously measured rainfall, stream discharge, soil temperature and depth to water table in the Great Marsh . A similar analysis indicated that 90% of the variability in measured E . coli concentrations at the outlet of Burns Ditch (n = 43) during storms could be accounted for by a combination of continuously measured water-quality variables including nitrate and ammonium . These models, which utilize data that can be collected on a real-time basis, could form part of an Early Warning System for predicting beach closures.

Mol Cell Biochem, 2003 Jan, 243(1-2), 15 - 22
Single amino acid substitution enhances bacterial expression of PARP-4D214A; Gagnon SN et al.; Poly(ADP-ribose) polymerase-1 (PARP-1) is the canonical member of the PARP family of enzymes and modulates many crucial nuclear functions . PARP-1 is involved in apoptosis and is the substrate of caspase-3, a protease that cleaves PARP-1 at the conserved sequence 211DEVD214 . To generate a caspase-3-uncleavable PARP-1, we introduced an amino acid substitution D214-->A214 at the site of cleavage . We observed that following over-expression in bacteria, the mutant protein HIS-PARP-1D214A was expressed several-fold more than a unmutated copy, HIS-PARP-1 . The specific activity of HIS-PARP-1 enzyme in total bacterial extracts was 6.94 U/mg and 4.61 U/mg for HIS-PARP-1D214A . This approach should provide new avenues for crystallographic study of PARP-1 as well as new information for drug design targeting PARP-1.

Mol Cell Biochem, 2003 Jan, 242(1-2), 65 - 70
Cloning and bacterial expression of postnatal mouse heart FGF-16; Sontag DP et al.; Fibroblast growth factor-16 (FGF-16) has been reported as the sixteenth member of the heparin sulphate proteoglycan binding growth factor family, which includes acidic and basic FGFs (FGF-1 and FGF-2), based on sequence similarity . The sequences of human (h) and rat (r) FGF-16 complimentary DNA (cDNA) sequences are known . Rat FGF-16 is expressed in brown adipose tissue during embryonic development but also shows some specificity for the postnatal heart . In spite of the importance of other FGF family members in cardiac physiology, there is scant information about FGF-16 function . As a first step towards exploiting mouse genetics in this regard, we have used reverse transcriptase-polymerase chain reaction and primers based on the rFGF-16 sequence to clone the adult mouse (m) FGF-16 cDNA . An mFGF-16 cDNA of 624 base pairs was generated . Based on sequence analysis, mFGF-16 and hFGF-16 share at least 95.2 and 99% nucleotide and amino acid similarity, respectively . In terms of other family members, FGF-16 is most closely related to FGF-9 . When used as a radiolabeled probe, the mFGF-16 cDNA detected a single 1.8 kilobase transcript in adult mouse heart RNA . The mFGF-16 cDNA was also used to generate an amino-terminal poly-histidine tagged FGF-16 protein in bacteria . Using sodium dodecyl sulphate-polyacrylamide gel electrophoresis and taking into account the poly-histidine tag, an FGF-16 protein of 26.3 kDa was detected . The generation of cardiac mFGF-16 cDNA and a purified FGF-16 protein preparation are seen as important tools in the further characterization of FGF-16 expression and function in the mammalian heart.

Dev Dyn, 2003 Mar, 226(3), 439 - 45
Development of transgenic chickens expressing bacterial beta-galactosidase; Mozdziak PE et al.; Replication-defective retroviral vectors are efficient vehicles for the delivery of exogenous genes, and they may be used in the generation of transgenic animals . The replication-defective retroviral SNTZ vector carrying the lacZ gene with a nuclear localized signal was injected into the subgerminal cavity of freshly laid eggs . Subsequently, the eggs were allowed to hatch, and the chickens were screened for the lacZ gene by using the polymerase chain reaction . Eight of 15 male chickens that survived to sexual maturity contained the lacZ gene in their semen . Subsequently, these males were mated with wild-type female chickens . From one of the eight lacZ-positive G(0) males, two lacZ-positive male chickens were produced from a total of 224 G(1) progeny for a germline transmission rate of 0.89% . Both G(1) male chickens carrying the lacZ gene were mated with wild-type female chickens and 46.5% of the G(2) progeny contained the lacZ gene, which is consistent with the expected Mendelian 50% ratio for a heterozygous dominant allele . The product of the lacZ gene, nuclear localized beta-galactosidase, was expressed in primary myoblast cultures derived from G(2) chickens, and it was also expressed in whole G(2) chicken embryos .

