Exp Neurol, 2001 Nov, 172(1), 137 - 52 Systemic LPS injection leads to granulocyte influx into normal and injured brain: effects of ICAM-1 deficiency; Bohatschek M et al.; The lipopolysaccharide (LPS) constituents of the gram-negative bacterial wall are among the most potent activators of inflammation . In the current study, we examined the effect of subcutaneous injection of Escherichia coli LPS on leukocyte influx into the normal and injured brain using endogenous peroxidase (EP) . Normal brain parenchyma does not contain granulocytes and this does not change after indirect trauma, in facial axotomy . However, systemic injection of 1 mg LPS led to a gradual appearance of EP-positive parenchymal granulocytes within 12 h, with a maximum at 1-4 days after injection . Facial axotomy (day 14) led to a further 50-300% increase in granulocyte number . Of the five mouse strains tested in the current study, four--Balb/C, FVB, C57Bl/6, and C3H/N--showed vigorous granulocyte influx (60-90 cells per 20-microm section in axotomized facial nucleus, 20-40 cells per section on the contralateral side) . The influx was an order of magnitude lower in the SJL mice . The peroxidase-positive cells were immunoreactive for neutrophil antigen 7/4 and alpha M beta 2 integrin, were negative for IBA1 (monocytes) and CD3 (T cells), and could be prelabeled by subcutaneous injection with rhodamine B isothiocyanate (RITC), confirming their origin as blood-borne granulocytes . All RITC-positive cells were IBA1 negative . This influx of granulocytes was accompanied by a disruption of the blood-brain barrier to albumin and induction of the cell adhesion molecule ICAM-1 on affected blood vessels . Transgenic deletion of ICAM-1 led to a more than 50% reduction in the number of infiltrating granulocytes compared to litter-matched wild-type controls, in normal brain as well as in axotomized facial motor nucleus . In summary, systemic injection of LPS leads to invasion of granulocytes into the mouse brain and a breakdown of the blood-brain barrier to blood-borne cells and to soluble molecules . Moreover, this mechanism may play a pathogenic role in the etiology of meningitis and in severe bacterial sepsis . Int J Med Microbiol, 2001 Sep, 291(4), 299 - 305 Host cell signaling in Helicobacter pylori infection; Naumann M; Helicobacter pylori represents a highly successful human microbial pathogen that has infected approximately half of the world's population . This gram-negative microorganism colonizes the human epithelial layer in the stomach and induces a state of chronic inflammation that does not resolve the underlying infection and often leads to gastric or duodenal ulcers, or more rarely to gastric cancer . Among the reactions in H . pylori-infected epithelial cells the induction of proinflammatory cytokines, cell spreading and movement, as well as a scattered phenotype appear strictly dependent on the expression of pathogenicity island-encoded proteins in H . pylori . This review will discuss the features of the H . pylori-induced signal transduction leading to changes in host cellular function . Topics discussed comprise the signaling and the phenotypes associated with the type IV secretion system, the activation of target genes involved in gastric physiology, and putative mechanisms leading to the development of gastric cancer. Med Clin North Am, 2001 Nov, 85(6), 1493 - 509 Guidelines for the management of community-acquired pneumonia . Current recommendations and antibiotic selection issues; Niederman MS; Numerous guidelines for CAP have been developed, and although each is different, many principles are common to all recommendations . A guideline should focus on a wide range of issues surrounding the delivery of care, including advice about when to admit patients to the hospital or ICU; which antibiotic regimens to select for specific patient populations; which pathogens to target in empiric therapy; which diagnostic tests to order; how to assess the importance of specific causative pathogens, such as drug-resistant pneumococci, atypical pathogens, and gram-negative pathogens; how to evaluate the response to therapy and when to switch responding patients to oral therapy; and how to prevent CAP effectively through appropriate use of immunization against pneumococcus and influenza . Currently, many new antibiotic choices have emerged in the macrolide, quinolone, beta-lactam, ketolide, and oxazolidinone classes, and specific issues surrounding selection of these agents must be considered . All of the available data can be synthesized into a disease management guideline, and current therapy in the United States generally is consistent with existing recommendations . This consistency not only has led to more uniformity in patient care, but also has led to measurable benefits in patient outcomes, including reduced mortality for hospitalized patients with CAP . Guidelines not only are a useful tool for managing patients with CAP, but also they serve the purpose of defining current issues in patient care and stimulating the search for new tools and management approaches for this important clinical problem. Appl Environ Microbiol, 2001 Nov, 67(11), 5010 - 6 Pathogenicity of Moraxella osloensis, a bacterium associated with the nematode Phasmarhabditis hermaphrodita, to the slug Deroceras reticulatum; Tan L et al.; Moraxella osloensis, a gram-negative bacterium, is associated with Phasmarhabditis hermaphrodita, a nematode parasite of slugs . This bacterium-feeding nematode has potential for the biological control of slugs, especially the grey garden slug, Deroceras reticulatum . Infective juveniles of P . hermaphrodita invade the shell cavity of the slug, develop into self-fertilizing hermaphrodites, and produce progeny, resulting in host death . However, the role of the associated bacterium in the pathogenicity of the nematode to the slug is unknown . We discovered that M . osloensis alone is pathogenic to D . reticulatum after injection into the shell cavity or hemocoel of the slug . The bacteria from 60-h cultures were more pathogenic than the bacteria from 40-h cultures, as indicated by the higher and more rapid mortality of the slugs injected with the former . Coinjection of penicillin and streptomycin with the 60-h bacterial culture reduced its pathogenicity to the slug . Further work suggested that the reduction and loss of pathogenicity of the aged infective juveniles of P . hermaphrodita to D . reticulatum result from the loss of M . osloensis from the aged nematodes . Also, axenic J1/J2 nematodes were nonpathogenic after injection into the shell cavity . Therefore, we conclude that the bacterium is the sole killing agent of D . reticulatum in the nematode-bacterium complex and that P . hermaphrodita acts only as a vector to transport the bacterium into the shell cavity of the slug . The identification of the toxic metabolites produced by M . osloensis is being pursued. Int J Dermatol, 2001 Jul, 40(7), 434 - 8 Cutaneous findings encountered in brucellosis and review of the literature; Metin A et al.; BACKGROUND: Human brucellosis is an infectious disease produced by Brucella species: small, coccoid or rod-like, aerobic, Gram-negative bacteria . The infection is common in developing countries, and can also affect the skin . Its prevalence is high in our region of Turkey, where stockbreeding is one of the main economic sources, compared with the industrially developed areas of Turkey, and dermatologic complaints due to brucellosis are fairly common . MATERIALS AND METHODS: One hundred and three patients with serologically and clinically confirmed brucellosis were studied in order to investigate the prevalence of cutaneous findings and their variability in brucellosis . Fifty-two (50.49%) were males and 51 (49.51%) were females with an age range of 4-70 years (mean, 30.45 +/- 15.08 years) . RESULTS: Of these patients, 14 (13.59%) had cutaneous findings probably related to brucellosis . These findings were more frequent in females (11 cases) than males, and most of the females (eight cases) were housewives; three were students . Urticaria-like papules and plaques were the most common findings; they were seen in six (35.3%) patients . One case had livedo reticularis and another palmar erythema, which have not been reported previously . No relationship was observed between the serologic values and the cutaneous findings . CONCLUSIONS: Cutaneous findings in our cases were more prevalent than in other reported studies . It is important to emphasize that cutaneous lesions are not specific to brucellosis and may be seen in a variety of other dermatologic diseases caused by many agents; therefore, these agents should be kept in mind in the differential diagnosis. Proteomics, 2001 May, 1(5), 705 - 20 Analysis of the outer membrane proteome of Caulobacter crescentus by two-dimensional electrophoresis and mass spectrometry; Phadke ND et al.; Caulobacter crescentus, a Gram negative alpha-purple bacterium that displays an invariant asymmetric cell division pattern, has become a key model system for the study of bacterial development . Membrane proteins play key roles in cell cycle events, both as components of landmark morphological structures and as critical elements in regulation of the cell cycle . Recent advances for the isolation and solubilization of bacterial membrane proteins prior to isoelectric focusing have significantly improved the separation of outer membrane proteins by two-dimensional (2-D) electrophoresis . In this work we describe the analysis of the outer membrane proteome of Caulobacter crescentus . Proteins were identified using 2-D gel electrophoresis and peptide mass fingerprinting by matrix-assisted laser desorption/ionization-time of flight mass spectrometry . We identified 54 unique proteins out of which 41 were outer membrane proteins . Of the outer membrane proteins, 16 were identified as TonB-dependent receptor proteins . These studies were executed simultaneously with the Caulobacter genome sequencing project and advantages and limitations of proteomic analysis of a nonannotated genome are discussed . Finally, protein levels between cells grown in rich and minimal media are compared which demonstrates that many of the TonB-dependent receptor proteins are found at higher levels in minimal medium. Gastroenterol Clin Biol, 2001 Jun-Jul, 25(6-7), 659 - 63 {UreI: a Helicobacter pylori protein essential for resistance to acidity and for the early steps of murine gastric mucosa infection}; Bury-Mone S et al.; Helicobacter pylori (H . pylori) is a Gram negative microaerophilic bacteria whose only known niche is the human gastric mucosa . The presence of H . pylori is associated with various pathologies ranging from peptic ulcer disease to gastric carcinoma . H . pylori virulence is dependent on its exceptional ability to resist to the stomach acidity by hydrolyzing urea into ammonia . Survival of H . pylori to acidity in the presence of urea relies on the activity of a membrane protein, UreI.AIMS: We decided to better characterize the role of UreI (i) in vitro in ammonia production through the action of urease, and (ii) in vivo in the colonization of the gastric mucosa.METHODS: Ammonia production by a wild type strain of H . pylori or by a UreI-deficient strain was measured as a function of extracellular pH . In addition, the kinetics of elimination of a UreI-deficient mutant in vivo were realized in the mouse model for colonization.RESULTS: UreI was associated with an increase of ammonia production in acidic conditions in vitro and was necessary for the initial steps of the mouse stomach colonization.CONCLUSION: UreI thus behaves as a sensor of extracellular pH . This protein activates urease at acidic pH; thereby, it probably allows H . pylori to resist to acidity in vivo during the first steps of infection. J Immunol, 2001 Nov 1, 167(9), 5231 - 9 Escherichia coli Braun lipoprotein induces a lipopolysaccharide-like endotoxic response from primary human endothelial cells; Neilsen PO et al.; All bacteria contain proteins in which their amino-terminal cysteine residue is modified with N-acyl S-diacylglycerol functions, and peptides and proteins bearing this modification are immunomodulatory . The major outer membrane lipoprotein of Escherichia coli, the Braun lipoprotein (BLP), is the prototypical triacylated cysteinyl-modified protein . We find it is as active as LPS in stimulating human endothelial cells to an inflammatory phenotype, and a BLP-negative mutant of E . coli was less inflammatory than its parental strain . While the lipid modification was essential, the lipidated protein was more potent than a lipid-modified peptide . BLP associates with CD14, but this interaction, unlike that with LPS, was not required to elicit endothelial cell activation . BLP stimulated endothelial cell E-selectin surface expression, IL-6 secretion, and up-regulation of the same battery of cytokine mRNAs induced by LPS . Quantitative microarray analysis of 4400 genes showed the same 30 genes were induced by BLP and LPS, and that there was near complete concordance in the level of gene induction . We conclude that the lipid modification of at least one abundant Gram-negative protein is essential for endotoxic activity, but that the protein component also influences activity . The equivalent potency of BLP and LPS, and their complete concordance in the nature and extent of endothelial cell activation show that E . coli endotoxic activity is not due to just LPS . The major outer membrane protein of E . coli is a fully active endotoxic agonist for endothelial cells. J Bacteriol, 2001 Nov, 183(22), 6499 - 508 A 12-amino-acid segment, present in type s2 but not type s1 Helicobacter pylori VacA proteins, abolishes cytotoxin activity and alters membrane channel formation; McClain MS et al.; Helicobacter pylori, a gram-negative bacterium associated with gastritis, peptic ulceration, and gastric adenocarcinoma in humans, secretes a protein toxin, VacA, that causes vacuolar degeneration of epithelial cells . Several different families of H . pylori vacA alleles can be distinguished based on sequence diversity in the "middle" region (i.e., m1 and m2) and in the 5' end of the gene (i.e., s1 and s2) . Type s2 VacA toxins contain a 12-amino-acid amino-terminal hydrophilic segment, which is absent from type s1 toxins . To examine the functional properties of VacA toxins containing this 12-amino-acid segment, we analyzed a wild-type s1/m1 