JAMA, 1995 Jul 26, 274(4), 338 - 45 Long-term survival and function after suspected gram-negative sepsis; Perl TM et al.; OBJECTIVE--To determine the long-term (> 3 months) survival of septic patients, to develop mathematical models that predict patients likely to survive long-term, and to measure the health and functional status of surviving patients . SETTING--A large tertiary care university hospital and an associated Veterans Affairs Medical Center . DESIGN--From December 1986 to December 1990, a total of 103 patients with suspected gram-negative sepsis entered a double-blind, placebo-controlled efficacy trial of monoclonal antiendotoxin antibody . Of these, we followed up 100 patients for 7667 patient-months . Beginning in May 1992, we reviewed hospital records and contacted all known survivors . We measured the health status of all surviving patients . MAIN OUTCOME MEASURES--The determinants of long-term survival (up to 6 years) were identified through two Cox proportional hazard regression models: one that included patient characteristics identified at the time of sepsis (bedside model) and another that included bedside, infection-related, and treatment characteristics (overall model) . RESULTS--Of the 60 patients in the cohort who died at a median interval of 30.5 days after sepsis, 32 died within the first month of the septic episode, seven died within 3 months, and four more died within 6 months . In the bedside multivariate model constructed to predict long-term survival, large hazard ratios (HRs) were associated with severity of underlying illness as classified by McCabe and Jackson criteria (for rapidly fatal disease, HR = 30.4, P < .001; for ultimately fatal disease, HR = 7.6, P < .001) and the use of vasopressors (HR = 2.5; P = .001) . In the overall model for long-term survival, severity of underlying illness (rapidly fatal disease, HR = 23.7, P < .001; ultimately fatal disease, HR = 6.5, P < .001), number of active comorbid illnesses (HR = 1.3; P = .04), use of vasopressors at the time of sepsis (HR = 2.0; P = .02), and development of adult respiratory distress syndrome (HR = 2.3; P = .02) predicted patients most likely to die . The Acute Physiology and Chronic Health Evaluation II score was not a significant predictor of outcome when either model included the simpler McCabe and Jackson classification of underlying disease severity . We compared the health status scores with norms for the general population and found that patients with resolved sepsis reported more physical dysfunction (P < .001), including problems with work and activities of daily living (P = .02), and more poorly perceived general health (P < .01) . In contrast, patients' scores for perceived emotional health were higher than those in the general population (P = .004) . The mean Barthel score of our patients was 85 (100 = total independence) and the mean Eastern Cooperative Oncology Group score was 0.7 (0 = normal, 4 = 100% bedridden), suggesting that the patients' physical function was not normal . CONCLUSIONS--At the onset of suspected gram-negative sepsis, severity of underlying illness and in-hospital use of vasopressors are strong and consistent predictors of short- and long-term survival . Our data validate the McCabe and Jackson severity of illness scoring system for predicting long-term survival after sepsis . Physical dysfunction and more poorly perceived general health occur commonly after sepsis. Proc Natl Acad Sci U S A, 1995 Jul 18, 92(15), 7085 - 9 Structure of the gene for porcine peptide antibiotic PR-39, a cathelin gene family member: comparative mapping of the locus for the human peptide antibiotic FALL-39; Gudmundsson GH et al.; PR-39 is a porcine 39-aa peptide antibiotic composed of 49% proline and 24% arginine, with an activity against Gram-negative bacteria comparable to that of tetracycline . In Escherichia coli, it inhibits DNA and protein synthesis . PR-39 was originally isolated from pig small intestine, but subsequent cDNA cloning showed that the gene is expressed in the bone marrow . The open reading frame of the clone showed that PR-39 is made as 173-aa precursor whose proregion belongs to the cathelin family . The PR39 gene, which is rather compact and spans only 1784 bp has now been sequenced . The coding information is split into four exons . The first exon contains the signal sequence of 29 residues and the first 37 residues of the cathelin propart . Exons 2 and 3 contain only cathelin information, while exon 4 codes for the four C-terminal cathelin residues and the mature PR-39 peptide extended by three residues . The sequenced upstream region (1183 bp) contains four potential recognition sites for NF-IL6 and three for APRF, transcription factors known to regulate genes for both cytokines and acute phase response factors . Genomic hybridizations revealed a fairly high level of restriction fragment length polymorphism and indicated that there are at least two copies of the PR39 gene in the pig genome . PR39 was mapped to pig chromosome 13 by linkage and in situ hybridization mapping . The gene for the human peptide antibiotic FALL-39 (also a member of the cathelin family) was mapped to human chromosome 3, which is homologous to pig chromosome 13. Gene, 1995 Jul 4, 160(1), 59 - 62 Mini-Tn10 transposon derivatives for insertion mutagenesis and gene delivery into the chromosome of gram-negative bacteria; Alexeyev MF et al.; Four mini-Tn10 derivatives bearing the nptII, cat, aacC1 and tet genes along with MCS of the pBluescriptII plasmid were constructed . These derivatives are embedded into the gamma-ori R6K-based suicide plasmid pBSL177, which can be mobilized into a broad range of hosts by the RP4 plasmid . The relatively small size and the presence of the mutant ATS Tn10 transposase gene, fused to Ptac promoter, as well as the versatile MCS of the pBluescriptII plasmid make these vectors a reliable tool for insertion mutagenesis and chromosomal insertion of the cloned DNA fragments. Proc Natl Acad Sci U S A, 1995 Jul 3, 92(14), 6625 - 9 Disparate responses to oxidative stress in saprophytic and pathogenic mycobacteria; Sherman DR et al.; To persist in macrophages and in granulomatous caseous lesions, pathogenic mycobacteria must be equipped to withstand the action of toxic oxygen metabolites . In Gram-negative bacteria, the OxyR protein is a critical component of the oxidative stress response . OxyR is both a sensor of reactive oxygen species and a transcriptional activator, inducing expression of detoxifying enzymes such as catalase/hydroperoxidase and alkyl hydroperoxidase . We have characterized the responses of various mycobacteria to hydrogen peroxide both phenotypically and at the levels of gene and protein expression . Only the saprophytic Mycobacterium smegmatis induced a protective oxidative stress response analogous to the OxyR response of Gram-negative bacteria . Under similar conditions, the pathogenic mycobacteria exhibited a limited, nonprotective response, which in the case of Mycobacterium tuberculosis was restricted to induction of a single protein, KatG . We have also isolated DNA sequences homologous to oxyR and ahpC from M . tuberculosis and Mycobacterium avium . While the M . avium oxyR appears intact, the oxyR homologue of M . tuberculosis contains numerous deletions and frameshifts and is probably nonfunctional . Apparently the response of pathogenic mycobacteria to oxidative stress differs significantly from the inducible OxyR response of other bacteria. Microb Drug Resist, 1995 Summer, 1(2), 111 - 4 beta-Lactamase induction in gram-negative bacteria is intimately linked to peptidoglycan recycling; Normark S; A number of Gram-negative organisms normally express a chromosomally mediated class C beta-lactamase that is inducible by beta-lactam antibiotics . Data have recently emerged suggesting a close link between beta-lactamase induction and the recycling of released muramyl peptides from the bacterial peptidoglycan . Thus the AmpG transporter is responsible for the uptake into the cell of GlcNAc-anhMurNAc-tripeptide . A mutant unable to express AmpG is therefore unable to recycle the cell wall and is at the same time not possible to induce by a beta-lactam . Once inside the cytosol the above muramyl peptide and its derivative anhMurNAc-tripeptide is degraded by the cytosolic AmpD amidase that specifically releases the tripeptide from cytosolic muramyl peptides brought into the cell via AmpG . Mutants unable to produce AmpD are blocked in a cytosolic step for cell wall recycling and accumulate large amounts of cytosolic anhMurNAc-tripeptide . It is believed that cytosolic muramyl peptides can act as ligands for the beta-lactamase regulator AmpR to activate expression of beta-lactamase . AmpD mutants, therefore, constitutively overproduce the chromosomal beta-lactamase and are beta-lactam resistant . In wild-type strains beta-lactams that result in an increased cell wall breakdown will cause an increase in the cytosol of muramyl peptides leading to beta-lactamase induction . Mutants affected in the ampD gene arise readily during treatment with third-generation cephalosporins . Since these mutants lack a functional cell wall recycling system they may be at a disadvantage in the absence of selection . However, since muramyl peptides may act as cytotoxins, especially for respiratory epithelial cells, ampD mutants due to their large accumulation of anhMurNAc-tripeptide may be altered in their pathogenic properties as compared to wild-type cells possessing a normal cell wall recycling system. Res Immunol, 1995 Jul-Aug, 146(6), 339 - 50 Human monocytes lacking the membrane-bound form of the bacterial lipopolysaccharide (LPS) receptor CD14 can mount an LPS-induced oxidative burst response mediated by a soluble form of CD14; Schutt C et al.; Monocytes and macrophages express a glycosyl phosphatidylinositol (GPI)-anchored lipopolysaccharide (LPS) receptor on the cell surface which enables them to detect minute amounts of LPS released from Gram-negative bacteria . A soluble form of CD14 is also found free in serum, though its physiological function is unknown . the interaction of LPS with CD14 on the monocyte surface leads to an activation of the cells which is manifested in the sudden release of reactive oxygen species, a process referred to as an oxidative burst . In patients suffering from the condition known as paroxysmal nocturnal haemoglobinuria (PNH), the synthesis of GPI anchors is blocked in haematopoietic cells which are therefore unable to express GPI-linked proteins on their surface . In severe cases, over 90% of monocytes lack membrane-bound CD14, though normal levels of the soluble form of the receptor-sCD14-are found in the serum . Despite this lack of membrane-bound CD14, monocytes from PNH patients can respond to low concentrations of LPS . Here we show that the LPS-induced oxidative burst of these PNH monocytes requires a component present in serum . The serum-dependent activation can be inhibited by monoclonal antibodies to CD14, can be removed from the serum by passage over a matrix to which an anti-CD14 antibody has been bound, and the depleted serum can be reconstituted by the addition of either purified natural or purified recombinant soluble CD14 . We conclude that an LPS-dependent oxidative burst in PNH monocytes can be mediated by soluble CD14. ASAIO J, 1995 Jul-Sep, 41(3), M430 - 4 Development of cationically modified cellulose adsorbents for the removal of endotoxins; Weber C et al.; The removal of endotoxins by extracorporeal adsorption processes seems the most promising therapeutic approach to Gram-negative sepsis and endotoxin shock . However, thus far adsorbents have failed to bind endotoxins efficiently or have shown adverse biocompatibility characteristics . To overcome these disadvantages, small particles of regenerated cellulose in the range of 1-8 microns in diameter were produced . Before use, the microspheres were cationically modified by substitution with polyethyleneimine (PEI) or diethylaminoethyl (DEAE) groups . A third kind of adsorbent was manufactured by (physically) coating the cellulose matrix with PEI . All three types of adsorbents exhibited a high adsorption capacity for endotoxins in human plasma, whereas activated charcoal and various anion exchange resins removed only small amounts of endotoxins under the same conditions . In addition, because the outer surface area is very large, adsorption takes place rapidly and diffusion becomes almost irrelevant . The adsorption process is primarily based on electrostatic interactions, which could be demonstrated by a significantly higher adsorption rate and binding capacity for lipid A-diphosphoryl, compared with lipid A-monophosphoryl . Use of these adsorbents in a newly developed plasma sorption system could be of great clinical interest because of the low production costs, the high adsorption efficiency, and the excellent biocompatibility data. J Infect Dis, 1995 Jul, 172(1), 161 - 72 Oral immunization with recombinant Helicobacter pylori urease induces secretory IgA antibodies and protects mice from challenge with Helicobacter felis; Lee CK et al.; Helicobacter pylori, a gram-negative spiral bacterium, is the cause of chronic superficial (type B) gastritis and peptic ulcer disease . The urease enzyme of H . pylori was expressed as an inactive recombinant protein in Escherichia coli, purified as particulate structures of 550-600 kDa molecular mass with a diameter of approximately 12 nm . Given orally, 5 micrograms of urease with an appropriate mucosal adjuvant, such as the labile toxin of E . coli, protected 60%-100% of mice against challenge with virulent Helicobacter felis . Protection correlated with the level of secretory IgA antibodies against urease . Oral administration of antigen was as effective or better than intragastric administration . Parenteral injection of antigen or intragastric administration of high-dose antigen without adjuvant elicited serum IgG but no IgA antibodies and did not confer protection . Recombinant urease as an oral vaccine candidate deserves further investigation as an approach to the prevention of Helicobacter-induced chronic gastroduodenal diseases in humans. J Exp Med, 1995 Jul 1, 182(1), 267 - 72 Triglyceride-rich lipoproteins prevent septic death in rats; Read TE et al.; Triglyceride-rich lipoproteins bind and inactive bacterial endotoxin in vitro and prevent death when given before a lethal dose of endotoxin in animals . However, lipoproteins