Proc Natl Acad Sci U S A, 1999 Oct 26, 96(22), 12345 - 9 A protein residing at the subunit interface of the bacterial ribosome; Agafonov DE et al.; Surface labeling of Escherichia coli ribosomes with the use of the tritium bombardment technique has revealed a minor unidentified ribosome-bound protein (spot Y) that is hidden in the 70S ribosome and becomes highly labeled on dissociation of the 70S ribosome into subunits . In the present work, the N-terminal sequence of the protein Y was determined and its gene was identified as yfia, an ORF located upstream the phe operon of E . coli . This 12.7-kDa protein was isolated and characterized . An affinity of the purified protein Y for the 30S subunit, but not for the 50S ribosomal subunit, was shown . The protein proved to be exposed on the surface of the 30S subunit . The attachment of the 50S subunit resulted in hiding the protein Y, thus suggesting the protein location at the subunit interface in the 70S ribosome . The protein was shown to stabilize ribosomes against dissociation . The possible role of the protein Y as ribosome association factor in translation is discussed. J Leukoc Biol, 1999 Oct, 66(4), 542 - 8 The actions of bacterial DNA on murine macrophages; Sester DP et al.; Murine macrophages are able to distinguish bacterial from mammalian DNA . The response is mimicked by single-stranded oligonucleotides containing unmethylated CG dinucleotides ("CpG" motifs) in specific sequence contexts . The dose-response curve for activation is influenced by variation in the sequence flanking the core CpG motif . CpG or bacterial DNA activates several signaling pathways in common with bacterial lipopolysaccharide (LPS), leading to induction of cytokine genes such as tumor necrosis factor alpha . Pretreatment with LPS causes desensitization to subsequent activation by CpG DNA . Both stimuli also cause cell cycle arrest in macrophages proliferating in response to the macrophage growth factor colony-stimulating factor-1 (CSF-1), but prevent apoptosis caused by growth factor removal . In part, cell cycle arrest by CpG DNA and LPS may be linked to rapid down-modulation of the CSF-1 receptor from the cell surface, a response that occurs in an all-or-nothing manner . The response of macrophages to CpG DNA has aspects in common with the DNA damage response in other cell types, which may provide clues to the underlying mechanism. Immunobiology, 1999 Sep, 201(1), 120 - 32 The role of intestinal bacterial flora in the tuning of the T cell repertoire; Sawamura SA et al.; The role of the intestinal bacterial flora on the Vbeta repertoire was examined using the gnotobiotic murine model . The ratio of Vbeta6-positive T cells in the periphery of DBA/2 mice under SPF conditions was only 2.2% (mean, n = 4), since the cells were eliminated by the endogenous superantigen Mls(a) . However, the ratio in germ-free (GF) mice was 31.7% . Similarly, the contamination of the GF Mice with the intestinal flora from SPF mice reduced the ratio of Vbeta6 in GF mice from 22.9% to 13.7% . In contrast, in BALB/c mice (Mls(b)) in which Vbeta6 cells do not react with this endogenous superantigen, the ratio of Vbeta6 cells do not react with this endogenous superantigen, the ratio of Vbeta6 of SPF mice (15.4%, mean, n = 3) was found to be comparable to that of GF mice (15.6%, n = 3) . These data suggested that the absence of intestinal flora deteriorated a part of the Mls(a) determinant, which reacted with the Vbeta6 T cells and thereby eliminated them, thus resulting in an increase of these cells in GF mice . Moreover, the alloantigenicity of minor histocompatible alloantigen(s) (mHAg) in SPF mice, which was detected in H-2 identical MLR experiments and a murine graft-versus-host (GVH) model, was reduced in GF and decontaminated SPF mice, thus indicating that the intestinal flora upregulated the mHAg including a part of Mls determinant . These results therefore suggest that the intestinal flora plays a role in the upregulation of mHAg including a part of endogenous superantigen and the consequent tuning of the Vbeta repertoire. Eur J Pediatr Surg, 1999 Aug, 9(4), 224 - 7 Bacterial translocation in short-bowel syndrome in rats; Schimpl G et al.; Massive intestinal resection results in short-bowel syndrome (SBS) and is associated with an increased risk of infectious complications mainly caused by the egress of intestinal bacteria to distant organs, a process termed bacterial translocation (BT) . The purpose of this experimental study in rats was to investigate in different models of SBS the impact of the type of intestinal resection on bacterial growth in the residual small bowel and on the occurrence of BT . SBS was created in 30 rats either by jejunal resection (JR), by ileal resection (IR) or by ileal resection including the ileocecal valve (IR+ICV) . 10 animals underwent only a sham laparotomy (SL) and served as controls . Two weeks after the operative procedure, intestinal bacterial colonization and BT to the portal vein, vena cava, mesenteric lymph nodes, liver and spleen were determined . All resected animals showed a decreased weight gain and a significant bacterial overgrowth in the residual small bowel compared to the SL group . BT occurred after SL in 12%, after JR in 70%, after IR in 58%, and was significantly less frequent (35%) after IR+ICV, respectively . These experimental findings suggest that BT in SBS might be promoted by the intestinal bacterial overgrowth in the residual bowel, and the incidence of BT seems to be related to the presence or absence of the ileocecal valve. Eur J Pediatr Surg, 1999 Aug, 9(4), 220 - 3 Bacterial translocation is favored by the preservation of the ileocecal valve in experimental short bowel with total parenteral nutrition; Eizaguirre I et al.; Sepsis in short-bowel syndrome (SBS) is in part due to bacterial translocation (BT) . Parenteral nutrition (PN) is often necessary in SBS and promotes BT . The aim of this study was to asses the effect of the presence or absence of ileocecal valve (ICV) on BT in parenterally-fed rats with massive intestinal resection . Sixty-five adult Wistar rats underwent central venous cannulations and were randomly assigned to one of five groups receiving for ten days five treatment regimes: Sham (n = 17) standard rat chow + i.v . saline . PN (n = 17) fasting + PN . Res-Sham (n = 10) standard rat chow + i.v . saline + 80% gut resection . Res-PN (n = 11) fasting, PN + 80% gut resection . Res-ICV-PN (n = 10) fasting, PN + 80% gut resection including ICV . At the end of the experiment they were euthanized and mesenteric lymph nodes (MLN), spleen and peripheral and portal blood specimens were recovered and cultured . BT was found in 47% of PN animals, 91% of Res-PN rats, 100% of Res-Sham group and 60% of Res-ICV-PN animals, but not in Sham ones . 97% of BT+ animals had positive cultures in MLN and/or portal blood, whereas germs beyond liver were detected in 30% of Res-Sham, 37% of PN, 50% of Res-PN and 0% of Res-ICV-PN rats . The present study confirms that both massive intestinal resection and PN promote BT . In addition, it shows that animals deprived of ICV have lower incidence of BT in this setting than those with it and that the germs do not reach in them peripheral blood in the same proportions as in ICV-intact animals . These results suggest that the presence of an intact ICV favor BT in parenterally-fed rats with massive intestinal resection. J Biol Chem, 1999 Oct 29, 274(44), 31236 - 44 Highly specific recognition of primer RNA structures for 2'-OH priming reaction by bacterial reverse transcriptases; Inouye S et al.; A minor population of Escherichia coli contains retro-elements called retrons, which encode reverse transcriptases (RT) to synthesize peculiar satellite DNAs called multicopy single-stranded DNA (msDNA) . These RTs recognize specific RNA structures in their individual primer-template RNAs to initiate cDNA synthesis from the 2'-OH group of a specific internal G residue (branching G residue) . The resulting products (msDNA) consist of RNA and single-stranded DNA, sharing hardly any sequence homology . Here, we investigated how RT-Ec86 recognizes the specific RNA structure in its primer-template RNA . On the basis of structural comparison with HIV-1 RT, domain exchanges were carried out between two E . coli RTs, RT-Ec86 and RT-Ec73 . RT-Ec86 (320 residues) and RT-Ec73 (316 residues) share only 71 identical residues (22%) . From the analysis of 10 such constructs, the C-terminal 91-residue sequence of RT-Ec86 was found to be essential for the recognition of the unique stem-loop structure and the branching G residue in the primer-template RNA for retron-Ec86 . Using the SELEX (systematic evolution of ligands by exponential enrichment) method with RT-Ec86 and primer RNAs containing random sequences, the identical stem-loop structure (including the 3-U loop) to that found in the retron-Ec86 primer-template RNA was enriched . In addition, the highly conserved 4-base sequence (UAGC), including the branching G residue, was also enriched . These results indicate that the highly diverse C-terminal region recognizes specific stem-loop structures and the branching G residue located upstream of the stem-loop structure . The present results with seemingly primitive RNA-dependent DNA polymerases provide insight into the mechanisms for specific protein RNA recognition. Comp Biochem Physiol C Pharmacol Toxicol Endocrinol, 1999 Jul, 123(3), 217 - 23 Eicosanoids mediate nodulation reactions to bacterial infections in larvae of the butterfly, Colias eurytheme; Stanley DW et al.; Nodulation is the first, and qualitatively predominant, cellular defense reaction to bacterial infections in insects . Treating larvae of the butterfly Colias eurytheme with the eicosanoid biosynthesis inhibitor dexamethasone, strongly impaired nodulation reactions to bacterial infections . The influence of dexamethasone was reversed by treating infected insects with arachidonic acid, an eicosanoid precursor . An eicosanoid biosynthesis system in C . eurytheme larvae is documented . Specifically, the presence of eicosanoid-precursor polyunsaturated fatty acids in tissue phospholipids was determined, an intracellular phospholipase A2 that can release arachidonic acid from tissue phospholipids was recorded, and eicosanoid biosynthesis, registered as conversion of exogenous radioactive 20:4n-6 into eicosanoids, was observed . These findings support the hypothesis that eicosanoids mediate cellular immune responses to bacterial infections in these butterfly larvae, and more broadly, in most, if not all, insects. Clin Infect Dis, 1999 Sep, 29(3), 536 - 43 Impact of bacterial pneumonia and Pneumocystis carinii pneumonia on human immunodeficiency virus disease progression . Pulmonary Complications of HIV Study Group; Osmond DH et al.; The course of pneumonia caused by pyogenic bacteria and Pneumocystis carinii was examined in a multicity cohort study of HIV infection . The median duration of survival among 150 individuals following initial bacterial pneumonia was 24 months, compared with 37 months among 299 human immunodeficiency virus (HIV)-infected control subjects matched by study site and CD4 lymphocyte count (P<.001) . For 152 subjects with P . carinii pneumonia, median survival was 23 months, compared with 30 months for 280 matched control subjects (P = .002) . Median durations of survival associated with the two types of pneumonia differed by only 47 days, despite a higher median CD4 lymphocyte count associated with bacterial pneumonia . These results suggest that both P . carinii pneumonia and bacterial pneumonia are associated with a significantly worse subsequent HIV disease course . The similarity of prognosis after one episode of bacterial pneumonia vs . an AIDS-defining opportunistic infection and the proportion of cases occurring in association with a CD4 lymphocyte count of >200 suggest that measures to prevent bacterial pneumonia should be emphasized. Biochemistry, 1999 Oct 19, 38(42), 14077 - 87 Kinetics and interactions of molybdenum and iron-sulfur centers in bacterial enzymes of the xanthine oxidase family: mechanistic implications; Canne C et al.; For isoquinoline 1-oxidoreductase (IsoOr), the reaction mechanism under turnover conditions was studied by EPR spectroscopy using rapid-freeze methods . IsoOr displays several EPR-active Mo(V) species including the "very rapid" component found also in xanthine oxidase (XanOx) . For IsoOr, unlike XanOx or quinoline 2-oxidoreductase (QuinOr), this species is stable for about 1 h in the absence of an oxidizing substrate {Canne, C., Stephan, I., Finsterbusch, J., Lingens, F., Kappl, R., Fetzner, S., and Huttermann, J . (1997) Biochemistry 36, 9780-9790} . Under rapid-freeze conditions in the presence of ferricyanide the very rapid species behaves as a kinetically competent intermediate present only during steady-state turnover . To explain the persistence of the very rapid species in IsoOr in the absence of an added oxidant, extremely slow product dissociation is required . This new finding that oxidative conditions facilitate decay of the very rapid signal for IsoOr supports the mechanism of substrate turnover proposed by Lowe, Richards, and Bray {Lowe, D . J., Richards, R . L., and Bray, R . C . (1997) Biochem . Soc . Trans . 