VacA and a chimeric s2/m1 VacA protein . Purified s1/m1 VacA from H . pylori strain 60190 induced vacuolation in HeLa and Vero cells, whereas the chimeric s2/m1 toxin (in which the s1 sequence of VacA from strain 60190 was replaced with the s2 sequence from strain Tx30a) lacked detectable cytotoxic activity . Type s1/m1 VacA from strain 60190 formed membrane channels in a planar lipid bilayer assay at a significantly higher rate than did s2/m1 VacA . However, membrane channels formed by type s1 VacA and type s2 VacA proteins exhibited similar anion selectivities (permeability ratio, P(Cl)/P(Na) = 5) . When an equimolar mixture of the chimeric s2/m1 toxin and the wild-type s1/m1 toxin was added to HeLa cells, the chimeric toxin completely inhibited the activity of the s1/m1 toxin . Thus, the s2/m1 toxin exhibited a dominant-negative phenotype similar to that of a previously described mutant toxin, VacA-(Delta6-27) . Immunoprecipitation experiments indicated that both s2/m1 VacA and VacA-(Delta6-27) could physically interact with a c-myc epitope-tagged s1/m1 VacA, which suggests that the dominant-negative phenotype results from the formation of heterooligomeric VacA complexes with defective functional activity . Despite detectable differences in the channel-forming activities and cytotoxic properties of type s1 and type s2 VacA proteins, the conservation of type s2 sequences in many H . pylori isolates suggests that type s2 VacA proteins retain an important biological activity. Immunol Lett, 2001 Sep 3, 78(2), 97 - 101 Toll-like receptor 2 and 4 surface expressions on human monocytes are modulated by interferon-gamma and macrophage colony-stimulating factor; Mita Y et al.; Human Toll-like receptor 2 (TLR2) and TLR4 are recently identified receptors . TLR4 was shown to be the main protein involved in recognizing Gram-negative bacteria, whereas TLR2 is apparently the key factor in responses to other types of microbial pathogens . We examined regulations of TLR2 and TLR4 surface expressions in human peripheral blood monocytes by interferon-gamma (IFN-gamma) and macrophage colony-stimulating factor (M-CSF) . IFN-gamma up-regulated both TLR2 and TLR4, but enhanced the surface expression, on human peripheral blood monocytes of TLR4 more than that of TLR2 . On the other hand, M-CSF up-regulated both TLR2 and TLR4 surface expression on human peripheral blood monocytes, with no change in the ratio of TLR2/TLR4 surface expression . These results indicate that IFN-gamma enhances receptors involved in the response to Gram-negative bacteria more than those involved in responses to other types of microbial pathogens, whereas M-CSF enhances the receptor response to Gram-negative bacteria in the same manner as to other types of microbial pathogens. Biochemistry, 2001 Oct 30, 40(43), 13041 - 50 Conformational change in the stator of the bacterial flagellar motor; Kojima S et al.; MotA and MotB are integral membrane proteins of Escherichia coli that form the stator of the proton-fueled flagellar rotary motor . The motor contains several MotA/MotB complexes, which function independently to conduct protons across the cytoplasmic membrane and couple proton flow to rotation . MotB contains a conserved aspartic acid residue, Asp32, that is critical for rotation . We have proposed that the protons energizing the motor interact with Asp32 of MotB to induce conformational changes in the stator that drive movement of the rotor . To test for conformational changes, we examined the protease susceptibility of MotA in membrane-bound complexes with either wild-type MotB or MotB mutated at residue 32 . Small, uncharged replacements of Asp32 in MotB (D32N, D32A, D32G, D32S, or D32C) caused a significant change in the conformation of MotA, as evidenced by a change in the pattern of proteolytic fragments . The conformational change does not require any flagellar proteins besides MotA and MotB, as it was still seen in a strain that expresses no other flagellar genes . It affects a cytoplasmic domain of MotA that contains residues known to interact with the rotor, consistent with a role in the generation of torque . Influences of key residues of MotA on conformation were also examined . Pro173 of MotA, known to be important for rotation, is a significant determinant of conformation: Dominant Pro173 mutations, but not recessive ones, altered the proteolysis pattern of MotA and also prevented the conformational change induced by Asp32 replacements . Arg90 and Glu98, residues of MotA that engage in electrostatic interactions with the rotor, appear not to be strong determinants of conformation of the MotA/MotB complex in membranes . We note sequence similarity between MotA and ExbB, a cytoplasmic-membrane protein that energizes outer-membrane transport in Gram-negative bacteria . ExbB and associated proteins might also employ a mechanism involving proton-driven conformational change. J Med Humanit, 1993 Spring, 14(1), 5 - 14 We have a prejudice against ourselves -- sentiment, ethics, and reason; Rosenberg LT; Briefly stated my point is that the well-being of each person in a community conceived abstractly may be all too easily sacrificed for the sake of the abstraction . Physicians may offer critically ill patients places in programs of experimental treatment, but there is commonly a catch to the offer . To take part in a program of clinical experiment a patient must not only risk a possible failure of a fresh drug and the chance of destructive side effects from the drug, but the patients must risk only getting the traditional treatment along with a placebo rather than the experimental drug . Placebo control, double blind critical protocols for testing effects of fresh drugs on critically ill patients are a commonplace . I question the scientific objectivity of the protocols and the underlying ethic, and suggest use of alternate protocols . Experimental tests in the treatment of gram-negative bacteria blood infections, muscular dystrophy, and AIDS and AIDS-related diseases are examples. Genetika, 2001 Aug, 37(8), 1160 - 4 {Tn5037-a Tn21-like mercury transposon, detected in Thiobacillus ferrooxidans}; Kaliaeva ES et al.; The 6645-bp mercury resistance transposon of the chemolithotrophic bacterium Thiobacillus ferrooxidans was cloned and sequenced . This transposon, named Tn5037, belongs to the Tn21 branch of the Tn21 subgroup, many members of which have been isolated from clinical sources . Having the minimum set of the genes (merRTPA), the mercury resistance operon of Tn5037 is organized similarly to most of the Gram-negative bacteria mer operons and is closest to that of Thiobacillus 3.2 . The operator-promoter region of the mer operon of Tn5037 also has the common (Tn21/Tn501-like) structure . However, its inverted, presumably MerR protein binding repeats in the operator/promoter element are two base pairs shorter than in Tn21/Tn501 . In the merA region, this transposon shares 77.4, 79.1, 83.2 and 87.8% identical bases with Tn21, Tn501, T . ferrooxidance E-15, and Thiobacillus 3.2, respectively . No inducibility of the Tn5037 mer operon was detected in the in vivo experiments . The transposition system (terminal repeats plus gene tnpA) of Tn5037 was inactive in Escherichia coli K12, in contrast to its resolution system (res site plus gene tnpR) . However, transposition of Tn5037 in this host was provided by the tnpA gene of Tn5036, a member of the Tn21 subgroup . Sequence analysis of the Tn5037 res site suggested its recombinant nature. Genes Immun, 2001 Oct, 2(6), 349 - 55 IFNs activate toll-like receptor gene expression in viral infections; Miettinen M et al.; Toll-like receptors (TLRs) mediate innate immune responses to microbes . TLR2, TLR5, TLR6, and TLR9 have been implicated in responses to bacterial components, and TLR4 is the receptor for Gram-negative bacteria . Recently, TLR4 was described to function in respiratory syncytial virus-induced NF-kappaB activation . Here we have analyzed TLR1-9 mRNA expression in human primary macrophages infected with influenza A and Sendai viruses . TLR1, TLR2, TLR4, TLR6, and TLR8 mRNAs were expressed at basal levels in macrophages . Viral infection enhanced TLR1, TLR2, TLR3, and TLR7 mRNA expression, and neutralizing anti-IFN-alpha/beta antibodies downregulated gene expression of these TLRs . Exogenously added IFN-alpha upregulated TLR1, TLR2, TLR3, and TLR7 mRNA expression in macrophages, as well as TLR3 mRNA expression in epithelial and endothelial cell lines . IFN-gamma enhanced the expression of TLR1 and TLR2 mRNA in macrophages, and TLR3 in epithelial and endothelial cells . The data suggests a novel role for IFNs in the activation of innate immunity. Int Immunopharmacol, 2001 Oct, 1(11), 1979 - 87 Immunoferon, a glycoconjugate of natural origin, inhibits LPS-induced TNF-alpha production and inflammatory responses; Brieva A et al.; We have analyzed the effect of a patented glycoconjugate (GC) of natural origin, Inmunoferon, in the development of the response to endotoxemia induced by administration of LPS in rodents . We have observed that oral treatment with the drug reduced the levels of serum TNF-alpha induced by an intravenous pulse of LPS . The serum of pretreated mice blocked TNF-alpha production by peritoneal macrophages . The drug increased the levels of TNF-alpha regulators such as IL-10 and corticosteroids, whereas it inhibited TNF-alpha-dependent IL-6 production . Further TNF-alpha-dependent responses, such as cell extravasation, was decreased in treated mice . According to these results, Inmunoferon is postulated as an inhibitor of the systemic response to LPS . Correlation of the observations made in mice with a rat model suggests the efficacy of this product in reducing TNF-alpha production in a species-independent fashion and opens the possibility of its trial as an adjuvant of antibiotics in treatment against gram-negative bacterial infection. Anal Chem, 2001 Oct 1, 73(19), 4566 - 73 Bioinformatics and mass spectrometry for microorganism identification: proteome-wide post-translational modifications and database search algorithms for characterization of intact H . pylori; Demirev PA et al.; MALDI-TOF mass spectrometry has been coupled with Internet-based proteome database search algorithms in an approach for direct microorganism identification . This approach is applied here to characterize intact H . pylori (strain 26695) Gram-negative bacteria, the most ubiquitous human pathogen . A procedure for including a specific and common posttranslational modification, N-terminal Met cleavage, in the search algorithm is described . Accounting for posttranslational modifications in putative protein biomarkers improves the identification reliability by at least an order of magnitude . The influence of other factors, such as number of detected biomarker peaks, proteome size, spectral calibration, and mass accuracy, on the microorganism identification success rate is illustrated as well. J Virol, 2001 Nov, 75(22), 10730 - 7 Involvement of toll-like receptor 4 in innate immunity to respiratory syncytial virus; Haynes LM et al.; The mammalian Toll-like receptor 4, TLR4, is an important component in the innate immune response to gram-negative bacterial infection . The role of TLR4 in antiviral immunity has been largely unexplored . In this study, the in vivo immune responses to respiratory syncytial virus (RSV) and influenza virus infection were examined in TLR4-deficient (C57BL/10ScNCr) and TLR4-expressing (C57BL/10Sn) mice . TLR4-deficient mice challenged with RSV, but not influenza virus, exhibited impaired natural killer (NK) cell and CD14(+) cell pulmonary trafficking, deficient NK cell function, impaired interleukin-12 expression, and impaired virus clearance compared to mice expressing TLR4 . These findings suggest that Toll signaling pathways have an important role in innate immunity to RSV. Appl Microbiol Biotechnol, 2001 Sep, 56(5-6), 736 - 41 The meroperon of a mercury-resistant Pseudoalteromonas haloplanktis strain isolated from Minamata Bay, Japan; Iohara K et al.; A mer operon of mercury-resistant Pseudoalteromonas haloplanktis strain M1, isolated from sea water of Minamata Bay, was cloned and analyzed . The mer genes were located in the chromosome and organized as merR-merT-merP-merC-merA-merD, the same order as that in Tn21 . However, the orientation of the merR gene is the same as that of other mer genes (opposite direction to Tn21), and merR was cotranscribed with other mer genes, a pattern that has not been previously seen with mer determinants from other Gram-negative bacteria . Furthermore, the amino acid similarities of the corresponding mer gene products between those from strain M1 and Tn21 were unusually low. Am J Physiol Cell Physiol, 2001 Nov, 281(5), C1587 - 95 Human endothelial cell response to gram-negative lipopolysaccharide assessed with cDNA microarrays; Zhao B et al.; To assess the feasibility of using cDNA microarrays to understand the response of endothelial cells to lipopolysaccharide (LPS) and to evaluate potentially beneficial agents in treatment of septic shock, human umbilical vein endothelial cells were exposed to Escherichia coli LPS for 1, 4, 7, 12, or 24 h . Total RNA was isolated and reverse-transcribed into (33)P-labeled cDNA probes that were hybridized to human GeneFilter microarrays containing approximately 4,000 genes . The mRNA levels of several genes known to respond to LPS changed after stimulation . In addition, a number of genes not previously implicated in the response of endothelial cells to LPS also appeared to be altered in expression . Nuclear factor-kappaB (NF-kappaB) was shown to play an important role in regulating genes identified from the microarray studies . Pretreatment of endothelial cells with a specific NF-kappaB translocation inhibitor eliminated most of the alterations in gene expression . Quantitative RT-PCR results independently confirmed the microarray results for monocyte chemotactic protein-1 and interleukin-8, and enzyme-linked immunosorbent assays demonstrated that augmented transcription was followed by translation and secretion. Infect Immun, 2001 Nov, 69(11), 6942 - 50 Dual role of lipopolysaccharide (LPS)-binding protein in neutralization of LPS and enhancement of LPS-induced activation of mononuclear cells; Gutsmann T et al.; The lipopolysaccharide (LPS)-binding protein (LBP) has a concentration-dependent dual role in the pathogenesis of gram-negative sepsis: low concentrations of LBP enhance