J Bacteriol, 2003 Mar, 185(6), 1987 - 94
Mass distribution and spatial organization of the linear bacterial motor of Spiroplasma citri R8A2; Trachtenberg S et al.; In the simple, helical, wall-less bacterial genus Spiroplasma, chemotaxis and motility are effected by a linear, contractile motor arranged as a flat cytoskeletal ribbon attached to the inner side of the membrane along the shortest helical line . With scanning transmission electron microscopy and diffraction analysis, we determined the hierarchical and spatial organization of the cytoskeleton of Spiroplasma citri R8A2 . The structural unit appears to be a fibril, approximately 5 nm wide, composed of dimers of a 59-kDa protein; each ribbon is assembled from seven fibril pairs . The functional unit of the intact ribbon is a pair of aligned fibrils, along which pairs of dimers form tetrameric ring-like repeats . On average, isolated and purified ribbons contain 14 fibrils or seven well-aligned fibril pairs, which are the same structures observed in the intact cell . Scanning transmission electron microscopy mass analysis and sodium dodecyl sulfate-polyacrylamide gel electrophoresis of purified cytoskeletons indicate that the 59-kDa protein is the only constituent of the ribbons.

J Biol Chem, 2003 May 9, 278(19), 16857 - 62 Epub 2003 Mar 03.
37-kDa laminin receptor precursor modulates cytotoxic necrotizing factor 1-mediated RhoA activation and bacterial uptake; Chung JW et al.; Cytotoxic necrotizing factor 1 (CNF1) is a bacterial toxin known to activate Rho GTPases and induce host cell cytoskeleton rearrangements . The constitutive activation of Rho GTPases by CNF1 is shown to enhance bacterial uptake in epithelial cells and human brain microvascular endothelial cells . However, it is unknown how exogenous CNF1 exhibits such phenotypes in eukaryotic cells . Here, we identified 37-kDa laminin receptor precursor (LRP) as the receptor for CNF1 from screening the cDNA library of human brain microvascular endothelial cells by the yeast two-hybrid system using the N-terminal domain of CNF1 as bait . CNF1-mediated RhoA activation and bacterial uptake were inhibited by exogenous LRP or LRP antisense oligodeoxynucleotides, whereas they were increased in LRP-overexpressing cells . These findings indicate that the CNF1 interaction with LRP is the initial step required for CNF1-mediated RhoA activation and bacterial uptake in eukaryotic cells.

J Biotechnol, 2003 Mar 20, 101(3), 219 - 28
DNA extraction using bacterial magnetic particles modified with hyperbranched polyamidoamine dendrimer; Yoza B et al.; A cascading hyperbranched polyamidoamine dendrimer was synthesized on the surface of bacterial magnetite from Magnetospirillum magneticum AMB-1 to allow enhanced extraction of DNA from fluid suspensions . Characterization of the synthesis revealed linear doubling of the surface amine charge from generations one through five starting with an amino silane initiator . Furthermore, transmission electron microscopy revealed clear dispersion of the single domain magnetite in aqueous solution . The dendrimer modified magnetic particles have been used to carry out magnetic separation of DNA . Binding and release efficiencies increased with the number of generations and those of bacterial magnetite modified with six generation dendrimer were 7 and 11 times respectively as many as those of bacterial magnetite modified with only amino silane.

Chem Commun (Camb), 2003 Feb 7, (3), 384 - 5
Conformational studies on phenyl thioglycosides: a remote effect on disaccharide linkage by phenyl aglycons attenuates recognition of galabiosides by a bacterial adhesin; Ohlsson J et al.; Phenyl S-galabiosides display altered conformational properties, as compared to phenyl O-galabiosides, characterised by a remote effect on the galabiose intersaccharidic glycoside bond by the phenyl aglycon, resulting in significantly lowered affinity for the PapG class II adhesin of uropathogenic E . coli.