25, 774-778} . Additional stopped-flow data reveal that alternative catalytic cycles occur in IsoOr and show that the product dissociates after transfer of a single oxidizing equivalent from ferricyanide . In rapid-freeze measurements magnetic interactions of the very rapid Mo(V) species and the iron-sulfur center FeSI of IsoOr and QuinOr were observed, proving that FeSI is located close to the molybdopterin cofactor in the two proteins . This finding is used to relate the two different iron-sulfur centers of the aldehyde oxidoreductase structure with the EPR-detectable FeS species of the enzymes. Biochemistry, 1999 Oct 19, 38(42), 14036 - 44 The FinO repressor of bacterial conjugation contains two RNA binding regions; Ghetu AF et al.; Conjugative transfer of F-like plasmids in Escherichia coli is repressed by a plasmid-encoded protein, FinO . FinO blocks the translation of TraJ, a positive activator of transcription of genes required for conjugation . FinO binds a traJ antisense RNA, FinP, thereby protecting it from degradation, and catalyzes FinP-traJ mRNA hybridization . Interactions between these two RNAs are predicted to block the traJ ribosomal binding site . In this paper, we use limited proteolysis, circular dichroism spectroscopy, and an electrophoretic mobility shift assay to map the regions within FinO that are required for interactions with RNA . Our results show that FinO is largely helical, binds to its highest affinity binding site within FinP as a monomer, and contains two distinct RNA binding regions, one of which is localized between residues 26 and 61, and a second which is localized between residues 62 and 186. Biochemistry, 1999 Oct 19, 38(42), 13780 - 6 A low-redox potential heme in the dinuclear center of bacterial nitric oxide reductase: implications for the evolution of energy-conserving heme-copper oxidases; Gronberg KL et al.; Bacterial nitric oxide reductase (NOR) catalyzes the two-electron reduction of nitric oxide to nitrous oxide . It is a highly diverged member of the superfamily of heme-copper oxidases . The main feature by which NOR is distinguished from the heme-copper oxidases is the elemental composition of the active site, a dinuclear center comprised of heme b(3) and non-heme iron (Fe(B)) . The visible region electronic absorption spectrum of reduced NOR exhibits a maximum at 551 nm with a distinct shoulder at 560 nm; these are attributed to Fe(II) heme c (E(m) = 310 mV) and Fe(II) heme b (E(m) = 345 mV), respectively . The electronic absorption spectrum of oxidized NOR exhibits a characteristic shoulder around 595 nm that exhibits complex behavior in equilibrium redox titrations . The first phase of reduction is characterized by an apparent shift of the shoulder to 604 nm and a decrease in intensity . This is due to reduction of Fe(B) (E(m) = 320 mV), while the subsequent bleaching of the 604 nm band represents reduction of heme b(3) (E(m) = 60 mV) . This separation of redox potentials (>200 mV) allows the enzyme to be poised in the three-electron reduced state for detailed spectroscopic examination of the Fe(III) heme b(3) center . The low midpoint potential of heme b(3) represents a thermodynamic barrier to the complete (two-electron) reduction of the dinuclear center . This may avoid formation of a stable Fe(II) heme b(3)-NO species during turnover, which may be an inhibited state of the enzyme . It would also appear that the evolution of significant oxygen reducing activity by heme-copper oxidases was not simply a matter of the substitution of copper for non-heme iron in the dinuclear center . Changes in the protein environment that modulate the midpoint redox potential of heme b(3) to facilitate both complete reduction of the dinuclear center (a prerequisite for oxygen binding) and rapid heme-heme electron transfer were also necessary. Plant Mol Biol, 1999 Aug, 40(6), 977 - 83 Construction and characterization of a common bean bacterial artificial chromosome library; Vanhouten W et al.; We have constructed a common bean (Phaseolus vulgaris L.) bacterial artificial chromosome (BAC) library consisting of 33,792 clones and an estimated 3- to 5-fold coverage of the common bean genome . Leaf nuclei were used as the source for high-molecular-weight DNA, and an endonuclease/methylase competition assay was employed to partially cleave the DNA . The library was screened with a number of nuclear and mitochondrial probes . Each nuclear probe identified at least two BACs with an average insert size of ca . 100 kb . Only 26 clones were identified after hybridizing with mitochondrial probes, indicating contamination with organellar sequences is low . Numerous clones could be identified after screening the library with two repetitive probes flanking the nuclear fertility restorer Fr . Intriguingly, 12 clones appeared to hybridize to both markers, and restriction analysis of these clones revealed that they can be assembled into maximally four contigs, suggesting that these repetitive probes may be useful for the physical mapping of the Fr locus. Biochim Biophys Acta, 1999 Oct 18, 1441(1), 51 - 60 cDNA cloning and bacterial expression of phospholipase A(2) inhibitor PLIalpha from the serum of the Chinese mamushi, Agkistrodon blomhoffii siniticus(1); Okumura K et al.; The cDNA encoding of a phospholipase A(2) inhibitor (PLIalpha) of the Chinese mamushi, Agkistrodon blomhoffii siniticus, was identified from a liver cDNA library by use of a probe prepared by polymerase chain reaction (PCR) on the basis of the amino acid sequence of PLIalpha . It encoded a polypeptide of 166 amino acid residues, including 19 residues of the signal sequence and 147 residues of the complete mature sequence of PLIalpha . The PLIalpha cDNA was subcloned into the expression vector pET-16b and used to transform Escherichia coli strain BL21(DE3)pLysS . The recombinant PLIalpha expressed as a fusion protein was solubilized and purified to homogeneity by use of a metal affinity resin . The purified PLIalpha fusion protein underwent folding to form a trimeric structure like the intact PLIalpha, and showed inhibitory activity against the group II acidic PLA(2) from A . blomhoffii siniticus venom; although its binding constant (1/K(i)) value was 30-fold lower than that of the natural PLIalpha . The elimination of the N-terminal additional peptide from the fusion protein resulted in a marked increase in the inhibition activity with a binding constant comparable to that of the natural PLIalpha against the acidic PLA(2) . Furthermore, the carbohydrate chains of the natural PLIalpha were found to play an important role in the inhibitory activity against the basic PLA(2). Tex Heart Inst J, 1999, 26(3), 192 - 4 Technical aspects of mitral valve replacement with an allograft for acute bacterial endocarditis; Conklin LD et al.; Mitral valve replacement with a mitral valve allograft is receiving a resurgence of interest . We discuss the technical aspects of this procedure as it applies to cases of acute bacterial endocarditis infecting the mitral valve. Genome Res, 1999 Oct, 9(10), 989 - 93 A resource of mapped human bacterial artificial chromosome clones; Cheung VG et al.; To date, despite the increasing number of genomic tools, there is no repository of ordered human BAC clones that covers entire chromosomes . This project presents a resource of mapped large DNA fragments that span eight human chromosomes at approximately 1-Mb resolution . These DNA fragments are bacterial artificial chromosome (BAC) clones anchored to sequence tagged site (STS) markers . This clone collection, which currently contains 759 mapped clones, is useful in a wide range of applications from microarray-based gene mapping to identification of chromosomal mutations . In addition to the clones themselves, we describe a database, GenMapDB , that contains information about each clone in our collection. Hepatogastroenterology, 1999 Jul-Aug, 46(28), 2159 - 64 Obstructive jaundice promotes bacterial translocation in humans; Kuzu MA et al.; BACKGROUND/AIMS: Significant bacterial translocation was demonstrated following experimental biliary obstruction, however very little is known about the importance and the prevalence of gut-origin sepsis in obstructive jaundice patients . Therefore, the aim of this study was to investigate the concept of gut-origin sepsis in obstructive jaundiced patients and its clinical importance . METHODOLOGY: Twenty-one patients requiring laparotomy for obstructive jaundice (group I) and thirty patients operated on electively mainly for chronic cholecystitis (group II) were studied . Peritoneal swab, mesenteric lymph node, portal venous blood, liver wedge biopsy and bile were sampled for culture immediately after opening the peritoneum . Additionally, peripheral blood samples were taken pre- and post-operatively from all patients . Post-operatively, patients were monitored for infectious complications . RESULTS: The mean serum bilirubin concentration, gamma glutamyl transferase and alkaline phosphatase levels in jaundiced patients before therapeutic intervention were significantly higher than in control patients . Five patients demonstrated bacterial translocation in group I (24%), whereas only one did so in group II (3.5%, p < 0.05) . Septic complications were detected in three patients, but only in two with bacterial translocation in group I . There was one patient with bacterial translocation who had septic complication in group II . CONCLUSIONS: The present study demonstrated that obstructive jaundice significantly promotes bacterial translocation in humans, however, its clinical importance has yet to be defined. Fertil Steril, 1999 Oct, 72(4), 730 - 2 Bacterial vaginosis and past chlamydial infection are strongly and independently associated with tubal infertility but do not affect in vitro fertilization success rates; Gaudoin M et al.; OBJECTIVE: To determine the incidence of active vaginal infection in women undergoing IVF, relate it to the cause of infertility, and investigate a relation with the outcome of fresh ET . DESIGN: Cross-sectional study . SETTING: Tertiary care infertility referral center . PATIENT(S): Two hundred eighty-six women who underwent 344 oocyte recovery procedures for IVF cycles between March 1997 and January 1998 . INTERVENTION(S): High vaginal swab specimens and endocervical swab specimens were obtained and ELISA serology was performed for detection of Chlamydia species on samples taken immediately before oocyte recovery . The results were related to the cause of infertility and the outcome parameters . MAIN OUTCOME MEASURE(S): Pregnancy rates . RESULT(S): Seropositivity for Chlamydia species and the presence of bacterial vaginosis both were strongly and independently associated with tubal disease . There was no difference in pregnancy rates in any of the groups regardless of their serologic status for chlamydial infection or current infection with bacterial vaginosis . CONCLUSION(S): This study provides further evidence of the pelvic pathogenicity of bacterial vaginosis . However, it shows that women who have bacterial vaginosis or who have been treated for chlamydial infection in the past achieve pregnancy rates with IVF treatment similar to those of women who have no evidence of such infections. Anaesth Intensive Care, 1999 Oct, 27(5), 493 - 6 Bacterial contamination of propofol in the operating theatre; Soong WA; There have been several reports of propofol becoming extrinsically contaminated with bacteria . These reports have usually related to infusions or delays in administration after the ampoule has been opened . This observational study was performed to examine bacterial contamination of propofol during usual practice in the operating theatres of a single large hospital group . One hundred samples of propofol were collected and cultured . Samples were taken immediately after administration in cases where the delay between opening the ampoule and administration was at least 15 minutes . The samples were classified according to whether the propofol was kept in the ampoule or a syringe after opening the ampoule and whether the intended use was for a single patient or multiple patients . The time between opening the ampoule and administration was recorded . There were three positive bacterial cultures . These samples all came from ampoules used for more than one patient, without the later dose (does) being drawn into a syringe at the time the ampoule was opened . This common clinical practice, especially in paediatric anaesthesia, does not comply with the manufacturer's recommendations . The clinical significance of the bacterial contamination detected is not clear . It is recommended that propofol should be handled in an aseptic fashion and measures taken to minimize the risk of bacterial contamination. Scand J Immunol, 1999 Oct, 50(4), 387 - 93 Participation of CD1 molecules in the presentation of bacterial protein antigens in humans; Ulanova M et al.; Human CD1 molecules, expressed on the surface of professional antigen-presenting cells (including dendritic cells, Langerhans' cells, B cells and activated monocytes) are structurally homologous to major histocompatibility complex (MHC) class I and class II molecules . CD1b and CD1c have been shown to present nonpeptide bacterial antigens to T cells . We hypothesized that CD1 molecules may also be involved in the presentation of bacterial protein antigens . Human peripheral blood mononuclear cells (PBMC) were exposed to two medically important proteins, tetanus toxoid (TT) and purified protein derivative (PPD), with and without murine monoclonal antibodies (MoAbs) specific for CD1a, CD1b and CD1c . All the MoAbs substantially inhibited the proliferative responses of PBMC to TT and PPD . Simultaneous interaction of CD1 and MHC class II molecules was even more inhibitory to these antigen-specific