have not yet been demonstrated to improve survival in polymicrobial gram-negative sepsis . We therefore tested the ability of triglyceride-rich lipoproteins to prevent death after cecal ligation and puncture (CLP) in rats . Animals were given bolus infusions of either chylomicrons (1 g triglyceride/kg per 4 h) or an equal volume of saline for 28 h after CLP . Chylomicron infusions significantly improved survival (measured at 96 h) compared with saline controls (80 vs 27%, P < or = 0.03) . Chylomicron infusions also reduced serum levels of endotoxin, measured 90 min (26 +/- 3 vs 136 +/- 51 pg/ml, mean +/- SEM, P < or = 0.03) and 6 h (121 +/- 54 vs 1,026 +/- 459 pg/ml, P < or = 0.05) after CLP . The reduction in serum endotoxin correlated with a reduction in serum tumor necrosis factor, measured 6 h after CLP (0 +/- 0 vs 58 +/- 24 pg/ml, P < or = 0.03), suggesting that chylomicrons improve survival in this model by limiting macrophage exposure to endotoxin and thereby reducing secretion of inflammatory cytokines . Infusions of a synthetic triglyceride-rich lipid emulsion (Intralipid; KabiVitrum, Inc., Alameda, CA) (1 g triglyceride/kg) also significantly improved survival compared with saline controls (71 vs 27%, P < or = 0.03) . These data demonstrate that triglyceride-rich lipoproteins can protect animals from lethal polymicrobial gram-negative sepsis. J Exp Med, 1995 Jul 1, 182(1), 147 - 54 Activation of Raf-1 and mitogen-activated protein kinase in murine macrophages partially mimics lipopolysaccharide-induced signaling events; Hambleton J et al.; Lipopolysaccharide (LPS), a highly conserved component of the outer membrane of gram-negative bacteria, stimulates macrophages to release various cytokine and eicosanoid mediators of the immune response . The mechanism by which LPS stimulates these cells is poorly characterized . One of the most rapid LPS-stimulated events is the phosphorylation and activation of the p42 and p44 isoforms of mitogen-activated protein (MAP) kinase . We wished to examine the role of MAP kinase in LPS-induced signaling in murine macrophages by activating MAP kinase independently of LPS . An expression vector encoding a Raf-1:estrogen receptor (ER) chimeric protein was transfected into the murine macrophage cell line RAW 264.7 . Activation of this chimeric protein (delta Raf-1:ER) by estradiol resulted in rapid and prolonged activation of MAP kinase, as expected from previous results implicating Raf-1 as an upstream activator of this signaling cascade . LPS stimulation induced accumulation of MAP kinase phosphatase 1 messenger RNA, whereas delta Raf-1:ER activation did not, perhaps accounting for the more prolonged activation of MAP kinase seen in response to delta Raf-1:ER activation . Similarly, activation of DNA binding by the transcription factor, nuclear factor (NF) kappa B, as assessed by electrophoretic mobility shift assay, occurred in response to LPS stimulation but not in response to delta Raf-1:ER activation or phorbol myristate acetate (PMA) stimulation . Using an enzyme-linked immunosorbent assay for murine tumor necrosis factor alpha (TNF-alpha), we found that LPS and PMA stimulation and delta Raf-1:ER activation induced secretion of TNF-alpha, although the amount of TNF-alpha secreted in response to delta Raf-1:ER activation and PMA stimulation was approximately 20-fold less than that secreted in response to LPS . Correspondingly, accumulation of TNF-alpha messenger RNA was weakly induced by delta Raf-1:ER activation or PMA stimulation, whereas strong induction was noted in response to LPS . These results suggest that Raf-1 or PMA activation of MAP kinase in murine macrophages is sufficient for a small amount of TNF-alpha production and secretion in the absence of NF-kappa B activation, but LPS stimulation involves additional signaling events, such as NF-kappa B activation, that augment the response seen with activation of MAP kinase alone. J Surg Res, 1995 Jul, 59(1), 153 - 8 PEG-BP-30 monotherapy attenuates the cytokine-mediated inflammatory cascade in baboon Escherichia coli septic shock; Espat NJ et al.; Septic shock following gram-negative infection is a leading cause of mortality in critically ill patients, accounting for nearly 200,000 deaths a year . The exaggerated production of tumor necrosis factor-alpha (TNF alpha) is known to contribute to hemodynamic collapse and the hematological dyscrasia associated with gram-negative sepsis . Although previous studies have shown TNF alpha antibodies and TNF immunoadhesins to be effective in experimental gram-negative sepsis, we postulated that administration of a novel construct of two modified soluble p55 receptors linked to polyethylene glycol (PEG-BP-30) would also attenuate the hemodynamic and hematologic alterations to lethal Escherichia coli septic shock in non-human primates . Nine adult female and male baboons (Papio anubis), weighing 10-17 kg, were anesthetized and invasively monitored . The nine animals were randomized to receive either 0.2 mg/kg body wt PEG-BP-30 (n = 3), 5.0 mg/kg body wt PEG-BP-30 (n = 3), or placebo (n = 3) . One hour after pretreatment, animals were infused with 5-10 x 10(10) CFU/kg of live E . coli iv and vital signs were recorded for the next 8 hr . Arterial blood was drawn for baseline parameters and throughout the study to obtain total and differential white blood cell and platelet counts and cytokine levels (TNF alpha, IL-1 beta, IL-6, IL-8) . E . coli bacteremic baboons receiving only placebo demonstrated a significant fall in mean blood pressure and leukopenia . Two of the three animals expired . In contrast, five of the six baboons receiving the PEG-BP-30 survived and these animals exhibited markedly attenuated declines in blood pressure and leukocyte numbers.(ABSTRACT TRUNCATED AT 250 WORDS) Ann Surg, 1995 Jul, 222(1), 57 - 65 Controlled clinical trial of selective decontamination for the treatment of severe acute pancreatitis; Luiten EJ et al.; OBJECTIVE: A randomized, controlled, multicenter trial was undertaken in 102 patients with objective evidence of severe acute pancreatitis to evaluate whether selective decontamination reduces mortality . SUMMARY BACKGROUND DATA: Secondary pancreatic infection is the major cause of death in patients with acute necrotizing pancreatitis . Controlled clinical trials to study the effect of selective decontamination in such patients are not available . METHODS: Between April 22, 1990 and April 19, 1993, 102 patients with severe acute pancreatitis were admitted to 16 participating hospitals . Patients were entered into the study if severe acute pancreatitis was indicated, on admission, by multiple laboratory criteria (Imrie score > or = 3) and/or computed tomography criteria (Balthazar grade D or E) . Patients were randomly assigned to receive standard treatment (control group) or standard treatment plus selective decontamination (norfloxacin, colistin, amphotericin; selective decontamination group) . All patients received full supportive treatment, and surveillance cultures were taken in both groups . RESULTS: Fifty patients were assigned to the selective decontamination group and 52 were assigned to the control group . There were 18 deaths in the control group (35%), compared with 11 deaths (22%) in the selective decontamination group (adjusted for Imrie score and Balthazar grade: p = 0.048) . This difference was mainly caused by a reduction of late mortality (> 2 weeks) due to significant reduction of gram-negative pancreatic infection (p = 0.003) . The average number of laparotomies per patient was reduced in patients treated with selective decontamination (p < 0.05) . Failure of selective decontamination to prevent secondary gram-negative pancreatic infection with subsequent death was seen in only three patients (6%) and transient gram-negative pancreatic infection was seen in one (2%) . In both groups of patients, all gram-negative aerobic pancreatic infection was preceded by colonization of the digestive tract by the same bacteria . CONCLUSION: Reduction of gram-negative colonization of the digestive tract, preventing subsequent pancreatic infection by means of selective decontamination, significantly reduces morbidity and mortality in patients with severe acute necrotizing pancreatitis. Clin Immunol Immunopathol, 1995 Jul, 76(1 Pt 1), 32 - 6 Serum concentrations of MIP-1 alpha and interleukin-8 in patients suffering from acute Plasmodium falciparum malaria; Burgmann H et al.; The chemokines are a superfamily of small proteins secreted primarily by leukocytes and related by a conserved four-cystein motif . In the present study we investigated the serum levels of macrophage inflammatory protein 1 alpha (MIP-1 alpha) and interleukin-8 (IL-8) . MIP-1 alpha is a neutrophil chemotactic protein important in acute and chronic inflammation . Recent studies demonstrated that MIP-1 alpha may also act as potent inhibitor of hemopoetic stem cell proliferation, which may be important in the development of prolonged anemia in patients suffering from Plasmodium falciparum malaria . IL-8 serum concentrations correlate with severity and outcome of infectious diseases . Moreover, recent reports indicate that IL-8 plays a major role in fatal gram-negative sepsis . It was the aim of this study to investigate the time course of MIP-1 alpha and IL-8 concentrations in patients suffering from acute P . falciparum infection . Blood samples of 20 patients suffering from severe P . falciparum malaria were investigated . MIP-1 alpha and IL-8 concentrations were determined using ELISA technique at admission, on Days 7, 14, 21, and 28 . Maximal concentrations of MIP-1 alpha and IL-8 were found on Day 14, at a time when parasites were not detected in the smears . The serum levels of IL-8 on the day of admission were correlated to the parasite count . No correlation was seen between the hematokrit values and the MIP-1 alpha concentrations at any time. J Bacteriol, 1995 Jul, 177(13), 3673 - 9 A Sorangium cellulosum (myxobacterium) gene cluster for the biosynthesis of the macrolide antibiotic soraphen A: cloning, characterization, and homology to polyketide synthase genes from actinomycetes; Schupp T et al.; A 40-kb region of DNA from Sorangium cellulosum So ce26, which contains polyketide synthase (PKS) genes for synthesis of the antifungal macrolide antibiotic soraphen A, was cloned . These genes were detected by homology to Streptomyces violaceoruber genes encoding components of granaticin PKS, thus extending this powerful technique for the identification of bacterial PKS genes, which has so far been applied only to actinomycetes, to the gram-negative myxobacteria . Functional analysis by gene disruption has indicated that about 32 kb of contiguous DNA of the cloned region contains genes involved in soraphen A biosynthesis . The nucleotide sequence of a 6.4-kb DNA fragment, derived from the region with homology to granaticin PKS genes, was determined . Analysis of this sequence has revealed the presence of a single large open reading frame beginning and ending outside the 6.4-kb fragment . The deduced amino acid sequence indicates the presence of a domain with a high level of similarity to beta-ketoacyl synthases that are involved in polyketide synthesis . Other domains with high levels of similarity to regions of known polyketide biosynthetic functions were identified, including those for acyl transferase, acyl carrier protein, ketoreductase, and dehydratase . We present data which indicate that soraphen A biosynthesis is catalyzed by large, multifunctional enzymes analogous to other bacterial PKSs of type I. Drugs, 1995 Jul, 50(1), 62 - 72 A guide to the treatment of lower respiratory tract infections; Vogel F; Acute bronchitis is usually a viral infection which, unless there is a special disposition, does not require antibiotic therapy . For the initial oral chemotherapy of bacterial infections of the lower respiratory tract (chronic bronchitis, pneumonia) the effective and well tolerated cephalosporins, macrolides and amoxicillin plus beta-lactamase-inhibitor are recommended . In complicated cases with severe underlying disease, longer history or frequent exacerbations, quinolones should be given if Gram-negative infections are suspected or if initial therapy with other substances has failed . If Legionella, Mycoplasma or Chlamydia spp., so-called 'atypical' pathogens, are involved, macrolide antibiotics are the therapy of first choice . Special attention should be given to the increase in resistance against cotrimoxazole (trimethoprim-sulfamethoxazole) and tetracyclines . In hospitals where primary pneumonias are treated preferentially by intravenous medication, therapy should be switched to oral antibiotics as soon as feasible (follow-up therapy) . For severely ill patients with secondary pneumonia and underlying disease, second generation cephalosporins with aminoglycosides, or monotherapy with third generation cephalosporins are recommended . In very severe, high-risk cases, third generation cephalosporins, combinations with high-dosage quinolones or ureidopenicillins plus beta-lactamase-inhibitors are suitable . Future development in the antibiotic treatment of respiratory infections will follow the current trend of lower dosages, with the clear objective of shortening treatment periods and achieving earlier discharge from hospital. Biokhimiia, 1995 Jul, 60(7), 1161 - 70 {Permeability of the Escherichia coli outer membrane for ethidium ions and periplasmic alkaline phosphatase during increased synthesis of it}; Mikhaleva NI et al.; During augmented synthesis of periplasmic alkaline phosphatase by various strains of Escherichia coli, the outer membrane of bacterial cells becomes permeable for both the enzyme and ethidium ions which do not generally penetrate inside the cells of gram-negative bacteria . In the absence of the lipoprotein in the outer membrane, its permeability for these compounds as well as its sensitivity to membranotropic agents increases, thus testifying to the influence of the lipoprotein upon certain properties of the outer membrane . A competitive interaction was found between the lipoprotein and lipopolysaccharide content in the outer membrane and their content and alkaline phosphatase secretion into the external medium . It is suggested that increased permeability of the E . coli outer membrane during augmented synthesis of the secreted protein is due to impaired biogenesis of membrane components. Trends Microbiol, 1995 