Arch Cardiol Mex, 2002 Oct-Dec, 72(4), 303 - 5
{Non-bacterial thrombotic endocarditis as paraneoplastic manifestation of pulmonary adenocarcinoma}; Kraft F et al.; Non bacterial thrombotic endocarditis is characterized by the presence of non infected vegetation in aortic or mitral valves associated with systemic arterial emboli . Non-bacterial thrombotic endocarditis is a common complication of neoplastic diseases: adenocarcinoma of the lung, another adenocarcinomas, myeloma, lymphoma, leukemia, carcinoma of the pancreas, breast, cervix, colon and stomach . We report a case of non-bacterial thrombotic endocarditis localized in the aortic and mitral valves and systemic emboli as the first manifestation of adenocarcinoma of the lung.

Obstet Gynecol Surv, 2003 Mar, 58(3), 209 - 20
Antenatal causes of cerebral palsy: associations between inherited thrombophilias, viral and bacterial infection, and inherited susceptibility to infection; Gibson CS et al.; Cerebral palsy rates of 2 in every 1,000 births have varied little over the last 40 years, despite improvements in obstetric care . In the past, cerebral palsy was thought to be due to poor obstetric care and management; however, epidemiological studies have refuted this, suggesting that there is usually an antenatal timing to the neuropathology of cerebral palsy . There are many known risk factors for cerebral palsy, including multiple gestation, prematurity, and low birth weight . Recently, intrauterine infection, maternal pyrexia, and the presence of thrombophilic disorders (thrombophilia) have been identified as major risk factors for subsequent cerebral palsy . This review examines the links between intrauterine infection, the fetal inflammatory response, and thrombophilia as possible causes of cerebral palsy . The interactions of viral or bacterial infections during pregnancy, normal or abnormal fetal cytokine responses, and hereditary fetal thrombophilias as antenatal causes of the neuropathology of cerebral palsy are now areas of research priority . TARGET AUDIENCE: Obstetricians & Gynecologists, Family Physicians LEARNING OBJECTIVES: After completion of this article, the reader will be able to describe the condition cerebral palsy, list the risk factors for the development of cerebral palsy, outline the ultrasound findings associated with cerebral palsy, and point out other conditions associated with cerebral palsy.

Rev Med Chil, 2002 Dec, 130(12), 1329 - 34
{Endogenous ethanol production in patients with liver cirrhosis, motor alteration and bacterial overgrowth}; Madrid AM et al.; BACKGROUND: Small intestinal bacterial overgrowth generates endogenous ethanol production both in experimental animals and humans . Patients with cirrhosis have small intestinal bacterial overgrowth, but endogenous ethanol production has not been studied in them . AIM: To investigate endogenous ethanol production in patients with cirrhosis, altered intestinal motility and small intestinal bacterial overgrowth . PATIENTS AND METHODS: Eight patients with cirrhosis of different etiologies and altered gastrointestinal motility, consisting in changes in the migrating motor complex, were studied . All had also small intestinal bacterial overgrowth, measured by means of the H2 breath test with lactulose . Plasma ethanol levels were measured by gas liquid chromatography in fasting conditions and 120 min after a carbohydrate rich meal . RESULTS: In fasting conditions, no patient had endogenous ethanol production . Alter the meal, ethanol in concentrations of 11.3 and 8.2 mg/del were detected in two patients . Negligible amounts of ethanol were detected in 4 patients and two patients had undetectable alcohol levels . CONCLUSIONS: A low endogenous production of ethanol was demonstrated in six of eight patients with cirrhosis.