the LPS-induced activation of mononuclear cells (MNC), whereas the acute-phase rise in LBP concentrations inhibits LPS-induced cellular stimulation . In stimulation experiments, we have found that LBP mediates the LPS-induced cytokine release from MNC even under serum-free conditions . In biophysical experiments we demonstrated that LBP binds and intercalates into lipid membranes, amplified by negative charges of the latter, and that intercalated LBP can mediate the CD14-independent intercalation of LPS into membranes in a lipid-specific and temperature-dependent manner . In contrast, prior complexation of LBP and LPS inhibited binding of these complexes to membranes due to different binding of LBP to LPS or phospholipids . This results in a neutralization of LPS and, therefore, to a reduced production of tumor necrosis factor by MNC . We propose that LBP is not only present as a soluble protein in the serum but may also be incorporated as a transmembrane protein in the cytoplasmic membrane of MNC and that the interaction of LPS with membrane-associated LBP may be an important step in LBP-mediated activation of MNC, whereas LBP-LPS complexation in the serum leads to a neutralization of LPS. Clin Infect Dis, 2001 Nov 15, 33(10), 1789 - 91 Epub 2001 Oct 05. Bacteremia caused by a Helicobacter pullorum-like organism; Tee W et al.; We report a case of bacteremia caused by a Helicobacter pullorum-like organism in a 35-year-old man with pyrexia of unknown origin . Culture of blood samples obtained at admission yielded a motile, spiral-shaped gram-negative rod, and 16S rRNA gene sequencing identified this organism as Helicobacter pullorum-like, showing 23 base differences compared with the recently described "Helicobacter canadensis" (a recently proposed group that had previously been classified within H . pullorum) . We believe that this is the first report of bacteremia caused by this organism. Toxicon, 2001 Nov, 39(11), 1729 - 36 The cytolethal distending toxins induce DNA damage and cell cycle arrest; Cortes-Bratti X et al.; The cytolethal distending toxins (CDTs) are a newly discovered family of bacterial protein toxins with the unique ability to interfere with the cell cycle, causing irreversible cell cycle arrest and consequently death of the target cells . CDTs are encoded by three linked genes (cdtA, cdtB and cdtC) and are produced by a variety of Gram negative bacteria . The mechanism of action of this toxin family only now begins to be elucidated . CDTs are internalized by endocytosis and require an intact Golgi complex to exert their cytotoxic activity . The CdtB component was shown to have functional homology with the mammalian deoxyribonuclease I (DNase I) and the induction of cell cycle arrest in mammalian cells mimicked that induced by DNA damaging agents, suggesting that DNA is the cellular target . Still there are many issues that need to be clarified, such as identification of the function(s) of CdtA and CdtC, characterization of the receptor(s), understanding of the final steps of the internalization pathway and localization of the active component . This review focuses mainly on the effect of CDTs on mammalian cells, highlighting the questions that remain to be answered regarding their molecular mode of action. Int J Syst Evol Microbiol, 2001 Sep, 51(Pt 5), 1911 - 6 Desulforegula conservatrix gen . nov., sp . nov., a long-chain fatty acid-oxidizing, sulfate-reducing bacterium isolated from sediments of a freshwater lake; Rees GN et al.; A novel sulfate-reducing bacterium, strain Mb1PaT, was isolated from the sediments of a freshwater floodplain lake . Cells of strain Mb1PaT were rod-shaped, 1-1.3 microm wide and 2.6-3 microm long, motile and Gram-negative . The bacterium grew on straight-chain carboxylic acids with 4-17 carbon atoms . Electron donors with an even number of carbon atoms were oxidized to acetate and electron donors with an odd number of carbon atoms were oxidized to acetate and propionate . No other compounds were found to be used as electron donors . No growth occurred in the absence of sulfate . The optimum temperature for growth was between 25 and 30 degrees C and the maximum temperature for growth was 32 degrees C . Strain Mb1PaT grew very slowly in medium with 5 g NaCl l(-1) with optimum growth occurring with up to 1.0 g NaCl l(-1) . Analysis of the 16S rRNA gene showed that strain Mb1PaT belonged to the delta-subclass of the Proteobacteria, was a member of the family Desulfobacteraceae, but lacked similarity with any currently described representatives . The combined phylogenetic analysis and physiological data indicate that strain Mb1PaT represents a new genus and the name Desulforegula conservatrix is proposed . The type strain is Mb1PaT (= DSM 13527T = ATCC BAA-134T). Int J Syst Evol Microbiol, 2001 Sep, 51(Pt 5), 1873 - 80 Methanofollis aquaemaris sp . nov., a methanogen isolated from an aquaculture fish pond; Lai MC et al.; A novel methanogen, designated strain N2F9704T, was isolated from an aquaculture fish pond near Wang-gong, Taiwan . The cells were irregular cocci, non-motile, 1.2-2.0 microm in diameter and stained gram-negative . Cells of strain N2F9704T lysed easily by SDS treatment (0.1 g l(-1)) and the S-layer protein had an Mr of 137000 . The catabolic substrates used included formate and H2+CO2, but not acetate, methanol, trimethylamine or secondary alcohols . The optimal growth parameters for strain N2F9704T were pH 6.5, 37 degrees C with 0.5% NaCl . Trace amounts of tungstate not only promoted growth but also extended the range of growth conditions . Analysis of the 16S rDNA sequence revealed a phylogenetic relationship to Methanofollis species and the name Methanofollis aquaemaris sp . nov . is therefore proposed for strain N2F9704T (= OCM 746T = CCRC 16166T) . Additionally, the strain was infected with a novel coccus-shaped, enveloped virus with a diameter of 200 nm. Int J Syst Evol Microbiol, 2001 Sep, 51(Pt 5), 1825 - 9 Natronobacterium nitratireducens sp . nov., a aloalkaliphilic archaeon isolated from a soda lake in China; Xin H et al.; Two novel haloalkaliphilic archaea, strains C231T and C42, were isolated from a soda lake in China . Cells of the two strains were rod-shaped and gram-negative and colonies were bright red . They required at least 2.5 M NaCl for growth, with an optimum at 3.5 M NaCl, and grew over a pH range from 8.0 to 10.5, with an optimum at pH 8.5 . Hypotonic treatment with less than 1.5 M NaCl caused cell lysis . They had similar polar lipid compositions, possessing the diphytanyl (C20:C20) and phytanyl-sesterterpanyl (C20:C25) diether derivatives of phosphatidylglycerol and phosphatidylglycerophosphate methyl ester and a minor phospholipid, PL1 . No glycolipids were detected . Comparison of 16S rDNA sequences and morphological features placed them in the genus Natronobacterium . Detailed phenotypic characterization and DNA-DNA hybridization studies revealed that the two strains belong to a new species in the genus Natronobacterium, for which the name Natronobacterium nitratireducens sp . nov . is proposed . The type strain is C231T (= AS 1.1980T = JCM 10879T). Int J Syst Evol Microbiol, 2001 Sep, 51(Pt 5), 1663 - 9 Methanoculleus chikugoensis sp . nov., a novel methanogenic archaeon isolated from paddy field soil in Japan, and DNA-DNA hybridization among Methanoculleus species; Dianou D et al.; A strictly anaerobic, irregularly coccoid, methanogenic archaeon, strain MG62T (= JCM 10825T = DSM 13459T), was isolated from paddy field soil in Chikugo, Fukuoka, Japan . The cells stained gram-negative, were 1.0-2.0 microm in diameter, were lysed by SDS and hypotonic solutions and were flagellated . Motility was not observed . The strain was able to use H2/CO2, 2-propanol/CO2, formate, 2-butanol/CO2 and cyclopentanol/CO2 as substrates for methanogenesis, but did not utilize acetate, ethanol, methanol or methylamines . The optimum temperature and pH were 25-30 degrees C and 6.7-7.2 . Analysis of lipid component parts (core lipids, phospholipid polar head groups and glycolipid sugar moieties) showed the characteristic pattern of members of the family Methanomicrobiaceae except for the absence of glucose as a glycolipid sugar moiety . The G+C content of the DNA was 62.2 mol % . Sequence analysis of the 16S rDNA revealed that the strain belonged to the genus Methanoculleus . The strain had DNA-DNA hybridization values of less than 50% with type strains of Methanoculleus species . On the basis of phenotypic, genotypic and phylogenetic characteristics, the name Methanoculleus chikugoensis sp . nov . is proposed for strain MG62T (= JCM 10825T = DSM 13459T) . The DNA hybridization study also revealed the close relationships of three species, Methanoculleus olentangyi, Methanoculleus bourgensis and Methanoculleus oldenburgensis, among Methanoculleus species. Dis Aquat Organ, 2001 Aug 22, 46(1), 57 - 66 Parasitism by the protozoan Perkinsus atlanticus favours the development of opportunistic infections; Montes JF et al.; It has been suggested that opportunistic pathogens could contribute to the mortality of Perkinsus atlanticus-infected clams . Examination of Tapes semidecussatus clams from the northern Mediterranean coast of Spain revealed that while 86% of the clams heavily infected with P . atlanticus were co-infected by bacteria and/or viruses, neither non-infected nor lightly P . atlanticus-infected specimens had bacterial or viral infections . The bacteria, which had a Gram-negative cell wall, were always located in the apical pole of gill epithelial cells and enclosed within membranous compartments . Bacteria-containing cells were hypertrophied and showed dysplasia with loss of cilia and microvilli . The viruses shared ultrastructural, morphologic and cytopathic characteristics of a polyomavirus . Viral particles with icosahedral symmetry were found in both the cytoplasm and the nucleus of numerous cell types . Virus-infected cells showed severe alterations, including hypertrophy, reduction of the intracellular compartments and extrusion of the nuclear envelope . Moreover, gill epithelial cells showed disorganization and swelling of the apical region, which affected the ciliary structure . Our findings show that P . atlanticus parasitism favours the development of opportunistic infections which have detrimental effects in this clam population. J Biol Chem, 2001 Dec 7, 276(49), 46568 - 74 Epub 2001 Oct 08. Amino acid sequence determinants of extended spectrum cephalosporin hydrolysis by the class C P99 beta-lactamase; Zhang Z et al.; Class C beta-lactamases are commonly encoded on the chromosome of Gram-negative bacterial species . Mutations leading to increased expression of these enzymes are a common cause of resistance to many cephalosporins including extended spectrum cephalosporins . Recent reports of plasmid- and integrin-encoded class C beta-lactamases are a cause for concern because these enzymes are likely to spread horizontally to susceptible strains . Because of their increasing clinical significance, it is critical to identify the determinants of catalysis and substrate specificity of these enzymes . For this purpose, the codons of a set of 21 amino acid residues that encompass the active site region of the P99 beta-lactamase were individually randomized to create libraries containing all possible amino acid substitutions . The amino acid sequence requirements for the hydrolysis of ceftazidime, an extended spectrum cephalosporin commonly used to treat serious infections, were determined by selecting resistant mutants from each of the 21 libraries . DNA sequencing identified the residue positions that are critical for ceftazidime hydrolysis . In addition, it was found that certain amino acid substitutions in the omega-loop region of the P99 enzyme result in increased ceftazidime hydrolysis suggesting the loop is an important determinant of substrate specificity. Trends Plant Sci, 2001 Oct, 6(10), 479 - 85 Molecular secrets of bacterial type III effector proteins; Lahaye T et al.; Most Gram-negative phytopathogenic bacteria are thought to inject effector proteins into the plant cell via a type III secretion system that is essential for pathogenicity . Plant targets and the mode of action of type III effector proteins, which include avirulence (Avr) proteins, are largely unknown . However, recent findings have shed light on the molecular mechanisms of Avr action . Here, we focus on two classes of Avr proteins (the AvrBs3 and AvrRxv/YopJ families) that have been suggested to act as transcription factors and proteases, respectively. Eur J Biochem, 2001 Oct, 268(19), 5092 - 7 Structural investigation on the lipooligosaccharide fraction of psychrophilic Pseudoalteromonas haloplanktis TAC 125 bacterium; Corsaro MM et al.; The core structure of the cell-wall lipooligosaccharide (LOS) fraction of an Antarctic Gram-negative bacterium, Pseudoalteromonas haloplanktis TAC 125 strain, was determined to be deacetylated alditols . These were obtained from native LOS fraction by O-deacylation, dephosphorylation, reduction and finally N-deacylation . Two novel structures were detected, the more highly represented molecule consisting of the following hexasaccharide chain: alpha-D-ManpNH(2)-(1-->3)-beta-D-Galp-(1-->4)-alpha-L-glycero-D-manno-Hepp-(1-->5)-alpha-D-Kdo-(2-->6)-beta-D-GlcpNH(2)-(1-->6)-D-GlcNH(2)(ol) while the corresponding pentasaccharide, lacking the ManpNH(2) residue, was less abundant . To the best of our knowledge, the structural investigation presented here, mainly performed by NMR and MS methods, is the first report of the lipopolysaccharide fraction of a psychrophilic bacterium. Cornea, 2001 Oct, 20(7), 765 - 8 An unusual case of fungal keratitis: Metarrhizium anisopliae; Jani BR et al.; PURPOSE: To report a case of fungal keratitis caused by Metarrhizium anisopliae, which to our knowledge is the first reported case in the United States . METHOD: Case report . RESULTS: A 36-year-old female librarian who wore extended-wear soft contact lenses was seen by an ophthalmologist on September 11 for an irritated right eye, and a corneal ulcer was diagnosed . Symptoms increased by September 27, and the patient was referred to another ophthalmologist who cultured the ulcer and had scrapings examined, which were Gram-negative for microorganisms . The patient was referred to one of the authors (W.J.R.) . Her exam on October 1 showed vision corrected to 20/25 OD, a 5-mm epithelial defect with a 2.5-mm anterior stromal