Jul, 3(7), 257 - 61 Virulence in Actinobacillus pleuropneumoniae and RTX toxins; Frey J; RTX toxins are pore-forming, cytolytic protein toxins that occur widely among pathogenic Gram-negative bacteria . RTX toxins appear to play a direct role in the virulence of Actinobacillus pleuropneumoniae, the etiological agent of porcine pleuropneumonia . This discovery has led to the development of new diagnostic and epidemiological tools, as well as vaccines, that are useful for a broad variety of serotypes. Microbiology, 1995 Jul, 141 ( Pt 7), 1647 - 54 Escherichia coli periplasmic protein FepB binds ferrienterobactin; Stephens DL et al.; Most high-affinity systems for iron uptake in Gram-negative bacteria are thought to employ periplasmic-binding-protein-dependent transport . In Escherichia coli, FepB is a periplasmic protein required for uptake of iron complexed to its endogenously-synthesized siderophore enterobactin (Ent) . Direct evidence that ferrienterobactin (FeEnt) binds to FepB is lacking because high background binding by FeEnt prevents use of the usual binding protein assays . Here the membrane localization vehicle LppOmpA {Francisco, J.A., Earhart, C.F . & Georgiou, G . (1992) . Proc Natl Acad Sci USA 89, 2713-2717} was employed to place FepB in the E . coli outer membrane . Plasmid pTX700 was constructed and shown to encode, under lac operator control, the 'tribrid' protein LppOmpAFepB; the carboxy-terminal FepB portion lacks at most two amino acids of mature FepB . After short induction periods, most of the tribrid was in the outer membrane . A number of LppOmpAFepB species could be detected; some were degradation products and some may be related to the multiplicity of FepB forms previously observed in minicells and maxicells . Outer membrane harbouring the tribrid and lacking FepA, the normal outer membrane receptor for FeEnt, bound approximately four times more FeEnt than outer membrane from uninduced cells, from cells lacking pTX700 and from cells expressing only an LppOmpA 'dibrid' . Similarly, whole UT5600(fepA)/pTX700 cells induced for tribrid synthesis bound FeEnt and this binding was not affected by energy poisons . The results demonstrated that FepB can bind FeEnt, thereby definitely placing FeEnt transport in the periplasmic permease category of transport systems, and that the LppOmpA localization vehicle can be used with periplasmic binding proteins. Clin Sci (Lond), 1995 Jul, 89(1), 83 - 9 Lipopolysaccharide induces upregulation of neutral endopeptidase 24.11 on human neutrophils: involvement of the CD14 receptor; Fagny C et al.; 1 . As lipopolysaccharide is a major stimulator of neutrophil responses during Gram-negative bacterial infections, we studied its effect on the membrane expression of neutral endopeptidase 24.11/CD10 on neutrophils in a model of endotoxaemia in vitro . Lipopolysaccharide added to human whole-blood induced a marked and sustained CD10/neutral endopeptidase upregulation that was already detectable at 0.1 ng/ml and was maximal at a lipopolysaccharide concentration of 10 ng/ml . 2 . We observed that neither tumour necrosis factor-alpha nor any newly synthesized protein was involved in the upregulation observed after 1 h incubation with 10 ng/ml lipopolysaccharide . 3 . We further studied whether the lipopolysaccharide-induced CD10/neutral endopeptidase upregulation was mediated by lipopolysaccharide binding to the neutrophil CD14 receptor . Incubation of whole blood with an anti-CD14 monoclonal antibody before the addition of 0.1 ng/ml or 0.5 ng/ml lipopolysaccharide resulted in complete inhibition of CD10/neutral endopeptidase upregulation . In contrast, at a lipopolysaccharide concentration of 10 ng/ml, the anti-CD14 monoclonal antibody had an incomplete blocking effect . 4 . The differential requirement for the CD14 receptor, depending on the lipopolysaccharide dose, was confirmed by the study of a patient suffering from paroxysmal nocturnal haemoglobinuria (in whom a complete defect in neutrophil CD14 expression was previously documented) . 5 . We finally confirmed these results using purified neutrophils, demonstrating that lipopolysaccharide-induced CD10/neutral endopeptidase upregulation depends on direct interaction with neutrophil CD14. J Clin Microbiol, 1995 Jul, 33(7), 1879 - 83 Identification of Bartonella (Rochalimaea) species among fastidious gram-negative bacteria on the basis of the partial sequence of the citrate-synthase gene; Joblet C et al.; The bacterial genus Bartonella (Rochalimaea) includes emerging human pathogens with five recognized species . These are fastidious gram-negative bacteria, exhibiting few phenotypic characteristics and whose identification relies upon serotyping, cellular fatty acid analysis, and molecular typing . Most of the isolates have been recovered from the blood of patients, and three of the four pathogenic Bartonella species are associated with infectious endocarditis . We performed PCR-restriction fragment length polymorphism (RFLP) analysis of the blood culture bottle supernatant for the routine identification of Bartonella species among fastidious gram-negative bacteria . The amplification of the citrate-synthase gene with primers previously reported (R . L . Regnery, C . L . Spruill, and B . D . Plikaytis, J . Bacteriol . 173:1576-1589, 1991) yielded a 379-bp product from Bartonella species and a 382-bp product for Capnocytophaga ochracea but no product from any of the other 15 genotypically or phenotypically related species tested . We determined the sequences of the citrate-synthase gene-amplified products for Bartonella species and C . ochracea in order to predict the optimal restriction enzyme to be used in RFLP analysis . TaqI and AciI allowed identification of Bartonella species and C . ochracea . We propose that acridine orange and Gram staining, followed by PCR-RFLP analysis of the blood bottle supernatant, be included in the examination of blood samples from patients with suspected infectious endocarditis. J Immunol, 1995 Jul 1, 155(1), 316 - 24 Lipopolysaccharide binding protein and CD14 modulate the synthesis of platelet-activating factor by human monocytes and mesangial and endothelial cells stimulated with lipopolysaccharide; Camussi G et al.; The biosynthesis of platelet-activating factor (PAF) during Gram-negative involves the interaction of LPS with the cells of the host . We have investigated the molecular mechanism that controls cell recognition and PAF biosynthetic response to LPS in human monocytes (MO), glomerular mesangial cells (MC), and HUVEC in culture . The synthesis of PAF by MO and MC involves two proteins, plasma LPS binding protein (LBP) and cell membrane CD14 (mCD14) . As MO, MC were shown to express the mCD14 molecule by several mAbs . MO and mCD14-positive MC were stimulated to synthesize PAF either by the 63D3 and IOM-2 mAbs or by the natural ligand LBP-LPS complex . Moreover, LeuM3, 28C5, and 18E12 mAbs that were themselves unable to stimulate the synthesis of PAF blocked PAF synthesis initiated by LBP-LPS complex . LBP was required for synthesis of PAF by MO . In MC, which synthesize PAF also after stimulation by LPS alone, the LBP was shown to speed and significantly enhance the synthesis of PAF . The soluble form of CD14 (sCD14), when added to MO stimulated with LBP-LPS complexes, inhibited the synthesis of PAF possibly by competing with mCD14 . In contrast, sCD14 was shown to be required for LPS-induced synthesis of PAF by HUVEC, which did not express mCD14 . Therefore, membrane receptors (mCD14) and plasma soluble proteins (LBP and sCD14) may enable different human cell types to synthesize PAF after LPS stimulation. Infect Immun, 1995 Jul, 63(7), 2576 - 80 Binding of lipopolysaccharide (LPS) to an 80-kilodalton membrane protein of human cells is mediated by soluble CD14 and LPS-binding protein; Schletter J et al.; Activation of cells by bacterial lipopolysaccharide (LPS) plays a key role in the pathogenesis of gram-negative septic shock . The 55-kDa glycoprotein CD14 is known to bind LPS and initiate cell activation . However, there must be additional LPS receptors because CD14 is linked by a glycosylphosphatidyl inositol anchor to the cell membrane and therefore unable to perform transmembrane signalling . Searching for potential LPS receptors, we investigated the binding of LPS to membrane proteins of the human monocytic cell line Mono-Mac-6 . Membrane proteins were electrophoretically separated under reducing conditions, transferred to nitrocellulose, and exposed to LPS, which was visualized with anti-LPS antibody . Smooth- and rough-type LPS, as well as free lipid A, bound to a variety of proteins in the absence of serum . However, in the presence of serum, additional or preferential binding to a protein of approximately 80-kDa was observed . Experiments with differently acylated lipid A structures showed that the synthetic tetraacyl compound 406 was still able to bind, whereas no binding was detected with the bisacyl compound 606 . The 80-kDa membrane protein was also detected on human peripheral blood monocytes and endothelial cells . The serum factors mediating the binding of lipid A to the 80-kDa membrane protein were identified as soluble CD14 and LPS-binding protein . From these results, we conclude that this 80-kDa protein is a candidate for the hypothetical molecule for LPS and/or LPS-CD14 recognition and signal transduction. J Pediatr Surg, 1995 Jul, 30(7), 1086 - 8; discussion 1088-9 Enteral feeding increases sepsis in infants with short bowel syndrome; Weber TR; Sepsis secondary to bacterial translocation is common in infants with short bowel syndrome (SBS) . Although early feeding is advocated to enhance adaptation in SBS, the effects of feeding on sepsis in SBS patients have not been examined . Twenty-one infants and children (aged 2 months to 3 years) with SBS (< 80 cm small bowel length) from a variety of causes (15 necrotizing enterocolitis, 2 atresia, 2 gastroschisis, 2 volvulus) had follow-up prospectively for septic episodes before and after feedings were initiated, while still receiving total parenteral nutrition . The incidence and number of septic episodes and microbiology (blood cultures) were tabulated and compared with those of 20 patients with similar ages, and diagnoses without SBS . Statistically significant differences among infants with SBS were noted with respect to sepsis incidence (6 of 21 {29%} NPO v 16 of 21 {76%} feeding) number of septic episodes (1.3 +/- .2 NPO v 4.2 +/- .4 feeding), and presence of gram-negative rods causing bacteremia (1 of 6 {17%} NPO v 13 of 16 {81%} feeding) (all: P < .05) . There were similar differences between SBS and non-SBS infants . These data show that enteral feeding increases the incidence and number of episodes of sepsis in SBS infants, but not in matched non-SBS patients . The predominance of gram-negative organisms in sepsis in SBS suggests increased gut bacterial translocation in these patients, implying that selective gut decontamination may reduce the episodes of bacteremia. J Biol Chem, 1995 Jun 16, 270(24), 14829 - 34 Hemolysin transport in Escherichia coli . Point mutants in HlyB compensate for a deletion in the predicted amphiphilic helix region of the HlyA signal; Sheps JA et al.; The alpha-hemolysin transporter of Escherichia coli, a member of the ATP-binding cassette transporter super-family, is responsible for secretion of the 107-kDa protein toxin HlyA across both membranes of the Gram-negative envelope in a single step . Secretion of HlyA is dependent on a signal sequence, which occupies the C-terminal 50-60 amino acids of HlyA . Previously, it was shown that point mutants in the transmembrane domain of the transporter HlyB could partially correct the transport defect caused by a deletion of the C-terminal 29 amino acids of HlyA . These suppressor mutations demonstrated a direct interaction between HlyA and HlyB . They also displayed suppressor effects on a broad spectrum of HlyA signal mutants . In the present study, we selected HlyB alleles that complemented an internal deletion of 29 amino acids in HlyA containing a predicted amphiphilic helix region immediately upstream from the previous deletion . This set of HlyB mutants identifies further sites in HlyB that modulate substrate specificity but display allele-specific effects on a range of HlyA signal mutants . The inability to isolate mutations with effects restricted to either half of the signal sequence suggests that the signal is not recognized in a modular fashion by the transporter but rather functions as an integrated whole . We also report the isolation of the first substrate specificity mutation, which lies within the ATP-binding domain of HlyB . This could support a model in which the region of the ATP-binding cassette between the two Walker consensus motifs involved in ATP binding interacts with either the substrate or the transmembrane domains. J Biol Chem, 1995 Jun 9, 270(23), 13698 - 705 Overproduction and one-step purification of Escherichia coli 3-deoxy-D-manno-octulosonic acid 8-phosphate synthase and oxygen transfer studies during catalysis using isotopic-shifted heteronuclear NMR; Dotson GD et al.; The enzyme 3-deoxy-D-manno-octulosonic acid 8-phosphate synthase catalyzes the condensation of D-arabinose 5-phosphate with phosphoenolpyruvate to give the unique 8-carbon acidic sugar 3-deoxy-D-manno-octulosonic acid 8-phosphate (KDO 8-P) found only in Gram-negative bacteria and required for lipid A maturation and cellular growth . The Escherichia coli gene kdsA that encodes KDO 8-P synthase has been amplified by polymerase chain reaction methodologies and subcloned into the expression vector, pT7-7 . A simple one-step purification yields 200 mg of homogeneous KDO 8-P synthase per liter of cell culture . {2-13C,18O}Phosphoenolpyruvate (PEP) was prepared by first, exchange of {2-13C}-3-bromopyruvate with 2H2 18O followed by reaction of the labeled bromopyruvate with trimethylphosphite . The fate of the enolic oxygen in this multilabeled PEP, during the course of the KDO 8-P synthase-catalyzed reaction with D-arabinose 5-phosphate, was monitored by 13C and 31P NMR spectroscopy . The inorganic phosphate formed during the reaction was further analyzed via mass spectral analysis of its trimethyl ester derivative . The 13C NMR spectrum of an incubation mixture of {2-13C}PEP and D-arabinose 5-phosphate in 2H2 18O in the presence of KDO 8-P synthase was also recorded . {2-13C}KDO 8-P was utilized to determine the extent