proliferative responses . In contrast, neither mixed lymphocyte reaction nor superantigen and mitogenic responses were affected by CD1-specific antibodies, indicating a certain restriction pattern in antigen presentation . Our findings suggest that, besides MHC class I and II molecules, there is a family of nonpolymorphic cell surface molecules that is able to present certain bacterial protein antigens to T cells. J Heart Valve Dis, 1999 Sep, 8(5), 591 - 2 Fatal bacterial endocarditis following aortic valve replacement in a patient being treated with methotrexate; Wilkinson NM; A 41-year-old man being treated with methotrexate for psoriasis underwent aortic valve replacement . He subsequently developed fulminating bacterial endocarditis . Bacterial endocarditis occurs in 1-2% of cases after prosthetic valve replacement and has a high mortality . The long-term use of methotrexate and similar drugs is increasing in conditions such as psoriasis, rheumatoid arthritis and inflammatory bowel disease . Thus, more patients undergoing heart valve surgery will be taking these preparations for coexisting disease . As methotrexate increases the risk of infection, its perioperative use in these patients requires further evaluation. Hum Gene Ther, 1999 Sep 20, 10(14), 2373 - 9 In vivo transfer of bacterial marker genes results in differing levels of gene expression and tumor progression in immunocompetent and immunodeficient mice; Lukacs KV et al.; To optimize gene delivery for the treatment of malignant mesothelioma, expression of the beta-galactosidase marker gene was examined in a murine model of intraperitoneal malignant mesothelioma . The beta-galactosidase gene was delivered to the peritoneal cavity of tumor-bearing mice by various plasmid-liposome complexes or by replication-incompetent retrovirus, used alone or complexed to liposomes . In tumor samples from immunodeficient nude mice, moderate levels of gene expression were achieved by liposome-complexed plasmids . Retroviral gene delivery was more effective, and was increased nearly 10-fold by complexing the retrovirus to liposomes . In contrast, in tumor samples from immunocompetent CBA mice treated with the same vectors, no marker gene expression was detected . In immunodeficient mice, tumor growth was not affected by beta-galactosidase gene transfer . However, immunocompetent mice showed a significant decrease in tumor size and increase in survival time after beta-galactosidase delivery . Induction of cytotoxic T cells capable of lysing beta-Gal-transfected tumor cells suggests that tumor cells transduced with the bacterial beta-galactosidase gene may be eliminated in immunocompetent hosts . Our findings also indicate that plasmid-liposome complexes, which achieve a low level of gene expression, and retrovirus-liposome complexes, which result in nearly 100 times higher levels of gene expression in tumor cells in vivo, are similarly effective in inducing an antitumor immune response. Biochem Biophys Res Commun, 1999 Oct 5, 263(3), 820 - 4 Spectral properties of Trp182, Trp194, and Trp250 on the alpha subunit of bacterial luciferase; Li Z et al.; The phosphorescence and fluorescence properties of bacterial luciferase (alphabeta) mutants from Xenorhabdus luminescens were investigated . All tryptophans in the alpha and beta subunits were replaced with tyrosines except for one or two tryptophans in the alpha subunit . Because one luciferase mutant (W250) retained only a single tryptophan in the alpha subunit while two other mutants (W182/250 and W194/250) each contained two tryptophans in the alpha subunit, it was possible to deduce the spectral properties of these specific tryptophans (Trp182, Trp194, Trp250) . Analyses of the phosphorescence properties were particularly revealing as only a single phosphorescence emission peak at 411-414 nm was observed for the W250 and W194/250 mutants while peaks at 409 and 414 nm could be clearly observed for the W182/250 mutant . Coupled with intrinsic fluorescence quenching experiments, these results show that alphaTrp182 is in a distinctly polar environment while alphaTrp250 is in a hydrophobic region and illustrate the advantages of using phosphorescence to recognize different microenvironments for tryptophan residues . J Mol Biol, 1999 Oct 8, 292(5), 1003 - 16 High-level expression in Escherichia coli of selenocysteine-containing rat thioredoxin reductase utilizing gene fusions with engineered bacterial-type SECIS elements and co-expression with the selA, selB and selC genes; Arner ES et al.; Mammalian thioredoxin reductase (TrxR) catalyzes reduction of thioredoxin and many other substrates, and is a central enzyme for cell proliferation and thiol redox control . The enzyme is a selenoprotein and can therefore, like all other mammalian selenoproteins, not be directly expressed in Escherichia coli, since selenocysteine-containing proteins are synthesized by a highly species-specific translation machinery . This machinery involves a secondary structure, SECIS element, in the selenoprotein-encoding mRNA, directing selenocysteine insertion at the position of an opal (UGA) codon, normally conferring termination of translation . It is species-specific structural features and positions in the selenoprotein mRNA of the SECIS elements that hitherto have hampered heterologous production of recombinant selenoproteins . We have discovered, however, that rat TrxR can be expressed in E . coli by fusing its open reading frame with the SECIS element of the bacterial selenoprotein formate dehydrogenase H . A variant of the SECIS element designed to encode the conserved carboxyterminal end of the enzyme (-Sec-Gly-COOH) and positioning parts of the SECIS element in the 3'-untranslated region was also functional . This finding revealed that the SECIS element in bacteria does not need to be translated for full function and it enabled expression of enzymatically active mammalian TrxR . The recombinant selenocysteine-containing TrxR was produced at dramatically higher levels than formate dehydrogenase O, the only endogenous selenoprotein expressed in E . coli under the conditions utilized, demonstrating a surprisingly high reserve capacity of the bacterial selenoprotein synthesis machinery under aerobic conditions . Co-expression with the selA, selB and selC genes (encoding selenocysteine synthase, SELB and tRNA(Sec), respectively) further increased the efficiency of the selenoprotein production and thereby also increased the specific activity of the recombinant TrxR to about 25 % of the native enzyme, with as much as 20 mg produced per liter of culture . These results show that with the strategy utilized here, the capacity of selenoprotein synthesis in E . coli is more than sufficient for making possible the use of the bacteria for production of recombinant selenoproteins . J Arthroplasty, 1999 Sep, 14(6), 677 - 81 Bacterial contamination rates during bone allograft retrieval; Journeaux SF et al.; A retrospective study of allograft bone retrieved from 401 donors between January 1987 and March 1996 was performed to determine the incidence of bacterial contamination . Contamination according to type of donor (live, multiorgan, cadaveric) was also determined . Live donors donating a femoral head demonstrated a contamination rate of 13%; multiorgan donors, 24%; and cadaveric donors, 35% . Donor contamination by type of bone (hemipelvis, femur, tibia) showed no significant difference in the multiorgan donors . In cadaveric donors, there was a significant increase in contamination of the hemipelves as compared to the femur and tibias . Recommendations for contamination control in allograft retrieval are given . Our findings are of great significance for musculoskeletal banks that do not secondarily irradiate and rely on screening of allograft bone for contamination alone. Curr Opin Chem Biol, 1999 Oct, 3(5), 607 - 13 Structure/function studies on enzymes in the diaminopimelate pathway of bacterial cell wall biosynthesis; Born TL et al.; Within the past 18 months work has continued on the structure and mechanisms of enzymes involved in the diaminopimelic acid/lysine biosynthetic pathway . A novel structure has been determined for a PLP-independent epimerase, and structures with bound substrates have been solved for two other enzymes . Additionally, new studies have appeared describing the chemical mechanisms of three enzymes in the pathway. Transfusion, 1999 Aug, 39(8), 808 - 17 The cost-effectiveness of autologous transfusion revisited: implications of an increased risk of bacterial infection with allogeneic transfusion; Sonnenberg FA et al.; BACKGROUND: Previous analyses have found autologous transfusion to be very expensive but have not considered avoidance of postoperative bacterial infections as one of its benefits . STUDY DESIGN AND METHODS: A cost-utility analysis using a Markov cohort simulation model compared autologous blood transfusion to allogeneic transfusion in a hypothetical cohort of patients undergoing elective total hip replacement with respect to discounted quality-adjusted life years (QALYs) and health-care system costs . RESULTS: Assuming a base case rate of serious infection of 3.7 percent, a relative risk of infection of 1.85, and additional costs of $12,980 per infection, autologous transfusion has a cost-effectiveness of $2,470 per QALY . If the relative risk of bacterial infection following allogeneic transfusion exceeds 1.1, the cost-effectiveness of autologous transfusion is less than $50,000 per QALY and if the relative risk exceeds 2.4, autologous transfusion is dominant, resulting in both lower costs and greater QALYs . If there were no increased risk of transfusion, the cost-effectiveness of autologous transfusion would be $3,400,000 per QALY . CONCLUSIONS: If there is only a modest increase in the risk of bacterial infection following allogeneic transfusion, autologous transfusion would result in improved outcomes at a cost of less than $50,000 per QALY . Autologous transfusion would be dominant above a relative risk of infection that is within the range of values observed in randomized controlled trials . However, if there is no increased risk of bacterial infection, autologous transfusion would be a very expensive strategy . Until more definitive data are available on the magnitude and costs of this risk, we advise against prematurely closing the debate about the cost-effectiveness of autologous transfusion. Int J Parasitol, 1999 Jul, 29(7), 1053 - 63 Hypergastrinaemia, abomasal bacterial population densities and pH in sheep infected with Ostertagia circumcincta; Simcock DC et al.; Serum gastrin and pepsinogen concentrations, food intake, abomasal pH and abomasal aerotolerant and anaerobic bacterial populations were measured in sheep infected with Ostertagia circumcincta to search for links between hypergastrinaemia, food intake and changes in the abomasal environment . Abomasal pH and serum gastrin and pepsinogen concentrations were elevated in each of five sheep infected via abomasal cannulae with 150000 exsheathed larval stage three, followed 11 days later by 100000 sheathed larvae given intraruminally . Unparasitised abomasa contained aerotolerant bacterial population densities of between 10(3) and 10(6) cells ml(-1) and these did not change significantly following parasitism . In contrast, anaerobic bacterial population densities increased markedly by about 10(4)-fold following parasitism . Anaerobic numbers changed rapidly when abomasal pH increased from 2.5 to 3.5 . At pH 4 and above, anaerobic bacterial numbers approached levels expected in rumen contents but parameters other than pH did not relate to bacterial numbers . Brief periods when serum gastrin was lower than expected, coinciding with raised abomasal pH, were not explicable by increased bacterial numbers . Food intake, which decreased for a variable period from around Day 5 p.i., correlated poorly with serum gastrin concentration, suggesting hypergastrinaemia is not the sole cause of anorexia in parasitised animals . The survival of substantial numbers of rumen bacteria in the abomasum at only slightly raised pH may significantly lower the bacterial protein available to the sheep. Adv Microb Physiol, 1999, 41, 291 - 337 The bacterial flagella motor; Berry RM et al.; The bacterial flagellum is probably the most complex organelle found in bacteria . Although the ribosome may be made of slightly more subunits, the bacterial flagellum is a more organized and complex structure . The limited number of flagella must be targeted to the correct place on the cell membrane and a structure with cytoplasmic, cytoplasmic membrane, outer membrane and extracellular components must be assembled . The process of controlled transcription and assembly is still not fully understood . Once assembled, the motor complex in the cytoplasmic membrane rotates, driven by the transmembrane ion gradient, at speeds that can reach many 100 Hz, driving the bacterial cell at several body lengths a second . This coupling of an electrochemical gradient to mechanical rotational work is another fascinating feature of the bacterial motor . A significant percentage of a bacterium's energy may be used in synthesizing the complex structure of the flagellum and driving its rotation . Although patterns of swimming may be random in uniform environments, in the natural environment, where cells are confronted with gradients of metabolites and toxins, motility is used to move bacteria towards their optimum environment for growth and survival . A sensory system therefore controls the switching