grayish-type infiltrate, and a quiet anterior chamber; the eye did not appear to be inflamed . The patient was reexamined on October 4 and was noted to have worsening vision . Because the initial cultures remained negative, the patient underwent a corneal biopsy, Gram stain, and cultures on October 6 . Scrapings at the time of the biopsy revealed septate hyphal elements, as did the biopsy specimen, and on October 7, the patient was started on a treatment of bacitracin ointment once a day and natamycin 5% every hour . The eye gradually quieted down . A mold growing from the biopsy culture, which had been sent to a reference laboratory in San Antonio, Texas, was identified as M . anisopliae var . anisopliae . The patient was subsequently fitted with a rigid gas permeable lens, which resulted in a best-corrected visual acuity of 20/20, although glare remained a major problem . CONCLUSION: Although not previously reported in the United States, M . anisopliae can cause a keratomycosis, and one must consider this common insect pathogen in the differential diagnosis of fungal keratitis. Curr Opin Microbiol, 2001 Oct, 4(5), 565 - 9 Integrons: natural tools for bacterial genome evolution; Rowe-Magnus DA et al.; Integrons were first identified as the primary mechanism for antibiotic resistance gene capture and dissemination among Gram-negative bacteria . More recently, their role in genome evolution has been extended with the discovery of larger integron structures, the super-integrons, as genuine components of the genomes of many species throughout the gamma-proteobacterial radiation . The functional platforms of these integrons appear to be sedentary, whereas their gene cassette contents are highly variable . Nevertheless, the gene cassettes for which an activity has been experimentally demonstrated encode proteins related to simple adaptive functions and their recruitment is seen as providing the bacterial host with a selective advantage . The widespread occurrence of the integron system among Gram-negative bacteria is discussed, with special focus on the super-integrons . Some of the adaptive functions encoded by these genes are also reviewed, and implications of integron-mediated genome evolution in the emergence of novel bacterial species are highlighted. Perit Dial Int, 2001 Jul-Aug, 21(4), 405 - 10 Late renal transplant failure: an adverse prognostic factor at initiation of peritoneal dialysis; Sasal J et al.; BACKGROUND: Early renal transplant failure necessitating a return to dialysis has been shown to be a poor prognostic factor for survival . Little is known about the outcome of patients with late transplant failure returning to dialysis . It was our clinical impression that late transplant failure (>2 months) carries an increased morbidity and mortality risk in patients returning to dialysis . OBJECTIVE: To determine whether patients with a failed renal transplant have an outcome different to those on dialysis who have never received a kidney transplant . SETTING: Peritoneal dialysis (PD) unit in a teaching hospital . PATIENTS AND DESIGN: All failed renal transplant patients (fTx) in the Toronto Hospital Peritoneal Dialysis program between 1989 and 1996 were identified . This cohort of 42 fTx patients was compared with a cohort of randomly selected never-transplanted PD patients (non-Tx).The PD program was selected because of the availability of well-documented patient archival material . The non-Tx group was matched for age and presence of diabetes . Data were collected until retransplantation, change of dialysis modality or center, death, or until June 1998 . RESULTS: There was no difference at initiation of PD between groups in serum albumin, residual renal function, or mean serum parathyroid hormone level . The mean low-density lipoprotein level was significantly higher in the fTx cohort . The duration of dialysis before Tx in fTx patients accounted for the increased total length of dialysis in fTx (mean 15 months) . However, post-Tx the duration of PD was similar for both groups (30.7 months for fTx vs 31.6 months for non-Tx) . The fTx group had a considerably worse outcome than the non-Tx group . The time to first peritonitis, subsequent episodes of peritonitis, catheter change, or transfer to hemodialysis occurred at a much faster rate in fTx patients.The most dramatic difference was in survival . There were 3 deaths in the non-Tx group and 12 in the fTx group (p < 0.01) . The mean age at time of death in the fTx group was 47.5 years . Deaths were due mainly to gram-negative peritonitis and cardiovascular disease . CONCLUSIONS: We conclude that late failed renal transplant patients starting dialysis are at increased risk of complications and have strikingly higher mortality rates than non-Tx patients . A previously failed kidney transplant can be considered an adverse prognostic factor for patients commencing PD; these patients need to be closely monitored . Although this study was limited to PD patients, the same principles likely apply to fTx patients returning to any form of renal replacement therapy. J Nutr, 2001 Oct, 131(10), 2789S - 2793S Helicobacter pylori: ulcers and more: the beginning of an era; Lacy BE et al.; Helicobacter pylori (H . pylori) may be found in up to 50% of the world's population, making it the most common infectious disease worldwide . H . pylori is a gram-negative, microaerophilic rod with flagella and normally resides in the stomach . It can be diagnosed endoscopically, via breath tests, or through a simple blood test . H . pylori can usually be eradicated with a combination of proton pump inhibitors and antibiotics . Although most people infected with this organism never develop a complication, H . pylori infection in others may produce significant mucosal inflammation leading to ulcers in the stomach and duodenum . H . pylori has also been recognized as a risk factor for the development of both gastric carcinoma and mucosal-associated lymphoid tumor . Although not widely known, H . pylori can also affect organ systems outside of the gastrointestinal tract . It is now apparent that H . pylori can infect the skin, liver and heart and that these infections may produce a number of different disease states . In addition, H . pylori infection can adversely affect the nutritional status of both children and adults . This article discusses the epidemiology and bacteriology of H . pylori, reviews the different methods of diagnosing and treating this common infection, and then focuses on the effects that H . pylori has on different organ systems within the body, including the nutritional status of those affected. FEMS Microbiol Lett, 2001 Sep 25, 203(2), 141 - 8 Cytolethal distending toxin (CDT): a bacterial weapon to control host cell proliferation? De Rycke J, Oswald E. Cytolethal distending toxins (CDT) constitute a family of genetically related bacterial protein toxins able to stop the proliferation of numerous cell lines . This effect is due to their ability to trigger in target cells a signaling pathway that normally prevents the transition between the G2 and the M phase of the cell cycle . Produced by several unrelated Gram-negative mucosa-associated bacterial species, CDTs are determined by a cluster of three adjacent genes (cdtA, cdtB, cdtC) encoding proteins whose respective role is not yet fully elucidated . The CDT-B protein presents sequence homology to several mammalian and bacterial phosphodiesterases, such as DNase I . The putative nuclease activity of CDT-B, together with the activation by CDT of a G2 cell cycle checkpoint, strongly suggests that CDT induces an as yet uncharacterized DNA alteration . However, the effective entry of CDT into cells and subsequent translocation into the nucleus have not yet been demonstrated by direct methods . The relationship between the potential DNA-damaging properties of this original family of toxins and their role as putative virulence factors is discussed. FEMS Microbiol Lett, 2001 Sep 25, 203(2), 131 - 9 Ubiquinone biosynthesis in microorganisms; Meganathan R; The quinoid nucleus of the benzoquinone, ubiquinone (coenzyme Q; Q), is derived from the shikimate pathway in bacteria and eukaryotic microorganisms . Ubiquinone is not considered a vitamin since mammals synthesize it from the essential amino acid tyrosine . Escherichia coli and other Gram-negative bacteria derive the 4-hydroxybenzoate required for the biosynthesis of Q directly from chorismate . The yeast, Saccharomyces cerevisiae, can either form 4-hydroxybenzoate from chorismate or tyrosine . However, unlike mammals, S . cerevisiae synthesizes tyrosine in vivo by the shikimate pathway . While the reactions of the pathway leading from 4-hydroxybenzoate to Q are the same in both organisms the order in which they occur differs . The 4-hydroxybenzoate undergoes a prenylation, a decarboxylation and three hydroxylations alternating with three methylation reactions, resulting in the formation of Q . The methyl groups for the methylation reactions are derived from S-adenosylmethionine . While the prenyl side chain is formed by the 2-C-methyl-D-erythritol 4-phosphate (non-mevalonate) pathway in E . coli, it is formed by the mevalonate pathway in the yeast. Arch Pediatr, 2001 Sep, 8 Suppl 4, 726s - 731s {Virulence factors associated with E . coli neonatal meningitis}; Bonacorsi S et al.; Escherichia coli K1 is the leading cause of gram-negative bacterial meningitis in neonates . It is associated with a mortality rate as high as 40%, and more than half of the survivors have neurologic sequelae . Bacterial meningitis is the result of bacterial translocation from gastrointestinal tract to the blood and from blood to the central nervous system . Successful crossing of the BBB by E . coli K1 requires (a) a high degree of bacteremia and (b) several E . coli determinants contributing to invasion of BMEC such as the K1 capsule, Sfa, Ibe proteins, and CNF1 . A better understanding for the molecular basis of E . coli K1 penetration of the BBB could potentially lead to the development of novel therapeutic and preventative strategies for E . coli K1 meningitis. J Endotoxin Res, 2001, 7(3), 167 - 202 Bacterial lipopolysaccharides and innate immunity; Alexander C et al.; Bacterial lipopolysaccharides (LPS) are the major outer surface membrane components present in almost all Gram-negative bacteria and act as extremely strong stimulators of innate or natural immunity in diverse eukaryotic species ranging from insects to humans . LPS consist of a poly- or oligosaccharide region that is anchored in the outer bacterial membrane by a specific carbohydrate lipid moiety termed lipid A . The lipid A component is the primary immunostimulatory centre of LPS . With respect to immunoactivation in mammalian systems, the classical group of strongly agonistic (highly endotoxic) forms of LPS has been shown to be comprised of a rather similar set of lipid A types . In addition, several natural or derivatised lipid A structures have been identified that display comparatively low or even no immunostimulation for a given mammalian species . Some members of the latter more heterogeneous group are capable of antagonizing the effects of strongly stimulatory LPS/lipid A forms . Agonistic forms of LPS or lipid A trigger numerous physiological immunostimulatory effects in mammalian organisms, but--in higher doses--can also lead to pathological reactions such as the induction of septic shock . Cells of the myeloid lineage have been shown to be the primary cellular sensors for LPS in the mammalian immune system . During the past decade, enormous progress has been obtained in the elucidation of the central LPS/lipid A recognition and signaling system in mammalian phagocytes . According to the current model, the specific cellular recognition of agonistic LPS/lipid A is initialized by the combined extracellular actions of LPS binding protein (LBP), the membrane-bound or soluble forms of CD14 and the newly identified Toll-like receptor 4 (TLR4)*MD-2 complex, leading to the rapid activation of an intracellular signaling network that is highly homologous to the signaling systems of IL-1 and IL-18 . The elucidation of structure-activity correlations in LPS and lipid A has not only contributed to a molecular understanding of both immunostimulatory and toxic septic processes, but has also re-animated the development of new pharmacological and immunostimulatory strategies for the prevention and therapy of infectious and malignant diseases. Acta Gastroenterol Latinoam, 2001, 31(3), 137 - 41 {Role of cytokines in chronic gastritis by Helicobacter pylori}; Jimenez FP et al.; Helicobacter pylori is a curved, gram negative bacterium that inhabits only the gastric mucous membrane . Since its discovery and characterization, it has been related to the physiopathology of gastroduodenal diseases, including gastritis, peptic ulcers, gastric carcinoma and MALT lymphoma . This has resulted in numerous hypotheses that try to explain the different events that take place during the inflammation . The bacterium Settler, characterized by a marked infiltration of inflammatory cells (neutrophils, monocytes, linfocytes, etc.) which, after being activated, liberate locally various chemical mediators, which cause tissue damage . Among these, the cytokines are important mediators in this process . We have revised the literature related to the various biological functions of cytokines in tissue damage of the gastric mucosa. Microbiology, 2001 Oct, 147(Pt 10), 2643 - 9 FindTarget: software for subtractive genome analysis; Chetouani F et al.; In silico subtractive/differential genome analysis is a powerful approach for identifying genus- or species-specific genes, or groups of genes that are responsible for a unique phenotype . By this method, one searches for genes present in one group of bacteria and absent in another group . A software package has been developed, named FindTarget, that has a user-friendly web interface to facilitate differential genome analysis . The user chooses the genomes to compare, the similarity criteria and the thresholds to decide if a gene has a counterpart in another genome . The searches are based on BLASTP comparisons of proteomes . FindTarget also includes access to sequences, coloured multiple alignments, phylogenetic trees of conserved proteins and links to public annotated databases which provide a means for validation of the results . To illustrate this approach, a FindTarget search