Bacteria

Am J Trop Med Hyg, 1999 May, 60(5), 799 - 805
Survey of Bartonella species infecting intradomicillary animals in the Huayllacallán Valley, Ancash, Peru, a region endemic for human bartonellosis; Birtles RJ et al.; The natural cycle of Bartonella bacilliformis remains uncertain, and the suspected existence of animal reservoirs for the bacterium has never been convincingly demonstrated . We conducted a survey of Bartonella species infecting intradomicillary animals in a bartonellosis-endemic region of Peru, obtaining blood from 50 animals living in the homes of 11 families whose children had recently had bartonellosis . Bartonella-like bacteria were recovered from four of nine small rodents included in the study, but from none of the 41 domesticated animals . Identification and comparison of these isolates, and two Bartonella-like isolates obtained from Phyllotis mice in a different endemic region of Peru using serologic and genotypic methods indicated that although none were strains of B . bacilliformis, five were probably representatives of three previously unrecognized Bartonella species and one was a likely strain of the pathogenic species B . elizabethae.

Appl Microbiol Biotechnol, 1999 Apr, 51(4), 491 - 7
Characterisation of cellulose-binding proteins that are involved in the adhesion mechanism of Fibrobacter intestinalis DR7; Miron J et al.; Cellulose-binding proteins (CBP) isolated from cell envelopes of the cellulolytic bacterium Fibrobacter intestinalis strain DR7 were studied in order to investigate the adhesion mechanism . The proteins were examined for their reaction with antibodies that specifically block bacterial adhesion, response to glycosylation staining and monosaccharide composition . To this end, the effect of some monosaccharides (CBP components) on blocking of DR7 adhesion to cellulose was determined . Previous study had shown the occurrence of 16 CBP in the outer membrane and periplasm of DR7, of which 6 had endoglucanase activity (Miron and Forsberg 1998) . Data from the present study show that most of the 16 CBP of DR7, except for the 38-, 90- and 180-kDa proteins, are glycosylated . Rabbit antibodies that specifically block DR7 adhesion were prepared by affinity preabsorption of antiserum against wild-type DR7 with bacterial cells of its adherence-defective mutant (DR7-M) . The preabsorbed antibodies reacted positively in Western blotting with glycosylated CBP of 225, 200, 150, 70, 45 and < 38 kDa from the DR7 outer membrane, and reacted weakly with CBP of DR7-M . Modification of glycosidic residues attached to the CBP of DR7 by periodate oxidation prevented any reaction with the preabsorbed antibodies . Monosaccharide analysis by HPLC of isolated CBP from the outer membrane and periplasm of DR7 cells, showed that galactosamine, glucosamine, galacturonic acid, and glucuronic acid were the predominant monosaccharide components of CBP that can block the adhesion of DR7 cells to cellulose . It is suggested that some glycosylated residues of CBP may have a predominant role in the adhesion of DR7 to cellulose.

FEBS Lett, 1999 Apr 23, 449(2-3), 183 - 6
Uptake of N-acetyl-D-mannosamine: an essential intermediate in polysialic acid biosynthesis by Escherichia coli K92; Revilla-Nuin B et al.; The N-acetyl-D-mannosamine (ManNAc) transport system of Escherichia coli K92 was studied when this bacterium was grown in a chemically defined medium containing ManNAc as carbon source . Kinetic measurements were carried out in vivo at 37 degrees C in 25 mM phosphate buffer, pH 7.5 . Under these conditions, the uptake rate was linear for at least 15 min and the calculated Km for ManNAc was 280 microM . The transport system was strongly inhibited by sodium arsenate (97%), potassium cyanide (84%) and 2,4-dinitrophenol (88%) added at final concentrations of 1 mM (each) . Analysis of bacterial ManNAc phosphotransferase activity revealed in vitro ManNAc phosphorylation activity only when phosphoenolpyruvate was present . These results strongly support the notion that ManNAc uptake depends on a specific phosphotransferase system . Study of specificities showed that N-acetylglucosamine and mannosamine specifically inhibited the transport of ManNAc in this bacterium . Analysis of expression revealed that the ManNAc transport system was induced by ManNAc, glucosamine, galactosamine, mannosamine and mannose but not by N-acetylglucosamine or N-acetylgalactosamine . Moreover, ManNAc permease was subject to glucose repression and cAMP stimulation . Full induction of the ManNAc transport system required the simultaneous presence of both cAMP and ManNAc.