of nonenzymatic incorporation of 18O into the C-2 position of KDO 8-P . The results indicate that the enolic oxygen of the PEP is recovered with the inorganic phosphate, and the C-2 oxygen of KDO 8-P originates from the solvent, H2O. Biochem Biophys Res Commun, 1995 Jun 6, 211(1), 183 - 9 Coordinate up- and down-modulation of inducible nitric oxide synthase, nitric oxide production, and tumoricidal activity in rat bone-marrow-derived mononuclear phagocytes by lipopolysaccharide and gram-negative bacteria; Keller R et al.; Simultaneous incubation of primary rat bone-marrow-derived mononuclear phagocytes (BMMo) and tumor cells with gram-negative agents triggers within 24 h interferon gamma (IFN gamma)- and tumor necrosis factor (TNF alpha)-independent tumoricidal activity . On the other hand, BMMo that had been incubated for 24 h with gram-negative agents prior to re-exposure to the same agent had largely lost their ability to generate tumoricidal activity, although their ability to bind lipopolysaccharide (LPS) was not diminished . Parallel measurements of the kinetics of inducible nitric oxide synthase (iNOS), nitrite secretion, and tumoricidal activity triggered in primary BMMo by LPS revealed that these parameters take a coordinate course, reaching a peak within 24 h and then rapidly decaying . Down-regulation of expression of NOS protein and iNOS activity could be attributed neither to down-regulation of LPS receptors nor to L-arginine depletion. Arch Latinoam Nutr, 1995 Jun, 45(2), 122 - 7 {Preservation and stability of corn tortillas at room temperature}; Higuera-Ciapara I et al.; Three treatments with chemical preservative (sodium propionate, potassium sorbate-methylparaben and hydrogen peroxidemethyl paraben) were tested to delay microbial spoilage and extend shelf-life of corn tortillas at room temperature (25 degrees C) . The treatment with the best results was selected for further studies using two types of packaging: Paper and high density polyethylene . Quality of corn tortillas during storage was assessed by measuring water content, microbial analysis (Total Plate Count, molds and yeast) and throguh sensory evaluation . Results were analyzed by covariance analysis and slope contrast between packaging materials at p<0.05 . Spoilage of tortilla without preservative occurred within 24 hours due to a large number of gram negative bacteria, molds and yeasts, which were responsible for offensive odors . Only the combination of hydrogen peroxide-methyl paraben had a significant effect on retarding bacterial yeast spoilage . In addition, hydrogen peroxide residues could not {correction of no} be chemically detected after 2 days of storage . Results from this study show that tortilla can be kept for up to six days at room temperature with acceptable sensory properties with proper preservative treatment and packaging. Zentralbl Hyg Umweltmed, 1995 Jun, 197(5), 398 - 407 {Air microbial burden at garbage sorting facilities}; Jager E et al.; At two german garbage sorting facilities, germ concentrations in the air were analyzed at different working environments (waste reception, manual sorting) . At plant A, maximum concentrations of total bacteria (14700 CFU/m3), gram-negative bacteria (7279 CFU/m3) and moulds (> 84806 CFU/m3) occurred in the air at the final manual sorting belts . Referring to concentrations of total bacteria and moulds, concentrations in the air at the final sorting belts were significantly higher (p < 0.001) than at the other working environments . At plant B, the counts of total bacteria reached 7173 CFU/m3 in the air at the waste reception site and 5512 CFU/m3 at the manual sorting belt . Maximum concentrations for gram-negative bacteria were 247 resp . 206 CFU/m3 and 70919 resp . 60848 CFU/m3 for moulds . At the waste reception site, the aerial concentrations of total bacteria lay highly significantly (p < 0.001) and of moulds significantly (p < 0.05) above aerial concentrations at the manual sorting belt . As causes, an intensive mechanical pretreatment of recycle material at plant A and whirling up of sedimentary dust of the floor during manual sweeping of the material on to the transportation at plant B are discussed. Res Microbiol, 1995 Jun, 146(5), 385 - 96 Phylogenetic analysis of the genus Rickettsia by 16S rDNA sequencing; Roux V et al.; Rickettsiae are Gram-negative bacteria which multiply only inside host cells and need arthropods either as reservoirs or as vectors . Using the polymerase chain reaction and an automated laser fluorescent DNA sequencer, we amplified and sequenced the 16S rRNA (rDNA) of all available bacteria of the genus Rickettsia . R . tsutsugamushi remained close to the other bacteria of the genus Rickettsia using this technique, contrary to previous conclusions based on the study of the Sta-58 protein antigen . We found that R . canada was not included in the typhus group, as is currently recognized, but was grouped with the rickettsiae of the spotted fever group (SFG) . All the SFG rickettsiae tested were grouped in the same cluster (R . conorii, Indian tick typhus rickettsia, Astrakhan fever rickettsia, Israeli tick typhus rickettsia, HA-91, R . sibirica, R . parkeri, "R . africae", "R . slovaca", R . rickettsii, Thai tick typhus rickettsia, R . japonica, R . massiliae, R . rhipicephali, R . montana, two recent isolates GS and Bar 29, R . australis, R . akari, R . bellii and R . helvetica) . The recently described ELB bacterium, the agent of the Californian murine typhus, and AB bacterium, a bacterium associated with male killing in the ladybird beetle, were found in this cluster . The sequences of R . conorii Moroccan strain/Indian tick typhus rickettsia, R . massiliae/GS and R . sibirica/HA-91 were identical . All the rickettsiae had a unique ancestor with bacteria also isolated in arthropods (Ehrlichia, Cowdria, Anaplasma, Wolbachia pipientis), eventually pathogenic for mammals and implicated in parthenogenesis and cytoplasmic incompatibility . We conclude that a unique bacterium started a stable association with arthropod ancestors and generated the observed diversity of the currently isolated members of the Rickettsiales. J Surg Res, 1995 Jun, 58(6), 739 - 45 Lymphocyte-derived cytokines augment macrophage tumor necrosis factor-alpha and interleukin-6 secretion during experimental gram-negative bacterial sepsis; Battafarano RJ et al.; Although lymphocyte-derived cytokines are known to augment macrophage cytokine production in vitro, their effect on macrophage tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) secretion during gram-negative bacterial sepsis has not been characterized . The purpose of this study was to examine the effect of lymphocyte-derived cytokines on macrophage TNF-alpha and IL-6 secretion during gram-negative bacterial peritonitis . To examine this problem, uninfected and infected mice were studied . Mice were infected with Escherichia coli O111:B4 and two subgroups were examined consisting of those pretreated iv 1 hr prior to bacterial challenge with either (1) saline or (2) anti-E . coli O111:B4 LPS mAb 2A3, the latter administered to abrogate the effects of LPS in vivo . Thus, three groups of mice were studied in relation to pretreatment and infectious challenges: (1) saline/saline (control); (2) saline/E . coli (saline); and (3) mAb 2A3/E . coli (mAb 2A3) . Nonadherent splenocytes (> 95% lymphocytes by histologic staining criteria) harvested 16 hr later from mice in each group were incubated in culture ex vivo for 3 hr to obtain supernatants containing lymphocyte-derived cytokines . These supernatants containing lymphocyte-derived cytokines then were incubated in vitro with naive splenic macrophages with or without E . coli O111:B4 LPS . Macrophage TNF-alpha and IL-6 levels were determined using L929 and B9 bioassays.(ABSTRACT TRUNCATED AT 250 WORDS) Crit Care Med, 1995 Jun, 23(6), 994 - 1006 A second large controlled clinical study of E5, a monoclonal antibody to endotoxin: results of a prospective, multicenter, randomized, controlled trial . The E5 Sepsis Study Group; Bone RC et al.; OBJECTIVE: To evaluate the safety and efficacy of E5, a murine, monoclonal antibody directed against endotoxin, in the treatment of patients with Gram-negative sepsis . DESIGN: A multicenter, randomized, double-blind, placebo-controlled trial . SETTING: Fifty-three hospitals across the United States, including university medical centers, Veterans Affairs Medical Centers, and community hospitals . PATIENTS: 847 patients were randomized into this study . Enrolled patients met criteria for three conditions: a) known or suspected Gram-negative infection; b) clinical evidence of sepsis; and c) signs of end-organ dysfunction . Patients with refractory shock were excluded from the study . INTERVENTIONS: Two doses of E5 (2 mg/kg/day by intravenous infusion 24 hrs apart), or placebo that was identical in appearance were administered . In addition, all patients received standard supportive therapy and broad-spectrum antibiotics . MEASUREMENTS AND MAIN RESULTS: The primary end point was mortality over 30 days . Secondary outcome measures included the resolution and prevention of organ failure in the same two populations . Additionally, the safety of E5 was evaluated . There was no significant improvement in survival over 30 days among patients with Gram-negative sepsis who received E5 compared with those patients who received placebo (n = 530; p = .21) . In addition, E5 did not improve survival for patients with Gram-negative sepsis and organ failure (n = 139; p = .3) . However, a significantly greater percentage of patients with Gram-negative sepsis experienced resolution of major organ failure if they received E5, compared with those patients who received placebo (n = 139; 48% E5 vs . 25% placebo; p = .005) . This result extended to all patients who entered the study with one or more major organ failures, regardless of the etiology of the infection (n = 225; 41% E5 vs . 27% placebo; p = .024) . E5 also provided protection against the development of some organ failures, but significant prevention was only observed for adult respiratory distress syndrome (p = .007) and central nervous system dysfunction (p = .050) . Hypersensitivity reactions attributable to E5 occurred in 2.6% of patients . An asymptomatic antibody response occurred in 44% of the E5-treated patients and in 12% of the patients who received placebo . CONCLUSIONS: In this study, E5 did not reduce mortality in nonshock patients with Gram-negative sepsis whether or not those patients also had organ failure . However, E5 did result in greater resolution of organ failure in patients with Gram-negative sepsis . This benefit extended to those patients with suspected Gram-negative etiology . This finding is important because patients with suspected Gram-negative sepsis and organ failure can be identified without waiting for culture results . In addition, E5 resulted in the prevention of adult respiratory distress syndrome and central nervous system organ failure . However, more studies are needed to determine if this result can be extended to organ failure in general . E5 is safe as a treatment for patients with Gram-negative sepsis. Crit Care Med, 1995 Jun, 23(6), 1033 - 9 Effect of the antiendotoxic agent, taurolidine, in the treatment of sepsis syndrome: a placebo-controlled, double-blind trial; Willatts SM et al.; OBJECTIVE: To assess the benefit gained from administration of the antiendotoxic drug, taurolidine, on outcome in critically ill patients with sepsis syndrome . DESIGN: A prospective, randomized, double-blind trial . SETTING: The general intensive therapy unit in a university teaching hospital . PATIENTS: One hundred patients admitted with sepsis syndrome over a 2-yr period . INTERVENTIONS: Patients were randomized to receive the amino-acid derivative, taurolidine, or an identically presented placebo . MEASUREMENTS AND MAIN RESULTS: Acute Physiology and Chronic Health Evaluation II (APACHE II), sepsis, and organ failure scores were measured daily . Blood for culture and endotoxin assay (using the limulus amoebocyte lysate assay) was sampled every 12 hrs for up to 5 days . Hemodynamic variables were recorded every 4 hrs . Forty-nine patients received taurolidine and 51 patients received placebo . There was no difference in APACHE II score, Sepsis Score, or presence of infections between the groups . The frequency of Gram-negative bacteremia was low at 12% . There was no difference in endotoxin activity, clinical or bacteriologic outcome, resolution of organ failure, or mortality rate between groups . Predicted risk of death for patients receiving taurolidine was 45%, and the actual mortality rate was 44% . In the group that received placebo, the predicted mortality rate was 38% and the actual mortality rate was 39% . CONCLUSION: Taurolidine had no beneficial therapeutic effect on the outcome of patients admitted to the intensive therapy unit with sepsis syndrome, using clinical, bacteriologic outcomes, progression of endotoxemia, resolution of organ failure, and 28-day mortality rate as end points. Am J Kidney Dis, 1995 Jun, 25(6), 928 - 33 Tumefactive megalocytic interstitial nephritis in a patient with Escherichia coli bacteremia; Krupp G et al.; Megalocytic interstitial nephritis is rare and primarily affects the cortex in an otherwise normal kidney . We recently encountered a patient with Escherichia coli bacteremia and oliguric acute renal failure who died of gram-negative septicemia . At autopsy, this patient's kidneys displayed typical features of megalocytic interstitial nephritis . We were able to perform special stains suggesting that the histiocytic interstitial cells originated from infiltrating macrophages . Our patient illustrates that macrophage proliferation can result in interstitial inflammation sufficiently severe to cause anuric acute renal failure. Infect Immun, 1995 Jun, 63(6), 2387 - 9 Electrophoretic analysis of the major outer membrane protein of Chlamydia psittaci reveals multimers which are recognized by protective monoclonal antibodies; McCafferty MC et al.; Purified major outer membrane protein, detergent solubilized and reduced with dithiothreitol but not heated, gave an apparent molecular weight in sodium dodecyl sulfate (SDS)-polyacrylamide gels almost three times that observed for the heat-denatured SDS-treated peptide . This is similar to the behavior of porin trimers from gram-negative bacteria . Two protective monoclonal antibodies