for genes putatively involved in the specificity of cell envelope synthesis of Gram-negative bacteria is presented . The results show that most of the identified genes are clearly involved in cell wall processes, underlining the power of such an approach in general and that of FindTarget in particular. J Clin Microbiol, 2001 Oct, 39(10), 3548 - 54 Aortic valve endocarditis in a dog due to Bartonella clarridgeiae; Chomel BB et al.; We report the first documented case of endocarditis associated with Bartonella clarridgeiae in any species . B . clarridgeiae was identified as a possible etiological agent of human cat scratch disease . Infective vegetative valvular aortic endocarditis was diagnosed in a 2.5-year-old male neutered boxer . Historically, the dog had been diagnosed with a systolic murmur at 16 months of age and underwent balloon valvuloplasty for severe valvular aortic stenosis . Six months later, the dog was brought to a veterinary hospital with an acute third-degree atrioventricular block and was diagnosed with infective endocarditis . The dog died of cardiopulmonary arrest prior to pacemaker implantation . Necropsy confirmed severe aortic vegetative endocarditis . Blood culture grew a fastidious, gram-negative organism 8 days after being plated . Phenotypic and genotypic characterization of the isolate, including partial sequencing of the citrate synthase (gltA) and 16S rRNA genes indicated that this organism was B . clarridgeiae . DNA extraction from the deformed aortic valve and the healthy pulmonic valve revealed the presence of B . clarridgeiae DNA only from the diseased valve . No Borrelia burgdorferi or Ehrlichia sp . DNA could be identified . Using indirect immunofluorescence tests, the dog was seropositive for B . clarridgeiae and had antibodies against Ehrlichia phagocytophila but not against Ehrlichia canis, Ehrlichia ewingii, B . burgdorferi, or Coxiella burnetii. Br J Anaesth, 2001 Apr, 86(4), 581 - 6 Review of management of purpura fulminans and two case reports; Nolan J et al.; Purpura fulminans (PF) is a haemorrhagic condition usually associated with sepsis or previous infection . Features include tissue necrosis, small vessel thrombosis and disseminated intravascular coagulation . Gram-negative organisms are the commonest cause of the acute infectious type, which is often associated with multi-organ failure . An idiopathic variety, however, is often confined to the skin . The mortality rate has decreased with better treatment of secondary infections, supportive care and new treatments, but it remains a disabling condition often requiring major amputations . We describe two cases and review the various treatments for this condition. J Pediatr Gastroenterol Nutr, 2001 Aug, 33(2), 165 - 70 Age-dependent changes in the regulation of cyclooxygenases in the gastrointestinal tract after gram-negative endotoxemia; Martinez FE et al.; BACKGROUND: Cyclooxygenases (COXs) modulate prostaglandin synthesis in the gastrointestinal tract . Prostaglandins have been shown to have a cytoprotective effect on bowel mucosa in adults, but no similar data are available in neonates . Thus, the purpose of the current study was to evaluate age-dependent changes in gastrointestinal tract COX regulation after Escherichia coli lipopolysaccharide exposure in rats . METHODS: Stomach, small bowel, and large bowel COX-1 and COX-2 mRNA levels (reverse transcription polymerase chain reaction technique) and protein content (Western blot) were obtained from neonates (younger than 3 days old) and adult rats 18 hours after exposure to E . coli O111:B4 lipopolysaccharide toxin . Untreated animals served as controls . RESULTS: Stomach, small bowel, and large bowel tissue COX-1 mRNA levels in the newborn were significantly lower (P < 0.01) than in the adult . No age-dependent differences were found for COX-2 mRNA levels . After lipopolysaccharide exposure, no significant changes in COX-1 levels were seen at either age, whereas COX-2 mRNA levels were increased only in the stomach for both ages . Western blot analysis of small bowel tissue for COX-1 and COX-2 showed no lipopolysaccharide-induced changes in protein content, but the COX-1 content was significantly lower in the newborn (P < 0.01) . CONCLUSIONS: In the rat, COX expression in the gastrointestinal tract is regulated in an age-dependent fashion . Lower COX-1 expression and a lack of observable increase in COX-2 mRNA levels in the newborn small bowel after endotoxemia may render the bowel more susceptible to bowel injury early in life. Vaccine, 2001 Oct 12, 20(1-2), 235 - 41 Antibodies to BrkA augment killing of Bordetella pertussis; Oliver DC et al.; BrkA is a Bvg-regulated Bordetella pertussis protein that mediates serum resistance and adherence . It shares sequence identity with another B . pertussis virulence factor called pertactin, and it is a member of the diverse group of proteins found in Gram-negative bacteria that are secreted by an autotransporter mechanism . Sera, either from individuals who have been vaccinated with acellular pertussis vaccines, or from individuals who have no re-collection of recent infection with B . pertussis fail to kill wild-type B . pertussis, but kill brkA mutant strains very well . We examined whether BrkA could be neutralised in serum fitting this profile . BrkA is synthesised as a 103kDa precursor that is processed into a surface-associated N-terminal 73kDa passenger domain, and an outer-membrane embedded C-terminal 30kDa transporter moiety . Polyclonal antibodies were raised to a recombinant, re-folded histidine-tagged fusion protein representing the 73kDa passenger region . These anti-BrkA antibodies were shown to boost the existing bactericidal capacity of human serum against B . pertussis by neutralising BrkA. EMBO J, 2001 Sep 17, 20(18), 5040 - 8 Subtilisin-like autotransporter serves as maturation protease in a bacterial secretion pathway; Coutte L et al.; Proteins of Gram-negative bacteria destined to the extracellular milieu must cross the two cellular membranes and then fold at the appropriate time and place . The synthesis of a precursor may be a strategy to maintain secretion competence while preventing aggregation or premature folding (especially for large proteins) . The secretion of 230 kDa filamentous haemagglutinin (FHA) of Bordetella pertussis requires the synthesis and the maturation of a 367 kDa precursor that undergoes the proteolytic removal of its approximately 130 kDa C-terminal intramolecular chaperone domain . We have identified a specific protease, SphB1, responsible for the timely maturation of the precursor FhaB, which allows for extracellular release of FHA . SphB1 is a large exported protein with a subtilisin-like domain and a C-terminal domain typical of bacterial autotransporters . SphB1 is the first described subtilisin-like protein that serves as a specialized maturation protease in a secretion pathway of Gram-negative bacteria . This is reminiscent of pro-protein convertases of eukaryotic cells. EMBO J, 2001 Sep 17, 20(18), 5033 - 9 Crystal structure of the outer membrane protease OmpT from Escherichia coli suggests a novel catalytic site; Vandeputte-Rutten L et al.; OmpT from Escherichia coli belongs to a family of highly homologous outer membrane proteases, known as omptins, which are implicated in the virulence of several pathogenic Gram-negative bacteria . Here we present the crystal structure of OmpT, which shows a 10-stranded antiparallel beta-barrel that protrudes far from the lipid bilayer into the extracellular space . We identified a putative binding site for lipopolysaccharide, a molecule that is essential for OmpT activity . The proteolytic site is located in a groove at the extracellular top of the vase-shaped beta-barrel . Based on the constellation of active site residues, we propose a novel proteolytic mechanism, involving a His-Asp dyad and an Asp-Asp couple that activate a putative nucleophilic water molecule . The active site is fully conserved within the omptin family . Therefore, the structure described here provides a sound basis for the design of drugs against omptin-mediated bacterial pathogenesis . Coordinates are in the Protein Data Bank (accession No . 1I78) J Biotechnol, 2001 Oct 4, 91(2-3), 197 - 209 Detection, purification and characterisation of quorum-sensing signal molecules in plant-associated bacteria; Brelles-Marino G et al.; Quorum sensing (also called autoinduction) is a term that describes an environmental sensing system that allows bacteria to monitor their own population density . Autoinduction relies upon the interaction of a small diffusible signal molecule (the autoinducer) with a transcriptional activator protein to couple gene expression with cell population density . These signal molecules diffuse from bacterial cells and accumulate in the environment as a function of cell growth . Once a threshold concentration is reached, these signals serve as co-inducers to regulate the transcription of (a) set(s) of target genes . In Gram-negative bacteria, most autoinducers belong to the family of N-acylhomoserine lactones (AHLs) . The detection of AHLs (or AHL-like activities) has been greatly facilitated by the development of sensitive bioassays that allow fast screening of microorganisms for diffusible signal molecules . AHL or diketopiperazine-mediated cell-cell signalling play roles in regulating different bacterial functions, such as antibiotic biosynthesis, production of virulence factors, exopolysaccharide biosynthesis, bacterial swarming, plasmid conjugal transfer and transition into the stationary phase . Several bacterial species that interact with plants produce AHL-like compounds . In this review, we will summarise the current knowledge about the detection, characterisation and purification of quorum-sensing molecules from plant-associated bacteria . We will also discuss some of the future prospects and biotechnological applications of autoinducers. Arzneimittelforschung, 2001, 51(8), 690 - 7 Results of an open, non-placebo controlled pilot study investigating the immunomodulatory potential of autovaccine; Rusch V et al.; Autovaccines are prepared from autologous, human, non-pathogenic, "rough" variants of E . coli derived from the stool flora of individuals according to a highly standardized procedure . As a fundamental concept within microbiological therapy, these autovaccines are mainly used to treat chronic inflammatory disorders associated with impaired immune reactions resistant to standard therapeutic treatments . Generally, immunomodulatory effects of outer membrane components or cell wall fragments of gram-negative bacteria on innate or adaptive immunity are widely accepted but nevertheless mechanisms of actions of these autovaccines remained obscure, despite some recent publication about other autovaccine preparations of different origin . Hence, immunomodulating properties of autovaccine were investigated in a pilot study with 78 outpatients with variable disorders ranging from recurrent respiratory infections to diffuse gastrointestinal complaints . Patients received their autologous bacteria parenterally in increasing doses . Before application and 4 to 6 weeks after application of autovaccine, blood samples of the patients were taken to investigate a range of immunological parameters such as acute phase proteins, serum antibodies and cytokines . The results revealed that autovaccines were able to modulate significantly the release of three potent immunoregulatory cytokines e.g . interferon-gamma, granulocyte-macrophage-colony stimulating factor and interleukin-1 beta, whereas specific humoral immunity remained largely unaffected . From these results it may be concluded that the autovaccine mainly act antigen non-specifically on the cytokine level rather than inducing a specific vaccination . Further studies with more detailed kinetic measurements of cytokines will have to verify these results. Cytokine, 2001 Aug 7, 15(3), 156 - 65 Induction of proinflammatory and chemokine genes by lipopolysaccharide and paclitaxel (Taxol) in murine and human breast cancer cell lines; Zaks-Zilberman M et al.; In murine macrophages, the anti-tumor agent, paclitaxel, induces expression of a wide variety of inflammatory and anti-inflammatory genes, and causes cytokine secretion via signaling pathways that overlap with those engaged by lipopolysaccharide (LPS), the endotoxic component of Gram-negative bacteria . Using semi-quantitative RT-PCR for detection of gene expression, coupled with ELISA for the detection of secreted gene products, we analyzed the responsiveness of an extensive panel of cytokine and non-cytokine genes to induction by paclitaxel and LPS in the murine DA-3 breast cancer line . A subset of the genes examined (e.g., G-CSF, MIP-2, iNOS, and IL-1 beta, and GM-CSF) was upregulated >3-20-fold by both LPS and paclitaxel in the DA-3 cell line, while IP-10 mRNA was induced by paclitaxel, but not by LPS . In the human MDA-MB-231 breast cancer cell line, LPS also increased mRNA levels for both GM-CSF and IP-10 significantly, while, paclitaxel increased IP-10 mRNA levels with delayed kinetics and failed to induce GM-CSF mRNA . Co-cultures of murine breast cancer cells and macrophages, stimulated with IFN-gamma plus either paclitaxel or LPS, resulted in augmented release of nitric oxide . As both GM-CSF and IP-10 have been implicated in tumor rejection in vivo through either indirect actions on the host immune system or by inhibiting tumor angiogenesis, our data strengthen the hypothesis that tumor cell-derived inflammatory mediators may, in part, underlie the anti-tumor efficacy of paclitaxel in breast cancer . J Mol Biol, 2001 Sep 7, 312(1), 143 - 55 The structure of CMP:2-keto-3-deoxy-manno-octonic acid synthetase and of its complexes with substrates and substrate analogs; Jelakovic S et al.; The enzyme CMP-Kdo synthetase (CKS) catalyzes the activation of the sugar Kdo (2-keto-3-deoxy-manno-octonic acid) by forming a monophosphate diester . CKS is a pharmaceutical target because CMP-Kdo is used in the biosynthesis of lipopolysaccharides that are vital for Gram-negative bacteria . We have refined the structure of the unligated capsule-specific CKS from Escherichia coli at 1.8 A resolution (1 A=0.1 nm) and we have established the structures of its complexes with the substrate CTP, with CDP and CMP as well as with the product analog CMP-NeuAc (CMP-sialate) by X-ray diffraction analyses at resolutions between 2.1 A and 2.5 A . The N-terminal domains of the dimeric enzyme bind CTP in a peculiar nucleotide-binding fold, whereas the C-terminal domains form the dimer interface . The observed binding geometries together with the amino acid variabilities during evolution and the locations of a putative Mg(2+) and of a very strongly bound water molecule suggest a pathway for the catalysis . The N-terminal domain shows sequence homology with the CMP-NeuAc synthetases . Moreover, the chain fold and the substrate-binding position of CKS resemble those of other enzymes processing nucleotide-sugars . Infection, 2001 Aug, 29(4), 205 - 8 Infections caused by Stenotrophomonas maltophilia--a prospective study; Schaumann R et al.; BACKGROUND: Stenotrophomonas maltophilia is an opportunistic microorganism, often highly resistant to routinely tested antibiotics . This microorganism is isolated in specimens from patients with nosocomial infections with increasing frequency . PATIENTS AND METHODS: During a 1-year period (1998/1999) S . maltophilia was isolated from 137 specimens (0.26% of all investigated specimens) from 80 patients who were treated in a 1,500 bed major tertiary care teaching hospital in Leipzig . The data of 76 patients (133 specimens) could be collected and analyzed completely . RESULTS: The pathogen was most frequently detected in specimens from the respiratory tract (54%) . In five patients (six cases) S . maltophilia was isolated from blood cultures (0.3% of all positive blood cultures; 1.4% of all gram-negative isolates from blood cultures) . 