frequency of the rotating flagellum . This review deals primarily with the structure and operation of the bacterial flagellum . There has been a great deal of research in this area over the past 20 years and only some of this has been included . We apologize in advance if certain areas are covered rather thinly, but hope that interested readers will look at the excellent detailed reviews on those areas cited at those points. Adv Microb Physiol, 1999, 41, 93 - 137 Bacterial viability and culturability; Barer MR et al.; Renewed interest in the relationships between viability and culturability in bacteria stems from three sources: (1) the recognition that there are many bacteria in the biosphere that have never been propagated or characterized in laboratory culture; (2) the proposal that some readily culturable bacteria may respond to certain stimuli by entering a temporarily non-culturable state termed 'viable but non-culturable' (VBNC) by some authors; and (3) the development of new techniques that facilitate demonstration of activity, integrity and composition of non-culturable bacterial cells . We review the background to these areas of interest emphasizing the view that, in an operational context, the term VBNC is self-contradictory (Kell et al., 1998) and the likely distinctions between temporarily non-culturable bacteria and those that have never been cultured . We consider developments in our knowledge of physiological processes in bacteria that may influence the outcome of a culturability test (injury and recovery, ageing, adaptation and differentiation, substrate-accelerated death and other forms of metabolic self-destruction, prophages, toxin-antitoxin systems and cell-to-cell communication) . Finally, we discuss whether it is appropriate to consider the viability of individual bacteria or whether, in some circumstances, it may be more appropriate to consider viability as a property of a community of bacteria. Proc Natl Acad Sci U S A, 1999 Sep 28, 96(20), 11346 - 51 Response tuning in bacterial chemotaxis; Jasuja R et al.; Chemotaxis of enteric bacteria in spatial gradients toward a source of chemoattractant is accomplished by increases in the length of swimming runs up the gradient . Biochemical components of the intracellular signal pathway have been identified, but mechanisms for achieving the high response sensitivity remain unknown . Binding of attractant ligand to its receptor inactivates a receptor-associated histidine kinase, CheA, which phosphorylates the signal protein CheY . The reduction in phospho-CheY, CheY-P, levels prolongs swimming runs . Here, the stimulus-response relation has been determined by measurement of excitation responses mediated by the Tar receptor to defined concentration jumps of the attractant, aspartate, administered within milliseconds by photolysis of a photolabile precursor . The bacteria responded to <1% changes in Tar occupancy when adapted to aspartate over concentrations spanning three orders of magnitude . Response amplitudes increased approximately logarithmically with stimulus strength, extending responsiveness over a greater stimulus range . The extent and form of this relation indicates that, in contrast to mechanisms for adaptive recovery, excitation signal generation involves amplification based on cooperative interactions . These interactions could entail inactivation of multiple receptor-CheA signaling complexes and/or simultaneous activation of CheY-P dephosphorylation. Trends Microbiol, 1999 Oct, 7(10), 420 - 4 Bacterial mechanosensitive channels: integrating physiology, structure and function; Blount P et al.; When confronted with hypo-osmotic stress, many bacterial species are able rapidly to adapt to the increase in cell turgor pressure by jettisoning cytoplasmic solutes into the medium through membrane-tension-gated channels . Physiological studies have confirmed the importance of these channels in osmoregulation . Mutagenesis of one of these channels, combined with structural information derived from X-ray crystallography, has given the first clues of how a mechanosensitive channel senses and responds to membrane tension. Burns, 1999 Sep, 25(6), 515 - 7 Bacterial endocarditis following a minor burn injury . Case report and review; Paterson P et al.; A case of bacterial endocarditis in a patient with a total burn area of less than 1% is presented . A high index of suspicion for bacterial endocarditis should exist for any burns patient, regardless of burn size, who becomes unwell and has positive blood cultures. Infect Immun, 1999 Oct, 67(10), 5275 - 81 Differential regulation of macrophage interleukin-1 (IL-1), IL-12, and CD80-CD86 by two bacterial toxins; Foss DL et al.; The ability of innate immune cells to differentially respond to various bacterial components provides a mechanism by which the acquired immune response may be tailored to specific pathogens . The response of innate immune cells to bacterial components provides regulatory signals to cognate immune cells . These signals include secreted cytokines and costimulatory molecules, and to a large extent they determine the quantitative and qualitative nature of the immune response . In order to determine if innate immune cells can differentially respond to bacterial components, we compared the responses of macrophages to two bacterially derived molecules, cholera toxin (CT) and lipopolysaccharide (LPS) . We found that CT and LPS differentially regulated the expression of interleukin-12 (IL-12) and CD80-CD86 but not that of IL-1beta . LPS and CT each induced IL-1beta expression in macrophages, while only LPS induced IL-12 and only CT induced CD80-CD86 . These differences were markedly potentiated in gamma interferon (IFN-gamma)-treated macrophages, in which LPS potently induced IL-12 and CD80-CD86 expression . In contrast, IFN-gamma treatment had no effect on the expression of IL-1beta . These results define a molecular basis for the differential pathogenicities of bacterial toxins and are relevant to the design of vaccine adjuvants able to selectively induce desired types of immunity. J Leukoc Biol, 1999 Sep, 66(3), 528 - 34 Mechanisms of regulation of the MacMARCKS gene in macrophages by bacterial lipopolysaccharide; Chang S et al.; Bacterial lipopolysaccharide (LPS) stably induced the protein kinase C substrate, MacMARCKS, in murine resident peritoneal macrophages; initial induction of MacMARCKS mRNA was detected within 15 min and was protein synthesis-independent . This response was observed in the macrophage cell line RAW264, and occurred also in response to plasmid DNA, a partial mimetic of other responses to LPS . In murine bone marrow-derived macrophages, MacMARCKS was expressed constitutively due to induction by macrophage colony-stimulating factor . Nuclear run-on transcription revealed that, like tumor necrosis factor alpha (TNF-alpha), MacMARCKS was transcribed constitutively in RAW264 cells . The MacMARCKS promoter was sequenced to -1.7 kb and the transcription start site determined . Transient transfections of RAW264 cells revealed that the 113-bp GC-rich proximal promoter contained all the elements required for both high basal activity and 15- to 20-fold activation by LPS. Biochem J, 1999 Oct 1, 343 Pt 1, 177 - 83 Bacterial lipolytic enzymes: classification and properties; Arpigny JL et al.; Knowledge of bacterial lipolytic enzymes is increasing at a rapid and exciting rate . To obtain an overview of this industrially very important class of enzymes and their characteristics, we have collected and classified the information available from protein and nucleotide databases . Here we propose an updated and extensive classification of bacterial esterases and lipases based mainly on a comparison of their amino acid sequences and some fundamental biological properties . These new insights result in the identification of eight different families with the largest being further divided into six subfamilies . Moreover, the classification enables us to predict (1) important structural features such as residues forming the catalytic site or the presence of disulphide bonds, (2) types of secretion mechanism and requirement for lipase-specific foldases, and (3) the potential relationship to other enzyme families . This work will therefore contribute to a faster identification and to an easier characterization of novel bacterial lipolytic enzymes. Biochem Biophys Res Commun, 1999 Sep 24, 263(2), 570 - 4 Bacterial lipopolysaccharide increases prostaglandin production by rat astrocytes via inducible cyclo-oxygenase: evidence for the involvement of nuclear factor kappaB; Pistritto G et al.; This study was set to investigate the mechanisms through which bacterial lipopolysaccharide (LPS) stimulates prostaglandin (PG) production in rat astrocytes . Primary cultures of rat hypothalamic astrocytes were established . Cells were treated with LPS alone or LPS plus antagonists of various pathways, and the subsequent changes in cyclo-oxygenase (COX) activity were monitored by measuring a COX end product, PGE2, released into the incubation medium . It was found that (i) LPS produced a concentration-dependent increase in PGE2 release from astrocytes . The potency of LPS was significantly increased by the addition of serum into the incubation medium; (ii) after 24 h of incubation, inducible COX (COX-2) accounts for most of the LPS-stimulated PG production, as the latter was markedly reduced by dexamethasone and the specific COX-2 inhibitor NS 398; and (iii) nuclear factor kappaB appears to play a role in the activation of COX-2 induced by LPS, since certain inhibitors of this transcription factor were able to antagonize, at least in part, the effects of LPS on PGE2 release . C R Acad Sci III, 1999 Jul, 322(7), 551 - 6 {Regulation of bacterial proteolytic activity at natural concentrations in dissolved organic matter in a maritime pond}; Crottereau C et al.; The regulation of the bacterial exoproteolytic activity, at natural substrate concentrations, was studied during the survey of an Atlantic coastal marine pond (France) . The regulation of this activity occurs at two different levels: on the one hand, at the cellular level, the ectoenzyme synthesis is regulated by hydrolysis substrates, dissolved combined amino acids (DCAA), and end products, dissolved free amino acids (DFAA), in terms of the relative amounts available to the cell, and on the other hand, at the ecosystem level, i.e . the hydrolytic activity, by the total amounts of DCAA and DFAA in situ . The DFAA acts as an inhibitor in enzymatic synthesis; in contrast, dissolved proteins induce the enzymatic synthesis and the exoproteolytic activity . These results, obtained in natural concentration conditions, confirm the functioning in situ of the ectoenzymatic activity regulation model of Chrost, until now only validated in an enriched experimental medium. J Clin Microbiol, 1999 Oct, 37(10), 3402 - 4 Elimination of bacterial DNA from Taq DNA polymerases by restriction endonuclease digestion; Carroll NM et al.; The incidence of false positives due to the presence of bacterial DNA in Taq DNA polymerase is an obstacle to the use of PCR in the diagnosis of infection . We describe a method that uses a restriction enzyme to destroy the ability of contaminating sequences to act as templates for a nested PCR which uses primers based on the 16S rRNA genes . The method was used prior to a PCR that amplified 10 fg of bacterial DNA . This method can be readily adapted to suit other sensitive PCRs required for clinical applications. J Neurol Neurosurg Psychiatry, 1999 Oct, 67(4), 468 - 73 Bacterial meningitis associated with lumbar drains: a retrospective cohort study; Coplin WM et al.; OBJECTIVES: The infective potential of lumbar drainage is an important topic deserving particular study . The aetiology, incidence, and clinical findings associated with bacterial meningitis are described in patients having continuous lumbar CSF drainage to treat communicating hydrocephalus after subarachnoid haemorrhage or CSF leaks after traumatic dural rents . METHODS: Retrospective review of the records of patients with a positive CSF bacterial culture who underwent lumbar drain placement over a 39 month period . RESULTS: Thirteen cases of bacterial meningitis occurred subsequent to the use of 312 lumbar drain kits (4.2%) . All meningitic patients had CSF pleocytosis, but not all had peripheral leukocytosis . Fever, peripheral leukocytosis, and CSF pleocytosis did not help to differentiate the presence of bacterial meningitis from other infections . Eight patients had prior CSF drainage procedures, including ventriculostomy (n=5) or lumbar drain (n=5) placements; two patients received both procedures . Six of 13 patients developed their CSF infection within 24 hours of lumbar drain insertion . Six of 13 patients developed meningitis while receiving antibiotics for other reasons . CONCLUSIONS: External lumbar drainage seems to carry a low risk of infectious meningitis and offers a safe alternative to ventriculostomy or serial lumbar punctures . Antibiotics do not seem to protect completely against developing the infection . The infection happens most often with skin organisms . The meningitis often appears within 24 hours after lumbar drain placement . Daily CSF samples should include bacterial cultures but cell counts may not offer any additional useful information in diagnosing the complication . Lumbar drain insertion and management need not be confined to the intensive care unit. Mol Gen Genet, 1999 Jul, 261(6), 933 - 40 Expression and biochemical characterisation