showed strong binding to the proposed trimer but not to denatured, monomeric major outer membrane protein. Infect Immun, 1995 Jun, 63(6), 2180 - 4 Cross-reacting lipopolysaccharide antigens in Legionella pneumophila serogroups 1 to 14; Jurgens D et al.; Immunological cross-reactions among Legionella species were investigated with sonicated, proteinase K-digested cell lysates . The antigens separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis were either analyzed for lipopolysaccharides (LPSs) by silver staining or transferred to nitrocellulose membranes for serological characterization with rabbit antibodies directed against Legionella pneumophila serogroups 1 and 5 . When antiserum prepared against serogroup 5 was used to probe the LPSs from L . pneumophila serogroups 1 to 14, the antibodies recognized a common epitope harbored by all L . pneumophila serogroups but not by other Legionella species or by the gram-negative bacteria tested as controls . Hence, the serogroup 5 antiserum correctly identified all serogroups of L . pneumophila tested in the LPS immunoblot assay . Moreover, the silver-stained profiles of the isolated LPSs revealed characteristic patterns allowing the identification of the individual serogroups of L . pneumophila. J Exp Med, 1995 Jun 1, 181(6), 2289 - 93 Human tumor necrosis factor receptor (p55) and interleukin 10 gene transfer in the mouse reduces mortality to lethal endotoxemia and also attenuates local inflammatory responses; Rogy MA et al.; Anticytokine therapies have been promulgated in gram-negative sepsis as a means of preventing or neutralizing excessive production of proinflammatory cytokines . However, systemic administration of cytokine inhibitors is an inefficient means of targeting excessive production in individual tissue compartments . In the present study, human gene transfer was used to deliver to organs of the reticuloendothelial system antagonists that either inhibit tumor necrosis factor-alpha (TNF-alpha) synthesis or block its interactions with cellular receptors . Mice were treated intraperitoneally with cationic liposomes containing 200 micrograms of either a pCMV (cytomegalovirus)/p55 expression plasmid that contains the extracellular domain and transmembrane region of the human p55 TNF receptor, or a pcD-SR-alpha/hIL-10 expression plasmid containing the DNA for human interleukin 10 . 48 h later, mice were challenged with lipopolysaccharide (LPS) and D-galactosamine . Pretreatment of mice with p55 or IL-10 cDNA-liposome complexes improved survival (p < 0.01) to LPS-D-galactosamine . In additional studies, intratracheal administration of IL-10 DNA-liposome complexes 48 h before an intratracheal LPS challenge reduced pulmonary TNF-alpha levels by 62% and decreased neutrophil infiltration in the lung by 55% as measured by myeloperoxidase activity (both p < 0.05) . Gene transfer with cytokine inhibitors is a promising option for the treatment of both the systemic and local sequelae of septic shock. Eur J Clin Invest, 1995 Jun, 25(6), 418 - 24 Polymorphonuclear leucocyte dysregulation in patients with gram-negative septicaemia assessed by flow cytometry; Wenisch C et al.; Flow cytometry was used to study phagocytic function and release of reactive oxygen intermediates (ROI) following phagocytosis by granulocytes in 14 patients (six female, eight male) with gram-negative septicaemia prior to, during, and after therapy compared with a group of healthy controls . Phagocytic capacity was assessed by measuring uptake of fluorescein isothiocyanate (FITC)-labelled bacteria . Reactive oxygen generation after phagocytosis was measured by the quantification of dihydrorhodamine 123 converted to rhodamine 123 intracellulary . Compared with results in healthy controls granulocytes of septicaemic patients exhibited a decreased capacity to phagocytize Escherichia coli and to generate reactive oxygen products . Both phagocytosis and ROI production increased after initiation of therapy and normalized within 7 days of treatment . The results suggest that granulocytes do not only participate in, but are also a target of, the septic host inflammatory response. Microsc Res Tech, 1995 Jun 1, 31(2), 141 - 58 Cellular functions during activation and damage by pathogens: immunogold studies of the interaction of bacterial endotoxins with target cells; Risco C et al.; Bacterial endotoxins (lipopolysaccharides or LPS) are active components of Gram-negative bacteria that act on numerous cellular functions through the processes of cell activation and damage . The molecular mechanisms involved in the "endotoxic phenomenon" are not defined yet, although extensive studies have been carried out . Immunogold and electron microscopy (EM) have contributed to identify the primary target cells of endotoxins and the subcellular systems that receive the direct action of these bacterial agents . Here, we review our studies on immunogold detection of endotoxins in cellular and subcellular systems . The analysis of the interaction between endotoxins and cells was focussed on the following aspects: (1) morphological characteristics of the LPS aqueous suspensions used in experimental work; (2) binding of endotoxins to the plasma membrane of type II pneumocytes and alveolar macrophages (two of their cellular targets), and influence of the state of aggregation of the LPS; (3) movement and distribution of endotoxins inside the cell, from the plasma membrane to the nucleoplasm; and (4) interaction of LPS with microtubules and its effects on the integrity of the microtubular network . These approaches provide information at the molecular level as well as data for the establishment of physiological models of endotoxicity. J Crit Care, 1995 Jun, 10(2), 64 - 71 Serum neopterin and soluble interleukin-2 receptor for prediction of a shock state in gram-negative sepsis; Delogu G et al.; PURPOSE: This study aimed to investigate the predictive value of neopterin and soluble interleukin-2 (IL-2) receptor for shock occurrence in gram-negative sepsis . METHODS: We examined 57 patients admitted to an intensive care unit with gram-negative sepsis diagnosed according to preestablished criteria . Blood samples were collected every 24 hours and neopterin and soluble IL-2 receptor were measured by using commercially available test kits . To judge the predictive significance of these analyses the Cox proportional hazards regression model was used . RESULTS: Both neopterin (P < .05) and soluble IL-2 receptor (P < .01) were identified as significant predictors of a shock state, but the prognostic strength of neopterin exceeded that of soluble IL-2 receptor . To further assess if other factors could interfere with the predictive significance of both compounds, we also investigated other clinical and laboratory variables but these candidate predictors did not contribute any additional significant predictive information . CONCLUSION: The measurement of serum neopterin and soluble IL-2 receptor concentrations has predictability for identifying patients with gram-negative sepsis at risk for progression toward the syndrome of septic shock. Acta Paediatr Jpn, 1995 Jun, 37(3), 331 - 5 Clinical patterns of X linked agammaglobulinemia in Malaysian children; Noh LM et al.; X linked agammaglobulinemia (XLA) is rarely reported from developing countries especially from South East Asia . It appears that X linked agammaglobulinemia is less common in certain ethnic groups . It is very uncommon in black people in USA and South Africa . In multiracial Malaysia we have documented five XLA in Malays and Indians but not in the Chinese that constitute about 31% of the population . First degree relatives afflicted with XLA or other primary immunodeficiencies occurred more often in our study . All showed lung involvement although the etiologic organisms involved were atypical, being Gram negative. Ther Umsch, 1995 Jun, 52(6), 367 - 73 {Prostatitis}; Ludwig M et al.; During the last years, diagnosis and therapy of prostatitis have advanced rapidly . A detailed standardized diagnostic procedure is mandatory to delimit inflammatory from noninflammatory variations of the prostatitis syndrome . Whereas Gram-negative pathogens are agreed to be etiologically responsible, the relevance of chlamydia and mycoplasma is still under debate . Further diagnostic work-up should comprise ejaculate analysis, screening of the bladder voiding and transrectal prostatic sonography . The therapy of the inflammatory prostatitis is aimed to the demonstration of relevant pathogens, antibiotic treatment must be suited to expected sensitivity of the pathogen . Surgical procedures may be discussed when antibiotic treatment has failed . If it is not possible to identify relevant pathogens, the therapy remains of experimental character . Prostatodynia as noninflammatory disease often requires psychodynamic exploration. Eur J Clin Microbiol Infect Dis, 1995 Jun, 14(6), 520 - 3 Capnocytophaga canimorsus septicemia: fifth report of a cat-associated infection and five other cases; Valtonen M et al.; Capnocytophaga canimorsus is a fastidious, slow-growing, gram-negative, rod-shaped bacterium that belongs to the normal oral flora of dogs and cats . Human septicemic infections are associated with a high mortality; most cases occur in immunocompromised patients with a history of dog bite . The fifth case of cat-associated septicemia caused by Capnocytophaga canimorsus is described . The six case reports presented here point out the characteristics reported previously: (a) cats are a source of human infection; (b) alcohol abuse is an important risk factor for the development of septicemic Capnocytophaga canimorsus infection; (c) septicemic infection often manifests with disseminated intravascular consumption coagulopathy or purpura; and (d) some cases of septicemia in humans result from pets that lick skin ulcers. J Am Soc Nephrol, 1995 Jun, 5(12), 2067 - 72 Expression and preferential inhibition of inducible nitric oxide synthase in aortas of endotoxemic rats; Weigert AL et al.; Septic shock is associated with high mortality . There is in vitro evidence that the induction of nitric oxide synthase (iNOS) in vascular smooth muscle cells may be an important mediator of the systemic vasodilation and hypotension associated with sepsis . In this study, an in vivo murine model of sepsis was used to further examine this important question . Lipopolysaccharide (LPS), the major wall component of gram-negative bacteria, was administered to rats . By the use of a selective cDNA probe for iNOS, mRNA for iNOS was demonstrated in the aortas of these rats . The functional significance of this iNOS was then examined with aminoguanidine, a preferential inhibitor of iNOS . Aminoguanidine reversed the blunted phenylephrine-evoked contraction of endothelium-denuded aortic rings from LPS-treated rats or rings exposed to LPS in vitro . Aminoguanidine did not impair the relaxation of aortic rings with endothelium to acetylcholine, a known stimulator of endothelial NOS . The reversal of LPS-induced vascular hyporesponsiveness by aminoguanidine therefore strongly supports the functional importance of iNOS mRNA expression in the aorta of endotoxemic rats . Future clinical trials in treating septic shock should therefore consider the preferential inhibition of iNOS while maintaining the integrity of endothelial NOS. J Hosp Infect, 1995 Jun, 30 Suppl, 218 - 22 Protective isolation: who needs it? Fenelon LE. Infection continues to be a major cause of morbidity and mortality in neutropenic patients following chemotherapy or bone marrow transplantation (BMT) . Concerted efforts have been made to protect these patients from infection during the neutropenic period . Elaborate protocols to protect the patient from both intrinsic and extrinsic pathogens have been devised, ranging from simple single room isolation to laminar air flow units (LAFs), in association with varying degrees of antibiotic decontamination of the digestive tract . Comparative rates of infection using these techniques have varied in different studies, and their use has been somewhat controversial . More recently, prophylactic quinolone administration to neutropenic patients has significantly decreased the incidence of both Gram-negative septicaemia and pyrexial episodes, probably superseding any advantages which may have been conferred by previous regimens . LAFs with high efficiency particulate air filtration still appear to be the best means of protection against aspergillosis, but are expensive and would not be available for the majority of neutropenic patients . They should probably be allocated to patients who are most at risk; BMT recipients or others who may be expected to have a prolonged neutropenic period. Brain Behav Immun, 1995 Jun, 9(2), 113 - 28 Spatial learning impairment in mice infected with Legionella pneumophila or administered exogenous interleukin-1-beta; Gibertini M et al.; The effect of interleukin-1 beta (IL 1 beta) on spatial learning was examined . In one experiment, C57BL/6 mice were given daily injections (100 ng/mouse) of recombinant murine IL1 beta prior to training on the Morris water maze . In another experiment, mice were infected with a sublethal dose of a gram-negative bacterium (Legionella pneumophila; Lp) . Mice rendered ill by the infection were given either anti-IL1 beta antibodies (100 micrograms/mouse) or saline and then trained on the water maze . Results indicated that (1) exogenous IL1 beta blocked acquisition of spatial learning, (2) Lp infection attenuated learning on this task, and (3) neutralizing circulating IL1 beta in Lp-infected mice normalized learning despite the continuation of the illness . The data indicate that cognitive impairment may be a component of cytokine-mediated sickness behavior. Cardiovasc Res, 1995 Jun, 29(6), 813 - 9 Nitric oxide synthesis in cardiac myocytes and fibroblasts by inflammatory cytokines; Shindo T et al.; OBJECTIVE: The aim was to investigate nitric oxide (NO) synthase activity in cultured neonatal rat cardiac myocytes and fibroblasts upon treatment with inflammatory cytokines interleukin 1 beta (IL-1 beta), tumour necrosis factor alpha (TNF-alpha), IL-2, IL-6, IL-8, transforming growth factor beta (TGF-beta) and gram negative bacterial lipopolysaccharide (LPS) . METHODS: NO and guanosine 3',5'-cyclic monophosphate (cGMP) synthesis was measured in cultured neonatal rat cardiac myocytes and fibroblasts, using Griess reagent and an enzyme immunoassay kit, respectively . The expression of inducible NO synthase (iNOS) mRNA and protein was assayed by northern and western blotting, respectively . RESULTS: Incubation of cardiac myocytes for 24 h with IL-1 beta (10 ng.ml-1) or LPS (1 microgram.ml-1) caused significant increases in NO and cGMP production . TNF-alpha, IL-2, IL-6, IL-8, and TGF-beta showed no significant effect on their production . IL-1 beta induced NO and cGMP production in a time and dose dependent manner . IL-1 beta also increased iNOS mRNA and protein accumulation in cardiac myocytes . Simultaneous incubation of IL-1 beta with NG-monomethyl-L-arginine, genistein, calphostin C, cycloheximide, or actinomycin D completely inhibited the IL-1 beta induced NO production by cardiac myocytes . TGF-beta, dexamethasone, or cyclosporin A also dose dependently inhibited the IL-1 beta induced NO production . Exposure to IL-1 beta for 12-24 h decreased the beating rate of cardiac myocytes, but addition of dexamethasone completely overcame this inhibition . In contrast to cardiac myocytes, incubation of cardiac fibroblasts for 24 h with IL-1 beta or LPS showed no significant effect on NO or cGMP production . CONCLUSIONS: These observations suggest that IL-1 beta/LPS responsive iNOS, which is an important regulator of contractile function of the heart, is present in cardiac myocytes but not in cardiac fibroblasts. Infect Immun, 1995 Jun, 63(6), 2147 - 53 Cloning, characterization, and antigen specificity of T-lymphocyte subsets extracted from gingival tissue of chronic adult periodontitis patients; Wassenaar A et al.; Chronic periodontitis is characterized by dense infiltrations of B and T lymphocytes within the gingival connective tissue . Distinct anaerobic gram-negative bacteria as well as autoimmunity to collagen have been reported to play a role in the etiology and the pathogenesis of this disease . Here we describe the cloning and characterization of CD4+ and CD8+ T lymphocytes isolated from inflamed gingival tissue obtained from four patients with chronic periodontitis . Clones were raised with phytohemagglutinin and interleukin-2 and tested for proliferation in response to whole-cell antigens of Porphyromonas gingivalis, Prevotella intermedia, Actinobacillus actinomycetemcomitans, human collagen type I, and two bacterial heat shock proteins . CD4+ T-cell clones reactive with collagen type I were obtained from all four patients . Eighty percent of these clones had phenotypes resembling the mouse type 2 T helper (Th) phenotype, i.e., they produced high levels of interleukin-4 and low levels of gamma interferon . No collagen-type-I-reactive CD8+ clones were obtained . Bacterial-antigen-reactive CD4+ and/or CD8+ T-cell clones were also obtained from each patient, and the majority of the clones showed a Th0-like cytokine pattern and produced equal amounts of interleukin-4 and gamma interferon . Although most clones were reactive with P . intermedia, it seems that the immune response is not strictly directed against this particular microorganism, as clones reactive with one of the other bacteria were also obtained from two patients . We propose that collagen-specific CD4+ Th2-like T cells contribute to the chronicity of periodontitis but that their modes of activation might be controlled by Th0-like T cells specific for periodontitis-associated bacteria. Biochem Biophys Res Commun, 1995 May 25, 210(3), 746 - 52 Direct vesicle-vesicle exchange of phospholipids mediated by polymyxin B; Cajal Y et al.; Direct and rapid intermembrane exchange of phospholipids without fusion is shown to occur across stable vesicle-vesicle contacts formed by stoichiometric amounts of polymyxin B . The exchange is selective for monoanionic, but not dianionic, glycerophospholipids irrespective of their chain length or phase properties . Selective transfer mediated by protein contacts between membranes could serve as a model for stable fusion intermediates and provide a structural and organizational basis for direct phospholipid transfer as implicated in entry of enveloped viruses, secretion, endocytosis, and intracellular transport and targeting of lipids . It is also proposed that the loss of the specificity of phospholipid composition in the inner and outer membranes of gram negative bacteria mediated by polymyxin B could be the basis for its bactericidal action. J Mol Biol, 1995 May 5, 248(3), 627 - 38 The products of gene I and the overlapping in-frame gene XI are required for filamentous phage assembly; Rapoza MP et al.; The class I filamentous bacteriophage are non-lytic single-stranded DNA phage, which are assembled at the cell envelope as they are extruded from the Gram-negative bacteria, Escherichia coli . The process requires the products of the phage genes I and IV, which reside in the inner and outer membrane, respectively, and are not present in the mature phage particle . Gene I encodes two proteins, the full length 348-residue pI and a smaller pI*, which this report shows is the result of an internal translation initiation event at methionine codon 241 . Both pI and pI* are shown to be required for phage assembly . Therefore, pI* can be considered the product of an additional phage gene, XI, which is a separate in-frame gene that overlaps gene I . Both proteins contain a 13-residue region adjacent to the cytoplasmic face of the inner membrane that probably exists as a positively charged amphiphilic helix . Although this region is not required for membrane insertion of pI and pI*, it is shown to be required for phage assembly . Oligonucleotide-directed mutagenesis of this region, which removes positive charges or alters the hydrophobic face of the putative helix, renders pI and pI* unable to function in phage assembly . This region of pI and pI* is highly homologous in structure to the carboxyl-terminal 11 amino acids of pVIII, the main coat protein, which also reside adjacent to the cytoplasmic face of the inner membrane. Immunol Today, 1995 May, 16(5), 231 - 6 An important role for intestinally derived T cells in respiratory defence; Dunkley M et al.; Margaret Dunkley, Reinhard Pabst and Allan Cripps discuss the role of intestinally derived T cells in protecting the lung against Gram-negative bacterial infection . They describe the factors directing T-cell migration from gut-associated lymphoid tissue to lung, and focus on the role of T cells and T-cell-derived cytokines in bacterial clearance from the lung. Microbiology, 1995 May, 141 ( Pt 5), 1201 - 10 Evolution of the korA-oriV segment of promiscuous IncP plasmids; Thomas CM et al.; Plasmids belonging to Escherichia coli incompatibility group P are of particular interest because they can transfer between, and be stably maintained in, almost all Gram-negative bacterial species . The segment of the IncP alpha plasmid genome between the key regulatory gene korA and the vegetative replication origin, oriV, encodes a series of operons co-regulated with replication and transfer functions by the KorA protein . To determine which of these genes are likely to have an important role in IncP plasmid survival the equivalent region of the distantly related IncP beta plasmid R751 was sequenced . Sequence comparisons show that the kla operon (formerly the kilA locus, which is also responsible for a cryptic tellurite-resistance determinant) is completely absent from R751 . Similarly in the kle region, which encodes genes associated with the KilE+ phenotype of unknown function, kleC and kleD, which we proposed arose by a duplication of kleA and kleB, are also completely absent . The genes that are conserved are klcA (formerly kilC, responsible for the KilC+, and recently proposed to be involved in overcoming restriction barriers during transfer), klcB (an ORF interrupted by Tn1 insertion in RK2), korC (a transcriptional repressor which controls the klcK and kle operons), and kleA, kleB, kleE and kleF . A striking feature of the organization in R751 is the lack of the strong transcriptional termination signals which are present in IncP alpha plasmids . The degree of divergence between the plasmids facilitates the identification of motifs of probable functional importance in the primary protein sequences. Am J Physiol, 1995 May, 268(5 Pt 1), C1104 - 13 TNF modulates endothelial properties by decreasing cAMP; Koga S et al.; Tumor necrosis factor-alpha (TNF-alpha), a monokine that contributes to vascular dysfunction accompanying the host response to gram-negative sepsis, has been shown to increase vascular permeability in vivo and to diminish the barrier function of cultured endothelial cell (EC) monolayers . The studies reported here indicate that a mechanism through which TNF alters EC barrier function involves a reduction in intracellular adenosine 3',5'-cyclic monophosphate (cAMP) content, due in part to increased cyclic nucleotide phosphodiesterase (CNPDE) activities . TNF increased the diffusional transit of {3H}sorbitol, {3H}inulin, and 125I-labeled albumin across confluent bovine aortic EC monolayers . This effect of TNF was both time and dose dependent and occurred in parallel with a fall in EC cAMP . cAMP analogues, such as dibutyryl cAMP (DBcAMP), prevented TNF-induced perturbation of EC barrier function . TNF also mediated another important alteration in the EC phenotype, in that both mRNA and activity of the anticoagulant cofactor thrombomodulin were reduced after exposure of EC to TNF and were normalized by the addition of DBcAMP . EC monolayers exposed to TNF-alpha showed increased cAMP levels when exposed to 3-isobutyl-1-methylxanthine, a nonspecific CNPDE inhibitor . Ion exchange chromatography of cytosol derived from TNF-treated EC consistently showed an approximately 245% increase in phosphodiesterase (PDE) IV (high-affinity, cAMP-specific PDE) activity as identified by rolipram inhibition . PDE II activity was increased by 150% after TNF-alpha treatment of early passage EC, which was identified by cGMP-activated hydrolysis of cAMP . Western and Northern analyses, as well as activity studies, revealed that TNF treatment did not change the amount of PDE IV protein or mRNA but rather increased the specific activity of the isozyme, suggesting that a posttranslational modification had occurred . These data indicate that activation of EC CNPDE activity and decreased intracellular cAMP may represent a mechanism by which TNF increases EC permeability and promotes a procoagulant EC phenotype. Med Clin North Am, 1995 May, 79(3), 619 - 49 Treatment of pyelonephritis in adults; Bergeron MG; Although prescribing an antibiotic for the treatment of pyelonephritis seems to be a relatively easy task, a close look at the available data is disturbing . Optimal therapies for the different clinical syndromes of pyelonephritis have not yet been defined . The high failure rate suggests that in pyelonephritis (bacteria protected in the medulla) as well as in bacterial endocarditis (bacteria sequestered in vegetations) and in infections in neutropenic patients (host defenses not necessarily operating in conjunction with antibiotics), it may be necessary to maintain bactericidal levels at the site of infection (infected medulla) to achieve cure . Pharmacodynamic studies suggest that TMP/SMX, quinolones, and aminoglycosides, which penetrate well the infected renal parenchyma and are not impaired by the local inflammatory process, should, with the exception of pyelonephritis in pregnancy, be preferred to beta-lactams as first-choice agents for the therapy of gram-negative pyelonephritis. Med Clin North Am, 1995 May, 79(3), 581 - 97 The antibiotic treatment of community-acquired, atypical, and nosocomial pneumonias; Cunha BA; Optimal antibiotic regimens and duration of treatment are not universally agreed on for community-acquired or nosocomial pneumonias . Experience suggests that community-acquired pneumonias may be treated for less than 2 weeks with a combination of intravenous and oral antibiotics of appropriate spectrum that penetrate the lung, have a good safety profile, do not foster the development of resistance, and are cost-effective . After initial intravenous therapy, oral switch therapy may be begun as soon as the patient defervesces clinically, which is usually 3 days after admission . Switching to oral therapy does not invariably lead to earlier hospital discharge . There is no "standard of care" for pneumonias, but guidelines for empiric use have existed for decades . The least expensive beta-lactamase stable antibiotic should be used as monotherapy for the empiric treatment of community-acquired pneumonia . Because community-acquired atypical pneumonias are clinically distinct from bacterial pneumonias owing to their extrapulmonary features, clinicians should be able to differentiate atypical pneumonias from bacterial pneumonias, which permits prompt and appropriate treatment . Nosocomial pneumonias remain a difficult diagnostic challenge . Therapeutically the most important principle in treating nosocomial pneumonia is to provide for double-drug coverage against P . aeruginosa . Differentiation of respiratory tract colonization from respiratory tract invasion remains the central key issue in patients with pulmonary infiltrates acquired during hospitalization . Most patients complete their course of intravenous therapy for nosocomial pneumonia leaving little or no time for completion of their therapy by oral antibiotics . Hospital-acquired atypical pneumonias are largely limited to legionnaires' disease, which is a more difficult diagnosis than in the community-acquired setting . Clinicians taking care of patients with pneumonia should employ a simplified therapeutic approach using a single drug for community-acquired infections . The use of additional antibiotics to increase gram-negative coverage is medically unjustified and not cost-effective and is to be discouraged . The most cost-effective strategy for the treatment of community-acquired pneumonias is to switch the patient from an intravenous to an oral antibiotic as soon as the patient clinically defervesces and is able to take oral medications . Antimediator therapies have no role in the treatment of community-acquired or nosocomial pneumonias. Ann Surg, 1995 May, 221(5), 572 - 7; discussion 577-8 Prolongation of skin allografts by recombinant tumor necrosis factor and interleukin-1; Parenteau GL et al.; OBJECTIVE: The hypothesis is that systemic administration of recombinant tumor necrosis factor-alpha (TNF-alpha) and/or recombinant interleukin-1 alpha (IL-1 alpha) can