70 of the infected patients were inpatients and 32 (42%) of them were treated on the internal medicine wards . Of these 32 patients only six (19%) were pretreated with imipenem . The Length of stay at the hospital resulted in an independent increased risk of infection with S . maltophilia . In addition, this organism was detected in six infected outpatients . CONCLUSION: S . maltophilia is not only a nosocomial pathogen . Pretreatment with a carbapenem is no longer an unequivocal risk factor for an infection with S . maltophilia. Nature, 2001 Sep 6, 413(6851), 78 - 83 Mal (MyD88-adapter-like) is required for Toll-like receptor-4 signal transduction; Fitzgerald KA et al.; The recognition of microbial pathogens by the innate immune system involves Toll-like receptors (TLRs), which recognize pathogen-associated molecular patterns . Different TLRs recognize different pathogen-associated molecular patterns, with TLR-4 mediating the response to lipopolysaccharide from Gram-negative bacteria . All TLRs have a Toll/IL-1 receptor (TIR) domain, which is responsible for signal transduction . MyD88 is one such protein that contains a TIR domain . It acts as an adapter, being involved in TLR-2, TLR-4 and TLR-9 signalling; however, our understanding of how TLR-4 signals is incomplete . Here we describe a protein, Mal (MyD88-adapter-like), which joins MyD88 as a cytoplasmic TIR-domain-containing protein in the human genome . Mal activates NF-kappaB, Jun amino-terminal kinase and extracellular signal-regulated kinase-1 and -2 . Mal can form homodimers and can also form heterodimers with MyD88 . Activation of NF-kappaB by Mal requires IRAK-2, but not IRAK, whereas MyD88 requires both IRAKs . Mal associates with IRAK-2 by means of its TIR domain . A dominant negative form of Mal inhibits NF-kappaB, which is activated by TLR-4 or lipopolysaccharide, but it does not inhibit NF-kappaB activation by IL-1RI or IL-18R . Mal associates with TLR-4 . Mal is therefore an adapter in TLR-4 signal transduction. J Immunol, 2001 Sep 15, 167(6), 3316 - 23 Predominant role of toll-like receptor 2 versus 4 in Chlamydia pneumoniae-induced activation of dendritic cells; Prebeck S et al.; Chlamydia pneumoniae is an obligate intracellular human pathogen causing diseases such as pneumonia, bronchitis, and pharyngitis . Because of its intracellular replication, cell-mediated immune responses are needed to mediate successful defenses of the host . Because dendritic cells play a central role in linking innate immunity and Ag-specific cell-mediated immune responses we asked whether dendritic cells are activated upon contact with C . pneumoniae and whether known Toll like receptors (TLR) are involved in this process . Here we show that C . pneumoniae was taken up by bone marrow-derived murine dendritic cells . Ingested C . pneumoniae appeared to be unable to develop mature inclusion inside dendritic cells . Furthermore, upon contact with C . pneumoniae dendritic cells were potently stimulated because NF-kappaB was activated and translocated to the nucleus, cytokines like IL-12p40 and TNF-alpha were secreted, and expression of MHC class II molecules, CD40, CD80, and CD86 was up-regulated . Importantly, secretion of cytokines as well as translocation of NF-kappaB were dependent on the presence of TLR2 and independent from TLR4 with the exception of IL-12p40 secretion, which was attenuated in the absence of either a functional TLR2 or 4 . In conclusion, we show here that recognition of the Gram-negative bacterium C . pneumoniae depends largely on TLR2 and only to a minor extent on TLR4. J Bacteriol, 2001 Oct, 183(19), 5639 - 44 Novel macrolide-specific ABC-type efflux transporter in Escherichia coli; Kobayashi N et al.; In the Escherichia coli genome, five putative open reading frame (ORF) clusters, mdlAB, ybjYZ, yddA, yojHI, and yhiH, have been assumed to be possible genes for ABC drug efflux transporters (I . T . Paulsen, M . K . Sliwinski, and M . H . Saier, Jr., J . Mol . Biol . 277:573-592, 1998) . We cloned all of these ORFs in multicopy plasmids and investigated the drug resistance of drug-supersensitive host cells lacking constitutive multidrug efflux transporter genes acrAB . Among them, only ybjYZ gave significant erythromycin resistance and significantly decreased the accumulation of {(14)C}erythromycin . Therefore, ybjYZ was renamed macAB (macrolide-specific ABC-type efflux carrier) . Plasmids carrying both the macA and -B genes conferred resistance against macrolides composed of 14- and 15-membered lactones but no or weak resistance against 16-membered ones . Neither of the two genes produced resistance alone . The DNA sequence suggests that MacB is an integral membrane protein with four transmembrane segments and one nucleotide-binding domain, while MacA belongs to a membrane fusion protein (MFP) family with a signal-like sequence at its N terminus . The expression of the histidine-tagged proteins confirmed that MacB is an integral membrane protein and MacA is a peripheral membrane protein . In addition, MacAB required TolC for its function in a way similar to that of most of the MFP-dependent transporters in E . coli . MacB is thus a novel ABC-type macrolide efflux transporter which functions by cooperating with the MFP MacA and the multifunctional outer membrane channel TolC . This is the first case of an experimentally identified ABC antibiotic efflux transporter in gram-negative organisms. JPEN J Parenter Enteral Nutr, 1991 Sep-Oct, 15(5), 503 - 8 Arginine supplementation improves histone and acute-phase protein synthesis during gram-negative sepsis in the rat; Leon P et al.; Mechanisms of nutrient alteration of hepatic protein synthesis during sepsis are unclear . In vitro, arginine downregulates endotoxin-stimulated hepatocyte protein synthesis but in vivo effects are unknown . This study evaluated the effects of supplemental arginine or glycine on fibrinogen (acute-phase protein), histone, albumin, and liver protein synthesis after Gram-negative sepsis in the rat . Adult rats (225 g, n=36) were randomized to receive isonitrogenous isocaloric total parenteral nutrition supplemented with 264 mg of N per kilogram per day as either arginine or glycine . On day 5, each group was further randomized to control or sepsis . Sepsis was induced by injection of 8 x 10(7) Escherichia coli per 100 g body weight, and then a continuous infusion of {1-14C} leucine was started . The rats were sacrificed 4 hours later . The fractional protein synthesis rates (percent per day) of histone, fibrinogen, albumin, and liver were determined . Supplemental arginine led to significantly increased histone (p < 0.05, analysis of variance) and fibrinogen (p < 0.01, analysis of variance) synthesis in the septic rats compared with all other groups . Histone and albumin synthesis were also significantly increased (p < 0.05) in the arginine-supplemented control group compared with the glycine-supplemented control group . Arginine supplementation during sepsis significantly increased (p < 0.05) albumin and liver protein synthesis compared with controls . Histones which are involved in DNA synthesis and are rich in arginine may play a role in the host response to stress and sepsis . These in vivo results appear to contradict hepatocyte-Kupffer cell coculture studies perhaps because of the hormonal and cytokine responses to nutrient substrate and acute septicemia. Anal Chem, 2001 Aug 15, 73(16), 3804 - 7 Direct microextraction and analysis of rough-type lipopolysaccharides by combined thin-layer chromatography and MALDI mass spectrometry; Therisod H et al.; A rapid method for the microscale extraction of lipopolysaccharides (endotoxins, LPSs) from rough-type Gram-negative bacteria was developed using thin-layer chromatography (TLC) combined with improved conditions for LPS analysis by mass spectrometry . TLC of intact bacteria on silica gel plates in an appropriate solvent selectively extracted and separated their LPS components . The bands of molecular species were scraped from the plates after nondestructive visualization, directly mixed with matrix, and analyzed by laser desorption time-of-flight mass spectrometry . Lipids A and Re-type LPSs were analyzed after transfer to a membrane . Adding citric acid to the matrix gave greatly improved mass spectra . The system allows characterization of bacterial LPS at the microscale level and is equally well applicable to heterogeneous LPS and lipid A preparations (Escherichia coli lipid A and Bordetella lipopolysaccharides were used) . The technique provides a rapid determination of the heterogeneity of unmodified preparations and the determination of the molecular weight of each separated component. Biol Blood Marrow Transplant, 2001, 7(7), 384 - 7 Association of TLR4 mutations and the risk for acute GVHD after HLA-matched-sibling hematopoietic stem cell transplantation; Lorenz E et al.; Lipopolysaccharide (LPS) has been implicated in the pathogenesis of graft-versus-host disease (GVHD) . The toll-like receptor (TLR)-4 has been recently identified as a major receptor for LPS . Mutations of TLR4 have been associated with LPS hyporesponsiveness . We hypothesized that TLR4 mutations reduce the risk of acute GVHD in allogeneic marrow transplant recipients . In a preliminary study to determine the frequency of TLR4 mutations and their possible association with GVHD, we tested 237 patients and their HLA-identical sibling donors for 2 TLR4 polymorphisms . All patients received methotrexate and cyclosporine for GVHD prophylaxis . One or more mutants were detected in 10.8% of patients and 10.6% of donors . Multivariable logistic regression models were used to analyze the association between TLR4 mutations and probability (1-sided) of GVHD . The odds ratio (adjusted for advanced disease, total body irradiation dose, and patient age) for development of grades II to IV GVHD when a mutation was present in the recipient was 0.63 (95% confidence interval {CI}, 0.25-1.60; P = .16) . When a mutation was present in the donor, the adjusted odds ratio was 0.88 (95% CI, 0.36-2.17; P = .40) . When a mutation was present in both recipient and donor, the odds ratio was 0.72 (95% CI, 0.22-2.32; P = .29) . Among 24 patients with TLR4 mutations in either donor or recipient, 4 (16.7%) developed gram-negative bacteremia . Among 213 patients without mutations, 14 (6.6%) developed gram-negative bacteremia (P = .09) . The data indicate that a reduced risk of acute GVHD is associated with TLR4 mutations and that TLR4 mutations may increase the risk for gram-negative bacteremia . However, these associations are not statistically significant in recipients of HLA-matched sibling marrow transplants who are prophylactically treated for infections and GVHD . A much larger study population would be needed to confirm the role of LPS in the pathogenesis of GVHD in humans. J Environ Monit, 1999 Apr, 1(2), 163 - 8 Use of quadrupole GC-MS and ion trap GC-MS-MS for determining 3-hydroxy fatty acids in settled house dust: relation to endotoxin activity; Saraf A et al.; Gas chromatography-mass spectrometry (GC-MS) using a quadrupole instrument and GC-tandem MS (GC-MS-MS) using an ion trap instrument were applied to determine 3-hydroxy fatty acids (3-OH FAs) with 10-18 carbon chain lengths, specific components of the endotoxin (lipopolysaccharide, LPS) of Gram-negative bacteria, in 30 house dust samples . The two methods provided similar detection sensitivity for methyl ester/trimethylsilyl derivatives of the 3-OH FAs and allowed these acids to be distinguished from co-eluting 2-OH FA derivatives . The correlation coefficients between endotoxin activity (Limulus test) and the combined amounts of 3-OH C10, 3-OH C12, and 3-OH C14 were 0.60 and 0.61 when using GC-MS and GC-MS-MS, respectively . The superior selectivity of GC-MS-MS was illustrated in analyses of sub-milligram amounts of dust, where the chromatograms achieved by GC-MS were difficult to interpret due to a high background and several closely eluting compounds . GC-MS-MS is therefore preferable to GC-MS for determining 3-OH FAs in minute (sub-milligram) amounts of dust. Genes Immun, 2001 Aug, 2(5), 258 - 62 Polymorphisms of Fc gamma-receptors RIIa, RIIIa, and RIIIb in patients with adult periodontal diseases; Meisel P et al.; Polymorphisms influencing the binding affinity between the Fcgamma receptors and IgG of different subclasses are thought to be of importance in the individual susceptibility to infections with Gram-negative bacteria contributing to periodontal disease . One hundred and fifty-four Caucasian subjects were clinically and radiographically examined for their periodontal status and genotyped for their allelic pattern of FcgammaRIIa, FcgammaRIIIa, and FcgammaIIIb polymorphism . In assessing periodontitis according to mean probing depth and attachment loss, no differences were found in allele frequencies or combined allotypes between the subjects with