of recombinant AceA, a bacterial alpha-mannosyltransferase; Geremia RA et al.; Biosynthesis of repeat-unit polysaccharides and N-linked glycans proceeds by sequential transfer of sugars from the appropriate sugar donor to an activated lipid carrier . The transfer of each sugar is catalysed by a specific glycosyltransferase . The molecular basis of the specificity of sugar addition is not yet well understood, mainly because of the difficulty of isolating these proteins . In this study, the aceA gene product expressed by Acetobacter xylinum, which is involved in the biosynthesis of the exopolysaccharide acetan, was overproduced in Escherichia coli and its function was characterised . The aceA ORF was subcloned into the expression vector pET29 in frame with the S.tag epitope . The recombinant protein was identified, and culture conditions were optimised for production of the soluble protein . The results of test reactions showed that AceA is able to transfer one alpha-mannose residue from GDP-mannose to cellobiose-P-P-lipid to produce alpha-mannose-cellobiose-P-P-lipid . AceA was not able to use free cellobiose as a substrate, indicating that the pyrophosphate-lipid moiety is needed for enzymatic activity. Biochim Biophys Acta, 1999 Aug 4, 1412(3), 273 - 81 Determination of Q(A)-content in bacterial reaction centers: an indispensable requirement for quantifying B-branch charge separation Ogrodnik A, Muller P, Hartwich G, Michel-Beyerle ME. We have been able to determine the occupancy of the quinone site at the A-branch (Q(A)) of a reaction center preparation with an accuracy of 2% . This is achieved by accumulating the P(+)Q(-)(A) state after multiple actinic excitation and monitoring the extent of the 30 ms ground state bleaching . This bleaching is corrected for deviations from complete saturation due to competing charge separation to the B-branch . On the other hand, knowledge of the Q(A) content is indispensable for determining the yield of B-branch charge separation from nanosecond transients associated with the recombination of P(+)H(-)(B), which have to be corrected for the nanosecond signal originating from P(+)H(-)(A) of RCs having lost Q(A). Lupus, 1999, 8(6), 449 - 55 Induction of antiphospholipid antibodies by immunization with synthetic viral and bacterial peptides; Gharavi EE et al.; We previously induced pathogenic antibodies against anionic phospholipids (PL) in experimental animals by immunization with lipid-free purified human beta2glycoprotein I (beta2GPI) . We hypothesized that antiphospholipid antibodies (aPL) are induced by in vivo binding of foreign beta2GPI to self-PL, thus forming an immunogenic complex against which aPL antibodies are produced . If this hypothesis is true, other PL-binding proteins that are products of ubiquitous viral/bacterial agents may also induce aPL . To test this hypothesis, groups of NIH/Swiss mice were immunized with synthetic peptides of viral and bacterial origin that share structural similarity with the putative PL-binding region of beta2GPI . Compared with the control groups, animals immunized with the peptides produced significantly higher levels of aPL and anti-beta2GPI antibodies . These findings demonstrate that some PL-binding viral and bacterial proteins function like beta2GPI in inducing aPL and anti-beta2GPI production, and are consistent with a role for such viral and bacterial proteins in inducing aPL antibody production in humans. J Virol, 1999 Oct, 73(10), 8320 - 9 Cloning of the human cytomegalovirus (HCMV) genome as an infectious bacterial artificial chromosome in Escherichia coli: a new approach for construction of HCMV mutants; Borst EM et al.; We have recently introduced a novel procedure for the construction of herpesvirus mutants that is based on the cloning and mutagenesis of herpesvirus genomes as infectious bacterial artificial chromosomes (BACs) in Escherichia coli (M . Messerle, I . Crnkovic, W . Hammerschmidt, H . Ziegler, and U . H . Koszinowski, Proc . Natl . Acad . Sci . USA 94:14759-14763, 1997) . Here we describe the application of this technique to the human cytomegalovirus (HCMV) strain AD169 . Since it was not clear whether the terminal and internal repeat sequences of the HCMV genome would give rise to recombination, the stability of the cloned HCMV genome was examined during propagation in E . coli, during mutagenesis, and after transfection in permissive fibroblasts . Interestingly, the HCMV BACs were frozen in defined conformations in E . coli . The transfection of the HCMV BACs into human fibroblasts resulted in the reconstitution of infectious virus and isomerization of the reconstituted genomes . The power of the BAC mutagenesis procedure was exemplarily demonstrated by the disruption of the gpUL37 open reading frame . The transfection of the mutated BAC led to plaque formation, indicating that the gpUL37 gene product is dispensable for growth of HCMV in fibroblasts . The new procedure will considerably speed up the construction of HCMV mutants and facilitate genetic analysis of HCMV functions. Immunol Lett, 1999 Aug 3, 69(2), 233 - 8 Bacterial lipopolysaccharide induced B cell activation is mediated via a phosphatidylinositol 3-kinase dependent signaling pathway; Venkataraman C et al.; Bacterial lipopolysaccharide (LPS) is a potent stimulant of B cells and macrophages . LPS induces B cell proliferation and differentiation into antibody secreting cells . In addition, LPS also stimulates IL-6 secretion in mature B cells and in immature B cell lines such as WEHI-231 . Although sufficient literature is available on LPS induced signaling events in monocytes and macrophages, the mechanisms involved in LPS induced B cell activation are not well understood . In this report, it is shown that both LPS mediated B cell proliferation and IL-6 secretion are dependent on phosphatidylinositol 3-kinase (PI 3-kinase) signaling pathways . The B cell specific co-receptor, CD19 is not tyrosine phosphorylated in LPS stimulated B cells . Thus, in contrast to B cell antigen receptor (BCR) signaling, the activation of PI 3-kinase appears not to be related to the recruitment of PI 3-kinase to tyrosine phosphorylated CD19 . This is the first demonstration of the importance of PI 3-kinase signaling pathway in LPS mediated B lymphocyte activation. FEMS Microbiol Lett, 1999 Oct 1, 179(1), 1 - 9 Regulation of bacterial photosynthesis genes by oxygen and light; Gregor J et al.; Most bacteria have the capability to adapt to changes in their environment . Facultatively phototrophic bacteria like Rhodobacter can switch from aerobic respiration to anoxygenic photosynthesis in the absence of oxygen . The formation of the photosynthetic apparatus is primarily regulated by oxygen tension . The amount of photosynthetic complexes is influenced by the light intensity in anaerobic cultures . This review focuses on the molecular mechanisms involved in the regulation of Rhodobacter photosynthesis genes by oxygen and light. Microbiol Immunol, 1999, 43(6), 543 - 50 Detection and nucleotide sequence analysis of human caliciviruses (HuCVs) from samples in non-bacterial gastroenteritis outbreaks in Hokkaido, Japan; Ohyama T et al.; Samples of feces and vomit collected from patients during 13 non-bacterial gastroenteritis outbreaks which occurred in Hokkaido between 1995 and 1998 were examined by electron microscopy (EM) and reverse-transcription polymerase chain reaction (RT-PCR) for evidence of infection with human caliciviruses (HuCVs) . In 6 food-borne outbreaks, oysters were the probable source of infection, while the origin of HuCVs was not found out for the other 7 outbreaks . One-hundred-eleven of 214 stool, vomit and oyster specimens examined gave positive results by RT-PCR, while HuCVs were detected by EM in 36 of 121 stool specimens examined . We determined the nucleotide sequences of 470-bp or 373-bp PCR products amplified from the RNA polymerase region of the HuCV genomes with primer sets MR3/4 and Yuri22F/R, respectively . The sequences of different strains revealed great heterogenicity, with a range of 60 to 100% homology among strains . In a few cases, a mixed genotype was found in the same patient or same outbreak by means of nested PCR and cloning of PCR products into an appropriate vector . Of the 19 different strains found, 4 strains could be classified as Norwalk virus (genogroup 1) and the other 15 strains as Snow Mountain agent (genogroup 2) based on genotyping with homology analysis . Furthermore, the strains belonging to genogroup 2 could be classified into 4 subgroups with more than 93% homology in amino acids among strains in the subgroup. Otolaryngol Clin North Am, 1999 Oct, 32(5), 793 - 811 Acute viral and bacterial infections of the salivary glands; McQuone SJ; Acute infection can involve any salivary gland, but it predominately affects the major salivary glands, especially the parotid gland . The anatomic and physiologic factors accounting for the parotid gland's predilection for infection are reviewed . Numerous conditions that are predisposed to acute bacterial sialadenitis and differ from risk factors associated with viral infection are also reviewed . The pathogenesis, diagnostic evaluation, treatment, complications, and prognosis of bacterial infections are discussed and contrasted with those of viral infections. Eur Cytokine Netw, 1999 Sep, 10(3), 373 - 82 Impairment of HIV polymorphonuclear leukocyte transmigration across T84 cell monolayers: an alternative mechanisms for increased intestinal bacterial infections in AIDS? Hofman P, Fischer F, Far DF, Selva E, Battaglione V, Bayle J, Rossi B. Our objective was to study the influence of HIV infection of polymorphonuclear leukocytes (PMN) on transepithelial migration . To date, reports of functional PMN chemotaxis in AIDS are contradictory . This is the first attempt to assess this function via an in vitro model allowing transmigration of neutrophils through an intestinal epithelial barrier . PMN were isolated from 45 HIV-infected patients and 45 healthy volunteers . PMN transmigration across T84 epithelial cells was initiated by applying either various concentrations of formyl-met-leu-phe peptide (f-MLP) or interleukin-8 and assayed by quantification of myeloperoxidase activity . CD11b, CD18, and CD47 expression on PMN was compared before and after transepithelial migration by flow cytometry analysis . CD11b expression was studied by electron microscopy . Apoptosis of transmigrated HIV PMN and control PMN was investigated by morphology and DNA fragmentation characterization . Compared to control PMN, HIV PMN exhibited a decrease in transepithelial migration that directly correlated with CD4+ counts . Basal and transepithelial migration-mediated expression of CD11b, CD18, and CD47 were unmodified in HIV PMN compared to control PMN . Electron microscopy labeling confirmed no difference in CD11b expression on HIV and control PMN . The index of apoptosis in transmigrated HIV PMN and control PMN was identical . These data provide evidence of a defect in the f-MLP-induced chemotaxis of PMN from HIV-infected patients across an intestinal epithelial barrier . This defective migration is not due to a quantitative modification of CD11b, CD18 and CD47 on HIV PMN suggesting a more subtle alteration . The impairment in the transmigration function may contribute in vivo to an increased susceptibility to intestinal bacterial infection in HIV-infected patients. Appl Environ Microbiol, 1999 Sep, 65(9), 3982 - 9 High bacterial diversity in permanently cold marine sediments; Ravenschlag K et al.; A 16S ribosomal DNA (rDNA) clone library from permanently cold marine sediments was established . Screening 353 clones by dot blot hybridization with group-specific oligonucleotide probes suggested a predominance of sequences related to bacteria of the sulfur cycle (43.4% potential sulfate reducers) . Within this fraction, the major cluster (19.0%) was affiliated with Desulfotalea sp . and other closely related psychrophilic sulfate reducers isolated from the same habitat . The cloned sequences showed between 93 and 100% similarity to these bacteria . Two additional groups were frequently encountered: 13% of the clones were related to Desulfuromonas palmitatis, and a second group was affiliated with Myxobacteria spp . and Bdellovibrio spp . Many clones (18.1%) belonged to the gamma subclass of the class Proteobacteria and were closest to symbiotic or free-living sulfur oxidizers . Probe target groups were further characterized by amplified rDNA restriction analysis to determine diversity within the groups and within the clone library . Rarefaction analysis suggested that the total diversity assessed by 16S rDNA analysis was very high in these permanently cold sediments and was only partially revealed by screening of 353 clones. Appl Environ Microbiol, 1999 Sep, 65(9), 3834 - 42 Spatial heterogeneity of bacterial populations along an environmental gradient at a shallow submarine hydrothermal vent near Milos Island (Greece); Sievert SM et al.; The spatial heterogeneity of bacterial populations at a shallow-water hydrothermal vent in the Aegean Sea close to the island of Milos (Greece) was examined at two different times by using acridine orange staining for total cell counts, cultivation-based techniques, and denaturing gradient gel electrophoresis (DGGE) analysis of PCR-amplified 16S rRNA gene fragments . Concurrent with measurements of geochemical parameters, samples were taken along a transect from the center of the vent to the surrounding area . Most-probable-number (MPN) counts of