Can J Gastroenterol, 1999 Apr, 13(3), 251 - 5
Determinants of ethnic or geographical differences in infectivity and transmissibility of Helicobacter pylori; Fallone CA; The prevalence of Helicobacter pylori infection is variable in different countries . There are two distinct patterns of H pylori prevalence with respect to age depending on the geographical region studied . The first pattern is widespread infection early in childhood with elevated prevalence rates of close to 80% throughout adulthood, and the second is increasing prevalence with age . This variability in pattern suggests a difference in infectivity or transmissibility of H pylori infection . Potential determinants of these differences are reviewed including environmental, bacterial and host factors . The most important determinant is likely socioeconomic class, which affects living conditions and sanitation, thus altering exposure to the bacterium . Host factors also play a role, perhaps via host receptors for H pylori . Bacterial factors may also contribute, although compelling evidence is lacking.

Can J Gastroenterol, 1999 Apr, 13(3), 218 - 23
Helicobacter pylori and its genome: lessons from the treasure map; Taylor DE; The availability of the complete genome sequence of Helicobacter pylori 26695 has opened new avenues for research in the molecular biology of this gastric pathogen . The present review gives a general overview of H pylori obtained from the complete genome sequence and compares this with data previously obtained from cloning and functional studies of H pylori . The cagA pathogenicity island of 40 kilobases, which encodes a type IV secretion system, is discussed . The diversity of H pylori genomes is well known, yet new data indicate that some aspects of the genome, particularly outer membrane protein genes, are conserved . Genes encoding proteins involved in molecular mimicry between bacterium and gastric epithelial tissue, specifically those encoding Lewis X and Lewis Y antigens, are discussed . The large number of DNA restriction and modification genes and their role in H pylori infection are considered . Finally, gene transfer is discussed . The availability of the complete genome sequence of H pylori 26695 and the soon to be available sequence of J99 will speed up and assist in the analysis of H pylori genes and their encoded proteins . The genomes of both strains will be useful as references with which other H pylori genomes can be compared.

J Mol Biol, 1999 May 14, 288(4), 753 - 63
The hyperthermostable indoleglycerol phosphate synthase from Thermotoga maritima is destabilized by mutational disruption of two solvent-exposed salt bridges; Merz A et al.; The recombinantly expressed protein indoleglycerol phosphate synthase from the hyperthermophilic bacterium Thermotoga maritima (tIGPS) was purified and characterized with respect to oligomerization state, catalytic properties and thermostability . This enzyme from the biosynthetic pathway of tryptophan is a monomer in solution . In contrast to IGPS from the hyperthermophilic archaeon Sulfolobus solfataricus, tIGPS shows high catalytic activity at room temperature and only weak product inhibition . In order to test the hypothesis that salt bridges in a critical context contribute to the high thermostability of tIGPS, two solvent-exposed salt bridges were selected, based on its three-dimensional structure, for individual disruption by site-directed mutagenesis . The first salt bridge fixes the N terminus to the core of the protein, and the second serves as a clamp between helices alpha1 and alpha8, which are widely separated in sequence but adjacent in the (betaalpha)8-barrel . Kinetics of irreversible heat inactivation reveal that the salt bridge crosslinking helices alpha1 and alpha8 stabilizes tIGPS more strongly than that tethering the N terminus .

J Bacteriol, 1999 May, 181(10), 3238 - 41
Rickettsia prowazekii transports UMP and GMP, but not CMP, as building blocks for RNA synthesis; Winkler HH et al.; Rickettsia prowazekii, the etiological agent of epidemic typhus, is an obligate intracellular bacterium and is apparently unable to synthesize ribonucleotides de novo . Here, we show that as an alternative, isolated, purified R . prowazekii organisms transported exogenous uridyl- and guanylribonucleotides and incorporated these labeled precursors into their RNA in a rifampin-sensitive manner . Transport systems for nucleotides, which we have shown previously and show here are present in rickettsiae, have never been reported in free-living bacteria, and the usual nucleobase and nucleoside transport systems are absent in rickettsiae . There was a clear preference for the monophosphate form of ribonucleotides as the transported substrate . In contrast, rickettsiae did not transport cytidylribonucleotides . The source of rickettsial CTP appears to be the transport of UMP followed by its phosphorylation and the amination of intrarickettsial UTP to CTP by CTP synthetase . A complete schema of nucleotide metabolism in rickettsiae is presented that is based on a combination of biochemical, physiological, and genetic information.