mild or moderate and those with severe signs of periodontitis . However, the extent and severity of bone loss were significantly associated with the genotype of the receptor FcgammaRIIIa . An increased risk of severe bone destruction was observed in individuals carrying the FcgammaRIIIa-VV genotype (OR = 5.3; 95% CI 1.4-26.2) . FcgammaRIIIb is in linkage disequilibrium with FcgammaRIIIa . Hence it is also related to periodontal disease . There is no indication of an association between the polymorphism of FcgammaRIIa and periodontitis . The results are evidence that the FcgammaRIIIa genotype coding for the high affinity receptor imposes an additional risk of bone loss as does the FcgammaRIIIb genotype coding for the low affinity receptor. Biochem Cell Biol, 2001, 79(4), 449 - 59 Helicobacter pylori from asymptomatic hosts expressing heptoglycan but lacking Lewis O-chains: Lewis blood-group O-chains may play a role in Helicobacter pylori induced pathology; Monteiro MA et al.; Helicobacter pylori is a widespread Gram-negative bacterium responsible for the onset of various gastric pathologies and cancers in humans . A familiar trait of H . pylori is the production of cell-surface lipopolysaccharides (LPSs; O-chain --> core --> lipid A) with O-chain structures analogous to some mammalian histo-blood-group antigens, those being the Lewis determinants (Lea, Leb, Lex, sialyl Lex, Ley) and blood groups A and linear B . Some of these LPS antigens have been implicated as autoimmune, adhesion, and colonization components of H . pylori pathogenic mechanisms . This article describes the chemical structures of LPSs from H . pylori isolated from subjects with no overt signs of disease . Experimental data from chemical- and spectroscopic-based studies unanimously showed that these H . pylori manufactured extended heptoglycans composed of 2- and 3-linked D-glycero-alpha-D-manno-heptopyranose units and did not express any blood-group O-antigen chains . The fact that another H . pylori isolate with a similar LPS structure was shown to be capable of colonizing mice indicates that H . pylori histo-blood-group structures are not an absolute prerequisite for colonization in the murine model also . The absence of O-chains with histo-blood groups may cause H . pylori to become inept in exciting an immune response . Additionally, the presence of elongated heptoglycans may impede exposure of disease-causing outer-membrane antigens . These factors may render such H . pylori incapable of creating exogenous contacts essential for pathogenesis of severe gastroduodenal diseases and suggest that histo-blood groups in the LPS may indeed play a role in inducing a more severe H . pylori pathology. Nat Immunol, 2001 Sep, 2(9), 882 - 8 Inducible IL-2 production by dendritic cells revealed by global gene expression analysis; Granucci F et al.; Dendritic cells (DCs) are strong activators of primary T cell responses . Their priming ability is acquired upon encounter with maturation stimuli . To identify the genes that are differentially expressed upon maturation induced by exposure to Gram-negative bacteria, a kinetic study of DC gene expression was done with microarrays representing 11,000 genes and ESTs (expressed sequence tags) . Approximately 3000 differentially expressed transcripts were identified . We found that functional interleukin 2 (IL-2) mRNA, which gave rise to IL-2 production, was transiently up-regulated at early time-points after bacterial encounter . In contrast, macrophages did not produce IL-2 upon bacterial stimulation . Thus, IL-2 is an additional key cytokine that confers unique T cell stimulatory capacity to DCs. Appl Environ Microbiol, 2001 Sep, 67(9), 4024 - 9 Enzymatic manganese(II) oxidation by a marine alpha-proteobacterium; Francis CA et al.; A yellow-pigmented marine bacterium, designated strain SD-21, was isolated from surface sediments of San Diego Bay, San Diego, Calif., based on its ability to oxidize soluble Mn(II) to insoluble Mn(III, IV) oxides . 16S rRNA analysis revealed that this organism was most closely related to members of the genus Erythrobacter, aerobic anoxygenic phototrophic bacteria within the alpha-4 subgroup of the Proteobacteria (alpha-4 Proteobacteria) . SD-21, however, has a number of distinguishing phenotypic features relative to Erythrobacter species, including the ability to oxidize Mn(II) . During the logarithmic phase of growth, this organism produces Mn(II)-oxidizing factors of approximately 250 and 150 kDa that are heat labile and inhibited by both azide and o-phenanthroline, suggesting the involvement of a metalloenzyme . Although the expression of the Mn(II) oxidase was not dependent on the presence of Mn(II), higher overall growth yields were reached in cultures incubated with Mn(II) in the culture medium . In addition, the rate of Mn(II) oxidation appeared to be slower in cultures grown in the light . This is the first report of Mn(II) oxidation within the alpha-4 Proteobacteria as well as the first Mn(II)-oxidizing proteins identified in a marine gram-negative bacterium. Mar Pollut Bull, 2001 Aug, 42(8), 689 - 95 The effect of TBT on the structure of a marine sediment comunity--a Boxcosm study; Dahllof I et al.; The effect of tri-n-butyl tin (TBT) on an intact marine sediment community after five months exposure was investigated . Changes in the structure of macro- and meiofauna communities were determined, as well as the functional diversity of the microbial community using BIOLOG microplates for Gram negative bacteria . Development of tolerance in the microbial community was investigated using Pollution Induced Community Tolerance (PICT) experiments with fluxes of nutrients as effect indicators . TBT affected the structure and recruitment of the macro- and meiofauna at nominal additions of 30-137 micromol TBT/m2 sediment . Number of species, diversity, biomass and community similarity was reduced at these concentrations compared to control . Species that molt seemed to be the most tolerant since they were predominant in boxes that had received the highest TBT addition and echinoderms were the most sensitive species . Renewed addition of TBT in PICT experiments with sediment from each boxcosm showed that TBT had an effect on individual nutrient fluxes from all sediments . Analyses of the flux patterns revealed a memory of previous TBT exposure, either due to induced tolerance or other community conditioning. Extremophiles, 2001 Aug, 5(4), 257 - 64 A novel replication element from an Antarctic plasmid as a tool for the expression of proteins at low temperature; Tutino ML et al.; Genetic manipulation of Antarctic bacteria has been very limited so far . This article reports the isolation and molecular characterization of a novel plasmid, pMtBL, from the Antarctic gram-negative bacterium Pseudoalteromonas haloplanktis TAC 125 . This genetic element, 4,081 bp long, appeared to be a multicopy cryptic replicon with no detectable transcriptional activity . By an in vivo assay, the pMtBL autonomous replication sequence was functionally limited to an AluI plasmid fragment of about 850 bp . This novel cold-adapted replication element showed quite a broad host range profile: it was cloned into a mesophilic genetic construction, obtaining a cold-adapted expression vector that was able to promote the production of P . haloplanktis A23 alpha-amylase in a psychrophilic bacterium . This study represents the first report of successful recombinant production of a cold-adapted protein in an Antarctic host. J Endotoxin Res, 2001, 7(2), 147 - 55 Influence of acyl chain fluidity on the lipopolysaccharide-induced activation of complement; Wiese A et al.; Lipopolysaccharides (LPSs, endotoxins) are the major amphiphilic constituents of the outer leaflet of the outer membrane of Gram-negative bacteria . They are known to activate the complement cascade to form lytic membrane pores . Here, we study the influence of the fluidity of the acyl chains of LPSs and lipid As on the formation of lytic pores . To this end, we have performed electrical measurements on asymmetric planar endotoxin/phospholipid bilayers as a reconstitution model of the outer membrane using two deep rough mutant LPSs (from Escherichia coli strains WBB01 and WBB25) and two lipid As (from E . coli WBB25 and Rhodobacter sphaeroides) . The two LPSs and the two lipid As each differ in their acylation pattern which is correlated with the fluidity . The addition of human serum to the endotoxin side of the bilayers led to the formation of membrane pores, and pore formation correlated in each case with acyl chain fluidity, i.e . time required for the first lytic pore to be formed was shorter for the more fluid endotoxin . Furthermore, in the case of LPSs, the activation rate was higher for the more fluid membrane and the respective bacteria had a higher susceptibility to the growth inhibitory action of serum. J Endotoxin Res, 2001, 7(1), 69 - 72 Outer membrane protein A (OmpA), peptidoglycan-associated lipoprotein (PAL), and murein lipoprotein (MLP) are released in experimental Gram-negative sepsis; Hellman J et al.; We previously showed that Escherichia coli bacteria incubated in normal human serum release complexes that contain three conserved Gram-negative bacterial outer membrane proteins (OMPs) and LPS . We have identified the OMPs as outer membrane protein A (OmpA), peptidoglycan-associated lipoprotein (PAL), and murein lipoprotein (MLP) . These OMPs are conserved among enteric Gram-negative bacteria and are bound by IgG in antisera raised to heat-killed rough bacteria such as E . coli J5 (J5 IgG) . The present experiments were performed to further analyze the release of these OMPs in a rat wound infection model of sepsis . Plasma was collected from thermally injured rats with E . coli O18 sepsis and filtered . LPS was affinity-purified from plasma filtrates using monoclonal antibody specific for the O-polysaccharide side chain of E . coli O18 LPS . Plasma filtrates were also incubated with J5 IgG conjugated to magnetic beads . Affinity-purified samples were analyzed for the OMPs by immunoblotting . OmpA, PAL, and MLP were released into septic rat blood in complexes with LPS . PAL was consistently present in samples affinity-purified using J5 IgG . The results indicate that OmpA, PAL, and MLP are released and circulate in experimental Gram-negative sepsis and suggest that a proportion of released OMPs are tightly associated with LPS. J Endotoxin Res, 2000, 6(6), 477 - 82 Plasma constituents regulate LPS binding to, and release from, the monocyte cell surface; Kitchens RL et al.; Innate immunity to Gram-negative bacteria involves regulated mechanisms that allow sensitive but limited responses to LPS . Two important pathways that lead to host cell activation and LPS deactivation involve: (i) LPS interactions with CD14 and Toll-like receptor 4 on cells (activation); and (ii) LPS sequestration by plasma lipoproteins (deactivation) . Whereas these pathways were previously thought to be independent and essentially irreversible, we found that they are connected by a third pathway: (iii) the movement of LPS from host cells to plasma lipoproteins . Our data show that, in the presence of human plasma, LPS binds transiently to monocyte surfaces and then moves from the cell surface to plasma lipoproteins . Soluble CD14 enhances LPS release from cells in the presence of lipoproteins, whereas LPS binding protein and phospholipid transfer protein do not . The transfer of cell-bound LPS to lipoproteins is accompanied by reduced cell responses to the LPS, suggesting that the movement of LPS from leukocytes into lipoproteins may attenuate host responses to LPS in vivo . Preliminary data suggest that changes that occur in the plasma after trauma or during sepsis decrease LPS binding to leukocytes while greatly increasing the rate of LPS release from cells. J Endotoxin Res, 2000, 6(5), 389 - 91 Innate recognition of lipopolysaccharide by Toll-like receptor 4/MD-2 and RP105/MD-1; Miyake K et al.; The Toll family of receptors has been implicated in innate recognition and subsequent activation of defense programs against pathogens such as bacteria and fungi . TLR4, for example, signals the presence of lipopolysaccharide (LPS), a membrane constituent of Gram-negative bacteria . LPS signaling via TLR4 is greatly enhanced by a molecule referred to as MD-2, which is associated with the extracellular domain of TLR4 . The TLR4/MD-2 complex, therefore, recognizes LPS . RP105, another member of the Toll family, has a striking similarity to TLR4 in that it is associated with an MD-2-like molecule MD-1 . B-cells lacking RP105 are severely impaired in LPS-induced proliferation and antibody production . Studies employing transfectants showed that RP105/MD-1, like MD-2, enhances the LPS signaling via TLR4 . RP105/MD-1 thus constitutes an LPS-signaling complex on B-cells . These results suggest that a variety of cell surface molecules regulate LPS recognition/signaling by TLR4. Immunity, 2001 Aug, 15(2), 313 - 21 Role of Janus kinase 3 in mast cell-mediated innate immunity against gram-negative bacteria; Malaviya R et al.; Mast cells play a pivotal role in innate host immune response to gram-negative bacteria . We report that Janus kinase 3 plays a role in mast cell-mediated bacterial clearance and neutrophil recruitment by regulating the release of tumor necrosis factor from mast cells . The role of JAK3 in mast cell-facilitated neutrophil recruitment and bacterial clearance was investigated by comparing the neutrophil influxes and bacterial clearance in mast cell-deficient W/W(v) mice reconstituted with JAK3(+/+) or JAK(-/-) mast cells . The neutrophil influx, bacterial clearance, and survival outcome in W/W(v) mice reconstituted with JAK3(+/+) mast cells was better than in W/W(v) mice reconstituted with JAK3(-/-) mast cells . These findings provide evidence that JAK3 is a key regulator of mast cell-mediated innate immunity against gram-negative bacteria. Antonie Van Leeuwenhoek, 2001 Jun, 79(2), 209 - 17 Methanotrophic diversity in an agricultural soil as evaluated by denaturing gradient gel electrophoresis profiles of pmoA, mxaF and 16S rDNA sequences; Fjellbirkeland A et al.; Molecular methods were used to characterize the diversity of a methanotrophic population in an agricultural soil . For this purpose we have used DGGE analysis of functional and phylogenetic markers . Functional markers utilised comprised the pmoA-gene coding for the alpha-subunit