decrease the rejection of a skin allograft . SUMMARY BACKGROUND DATA: Tumor necrosis factor and IL-1 are pluripotent cytokine hormones that are central to the host immunologic response to foreign substances . Cytokine effects and toxicity may be reduced by systemic administration of recombinant cytokines . The authors previously have demonstrated that pretreatment with cytokines such as IL-1 or TNF can reduce the lethality of endotoxin (lipopolysaccharide), gram-negative sepsis, cancer cachexia, and oxygen toxicity . METHODS: Skin grafts from the tails of Balb/c mice were placed on the backs of C57Bl/6 mice . Mice were treated with daily intraperitoneal saline, recombinant m-TNF (Genentech, South San Francisco, CA) or h-IL-1 (Hoffman LaRoche, Nutley, NJ) from postgraft day 1 to postgraft day 28 . Tumor necrosis factor and IL-1 high doses were chosen because they protected mice from the lethality of lipopolysaccharide . Animals were examined daily for toxicity and graft rejection . Graft survival was plotted in a Kaplan-Meier plot and analyzed by the log-rank test . Comparison of proportions was done using the Fisher's exact test . RESULTS: Either TNF or IL-1 alone significantly prolonged skin graft survival compared with saline control . Furthermore, the combination of TNF and IL-1 prolonged skin graft survival longer than either cytokine alone . Mice on the highest dose TNF and IL-1 combination did not reject skin grafts during the 28-day treatment period . Significant toxicity was associated with cytokine treatment . Similar significant proportions of death occurred with IL-1 alone and the highest combination of TNF and IL-1 . CONCLUSION: Both TNF and IL-1 can be effective as suppressors of skin allograft rejection in mice. J Cell Physiol, 1995 May, 163(2), 328 - 38 Lipopolysaccharides stimulate Na-dependent transport in alveolar cells and protect against oxidant injury; Azarian R et al.; We have evaluated the effect of lipopolysaccharides (LPS), endotoxins from gram negative bacteria, on sodium-coupled amino acid and phosphate transport by alveolar epithelial type II cells and on their alteration induced by oxidants . Alveolar type II cells were obtained by enzymatic digestion of rat lung and grown for 24 h prior to incubation with LPS and then exposed or not exposed to H2O2 (2.5 mM; 20 min) . LPS (10 micrograms/ml, 24 h) induced a significant increase in the Na-dependent component of alanine and phosphate uptake while they decreased Na,K-ATPase activity measured by ouabain-sensitive 86Rb influx . We showed that this stimulatory effect i) was independent from macrophage products since it was not mimicked either by supernatant of LPS-treated alveolar macrophages or by pretreatment with tumor necrosis factor and/or interleukin 1 and ii) was dependent on protein synthesis since it was abolished by protein synthesis inhibitors cycloheximide and actinomycin D . Moreover, LPS blunted H2O2-induced decrease of Na-dependent alanine and phosphate uptake . This protective effect of LPS against H2O2 injury i) was independent of macrophage products, ii) was abolished by cycloheximide, and iii) was not associated with either changes in extracellular H2O2 clearance or catalase and glutathione peroxidase activities . We conclude that, in alveolar type II cells, LPS stimulate sodium-coupled transport by a process involving protein synthesis and partially prevent H2O2-induced decrease of Na-coupled transport without discernible change in antioxidant activities. Inflamm Res, 1995 May, 44(5), 187 - 97 Modulins: a new class of cytokine-inducing, pro-inflammatory bacterial virulence factor; Henderson B et al.; Despite the fact that the inflammatory and immune responses have evolved to combat microorganisms, the present generation of inflammation researchers has evinced relatively little interest, with the exception of septic shock, in microbially-induced inflammation . This in spite of the fact that the Gram-negative cell wall constituent, lipopolysaccharide, has been widely used as a tool in inflammation research . The reason for such lack of interest has been due to the therapeutic efficacy of antibiotics which are the treatment of choice for infections and their inflammatory sequelae . However, this is likely to change within the next decade or so, with the relentless increase in the incidence of antibiotic-resistant strains of bacteria . This will return therapy to the stage where clinicians will have to treat the inflammatory symptoms of infection . Many of these symptoms are due to the stimulation of cytokine synthesis . The capacity of bacteria to induce cytokine synthesis has, until the past few years, centred exclusively on lipopolysaccharide . However, it has been established during the past 5-10 years that a range of other molecules, mainly associated with the surface of bacteria, have the capacity to induce cytokine production . Some of these are exquisitely potent stimulators of pro-inflammatory cytokine synthesis . The nature and mechanism of action of these various cytokine-inducing molecules, for which we have devised the name modulins, is the subject of this review . It is clear that bacteria still have many surprises for us, as exemplified by the recent discovery of the role played by Helicobacter pylori in gastritis, gastric ulceration and gastric cancer. Support Care Cancer, 1995 May, 3(3), 198 - 202 Epidemiology of influenza A virus infection in patients with acute or chronic leukemia; Elting LS et al.; Influenza infection is a significant cause of morbidity and mortality in immunocompromised hosts, but its importance in adult cancer patients is largely undescribed . We therefore conducted a prospective study of the incidence and clinical features of influenza infection in patients with acute or chronic leukemia . The cohort, which consisted of all adult leukemia patients undergoing remission-induction chemotherapy during the 1991-1992 influenza epidemic, was followed prospectively for development of signs and symptoms of acute infection of the upper or lower respiratory tract . Of these 294 patients, 111 received chemotherapy as inpatients and 183 as outpatients . Throat swabs and nasal washes for viral culture were obtained from all symptomatic patients, who were then followed until all signs and symptoms resolved . Symptoms of respiratory tract infection developed in 37 leukemia patients (13%) . Among these, influenza (A/Beijing/ H3N2) caused 3 (21%) of the 14 infections that developed during hospitalization but only 1 (4%) of the 23 that developed in the community (P = 0.14) . Influenza patients presented with fever, rhinorrhea, nasal congestion, headache, and myalgia; those with other infections presented with signs and symptoms of lower respiratory tract infection (productive cough, rales, or rhonchi) . Development of pneumonia was common in influenza patients, 1 of whom died from secondary fungal and gram-negative pneumonia . Influenza A virus infections accounted for a substantial portion of acute respiratory infections among adult leukemia patients during a community epidemic . Most infections appeared to be nosocomial and the most likely sources were visitors or hospital personnel . Immunization of household contacts and hospital staff may reduce the risk of influenza infection and its pulmonary complications in leukemia patients. Rev Clin Esp, 1995 May, 195(5), 289 - 93 {Psoas abscess: report of 8 cases and review of the literature}; Garcia Vazquez E et al.; Eight cases of psoas abscesses (PA) are reported and a comparative study is made with other series as well as a review of the literature . Cases in our series are analysed and an assessment is made of microbiological etiologies, the presence of an origin and its spreading into the surrounding tissues or their primary character, and particularly the diagnostic and therapeutic usefulness of CT-guided percutaneous drainage . Diagnosis was ascertained by CT in all seven cases in which it was performed; percutaneous drainage was performed in five cases and allowed the microbiological categorization of the abscess and an appropriate antibiotic therapy . Laparotomy was performed in only one case; and in two other cases only antibiotics were administered . A favourable outcome occurred in seven out of the eight cases . One of the two patients treated with antibiotics only died . Blood cultures were positive in only three patients . In summary, PA in our series were secondary to bone, urologic or digestive origins in 87.5% of cases . With regard to the causative agent, 37.5% of cases were caused by S . aureus and 50% by gram negative and/or anaerobic organisms . The diagnostic yields of echocardiography and CT were 40% and 100%, respectively . Percutaneous drainage ascertained the microbiological diagnosis in 100% of cases when it was performed, as well as a therapeutic regimen associated with the use of antibiotics . No relapses were recorded in the follow-up of patients . Therefore, we believe that therapy of PA should be based on the association of percutaneous drainage and antibiotics. J Clin Microbiol, 1995 May, 33(5), 1278 - 82 Reappraisal with meta-analysis of bacteremia, endotoxemia, and mortality in gram-negative sepsis; Hurley JC; Among patients with suspected sepsis, endotoxemia is variably present in association with gram-negative bacteremia . A total of 738 patients with suspected sepsis from 11 studies could be classified into four groups: 131 (18%) patients had both endotoxemia and gram-negative bacteremia (group 1), 87 (12%) had only gram-negative bacteremia (group 2), and 143 (19%) had only endotoxemia (group 3); in 377 (51%) patients neither could be detected (group 4) . By the statistical techniques of meta-analysis, the fatality risk for patients with either endotoxemia or gram-negative bacteremia or both was estimated with group-specific case fatality rates from these studies and expressed as an odds ratio (OR; 95% confidence interval {95% CI}) versus patients with these factors absent . This risk was increased marginally when endotoxemia was detected without gram-negative bacteremia (OR, 2.3; 95% CI, 1.3 to 4.0) or the converse (OR, 2.0; 95% CI, 1.0 to 3.8), in contrast to the striking increase when both endotoxemia and gram-negative bacteremia were detected (OR, 6.3; 95% CI, 4.0 to 10.0) . Alone, neither endotoxemia nor gram-negative bacteremia is a strong predictor of outcome in patients with suspected sepsis . However, in combination, the two identify a subpopulation with a substantially increased risk of mortality. Clin Sci (Lond), 1995 May, 88(5), 587 - 94 Plasminogen activator and plasminogen activator inhibitor I release during experimental endotoxaemia in chimpanzees: effect of interventions in the cytokine and coagulation cascades; Biemond BJ et al.; 1 . Disseminated intravascular coagulation frequently accompanies Gram-negative sepsis and may contribute to widespread deposition of microthrombi . Besides the endotoxin-induced activation of coagulation, an important role for the fibrinolytic system has been postulated . The precise mechanisms underlying these fibrinolytic changes during endotoxaemia are not known but have been suggested to be mediated directly by cytokines or secondary to thrombin generation . 2 . In the present study we have delineated in detail the fibrinolytic response to a bolus injection of endotoxin in non-human primates and analysed the contribution of cytokines and thrombin generation to the endotoxin-induced release of tissue-type plasminogen activator and plasminogen activator inhibitor 1 . Chimpanzees received a bolus injection of endotoxin alone or in combination with blocking monoclonal antibodies directed against tumour necrosis factor or interleukin 6 or in combination with pentoxifylline . Furthermore, to assess the effect of coagulation activation on the activation of fibrinolysis, another group of chimpanzees received endotoxin in combination with either anti-tissue factor antibodies or recombinant hirudin . 3 . Infusion of endotoxin induced a rapid increase in plasminogen activator activity and tissue-type plasminogen activator antigen levels and subsequent plasmin generation, reaching peak levels 2h after endotoxin administration . Plasminogen activator inhibitor 1 levels remained constant for the first 2 h, after which time a steep increase was observed . Plasminogen activator activity and plasmin generation decreased simultaneously with the rise in plasminogen activator inhibitor 1 levels . Fibrinolytic activity remained suppressed during the remainder of the study owing to sustained increased levels of plasminogen activator inhibitor 1 . The administration of pentoxifylline strongly attenuated the release of tissue-type plasminogen activator and plasminogen activator inhibitor 1, whereas the antitumour necrosis factor antibodies blocked the fibrinolytic response entirely . In contrast, interleukin 6-neutralizing antibodies did not affect the fibrinolytic response . Although endotoxin-induced generation of thrombin was completely prevented by the administration of tissue factor-neutralizing antibodies or by hirudin, no effect on the fibrinolytic response was observed.