J Bacteriol, 1999 May, 181(10), 3033 - 8
The A modules of the Azotobacter vinelandii mannuronan-C-5-epimerase AlgE1 are sufficient for both epimerization and binding of Ca2+; Ertesvag H et al.; The industrially important polysaccharide alginate is composed of the two sugar monomers beta-D-mannuronic acid (M) and its epimer alpha-L-guluronic acid (G) . In the bacterium Azotobacter vinelandii, the G residues originate from a polymer-level reaction catalyzed by one periplasmic and at least five secreted mannuronan C-5-epimerases . The secreted enzymes are composed of repeats of two protein modules designated A (385 amino acids) and R (153 amino acids) . The modular structure of one of the epimerases, AlgE1, is A1R1R2R3A2R4 . This enzyme has two catalytic sites for epimerization, each site introducing a different G distribution pattern, and in this article we report the DNA-level construction of a variety of truncated forms of the enzyme . Analyses of the properties of the corresponding proteins showed that an A module alone is sufficient for epimerization and that A1 catalyzed the formation of contiguous stretches of G residues in the polymer, while A2 introduces single G residues . These differences are predicted to strongly affect the physical and immunological properties of the reaction product . The epimerization reaction is Ca2+ dependent, and direct binding studies showed that both the A and R modules bind this cation . The R modules appeared to reduce the Ca2+ concentration needed for full activity and also stimulated the reaction rate when positioned both N and C terminally.

Int J Syst Bacteriol, 1999 Apr, 49 Pt 2, 859 - 65
Desulfovibrio zosterae sp . nov., a new sulfate reducer isolated from surface-sterilized roots of the seagrass Zostera marina; Nielsen JT et al.; A sulfate-reducing bacterium, designated strain lacT, was isolated from surface-sterilized roots of the benthic macrophyte Zostera marina . Cells were motile by means of a single polar flagellum . Strain lacT utilized lactate, pyruvate, malate, ethanol, L-alanine, fumarate, choline and fructose with sulfate as electron acceptor . In addition, fumarate, pyruvate and fructose were also degraded without an external electron acceptor . Sulfate could be substituted with thiosulfate, sulfite and elemental sulfur . Optimal growth was observed between 32.5 and 34.5 degrees C, at an NaCl concentration of 0.2 M and in a pH range between 6.8 and 7.3 . The G + C content of the DNA was 42.7 +/- 0.2 mol% . Desulfoviridin and catalase were present . Strain lacT contained c-type cytochromes . Comparative 16S rRNA gene sequence analysis and the fatty acid pattern grouped this isolate into the genus Desulfovibrio . However, strain lacT differs from all other described Desulfovibrio species on the bases of its 16S rRNA gene sequence, the G + C content, its cellular lipid pattern and the utilization pattern of substrates . These characteristics establish strain lacT (= DSM 11974T) as a novel species of the genus Desulfovibrio, for which the name Desulfovibrio zosterae sp . nov . is proposed.

Int J Syst Bacteriol, 1999 Apr, 49 Pt 2, 815 - 20
Simkania negevensis strain ZT: growth, antigenic and genome characteristics; Kahane S et al.; Simkania negevensis is the type species of Simkaniaceae, a recently proposed family in the order Chlamydiales . In the current study, growth, antigenic and genomic characteristics of this intracellular bacterium were investigated and compared to those of members of the family Chlamydiaceae . Growth of the organism, as assessed by infectivity assays, reached a plateau in 2-3 d although by light microscopy the cytopathic effect on the host cells increased for 12 or more days after infection . S . negevensis growth was unaffected by sulfadiazine . Cells infected by S . negevensis strain ZT were not recognized by either of two monoclonal antibodies specific for Chlamydiaceae LPS and several specific Chlamydiaceae ompA primers were unable to PCR amplify a S . negevensis gene . The S . negevensis genome contained one copy of the ribosomal operon . The genome size of S . negevensis strain ZT was determined by PFGE to be 1.7 Mbp, and the G + C content was 42.5 mol% . These data, taken together with other published data, are consistent with the proposal that S . negevensis belongs to a distinct family in the order Chlamydiales.

Int J Syst Bacteriol, 1999 Apr, 49 Pt 2, 503 - 6
Abiotrophia balaenopterae sp . nov., isolated from the minke whale (Balaenoptera acutorostrata); Lawson PA et al.; Phenotypic and phylogenetic studies were performed on a hitherto undescribed micro-organism isolated from a minke whale (Balaenoptera acutorostrata) . Comparative 16S rRNA gene sequencing studies demonstrated that the unknown strain constituted a new subline close to, but distinct from, Abiotrophia adiacens and Abiotrophia elegans . The unknown bacterium was readily distinguished from these two Abiotrophia species by biochemical tests and electrophoretic analysis of whole-cell proteins . On the basis of phylogenetic and phenotypic evidence, it is proposed that the unknown bacterium be classified as Abiotrophia balaenopterae sp . nov., the type strain of which is M1975/96/1T (= CCUG 37380T).