metabolically defined subpopulations generally constituted a minor fraction of the total cell counts; both counting procedures revealed the highest cell numbers in a transition zone from the strongly hydrothermally influenced sediments to normal sedimentary conditions . Total cell counts ranged from 3.2 x 10(5) cells ml(-1) in the water overlying the sediments to 6.4 x 10(8) cells g (wet weight) of sediment(-1) . MPN counts of chemolithoautotrophic sulfur-oxidizing bacteria varied between undetectable and 1.4 x 10(6) cells g(-1) . MPN counts for sulfate-reducing bacteria and dissimilatory iron-reducing bacteria ranged from 8 to 1.4 x 10(5) cells g(-1) and from undetectable to 1.4 x 10(6) cells g(-1), respectively . DGGE revealed a trend from a diverse range of bacterial populations which were present in approximately equal abundance in the transition zone to a community dominated by few populations close to the center of the vent . Temperature was found to be an important parameter in determining this trend . However, at one sampling time this trend was not discernible, possibly due to storm-induced disturbance of the upper sediment layers. AJNR Am J Neuroradiol, 1999 Aug, 20(7), 1359 - 64 Sonographic nomogram of the leptomeninges (pia-glial plate) and its usefulness for evaluating bacterial meningitis in infants; Jequier S et al.; BACKGROUND AND PURPOSE: To our knowledge, the upper limits of the thickness of normal meninges on neurosonograms are not known . We therefore established a nomogram for sonographic measurements of the leptomeninges (pia-glial plate) and assessed its usefulness in neurosonographic examinations of children with bacterial meningitis . METHODS: The pia mater-cortical glia limitans complex on the surface of the brain and in the sulcus of a frontal gyrus was measured on neurosonograms in 100 infants without meningeal disease in order to establish a nomogram of the thickness of this pia-glial plate, referred to as the leptomeninx . Effects of prematurity, age, sex, and single-layer (surface) versus double-layer (sulcus) measurements were analyzed . Meningeal thicknesses derived from a retrospective analysis of the neurosonograms of 33 patients with purulent meningitis and a prospective study of 22 patients with bacterial meningitis were compared with the nomograms . Clinical outcomes of children with meningeal thickening were compared with those of affected children with normal meninges . RESULTS: The distribution of sulci measurements was significantly asymmetrical around the mean . Statistical data showed no influence of prematurity and sex, but showed surface measurements to be more consistent than sulcal measurements . Older chronological age was related to slightly larger sulci, but did not influence the surface measurements . In children with bacterial meningitis, the surface meninges were less frequently thickened than were the sulci . Sulcal enlargement occurred often in combination with echogenic deposits in the sub-arachnoid space . CONCLUSION: Leptomeninges are best measured on the surface of a gyrus rather than in a sulcus, as the normal thickness of the sulci shows much more variability . Clinical outcome of bacterial meningitis cannot be predicted by presence or absence of meningeal thickening as the only sonographic abnormality. Infect Dis Clin North Am, 1999 Sep, 13(3), 647 - 59, viii Supportive management in bacterial meningitis; Rauf SJ et al.; The central nervous system and systemic complications of bacterial meningitis cause significant morbidity and mortality . This article offers insight into the clinical features, pathogenesis, and management of these complications . In many instances, the improved outcome of intervention is based on clinical suspicion and early recognition . The management of complications is evolving and is presently based mainly on supportive care. Infect Dis Clin North Am, 1999 Sep, 13(3), 711 - 33, viii Bacterial meningitis and the newborn infant; Pong A et al.; Bacterial meningitis in the neonate differs from meningitis in the older infant and child in a number of ways . Bacterial pathogens primarily are associated with the maternal genitourinary tract . Symptoms and physical findings may be nonspecific, and a high index of suspicion is needed . Management may vary depending on the maturity of the infant and the bacterial pathogen that is isolated. Infect Dis Clin North Am, 1999 Sep, 13(3), 579 - 94, vi-vii Clinical presentations, diagnosis, and prognostic factors of bacterial meningitis; Kaplan SL; The clinical presentations of children and adults with bacterial meningitis have not changed over the past several decades, and a high index of suspicion remains critical for timely identification of infected patients . With the virtual disappearance of H . influenzae type B meningitis (Hib) in areas of the world where Hib conjugate vaccine is administered routinely, the utility of commercially available tests for rapid detection of bacterial polysaccharides has diminished . Detection of gene products of meningeal pathogens in cerebrospinal fluid or blood is still experimental . The prognostic findings of recent studies are not different from those previously described, despite advances in the supportive care of critically ill patients. Infect Dis Clin North Am, 1999 Sep, 13(3), 527 - 48, v-vi Pathogenesis of bacterial meningitis; Leib SL et al.; Bacterial meningitis is fatal in 5% to 40% of patients and causes neurologic sequelae in up to 30% of survivors . Much has been learned recently about the mechanisms that lead to brain injury during meningitis . Once bacteria have gained access to the central nervous system, their multiplication triggers a complex host response consisting of humoral and cellular immune mediators, reactive oxygen intermediates, matrix-metalloproteinases, and other host-derived factors . Alterations of the cerebral vasculature, with disruption of the blood brain barrier and global and focal ischemia, ultimately lead to functional and structural brain damage . This article reviews current concepts of the pathophysiology of bacterial meningitis and emphasizes possible therapeutic strategies to prevent its harmful consequences. J Control Release, 1999 Aug 27, 61(1-2), 51 - 63 Bacterial ghosts as drug carrier and targeting vehicles; Huter V et al.; A novel system for the packaging of drugs as well as vaccines is presented . Bacterial ghosts are intact, non-denatured bacterial envelopes that are created by lysis of bacteria through the expression of cloned phage PhiX174 gene E . Inhibition of induced E-mediated lysis by MgSO(4), harvesting of cells by centrifugation, and resuspension in low-ionic-strength buffers leads to rapid, violent lysis and results in empty bacterial envelopes with large (approximately 1 microm in diameter) openings . The construction of plasmid pAV1, which encodes a streptavidin fusion protein with an N-terminal membrane anchor sequence, allows the loading of the inner side of the cytoplasmic membrane with streptavidin . The functionality and efficacy of binding of even large biotinylated compounds in such streptavidin ghosts (SA-ghosts) was assessed using the enzyme alkaline phosphatase . The successful binding of biotinylated fluorescent dextran, as well as fluorescent DNA complexed with biotinylated polylysine, was demonstrated microscopically . The display by bacterial ghosts of morphological and antigenic surface structures of their living counterparts permits their attachment to target tissues such as the mucosal surfaces of the gastrointestinal and respiratory tract, and their uptake by phagocytes and M cells . In consequence, SA-ghosts are proposed as drug carriers for site-specific drug delivery. Proc Natl Acad Sci U S A, 1999 Aug 31, 96(18), 10344 - 8 Bacterial inactivation by using near- and supercritical carbon dioxide; Dillow AK et al.; The three most common methods of sterilization in use today are ethylene oxide exposure, gamma-irradiation, and steam sterilization . Each of these methods has serious limitations for the sterilization of some materials used in medicine, especially thermally and hydrolytically sensitive polymers by themselves and in combination with proteins . In this work, we demonstrate a potential new method of sterilization by using supercritical fluid carbon dioxide . Using this method we achieve complete inactivation of a wide variety of bacterial organisms at moderate temperatures and in the absence of organic solvents or irradiation . Sterilization is a function of both the proximity to the fluid's critical point and the chemical nature of the fluid itself . When biodegradable polymers poly(lactic-co-glycolic) acid and polylactic acid were included in the sterilization process, there was no effect on the inactivation efficiency, yet no physical or chemical damage to these thermally and hydrolytically labile materials was observed. J Infect, 1999 Jul, 39(1), 55 - 60 Cerebrospinal fluid cytokine levels and dexamethasone therapy in bacterial meningitis; Ohga S et al.; OBJECTIVES: cerebrospinal fluid (CSF) levels of interleukin (IL)-1 beta and tumor necrosis factor (TNF) alpha were measured to assess the effect and application of dexamethasone (Dex) therapy for bacterial meningitis . METHODS: associations between clinical findings and CSF parameters were first investigated, and prognosis was compared between 25 patients with Dex and 12 without Dex therapy . RESULTS: patients with the presence of disturbed consciousness showed higher CSF levels of TNF alpha (mean: 3015 pg/ml) or protein (mean: 215 mg/dl) than those without it (both, P < 0.O5) . Simultaneous increase of TNF alpha (> 1000 pg/ml) and protein (> 100 g/dl) was observed in 80%, of patients with profoundly disturbed consciousness . Patients with Dex therapy presented higher TNF alpha/protein levels at diagnosis than those without Dex therapy (P < 0.05) . Despite worse conditions at diagnosis, only one of 14 Dex-treated patients whose initial CSF TNF alpha levels exceeded 1000 pg/ml developed deafness . On the other hand, two of four patients without Dex therapy who had the same TNF alpha level suffered from psychomotor retardation . The differences in the frequency of sequelae between those with and without Dex therapy were significant in patients showing high TNF alpha level (P < O.05), but not in those showing high CSF levels of IL-1 beta or protein . The logistic regression analysis indicated that high CSF protein level (P < O.0001), or no Dex therapy (P=0.0001) was the independent risk factor for sequelae . CONCLUSIONS: although the study number was small, our observations suggested that CSF TNF alpha/protein levels reflected the neurologic severity, and implied that early Dex therapy might be beneficial for patients with prominently high TNF alpha levels. Biotechnol Appl Biochem, 1999 Aug, 30 ( Pt 1), 59 - 64 Expression and purification of a secreted functional mouse/human chimaeric antibody against bacterial endotoxin in baculovirus-infected insect cells; Tan W et al.; We have created a mouse/human chimaeric antibody by taking antigen-binding fragment (Fab) genes of a mouse antibody-producing hybridoma with specificity for bacterial endotoxin and joining them to human Ig crystallizing-fragment (Fc) genes using recombinant DNA techniques . This chimaeric antibody has been expressed in Sf21 and High Fivetrade mark (BTI-TN-5B1-4) insect cells using the baculovirus expression system, which may allow the mass production of secretory recombinant antibodies . This was achieved by using infection with a double-recombinant virus containing cDNAs of both the Ig heavy-chain (HC) and light-chain (LC) genes . Prior to recombination, each gene was cloned into the dual-expression baculovirus transfer vector pPLSP2, which permitted the insertion of the LC gene in-frame with the signal peptide of honey bee melittin downstream of the polyhedrin promoter, and of the HC gene in-frame with the signal peptide of Bombyx mori larval serum protein downstream of the p10 promoter . Our results showed that the polypeptide chains were secreted by insect cells and correctly assembled into H(2)L(2) heterodimers containing N-linked carbohydrate at the heavy chain . Furthermore, the recombinant chimaeric antibody exhibited a similar antigen specificity to that of the monoclonal antibody . More importantly, it provides a generic method for the high-level expression of antibodies. Genes Dev, 1999 Aug 15, 13(16), 2134 - 47 Bacterial promoter architecture: subsite structure of UP elements and interactions with the carboxy-terminal domain of the RNA polymerase alpha subunit; Estrem ST et al.; We demonstrate here that the previously described bacterial promoter upstream element (UP element) consists of two distinct subsites, each of which, by itself, can bind the RNA polymerase holoenzyme alpha subunit carboxy-terminal domain (RNAP alphaCTD) and stimulate transcription . Using binding-site-selection experiments, we identify the consensus sequence for each subsite . The selected proximal subsites (positions -46 to -38; consensus 5'-AAAAAARNR-3') stimulate transcription up to 170-fold, and the selected distal subsites (positions -57 to -47; consensus 5'-AWWWWWTTTTT-3') stimulate transcription up to 16-fold . RNAP has subunit composition alpha(2)betabeta'sigma and thus contains two copies of alphaCTD . Experiments with RNAP derivatives containing only one copy of alphaCTD indicate, in contrast to a previous report, that the two alphaCTDs function interchangeably with respect to UP element recognition . Furthermore, function of the consensus