(ABSTRACT TRUNCATED AT 250 WORDS) Nippon Geka Gakkai Zasshi, 1995 May, 96(5), 277 - 85 {Endotoxin eliminating therapy in patients with severe sepsis--direct hemoperfusion using polymyxin B immobilized fiber column}; Kodama M et al.; BACKGROUND AND METHODS: PMX is a blood purifier containing chemically immobilized polymyxin B fiber (PMX-F) . To evaluate its effectiveness on the severe septic human, direct hemoperfusion (DHP) using a PMX-F was performed for 2 hours . The changes in various circulatory parameters, symptoms of septic shock, and blood endotoxin concentration and the survival rate, were evaluated . RESULTS: DHP was performed 61 times in 42 patients, of whom 38 had septic MOF, 25 with gram-negative bacterial infection . At the initiation of this treatment, 33 patients were receiving vasoactive agents, and 36 were under artificial ventilation via endotracheal intubation . The mean septic severity score (SSS) in all patients was about 46.6 . Twenty-two of the 42 survived . The endotoxin concentration (mean +/- S.E.; pg/ml) was 85.0 +/- 27.2 immediately before treatment but significantly decreased to 57.5 +/- 28.4 after treatment (n = 50) and to 28.2 +/- 4.4 on the next day (n = 23) (p < 0.01) . The endotoxin concentrations at the inlet and outlet of the PMX also significantly decreased 30 minutes after the initiation of DHP . Circulatory parameters, BP, CI, SVR and Vo2I demonstrated significant improvement . Body temperature also showed the same results . The removal of endotoxin in the blood using PMX was effective for severe sepsis or septic MOF . Various symptoms due to endotoxin was alleviated after this treatment. Diagn Microbiol Infect Dis, 1995 May-Jun, 22(1-2), 159 - 61 Prospective evaluation of twice-daily cefotaxime in the treatment of hospitalized patients with severe infections; Keller C; Cefotaxime 2 g every 12 h was administered to adults with severe nosocomial pneumonia in a prospective noncomparative study . The results confirmed that this regimen is adequate and appropriate therapy for nosocomial pneumonia, with the combination of cefotaxime and an aminoglycoside reserved for cases where multiresistant, Gram-negative bacterial infections are strongly suspected. New Horiz, 1995 May, 3(2), 342 - 51 Signaling events in monocytes and macrophages; Chow CW et al.; The cellular signaling events leading to the systemic inflammatory response syndrome and sepsis in monocytes/macrophages activated by lipopolysaccharide (LPS) are well understood . LPS is a glycolipid component of Gram-negative bacterial cell wall . It exerts its effect through the lipid A moiety . LPS binds to monocytes/macrophages via a membrane-bound receptor, CD14, an interaction which is optimized in the presence of plasma factors, LPS-binding protein, and septin . Although LPS is known to bind to other receptors, the roles of these receptors in transmembrane signaling and activation of monocytes/macrophages are not as well understood as is that of the CD14 receptor . Intracellular events in response to LPS stimulation are mediated by phospholipase (PL) C, protein kinases, PLA2, and PLD . Activation of PLC by LPS results in the release of diacylglycerol and inositol 1,4,5-trisphosphate . The former mediates the stimulation of protein kinase C, and the latter induces an increase in intracellular calcium concentration . LPS stimulation of monocytes/macrophages also results in the phosphorylation and activation of several protein kinases, including protein tyrosine kinases which mediate cytokine production, and mitogen-activated protein kinase which activates cytosolic PLA2 to release arachidonate . LPS also plays a role in cellular proliferation and differentiation . Upregulation of the secretory form of PLA2 has also been documented in response to LPS . PLD is stimulated by LPS to release phosphatidic acid (PA) . PA can activate the respiratory burst by increasing diacylglycerol production and by modulating the effects of guanine nucleotide-binding proteins . Therapeutic strategies to decrease the clinical effects of sepsis would logically include agents which block at initial receptor-ligand interaction, as well as those which attenuate the intracellular events that follow LPS stimulation . Early in vivo studies are promising, but clearly much work remains to be done. New Horiz, 1995 May, 3(2), 267 - 75 Pathogenic effects of endotoxin; Zivot JB et al.; Endotoxin is a lipopolysaccharide contained within the cell wall of Gram-negative bacteria . This molecule initiates a host inflammatory response to Gram-negative bacterial infection . An adequate inflammatory response likely enhances host survival by mediating clearance of infection and bacterial toxins . Unfortunately, this same host response can also produce dysfunction of multiple organ systems and mortality . This article focuses on the history of our understanding of the role of endotoxin in human septic shock . These pathophysiologic connections have led to therapies directed at endotoxin . Unfortunately, antiendotoxin therapy has not achieved significantly improved outcome in humans with severe sepsis . This may represent lack of antiendotoxin efficacy in the compounds used, or a failure of the investigative approach . Interest in antiendotoxin therapies persists, while investigators express more humility in their understanding of endotoxin's role in the pathophysiology of septic shock. J Surg Res, 1995 May, 58(5), 496 - 502 Postoperative selective bowel decontamination prevents gram-negative bacterial translocation in small-bowel graft recipients; Lee TK et al.; Gram-negative septic episodes are a potential risk of small-bowel transplantation; bacterial translocation through the graft is considered the mechanism . As a measure to prevent this complication, we evaluated postoperative selective bowel decontamination (SBD) in the rat model of orthotopic small-bowel transplantation {Lewis (LEW) and Brown-Norway (BN) rats as donors and recipients} . For 4 days after transplantation we gave FK 506, 2 mg/kg, which prevents rejection and results in indefinite recipient survival . For SBD, 24 mg/kg/day polymyxin E and 20 mg/kg/day tobramycin were administered via orogastric gavage to allograft recipients, both with and without FK 506 therapy . On Day 9, all rats were sacrificed, the peritoneal cavity was swabbed, and mesenteric lymph nodes (MLN), spleen, liver, and ileum were harvested for microbial qualitative and quantitative analysis . Animals with positive peritoneal swab cultures were excluded . SBD resulted in a significant reduction of the quantitative gram-negative bacterial flora in the ileum and cecum and of bacterial translocation to the MLN {0% versus 50% (no FK 506 therapy) and 8% versus 50% (FK 506 treated)} . In the allograft groups not treated with FK 506, SBD failed to significantly prolong survival, suggesting that acute rejection is not hastened by infection (bacterial translocation) . We conclude that SBD in small-bowel-graft recipients prevents bacterial translocation by reducing intestinal gram-negative bacterial flora; this may reduce local and systemic infections by gut-derived organisms. Microb Pathog, 1995 May, 18(5), 307 - 21 Mutational analysis of the putative leukotoxin transport genes in Actinobacillus actinomycetemcomitans; Guthmiller JM et al.; The periodontal pathogen, Actinobacillus actinomycetemcomitans, produces leukotoxin, a protein that specifically lyses host defense cells . The leukotoxin is similar in sequence and operon organization to the Escherichia coli alpha-hemolysin and other members of the RTX family of toxins . However, unlike the other RTX toxins, the A . actinomycetemcomitans leukotoxin is not secreted from the cell and instead remains associated with the outer membrane . Nonetheless, the A . actinomycetemcomitans Ikt operon contains two genes, IktB and IktD, that appear analagous to the toxin localization genes found in the other Gram-negative bacteria . Thus, to determine the roles of these putative transport genes in A . actinomycetemcomitans, we have used insertional mutagenesis to generate mutant strains lacking functional LktB and/or LktD . When either IktD or both IktB and IktD were inactivated, the level of detectable leukotoxin protein in the cell decreased significantly . However, the IktB and IktD mutations had no effect on the levels of leukotoxin RNA . Thus, the lack of LktB and LktD proteins must affect LktA synthesis post-transcriptionally . It is proposed that this is an indirect effect of leukotoxin mislocalization in IktB- and IktD- mutants . Finally, analysis of the mutants revealed that LktB and LktD are not essential for the formation of extracellular membrane vesicles in A . actinomycetemcomitans. Genes Dev, 1995 Apr 15, 9(8), 972 - 83 Myxococcus xanthus, a gram-negative bacterium, contains a transmembrane protein serine/threonine kinase that blocks the secretion of beta-lactamase by phosphorylation; Udo H et al.; A gene, pkn2, encoding a Myxococcus xanthus protein with significant similarities to eukaryotic protein serine/threonine kinases, was cloned using the polymerase chain reaction . The open reading frame for the protein, beginning with a GUG initiation codon, consists of 830 amino acids . The amino-terminal 279 residues show 37% identity to catalytic domain of Pkn1, another protein serine/threonine kinase expressed during the development at the onset of sporulation . The catalytic domain of Pkn2 contains 27% and 25% identity to rat Ca2+/calmodulin-dependent protein kinase and Bos taurus rhodopsin kinase, respectively . In the middle of the carboxy-terminal regulatory domain, there is a typical transmembrane domain consisting of 18 hydrophobic residues . The gene product, Pkn2, produced in Escherichia coli under a T7 promoter was phosphorylated at both serine and threonine residues . TEM-beta-lactamase produced in E . coli was found to serve as an effective substrate for Pkn2, phosphorylated only at threonine residues, shifting its apparent molecular mass from 29 to 44 kD . The phosphorylated beta-lactamase was unable to be secreted into the periplasmic space and localized in the cytoplasmic and membrane fractions . Analysis of phoA fusions with pkn2 demonstrated that Pkn2 is a transmembrane protein with the kinase domain in the cytoplasm and the 207-residue carboxy-terminal domain outside the cytoplasmic membrane . Disruption of pkn2 showed no effect on vegetative growth but reduced the yield of myxospores by 30%-50% . On the basis of the present results, we propose that Pkn2 is a transmembrane protein serine/threonine kinase that regulates the activity of endogenous beta-lactamase or related enzymes in response to an external signal yet to be identified. J Immunol, 1995 Apr 15, 154(8), 4032 - 8 Infection of bovine brain microvessel endothelial cells with Cowdria ruminantium elicits IL-1 beta, -6, and -8 mRNA production and expression of an unusual MHC class II DQ alpha transcript; Bourdoulous S et al.; Cowdria ruminantium is a bacterial parasite that infects ruminants, causing an acute and often fatal disease . These obligate intracellular Gram-negative bacteria preferentially infect neutrophils and vascular endothelial cells, especially in the brain . The present study was performed with bovine brain microvessel endothelial cells in culture, infected by C . ruminantium in the presence or absence of IFN-gamma . Infection induced the production of IL-1 beta, -6, and -8 mRNAs, and this effect was potentiated by IFN-gamma . A semi-quantitative PCR analysis indicated that similar amounts of IL-1 beta and IL-6 mRNAs were produced in response to C . ruminantium infection and to treatment with 30 to 40 ng/ml LPS . In addition, although IFN-gamma induced the synthesis of an MHC class II DQ alpha transcript (1.3 kb), an unusual transcript (1.5 kb) was induced by infection and not after LPS treatment . Infection did not affect MHC class I, class II DQ beta, and invariant chain mRNA levels . The present results suggest that C . ruminantium infection raises the immune activity of brain endothelial cells in vitro and that only part of this response can be attributed to LPS . One can hypothesize that cerebral endothelium in vivo efficiently contributes, by MHC Ag expression and production of ILs, to the activation and/or recruitment of leukocytes to the brain and thus plays an active role in the pathogenesis of cowdriosis and in the immune response to this pathogen. Gene, 1995 Apr 14, 156(1), 85 - 8 An enhanced broad-host-range vector for gram-negative bacteria: avoiding tetracycline phototoxicity during the growth of photosynthetic bacteria; Mather MW et al.; A mobilizable, broad-host-range (bhr) plasmid was derived from the widely used IncP1 vector pRK415 . The new vector, pRKD418, contains an additional resistance gene and an enlarged multiple cloning site (MCS) region . The optimal growth of pRK415-containing bacteria under photosynthetic conditions generally requires the use of optical filters to protect the selective antibiotic tetracycline (Tc) from photooxidation with the resulting production of toxic photoproducts; pRK415 is not stably maintained in the absence of selective pressure . The addition of a trimethoprim-resistant dihydrofolate reductase-encoding gene provided for optimal photosynthetic growth in the presence of a selective antibiotic without any special apparatus . The presence of an antibiotic marker not found in commonly used cloning vectors in many cases facilitates the subcloning of inserts into the bhr plasmid . The new MCS region provides further cloning flexibility with at least sixteen available restriction sites . Easily constructed derivative plasmids, exemplified by pRKD418KmE, provide a convenient screening procedure for the detection of recombinants during subcloning. Pharmacoeconomics, 1995 May, 7(5), 393 - 402 Once-daily aminoglycoside administration in gram-negative sepsis . Economic and practical aspects; Parker SE et al.; A fuller understanding of the pharmacodynamics of aminoglycoside antibiotics now exists compared with when they were introduced . Recent findings have shown that once-daily dosage regimens of aminoglycosides are as effective as bd or tid regimens in the treatment of Gram-negative sepsis . However, radical changes in dosage frequency based on this knowledge are resisted by some physicians because of fears about the peak concentration toxicity of aminoglycosides . These fears have been shown to be misplaced . The delay in the translation of research findings into practice may be attributable to the sheer quantity of medical literature and the limited time that clinicians have available to read it . Because healthcare resources are finite, physicians are increasingly becoming aware of the need to use drug therapy in the most cost-effective way . An important component of aminoglycoside therapy that may persuade clinicians to change their practice is the organised consideration of the various costs associated with different administration regimens . This review examines the source of those costs, and endorses once-daily dosage of aminoglycosides from both an economic and practical viewpoint. Science, 1995 Apr 7, 268(5207), 80 - 3 E5531, a pure endotoxin antagonist of high potency; Christ WJ et al.; Shock due to Gram-negative bacterial sepsis is a consequence of acute inflammatory response to lipopolysaccharide (LPS) or endotoxin released from bacteria . LPS is a major constituent of the outer membrane of Gram-negative bacteria, and its terminal disaccharide phospholipid (lipid A) portion contains the key structural features responsible for toxic activity . Based on the proposed structure of nontoxic Rhodobacter capsulatus lipid A, a fully stabilized endotoxin antagonist E5531 has been synthesized . In vitro, E5531 demonstrated potent antagonism of LPS-me