Eur J Gastroenterol Hepatol, 2003 Mar, 15(3), 267 - 73
Ca2+ response in neutrophils after exposure to bacterial N-formyl-methionyl-leucyl-phenylalanine: delayed response in ulcerative colitis; Vainer B et al.; OBJECTIVE: In acute stages of ulcerative colitis (UC), neutrophils migrate from the circulation into inflamed colonic tissue, initiated by yet unknown stimuli . The bacterial peptide N-formyl-methionyl-leucyl-phenylalanine (FMLP) is a component of the surface membrane of colonic bacteria such as Escherichia coli and stimulates Ca2+ influx into neutrophils, reflecting the fact that ionized calcium is an important secondary messenger for several neutrophil functions, including locomotion, phagocytosis and free oxygen radical production . Recent studies have revealed that Ca2+ dependent ICAM-1/beta 2-integrin mediated neutrophil migration is impaired in UC patients . The aim of the present work was to study the influx of Ca2+ into peripheral blood neutrophils of UC patients after exposure to FMLP and after binding of either beta 2-integrins or intercellular adhesion molecule-1 (ICAM-1) . METHODS: The relative intracellular Ca2+ levels ({Ca2+}i ) were measured spectrofluorometrically in neutrophils isolated from eight UC patients and eight controls . The cells were exposed to 1 nm FMLP, 5 pm free ICAM-1, or antibodies binding ICAM-1 or the beta 2-integrins CD11a, CD11b, CD11c and CD18 . RESULTS: A pronounced increase in {Ca2+}i was observed by exposure of cells to FMLP, and neutrophils from UC patients showed a consistent and significant delayed response as compared to cells from control subjects (P < 0.01) . Antibody mediated cross-linking of CD18 triggered a small but detectable increase in {Ca2+}i, which did not differ between patients and controls . CONCLUSION: A delayed response to bacterial peptides appears to be a phenotypic trait for neutrophils of UC patients . A connection between FMLP stimulated Ca2+ influx and CD11/CD18 upregulation is discussed.

J Microbiol Methods, 2003 Apr, 53(1), 43 - 50
Statistical analysis and biological interpretation of the flow cytometric heterogeneity observed in bacterial axenic cultures; Vives-Rego J et al.; Histogram comparison and meaningful statistics in flow cytometry is probably the most widely encountered mathematical problem in flow cytometry . Ideally, a test for determining the statistical equality or difference of flow cytometric distributions will identify the significant differences or similarities of the obtained histograms . This situation is of particular interest when flow cytometry is used to study the heterogeneity of axenic bacterial populations . We have statistically measured the heterogeneity of successive cytometric measures, the modifications produced after 20 transfers from the same culture, and the differences between 20 subcultures of identical origin . The heterogeneity of the bacterial populations and the similarity of the obtained 360 histograms were analysed by standard statistical methods . We have studied bacterial axenic cultures in order to detect, quantify and interpret their cytometric heterogeneity, and to assess intrinsic differences and differences produced by laboratory manipulations . We concluded that the standard axenic cultures have a considerable intrinsic cellular and molecular heterogeneity . We suggest that the heterogeneity we have detected basically has two origins: cell size diversity and cell cycle variations.

Acta Paediatr Taiwan, 2002 Sep-Oct, 43(5), 288 - 90
Bacterial meningitis of an infant with Currarino triad; Chou IC et al.; Currarino triad is a rarely hereditary condition including: (1) an anorectal malformation, (2) an anterior sacral defect, and (3) a presacral mass . Autosomal dominant transmission is suggested . We reported one case of Currarino triad, who was a 3-month-old male with sacral dysgenesis, imperforated anus and enteric cyst . This case presented with acute lower limbs paralysis due to bacterial meningitis complicated with acute arachnoiditis . The diagnosis of this condition led to a work-up of his sibling, who was found to have an incomplete type . The purpose of this case is to emphasize that a high index of suspicion for timely diagnosis and treatment of Currarino triad could prevent devastating complications.

Kaku Igaku, 2002 Nov, 39(4), 543 - 8
{Short time bacterial endotoxins test for positron emission tomography by means of positively charged filters}; Nakazawa N et al.; Positron emission tomography (PET) radiotracers have very short physical half-lives . It is hard to complete a bacterial endotoxins test prior to release from medical institutes . For endotoxin quantitative determination, limulus amebocyte lysate (LAL) reagent and kinetic-turbidimetry system were previously developed . We investigated the possibility of a short time test by means of positively charged filters . As a result of this study, the effects of positively charged filters on endotoin removal were over 99.5% for {18F}FDG and {18F}NaF, which were contaminated with the indicated concentration of endotoxin . Combining this filter and the kinetic-turbidimetric method, it was possible to complete a bacterial endotoxins test in 5 min prior to the patient's administration . This test should be required prior to release for PET radiopharmaceutical quality control . It has been suggested that this combination is a good method for this purpose.