proximal subsite requires only one copy of alphaCTD, whereas function of the consensus distal subsite requires both copies of alphaCTD . We propose that each subsite constitutes a binding site for a copy of alphaCTD, and that binding of an alphaCTD to the proximal subsite region (through specific interactions with a consensus proximal subsite or through nonspecific interactions with a nonconsensus proximal subsite) is a prerequisite for binding of the other alphaCTD to the distal subsite. Clin Ther, 1999 Jul, 21(7), 1158 - 70 Comparison of cefuroxime axetil and amoxicillin/clavulanate in the treatment of acute bacterial sinusitis; Henry DC et al.; This double-masked, multicenter, randomized clinical trial compared the efficacy and tolerability of cefuroxime axetil and amoxicillin/clavulanate in the treatment of acute bacterial maxillary sinusitis . A total of 263 patients with acute bacterial maxillary sinusitis were randomly assigned to receive 10 days of treatment with either cefuroxime axetil 250 mg twice daily (n = 132) or amoxicillin/clavulanate 500/125 mg 3 times daily (n = 131) . Patients' responses to treatment were assessed once during treatment (6 to 8 days after the start of treatment), at the end of treatment (1 to 3 days posttreatment), and at follow-up (26 to 30 days after cessation of treatment) . Clinical success, defined as cure or improvement, was equivalent in the cefuroxime axetil and amoxicillin/ clavulanate groups at the end-of-treatment and follow-up assessments . Patients in both groups showed improvements in symptoms of acute sinusitis at the during-treatment visit . Treatment with amoxicillin/clavulanate was associated with a significantly higher incidence of drug-related adverse events than treatment with cefuroxime axetil (29% vs 17%), primarily reflecting a higher incidence of gastrointestinal adverse events (23% vs 11%), particularly diarrhea . Two patients in the cefuroxime axetil group and 8 patients in the amoxicillin/clavulanate group withdrew from the study due to adverse events (P = 0.06) . These results indicate that cefuroxime axetil 250 mg twice daily is as effective as amoxicillin/clavulanate 500 mg 3 times daily in the treatment of acute sinusitis and produces fewer gastrointestinal adverse events . cefuroxime axetil, amoxicillin/clavulanate, acute sinusitis. Pediatr Infect Dis J, 1999 Aug, 18(8), 666 - 71 Comparison of procalcitonin with interleukin 8, C-reactive protein and differential white blood cell count for the early diagnosis of bacterial infections in newborn infants; Franz AR et al.; OBJECTIVE: To evaluate procalcitonin (PCT) as a test for early diagnosis of bacterial infections (BI) in newborn infants and to compare the results of PCT with those of interleukin 8 (IL-8), C-reactive protein (CRP) and differential white blood cell count . STUDY DESIGN: PCT was prospectively measured along with IL-8, CRP and differential white blood cell counts and blood cultures in 197 newborn infants at the first suspicion of bacterial infection . PCT, IL-8, CRP and differential white blood cell counts were analyzed for sensitivity, specificity and positive and negative predictive values after receiver operating characteristic curve analysis for best thresholds . The kinetics of PCT was determined in infants with and without BI . RESULTS: Forty-six infants were diagnosed clinically as having BI, of whom 9 had BI with positive blood cultures . At a cutoff value of 0.50 microg/l, PCT detected combined culture-proved and clinical BI with a sensitivity of 57% (95% confidence interval, 41%, 71%) and a specificity of 66% (95% confidence interval, 57%, 74%) . The combination of IL-8 > or =70 ng/l and/or CRP >10 mg/l achieved a sensitivity of 91% (95% confidence interval, 79%, 98%) and a specificity of 73% (95% confidence interval, 64%, 81%) . PCT values of infected and not infected infants tended to rise for 24 h after initial evaluation and then decreased . CONCLUSION: The combination of IL-8 and CRP is more reliable than PCT as a test for early diagnosis of BI in newborn infants. Drug Metab Dispos, 1999 Sep, 27(9), 999 - 1004 Enhancement of cytochrome P-450 3A4 catalytic activities by cytochrome b(5) in bacterial membranes; Yamazaki H et al.; Activities of testosterone, nifedipine, and midazolam oxidation by recombinant cytochrome P-450 (P-450) 3A4 coexpressed with human NADPH-P-450 reductase (NPR) in bacterial membranes (CYP3A4/NPR membranes) were determined in comparison with those of other recombinant systems and of human liver microsomes with high contents of CYP3A4 . Growth conditions for Escherichia coli transformed with the bicistronic construct affected expression levels of CYP3A4 and NPR; an excess of NPR over P-450 in membrane preparations enhanced CYP3A4-dependent testosterone 6beta-hydroxylation activities of the CYP3A4/NPR membranes . Cytochrome b(5) (b(5)) and apolipoprotein b(5) further enhanced the testosterone 6beta-hydroxylation activities of CYP3A4/NPR membranes after addition to either bacterial membranes or purified enzymes . NPR was observed to enhance catalytic activity when added to the CYP3A4/NPR membranes, either in the form of bacterial membranes or as purified NPR (in combination with cholate and b(5)) . Apparent maximal activities of testosterone 6beta-hydroxylation in CYP3A4/NPR membranes were obtained when the molar ratio of CYP3A4/NPR/b(5) was adjusted to 1:2:1 by mixing membranes containing each protein . Testosterone 6beta-hydroxylation, nifedipine oxidation, and midazolam 4- and 1'-hydroxylation activities in CYP3A4/NPR membranes plus b(5) systems were similar to those measured with microsomes of insect cells coexpressing CYP3A4 with NPR and/or of human liver microsomes, based on equivalent CYP3A4 contents . These results suggest that CYP3A4/NPR membrane systems containing b(5) are very useful models for prediction of the rates for liver microsomal CYP3A4-dependent drug oxidations. Br J Ophthalmol, 1999 Sep, 83(9), 1050 - 5 Intravitreal dexamethasone in exogenous bacterial endophthalmitis: results of a prospective randomised study; Das T et al.; AIM: To evaluate the efficacy of intravitreal dexamethasone co-administered with intravitreal antibiotics along with vitrectomy in the management of exogenous bacterial endophthalmitis . METHODS: In a prospective randomised clinical trial, 63 patients (63 eyes) with suspected bacterial endophthalmitis (postoperative and post-traumatic) were treated with vitrectomy and intravitreal antibiotics and randomised to intravitreal dexamethasone (IOAB with = 29 eyes) and no dexamethasone (IOAB without = 34 eyes) . Inflammation score (IS) and visual acuity were measured by two masked observers before surgery, and at 1, 4, and 12 weeks after surgery in both the groups . RESULTS: There was significant reduction (p <0.0001) in IS at 1, 4, and 12 weeks after the surgery in the "IOAB with" group; there was temporary but significant increase (p <0.01) in IS at 1 week in the "IOAB without" group, before decline (p <0.001) of IS at 4 and 12 weeks . The magnitude and relative percentage change in IS between the two groups were found to be significant at 1 (p <0.0001), and 4 (p <0.01) weeks, and not at 12 weeks . The visual acuity at 12 weeks was comparable in both the IOAB with and IOAB without groups . CONCLUSION: Intravitreal dexamethasone helps in early reduction of inflammation in exogenous bacterial endophthalmitis, but has no independent influence on the visual outcome . In selected patients with endophthalmitis where oral corticosteroids cannot be given for medical reasons intravitreal corticosteroids could be beneficial; in other situations they could be complementary to oral corticosteroid therapy. Br J Surg, 1999 Aug, 86(8), 1020 - 4 Bacterial infection and extent of necrosis are determinants of organ failure in patients with acute necrotizing pancreatitis; Isenmann R et al.; BACKGROUND: The risk factors predisposing to organ failure in patients with necrotizing pancreatitis remain unclear . The relationship between the extent of pancreatic necrosis, the presence of infection and the incidence of organ failure was analysed . METHODS: In a retrospective review, the occurrence of pulmonary insufficiency, renal insufficiency, shock, sepsis/sepsis-like syndrome (SLS) and coagulopathy was evaluated in 273 patients with necrotizing pancreatitis, and a comparison was made between patients with sterile or infected necrosis . Additionally, the relation between the incidence of organ failure and extent of pancreatic parenchymal necrosis was investigated by classifying the patients into three groups according to the amount of necrotic tissue found by contrast-enhanced computed tomography (group 1, extent less than 30 per cent; group 2, 30-50 per cent; group 3, more than 50 per cent) . RESULTS: Organ failure was more frequent in patients with infected necrosis than in those with sterile necrosis . Differences were found in the incidence of pulmonary insufficiency, sepsis/SLS and coagulopathy . Organ failure occurred more frequently in group 3 than in group 2 or 1 (95 versus 79 and 66 per cent; P = 0.0004) . The extent of infected necrosis was not related to the incidence of organ failure (group 1, 88 per cent; group 2, 86 per cent; group 3, 96 per cent) . However, there was a relation between the incidence of organ failure and the extent of sterile necrosis (group 1, 59 per cent; group 2, 74 per cent; group 3, 94 per cent; P = 0.0001) . Multivariate analysis confirmed the presence of infection and the extent of necrosis as independent determinants of organ failure . CONCLUSION: The incidence of organ failure is determined by both bacterial infection and extent of necrosis . The incidence of organ failure is determined by the extent of necrotic parenchyma in patients with sterile necrosis . Infected necrosis is associated with a high incidence of organ failure irrespective of the extent of necrosis. Surgery, 1999 Aug, 126(2), 349 - 57 Discordant tumor necrosis factor-alpha superfamily gene expression in bacterial peritonitis and endotoxemic shock; Tannahill CL et al.; BACKGROUND: Tumor necrosis factor-alpha (TNF-alpha) is a member of a large family of predominantly homotrimeric type II membrane-associated proteins with both proinflammatory and apoptosis-inducing properties . Although TNF-alpha expression has been studied extensively, little is known about the expression of other members of the TNF-alpha superfamily during acute inflammatory processes . METHODS: TNF-alpha, Fas ligand (FasL), and TRAIL (tumor necrosis factor-related apoptosis-inducing ligand) messenger RNA (mRNA) expression were examined in liver, lung, spleen, and kidney after either a cecal ligation and puncture or endotoxemic shock with use of semiquantitative reverse transcriptase-polymerase chain reaction . RESULTS: Cecal ligation and puncture increased TNF-alpha mRNA in lung and liver (both P < .05) within 3 hours, which was paralleled by increased FasL mRNA . In the spleen TNF-alpha and FasL mRNA significantly declined (both P < .05) . In contrast to TNF-alpha and FasL, TRAIL mRNA levels were unchanged in all organs except lung, where it was reduced at 24 hours (P < .05) . Endotoxemic shock also increased lung TNF-alpha and FasL mRNA levels (both P < .05) . CONCLUSIONS: In acute inflammatory processes TNF-alpha and FasL mRNA increase concordantly in several solid organs . In contrast, TRAIL mRNA levels do not consistently change during these acute inflammatory processes, suggesting that its expression is under independent and discordant regulatory control. Arch Pathol Lab Med, 1999 Sep, 123(9), 778 - 81 Endoscopic biopsy pathology of Helicobacter pylori gastritis . Comparison of bacterial detection by immunohistochemistry and Genta stain; Toulaymat M et al.; OBJECTIVES: To describe the endoscopic biopsy pathology of Helicobacter pylori gastritis, compare bacterial detection by immunohistochemistry using a specific antibody with the Genta stain, and to compare the relative costs of the 2 techniques . DESIGN: One hundred cases of gastritis identified as positive for H pylori by Genta stain and 100 cases considered negative by the same technique were stained using an anti-H pylori-specific polyclonal antibody . Laboratory reagent and labor costs for the 2 methods were compared . RESULTS: Chronic active gastritis with lymphoid follicles was significantly associated with H pylori infection (P <.0001) . The immunohistochemical method had a sensitivity of 97% and a specificity of 98% compared with the Genta stain, with strong agreement for grading density of organisms (kappa = 0.85; P <.001) . Reagent costs were similar for both methods, but immunohistochemistry using an autoimmunostainer required less dedicated technical time and hence was less expensive than the Genta stain . CONCLUSIONS: Immunohistochemistry using a specific antibody is an accurate and cost-effective method for H pylori detection in gastric biopsies. Sex Transm Dis, 1999 Aug, 26(7), 404 - 9 Bacterial vaginosis, ethnicity, and the use of genital cleaning agents: a case control study; Rajamanoharan S et al.; BACKGROUND AND OBJECTIVES: Bacterial vaginosis and vaginal douching are both reported to be more common in African-American and Caribbean than white women . It is also thought that douching alters the vaginal milieu . This study was conducted to examine associations between genital cleaning practices, bacterial vaginosis, and ethnic group . STUDY DESIGN: Case-control study of 100 women with bacterial vaginosis, diagnosed by Nugent's criteria, and 100 women without bacterial vaginosis