of the particulate methane monooxygenase (pMMO) present in all known methanotrophs and the mxaF-gene coding for the alpha-subunit of methanol dehydrogenase (MDH) present in all gram-negative methylotrophs . In addition, we have used 16S rDNA as a phylogenetic marker . DGGE patterns of an enrichment culture, and sequencing of major DGGE bands obtained with the bacterial specific primers showed that the community structure was dominated by methanotrophic populations related to Methylobacter sp . and Methylomicrobium sp . The PCR products amplified with the functional primer sets were related to both type I and type II methanotrophs . We also designed a new pmoA-targeting primer set which could be used in a nested protocol to amplify PCR-products from DNA extracted directly from the soil. Handchir Mikrochir Plast Chir, 2001 Jul, 33(4), 262 - 6 {Plasma endotoxin, procalcitonin, C-reactive protein, and organ functions in patients with major burns}; Ulrich D et al.; Sepsis is one of the most frequent causes of death after major burn injury . Usually, sepsis appears as a consequence of a gram-negative bacteriaemia with release of endotoxins.In this study, the plasma endotoxin levels of seven patients (three female, four male; average age 51.3 +/- 23.8 years) with burns between 43.5 and 78 % Total Body Surface Area (Abbreviated Burn Severity Index 8 - 12) were determined for five days after thermal trauma every three hours by ELISA and compared with the concentration of procalcitonin (PCT) and C-reactive protein (CRP) . A calculation of the Horrowitz-Index (PaO(2)/FiO(2)) and the Pressure-Adjusted Heart Rate (HR x CVP/MAP) took place to show a possible correlation between the endotoxin concentration and the cardiopulmonary organ function . Additionally, we analysed whether operative treatment can influence the level of plasma endotoxin in the early phase after burn injury.At any time after burn trauma, endotoxins could be detected in the plasma of all patients . Between the second and third day, there was a considerable increase in the endotoxin concentration with a maximum after 57 hours of 0.48 +/- 0.32 EU/ml . Two patients with sepsis and death in the further course had a rather distinctive increase . From the fourth day on, occasional episodes of increases in endotoxin concentration were noted . Postoperatively, there was a short increase in plasma endotoxin on the second and fourth day . The plasma endotoxin level showed no correlation with the PCT and CRP or with the oxygenation in the patients' blood . However, a positive correlation could be observed with the Pressure-Adjusted Heart Rate (p = 0.0061; r(2) = 0.212).An explanation for the endotoxin increase after 57 hours could be the translocation of intestinal bacteria, the beginning of bacterial colonisation or decomposition products of the burn wound with protein-protein complexes . Later on, infectious diseases such as pneumonia with gram-negative bacteria are of importance, too.According to the Two-Hit Model, the increase of plasma endotoxin can serve as a trigger and cause a recurrence of systemic inflammation with the changes observed in cardiac organ function, multiple organ dysfunction, and multiple organ failure. Sheng Wu Gong Cheng Xue Bao, 2001 May, 17(3), 241 - 5 {The new antiendotoxin strategies}; Wang DN et al.; Lipopolysaccharide(LPS) is important in the pathogenesis of sepsis infected by gram-negative bacteria in humans, and part or all of this pathophysiology is mediated by a complex secondary inflammatory response . Administration of purified LPS or a variety of LPS-free recombinant mediators can reproduce much of the pathophysiology of gram-negative sepsis . There has been an explosion of new information over the last several years regarding the interactions of LPS with proteins and cells . Blocking sepsis at the bacterial toxin level is appealing because it is upstream, so that the secondary pathologic inflammatory cascade may be limited or prevented . A variety of antiendotoxin strategies have been proposed . Therapies under investigation include agents that bind and neutralize LPS, agents or systems that enhance LPS clearance, and agents that inhibit LPS interaction with serum elements or cellular receptors. Scand J Infect Dis, 2001, 33(7), 533 - 7 Risk factors and prognostic indicators of bacterial meningitis in a cohort of 3580 postneurosurgical patients; Federico G et al.; In order to identify the incidence, risk factors and prognostic indicators of postneurosurgical bacterial meningitis, a 9-y retrospective cohort study was performed . The cohort comprised 3580 individuals who underwent neurosurgical operations at the Department of Neurosurgery of Catholic University, Rome, a 1,700-bed university hospital . We observed 52 episodes of postneurosurgical meningitis, with infection rates of 1.4/100 patients, 0.8/100 operations for craniotomies and 2.6/100 operations for internal cerebrospinal fluid (CSF) shunt or ventriculostomy with external drainage . The overall infection rate decreased from 2.5/100 operations in 1989 to 0.9/100 in 1997 (p = 0.03) . Logistic regression analysis indicated that duration of ventriculostomy with external drainage {p < 0.01; odds ratio (OR) = 9.67; 95% confidence interval (CI) = 2.44-38.321 and increasing value of the APACHE III score (p <0.01; OR = 8.51; 95% CI = 2.15-33.68) were independent risk factors for development of meningitis . The overall case fatality rate was 8% . Predictors of mortality were low ( < 1.66 mmol/l) CSF glucose concentration (p = 0.001), increasing value of the APACHE IIl score (p = 0.002) and Gram-negative aetiology (p = 0.003). Genetics, 2001 Aug, 158(4), 1657 - 64 Excess of rare amino acid polymorphisms in the Toll-like receptor 4 in humans; Smirnova I et al.; The Toll-like receptor 4 protein acts as the transducing subunit of the lipopolysaccharide receptor complex and assists in the detection of Gram-negative pathogens within the mammalian host . Several lines of evidence support the view that variation at the TLR4 locus may alter host susceptibility to Gram-negative infection or the outcome of infection . Here, we surveyed TLR4 sequence variation in the complete coding region (2.4 kb) in 348 individuals from several population samples; in addition, a subset of the individuals was surveyed at 1.1 kb of intronic sequence . More than 90% of the chromosomes examined encoded the same structural isoform of TLR4, while the rest harbored 12 rare amino acid variants . Conversely, the variants at silent sites (intronic and synonymous positions) occur at both low and high frequencies and are consistent with a neutral model of mutation and random drift . The spectrum of allele frequencies for amino acid variants shows a significant skew toward lower frequencies relative to both the neutral model and the pattern observed at linked silent sites . This is consistent with the hypothesis that weak purifying selection acted on TLR4 and that most mutations affecting TLR4 protein structure have at least mildly deleterious phenotypic effects . These results may imply that genetic variants contributing to disease susceptibility occur at low frequencies in the population and suggest strategies for optimizing the design of disease-mapping studies. Am J Surg, 2001 Jun, 181(6), 571 - 5 The effect of glucagon-like peptide 2 on intestinal permeability and bacterial translocation in acute necrotizing pancreatitis; Kouris GJ et al.; BACKGROUND: Acute pancreatitis (AP) initiates a generalized inflammatory response that increases intestinal permeability and promotes bacterial translocation (BT) . Impairment of the intestinal epithelial barrier is known to promote BT . Glucagon-like peptide 2 (GLP-2), a 33 residue peptide hormone, is a key regulator of the intestinal mucosa by stimulating epithelial growth . The purpose of this study was to determine whether GLP-2 decreases intestinal permeability and BT in AP . METHODS: To examine whether GLP-2 can decrease intestinal permeability and thereby decrease BT in acute necrotizing pancreatitis, 34 male Sprague-Dawley rats (200 to 300 g) were studied . AP was induced in group I and group II by pressure injection of 3% taurocholate and trypsin into the common biliopancreatic duct (1 mg/kg of body weight) . The potent analog to GLP-2 called ALX-0600 was utilized . Group I rats received GLP-2 analog (0.1 mg/kg, SQ, BID) and group II rats received a similar volume of normal saline as a placebo postoperatively for 3 days . Group III and group IV received GLP-2 analog and placebo, respectively . At 72 hours postoperatively, blood was drawn for culture of gram-negative organisms . Specimens from mesenteric lymph nodes (MLN), pancreas and peritoneum were harvested for culture of gram-negative bacteria . Intestinal resistance as defined by Ohm's law was determined using a modified Ussing chamber to measure transepithelial current at a fixed voltage . A point scoring system for five histologic features that include intestinal edema, inflammatory cellular infiltration, fat necrosis, parenchymal necrosis, and hemorrhage was used to evaluate the severity of pancreatitis . Specimens from MLN, pancreas, jejunum, and ileum were taken for pathology . RESULTS: All group I and group II rats had AP . The average transepithelial resistance in group I was 82.8 Omega/cm(2) compared with 55.9 Omega/cm(2) in group II (P <0.01) . Gram-negative BT to MLN, pancreas, and peritoneum was 80%, 0%, and 0%, respectively in group I compared with 100%, 30%, and 20% translocation in group II . CONCLUSION: GLP-2 treatment significantly decreases intestinal permeability in acute pancreatitis. Clin Transpl . 2000;:297-310. Sixteen-year experience with 1,000 heart transplants at UCLA; Marelli D et al.; 1 . The consecutive pre- and post-1994 eras have demonstrated improved survival for all age groups . This is linked to improved preservation methods, surgical technique and immunosuppression agents . 2 . The use of marginal donor hearts for Status I and alternate elderly patients has followed the model of matching donor and recipient risk without affecting patient outcome and minimized the use of implantable assist devices . 3 . A donor history of systemic gram-negative infection, hypertension, or traumatic intracranial bleeds was an important marker for risk . Younger age and shorter ischemia time could compensate for other hazards . 4 . Heart transplantation in carefully selected elderly recipients yielded clinical results similar to those of younger patients with less rejection . 5 . An adult alternate recipient list proved useful to prevent diversion of standard donors away from younger recipients . 6 . Retransplantation for TCAD is acceptable but much less satisfactory for acute graft failure . 7 . Trends show an increase in the use of implantable devices; refinement in technology for mechanical assist and replacement is forthcoming. Intensive Care Med, 2001 Aug, 27(8), 1274 - 80 Extremely low doses of tissue factor pathway inhibitor decrease mortality in a rabbit model of septic shock; Matyal R et al.; OBJECTIVE: We sought to determine the lowest dose of recombinant human tissue factor pathway inhibitor (TFPI) that can provide protection from lethality in a rabbit model of septic shock . METHODS: Sepsis was induced in New Zealand white rabbits by intraperitoneal implantation of 7.0 ml of a solution containing hemoglobin (4.8 g/dl), porcine mucin (6 g/dl), and 0.8-1.4 x 10(4) viable Escherichia coli (strain O:18 K+) . Gentamicin (5 mg/kg) was administered 4 h following surgery, and this dose was repeated every 12 h for 3 days . Beginning 4 h following the induction of sepsis, animals were treated with a bolus (1 ml) plus a continuous infusion (100 ml over 24) of either TFPI (various doses) or its vehicle . Four different doses of TFPI were studied, and each experiment included a contemporaneous control group . The primary outcome parameter was survival time . Results were analyzed using the Wilcoxen log rank test . RESULTS: The average survival time for rabbits treated with the highest dose of TFPI tested (50 microg/kg bolus and 0.5 microg/kg per minute infusion) was 118 h, as compared to 81 h in vehicle-treated controls) . The average survival time for septic rabbits treated with a much lower dose of TFPI (100 ng/kg bolus and 1.0 ng/kg per minute infusion) was 119 h as compared to 57 h in surviving vehicle-treated controls . Treatment with an even lower dose of TFPI (10 ng/kg bolus and 0.1 ng/kg per minute infusion) still produced a marginally significant prolongation of average survival time (80 h) relative to contemporaneously studied controls (47 h) . When the dose of TFPI was decreased still further (1.0 ng/kg bolus and 0.01 ng/kg per minute infusion), average survival times were not significantly different between TFPI-treated and vehicle-treated rabbits (77 and 51 h, respectively) . CONCLUSIONS: Delayed infusion with remarkably low doses of recombinant human TFPI prolongs survival in a rabbit model of antibiotic-treated Gram-negative bacterial sepsis . In planning human trials of TFPI as an adjuvant treatment for sepsis it may be reasonable to use much lower doses of the agent than were heretofore contemplated. J Immunol, 2001 Sep 1, 167(5), 2895 - 901 Differential role of CD18 integrins in mediating lung neutrophil sequestration and increased microvascular permeability induced by Escherichia coli in mice; Gao X et al.; The in vivo contributions of CD18 integrin-dependent and -independent mechanisms in mediating the increases in lung neutrophil (polymorphonuclear leukocyte; PMN) sequestration and microvascular permeability are not well understood . We determined the time course of these responses to Gram-negative sepsis in the mouse lung and addressed the specific contributions of CD18 integrins and ICAM-1 . PMN sequestration in the lung was assessed by morphometric analysis, and transalveolar PMN migration was assessed by bronchoalveolar lavage . Lung tissue PMN number increased by 6-fold within 1 h after i.p . Escherichia coli challenge; this value peaked at 3 h (7-fold above control) and decreased at 12 h (3.5-fold above control) . PMN migration into the airspace was delayed; the value peaked at 6 h and remained elevated up to 12 h . Saturating concentrations of anti-CD18 and anti-ICAM-1 mAbs reduced lung tissue PMN sequestration and migration; however, peak responses at 3 and 6 h were inhibited by 40%, indicating that only a small component of PMN sequestra