Toxicol Sci, 2003 Mar, 72(1), 43 - 56
Role of neutrophils in the synergistic liver injury from monocrotaline and bacterial lipopolysaccharide exposure; Yee SB et al.; Synergistic liver injury develops in Sprague-Dawley rats from administration of a small, noninjurious dose (7.4 x 10(6) EU/kg) of bacterial lipopolysaccharide (LPS) given 4 h after a nontoxic dose (100 mg/kg) of the pyrrolizidine alkaloid, monocrotaline (MCT) . Previous studies demonstrated that liver injury is mediated through inflammatory factors, such as Kupffer cells and tumor necrosis factor alpha (TNF-alpha), rather than through simple interaction between MCT and LPS . In the present study, the hypothesis that neutrophils (polymorphonuclear leukocytes or PMNs) are causally involved in this injury model is tested, and the interdependence between PMNs and other inflammatory components is explored . Hepatic PMN accumulation and the appearance of cytokine-induced neutrophil chemoattractant-1 in plasma preceded the onset of liver injury, suggesting that PMNs contribute to toxicity . Hepatic PMN accumulation was partially dependent on TNF-alpha . Prior depletion of PMNs in MCT/LPS-cotreated animals resulted in attenuation of both hepatic parenchymal cell (HPC) and sinusoidal endothelial cell (SEC) injury at 18 h . PMN depletion did not, however, protect against early SEC injury that occurred before the onset of HPC injury at 6 h . This observation suggests that SEC injury is not entirely dependent on PMNs in this model . In vitro, MCT caused PMNs to degranulate in a concentration-dependent manner . These results provide evidence that PMNs are critical to the HPC injury caused by MCT/LPS cotreatment and contribute to the progression of SEC injury.

Metab Brain Dis, 2002 Dec, 17(4), 303 - 9
Nitric oxide in patients with cirrhosis and bacterial infections; Such J et al.; Nitric oxide (NO) is a powerful vasodilator agent that has been found to be elevated in patients with cirrhosis, and that plays a key role in the pathogenesis of the hemodynamic abnormalities found in these patients . The reasons for this increased NO synthesis are not entirely known, but at least two main mechanisms are involved: shear stress and bacterial-induced NO synthesis . This review focuses on bacterial-induced NO synthesis . Induction of NO synthesis by different cellular populations occurs when proinflammatory cytokines act synergistically, and also by endotoxin . Spontaneous bacterial peritonitis (SBP) is the most dangerous infectious complication arising in patients with cirrhosis and ascites, and it is associated with high serum and ascitic fluid levels of proinflammatory cytokines . A subset of patients in this situation show high levels of serum and ascitic fluid NO levels when SBP is diagnosed, and these patients seem to be predisposed to the development of renal impairment . The increased NO synthesis and associated aggravated vasodilatation may be the reason why patients with SBP show high levels of plasma renin activity, in an attempt to counterbalance this new situation, and that the administration of albumin during the SBP episode significantly reduces the episode-related mortality.

Zhejiang Da Xue Xue Bao Yi Xue Ban, 2002 Aug, 31(6), 448 - 452
{Study on the bacterial identification using 16S-23S rRNA gene spacer regions}; Fu JF et al.; OBJECTIVE: To examine the use of PCR utilizing 16S-23S rRNA gene spacer regions in the identification of bacteria . METHODS Primers used in PCR were designed by using the target sequences from the genes encoding 16S-23S rRNA spacer regions . PCR was used for the detection of different standard and clinical bacterial strains . RESULTS Characteristic DNA maps were present after using the PCR to identify 27 standard strains from 27 species . The maps could be directly used for classification of the tested bacterial strains or further differentiated by RFLP . The sensitivity of the PCR may be as high as 2.5 CFU/ml . No non-specific amplification products were observed when using DNA from human PBMC funguses or viruses as templates . Thirty-two strains of bacteria isolated from clinical strains showed DNA maps similar to the DNA maps amplified from standard strains . CONCLUSION The PCR detection of bacteria using 16S-23S rRNA gene spacer regions is sensitive, rapid, specific and accurate for identification of bacteria and provides a new rapid method for determining the clinical diagnosis and the etiology of sepsis.