attending a sexually transmitted diseases clinic in an ethnically heterogeneous inner-city area in London, England . RESULTS: Bacterial vaginosis was more common among black Caribbean than white women (OR, 2.1; 95% CI, 1.1-4.1) . Vulval use of bubble bath or antiseptic solutions and douching with proprietary or homemade solutions were significantly more common in women with bacterial vaginosis than without . After controlling for use of vulval and vaginal antiseptics and bubble bath, douching, and a history of bacterial vaginosis, there was no ethnic difference in the occurrence of the condition (adjusted OR, 1.1; 95% CI, 0.5-2.5) . CONCLUSIONS: Ethnic differences in genital hygiene behaviors can explain a twofold increase in the risk of bacterial vaginosis in black Caribbean compared with white women . The role of vulval and vaginal cleaning practices in the development of bacterial vaginosis should be examined further in longitudinal or randomized controlled studies. Biotechniques, 1999 Aug, 27(2), 321 - 4, 326 Differentiation of hard-to-type bacterial strains by RNA mismatch cleavage; Bricker BJ; Many bacteria are difficult to subtype due to high genetic relatedness . In the cases of pathogens of medical or veterinary importance, subtyping is an essential tool of epidemiologists . This report describes a method for molecular subtyping based on the detection of point mutations without DNA sequencing or specialized equipment . The method, known as RNA mismatch cleavage, hybridizes RNA transcripts derived from PCR-amplified DNA, with a control RNA transcript followed by RNase cleavage at point-mutation mismatches . The method was successful in distinguishing all six Brucella species tested and was able to distinguish 11 of the 18 biovars studied . Of the remaining seven biovars (all of which are Brucella abortus strains), three subgroups were identified . The method should be applicable to all hard-to-subtype bacterial strains. Infect Immun, 1999 Sep, 67(9), 4834 - 42 The EspD protein of enterohemorrhagic Escherichia coli is required for the formation of bacterial surface appendages and is incorporated in the cytoplasmic membranes of target cells; Kresse AU et al.; The formation of EspA-containing surface appendages in pathogenic Escherichia coli strains, both enteropathogenic E . coli (EPEC) and Shiga toxin-producing E . coli strains, is essential for critical events in the infective process, e.g., localized bacterial adherence to host cells with formation of microcolonies and induction of attaching and effacing lesions . It has been reported that EPEC mutants deficient in the production of EspD, which is encoded by the esp operon, are unable to accumulate actin underneath adherent bacteria but exhibit an attachment similar to that of the wild type . Here, we report the construction and characterization of an in-frame espD deletion mutant of the enterohemorrhagic E . coli (EHEC) strain EDL933 . In contrast to what was observed in EPEC mutants, the EDL933 espD mutant not only lacked the capacity to accumulate actin but also exhibited an impaired attachment to HeLa cells . The synthesis of the EspD protein was also essential for the formation of EspA-containing filaments . Finally, localization studies demonstrated that the EspD protein is transferred to the cytoplasm and integrated into the cytoplasmic membranes of infected cells . These results help to elucidate the underlying molecular events in infections caused by EHEC. Infect Immun, 1999 Sep, 67(9), 4594 - 602 Murine splenocytes induce severe gastritis and delayed-type hypersensitivity and suppress bacterial colonization in Helicobacter pylori-infected SCID mice; Eaton KA et al.; The goal of this study was to evaluate the role of host immunity in gastritis and epithelial damage due to Helicobacter pylori . Splenocytes from H . pylori-infected and uninfected C57BL/6 mice were adoptively transferred to H . pylori-infected and uninfected severe combined immunodeficient (SCID) mice . Transfer was verified by flow cytometry, and all mice were evaluated for the presence of delayed-type hypersensitivity (DTH) by footpad inoculation with sterile H . pylori sonicate and for humoral immunity by enzyme-linked immunosorbent assay . The severity of gastritis and gastric epithelial damage was quantified histologically, epithelial proliferation was determined by proliferating cell nuclear antigen staining, and colonization was quantified by culture . C57BL/6 mice, but not nonrecipient SCID mice, developed moderate gastritis in response to H . pylori . In contrast, recipient SCID mice developed severe gastritis involving 50 to 100% of the gastric mucosa and strong DTH responses not present in C57BL/6 mice . DTH, but not serum anti-H . pylori immunoglobulin G, correlated with adoptive transfer, gastritis, and bacterial clearance . Severe gastritis, but not bacterial colonization, was associated with epithelial metaplasia, erosions, and an elevated labeling index . This study demonstrates that (i) adaptive immunity is essential for development of gastritis due to H . pylori in mice, (ii) T-cell-enriched lymphocytes in SCID mice induce DTH and gastritis, which is more severe than donor gastritis, and (iii) the host inflammatory response, not direct bacterial contact, causes epithelial damage . The greater severity of gastritis in recipient SCID mice than in donor C57BL/6 mice suggests that gastritis is due to specific T-cell subsets and/or the absence of regulatory cell subsets in the transferred splenocytes. J Exp Med, 1999 Aug 16, 190(4), 523 - 34 Transport of bacterial lipopolysaccharide to the golgi apparatus; Thieblemont N et al.; Addition of lipopolysaccharide (LPS) to cells in the form of LPS-soluble (s)CD14 complexes induces strong cellular responses . During this process, LPS is delivered from sCD14 to the plasma membrane, and the cell-associated LPS is then rapidly transported to an intracellular site . This transport appears to be important for certain cellular responses to LPS, as drugs that block transport also inhibit signaling and cells from LPS-hyporesponsive C3H/HeJ mice fail to exhibit this transport . To identify the intracellular destination of fluorescently labeled LPS after its delivery from sCD14 into cells, we have made simultaneous observations of different organelles using fluorescent vital dyes or probes . Endosomes, lysosomes, the endoplasmic reticulum, and the Golgi apparatus were labeled using Texas red (TR)-dextran, LysoTrackertrade mark Red DND-99, DiOC6(3), and boron dipyrromethane (BODIPY)-ceramide, respectively . After 30 min, LPS did not colocalize with endosomes, lysosomes, or endoplasmic reticulum in polymorphonuclear leukocytes, although some LPS-positive vesicles overlapped with the endosomal marker, fluorescent dextran . On the other hand, LPS did appear to colocalize with two markers of the Golgi apparatus, BODIPY-ceramide and TRITC (tetramethylrhodamine isothiocyanate)-labeled cholera toxin B subunit . We further confirmed the localization of LPS in the Golgi apparatus using an epithelial cell line, HeLa, which responds to LPS-sCD14 complexes in a CD14-dependent fashion: BODIPY-LPS was internalized and colocalized with fluorescently labeled Golgi apparatus probes in live HeLa cells . Morphological disruption of the Golgi apparatus in brefeldin A-treated HeLa cells caused intracellular redistribution of fluorescent LPS . These results are consistent with the Golgi apparatus being the primary delivery site of monomeric LPS. Curr Opin Struct Biol, 1999 Aug, 9(4), 455 - 61 Beta-barrel proteins from bacterial outer membranes: structure, function and refolding; Buchanan SK; Recently solved outer membrane protein structures include the smallest and largest known beta-barrel structures, with functions distinct from the general and specific porins . Both protein expressed in outer membranes and protein deposited as cytoplasmic aggregates have been used for the structure determinations . As most beta-barrel proteins can be overexpressed in an aggregated form (inclusion bodies) and refolded to the native state, this provides an alternative to membrane-targeted expression strategies and yields sufficient quantities of protein for future structural studies. Infect Dis Obstet Gynecol, 1999, 7(4), 190 - 4 Bacterial isolates from patients with preterm labor with and without preterm rupture of the fetal membranes; Mikamo H et al.; OBJECTIVE: The aim of this study is to describe the bacterial flora of women in preterm labor with or without premature rupture of membranes . METHODS: Retrospective studies of 239 patients with preterm labor were performed . RESULTS: One hundred and twenty-three of 239 patients with preterm labor (51.5%) had bacterial vaginosis . Seventy of the 239 patients with preterm labor (29.3%) developed premature rupture of the membranes (preterm PROM) . Of the 70 patients with preterm PROM, 51 (72.9%) had bacterial vaginosis . Therefore, 51 of the 123 patients with bacterial vaginosis (41.5%) developed preterm PROM . An increased number of organisms detected from the vaginal discharge in patients with preterm labor was associated with preterm PROM by Cochran-Armitage test . An increased number of organisms detected from the vaginal discharge in patients with preterm labor complicated with bacterial vaginosis was significantly associated with preterm PROM by Cochran-Armitage test . CONCLUSIONS: In preterm labor, the number of different species detected in the vagina provide sensitive and specific prediction of preterm PROM in patients with preterm labor. Arch Intern Med, 1999 Aug 9-23, 159(15), 1807 - 10 Wegener granulomatosis simulating bacterial endocarditis; Anthony DD et al.; Cardiac involvement in Wegener granulomatosis is uncommon . We report a case of Wegener granulomatosis that presented as culture-negative endocarditis with aortic valvular vegetation . The clinical manifestations included gingival hyperplasia, gangrenous digital infarcts, mononeuritis multiplex, high fever, inflammatory arthritis, pansinusitis, splenic infarct, and aortic valvular vegetation, which underscore the difficulty of distinguishing systemic vasculitis from bacterial endocarditis . Contrary to the common notion that valvular vegetation is invariably associated with bacterial endocarditis, this case proves that such findings can occur in Wegener granulomatosis as well . Clinicians are guided toward early treatment with corticosteroids and cyclophosphamide to prevent fatal complications. Curr Opin Immunol, 1999 Aug, 11(4), 400 - 5 T cell responses to bacterial infection; Kerksiek KM et al.; Many exciting advances in our understanding of T cell mediated immunity to bacterial infection have occurred in the past several years . T cell responses have been more fully characterized, due in part to the development of MHC class I tetramers . The importance of cytokines and various effector molecules in defense against infection has come to light . Finally, intracellular bacteria are being exploited to deliver antigens and DNA in an effort to induce immunity to pathogens. Cancer Res, 1999 Aug 1, 59(15), 3641 - 5 Quinol-glutathione conjugate-induced mutation spectra in the supF gene replicated in human AD293 cells and bacterial MBL50 cells; Jeong JK et al.; Hydroquinone is a nephrocarcinogen in rats but generally tests negative in standard mutagenicity assays . However, 2,3,5-tris-(glutathion-S-yl)hydroquinone, a potent nephrotoxic metabolite of hydroquinone, and 2-bromo-bis-(glutathion-S-yl)hydroquinone, another cytotoxic quinol-glutathione (GSH) conjugate, cause extensive single strand breaks in DNA in a manner that is dependent on the formation of reactive oxygen species . We, therefore, investigated whether quinol-GSH conjugates have the potential to behave as genotoxicants . The shuttle vector pSP189, containing the supF gene, was treated with 2,3,5-tris-(glutathion-S-yl)hydroquinone and replicated in both human AD293 cells and Escherichia coli MBL50 cells . The mutation frequency increased 4.6- and 2.6-fold in human AD293 and bacterial MBL50 cells, respectively . Base substitutions were the major type of mutations, and they occurred predominantly at G:C sites in both cell types . A high frequency of deletions (30%), including < 10- and > 10-bp deletions, were observed in AD293-replicated plasmids . The most common types of mutations in AD293 cells were G:C to A:T transitions (33.8%) and G:C to T:A (29.4%) and G:C to C:G (19.1%) transversions . In MBL50 cells, the major mutations were G:C to T:A (33.8%) and G:C to C:G (31.3%) transversions and G:C to A:T transitions (27.5%) . The mutation spectra were similar to those reported for *OH-induced mutations, suggesting that *OH generated from polyphenolic-GSH conjugates not only plays a role in cytotoxicity but also provides a basis for their mutagenicity and carcinogenicity. J Microbiol Methods, 1999 Aug, 37(2), 139 - 54 Development of radiographic and microscopic techniques for the characterization of bacterial transport in intact sediment cores from Oyster, Virginia; Dong H et al.; The objective of this study was to ascertain the physical and mineralogical properties responsible for the retention of bacteria in subsurface sediments . The sediment core chosen for this study was a fine-grained, quartz-rich sand with minor amounts of Fe and Al hydroxides . A bacterial transport experiment was performed using an intact core collected from a recent excavation of the Butler's Bluff member of the Nassawadox formation in the borrow pit at Oyster, VA . and a 14C-labeled bacterial strain OYS2-A was se