Hepatology, 2003 Mar, 37(3), 551 - 7
Oral bile acids reduce bacterial overgrowth, bacterial translocation, and endotoxemia in cirrhotic rats; Lorenzo-Zuniga V et al.; Experiments were performed to test whether conjugated bile acid administration would decrease bacterial overgrowth, bacterial translocation, and endotoxemia in ascitic cirrhotic rats . Cholylsarcosine, a deconjugation-dehydroxylation resistant and cholylglycine, a deconjugation-dehydroxylation susceptible bile acid were used . Rats with CCl(4)-induced cirrhosis and ascites were fed cholylsarcosine, cholylglycine (both at 70 mg/kg/d), or placebo for 2 weeks . Healthy rats, as controls, were treated similarly . In cirrhotic rats receiving placebo, bile secretion from an acute biliary fistula was lower than in healthy rats (27.2 +/- 6.5 vs . 53.0 +/- 3.1 microL/kg/min; mean +/- SE, P<.05) . The administration of conjugated bile acids to cirrhotic rats normalized bile secretion (cholylsarcosine, 51.8 +/- 6.29; cholylglycine, 52.72 +/- 8.9 microL/kg/min) . Total ileal bacterial content was 6-fold higher in ascitic cirrhotic rats than in healthy rats . Conjugated bile acid administration reduced bacterial content to normal levels . Bacterial translocation was less in cirrhotic animals receiving conjugated bile acids (cholylsarcosine, 33%; cholylglycine, 26%) than in animals receiving placebo (66%) . Endotoxemia was decreased in cirrhotic rats by conjugated bile acid feeding (cholylsarcosine, 0.098 +/- 0.002; cholylglycine 0.101 +/- 0.007 EU/mL) compared with placebo (0.282 +/- 0.124, P <.001) . Survival was greater in animals receiving conjugated bile acids (cholylsarcosine, 10/15; cholylglycine, 11/15; placebo, 5/15) . In conclusion, the administration of conjugated bile acids to ascitic cirrhotic rats increased bile acid secretion, eliminated intestinal bacterial overgrowth, decreased bacterial translocation, decreased endotoxemia, and increased survival . Oral conjugated bile acids may be useful in preventing bacterial translocation, endotoxemia, and spontaneous bacterial perotonitis in cirrhotic patients.

Proc Natl Acad Sci U S A, 2003 Mar 4, 100(5), 2789 - 94 Epub 2003 Feb 24.
Pathogen DNA as target for host-generated oxidative stress: role for repair of bacterial DNA damage in Helicobacter pylori colonization; O'Rourke EJ et al.; Helicobacter pylori elicits an oxidative stress during host colonization . This oxidative stress is known to cause lesions in the host DNA . Here we addressed the question as to whether the pathogen DNA is subject to lethal or mutational damage by the host-generated oxidative response . H . pylori Hpnth mutants unable to repair oxidized pyrimidines from the bacterial DNA were generated . H . pylori strains lacking a functional endonuclease III (HpNth) showed elevated spontaneous and induced mutation rates and were more sensitive than the parental strain to killing by exposure to oxidative agents or activated macrophages . Although under laboratory conditions the Hpnth mutant strain grows as well as the wild-type strain, in a mouse infection the stomach bacterial load gradually decreases while the population in the wild-type strain remains stable, showing that endonuclease III deficiency reduces the colonization capacity of the pathogen . In coinfection experiments with a wild-type strain, Hpnth cells are eradicated 15 days postinfection (p.i.) even when inoculated in a 1:9 wild-type:mutant strain ratio, revealing mutagenic lesions that are counterselected under competition conditions . These results show that the host effectively induces lethal and premutagenic oxidative DNA adducts on the H . pylori genome . The possible consequences of these DNA lesions on the adaptability of H . pylori strains to new hosts are discussed.

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