Amino Acids, 1999, 16(2), 113 - 31
Molecular cloning and expression analyses of mitochondrial and plastidic isoforms of cysteine synthase (O-acetylserine(thiol)lyase) from Arabidopsis thaliana; Hesse H et al.; Cysteine synthase, the key enzyme for fixation of inorganic sulfide, catalyses the formation of cysteine from O-acetylserine and inorganic sulfide . Here we report the cloning of cDNAs encoding cysteine synthase isoforms from Arabidopsis thaliana . The isolated cDNA clones encode for a mitochondrial and a plastidic isoform of cysteine synthase (O-acetylserine (thiol)-lyase, EC 4.2.99.8), designated cysteine synthase C (AtCS-C, CSase C) and B (AtCS-B; CSase B), respectively . AtCS-C and AtCS-B, having lengths of 1569-bp and 1421-bp, respectively, encode polypeptides of 430 amino acids (approximately 45.8 kD) and of 392 amino acids (approximately 41.8 kD), respectively . The deduced amino acid sequences of the mitochondrial and plastidic isoforms exhibit high homology even with respect to the presequences . The predicted presequence of AtCS-C has a N-terminal extension of 33 amino acids when compared to the plastidic isoform . Northern blot analysis showed that AtCS-C is higher expressed in roots than in leaves whereas the expression of AtCS-B is stronger in leaves . Furthermore, gene expression of both genes was enhanced by sulfur limitation which in turn led to an increase in enzyme activity in crude extracts of plants . Expression of the AtCS-B gene is regulated by light . The mitochondrial, plastidic and cytosolic (Hesse and Altmann, 1995) isoforms of cysteine synthase of Arabidopsis are able to complement a cysteine synthase-deficient mutant of Escherichia coli unable to grow on minimal medium without cysteine, indicating synthesis of functional plant proteins in the bacterium . Two lines of evidence proved that AtCS-C encodes a mitochondrial form of cysteine synthase; first, import of in vitro translation products derived from AtCS-C in isolated intact mitochondria and second, Western blot analysis of mitochondria isolated from transgenic tobacco plants expressing AtCS-C cDNA/c-myc DNA fusion protein.

FEMS Microbiol Lett, 1999 May 1, 174(1), 173 - 8
Isolation of a psychrotrophic Azospirillum sp . and characterization of its extracellular protease; Oh KH et al.; A novel psychrotrophic bacterium secreting a protease was isolated from a mountain soil in Korea . On the basis of a 16S rDNA sequence analysis and physiological properties, the isolate was identified as an Azospirillum sp . The protease purified from the culture supernatant was a monomer in its native form with an apparent molecular mass of 48.6 kDa on SDS-PAGE . The protease was active in a broad pH range around 8.5 and at temperatures up to 40 degrees C and stable at temperatures below 30 degrees C for 3 days . The proteolytic activity was inhibited by iodoacetamide and EDTA . The Mg2+ ion did not activate the enzyme much but reversed the inhibition by EDTA, suggesting that the protease belongs to a cysteine protease stabilized by the Mg2+ ion.

Aliment Pharmacol Ther, 1999 May, 13(5), 643 - 9
Head-to-head comparison of 1-week triple regimens combining ranitidine or omeprazole with two antibiotics to eradicate Helicobacter pylori; Savarino V et al.; BACKGROUND: Triple therapies containing omeprazole and ranitidine have been shown to be equivalent in eradicating H . pylori infection, but have been assessed either separately or head-to-head, only in small trials . AIM: To carry out a large randomized controlled study comparing omeprazole and ranitidine combined with two antibiotic combinations for 1 week . METHODS: Three hundred and twenty H . pylori-positive patients were randomly subdivided into four equal-sized groups and received one of the following treatments: OAM = omeprazole 20 mg b.d . + amoxycillin 1 g b.d . + metronidazole 500 mg b.d.; RAM = ranitidine 300 mg b.d . + amoxycillin 1 g b.d . + metronidazole 500 mg b.d.; OAC = omeprazole 20 mg b.d . + amoxycillin 1 g b.d . + clarithromycin 250 mg t.d.s.; RAC = ranitidine 300 mg b.d . + amoxycillin 1 g b.d . + clarithromycin 250 mg t.d.s . The assessment of H . pylori status was performed before and 4 weeks after the end of therapy by means of CLO-test and histology . H . pylori infection was considered to be eradicated when both tests were negative . RESULTS: OAM and RAM eradicated H . pylori in 89% and 85% of cases on per protocol (P = 0.48) and in 77% and 75% of cases on intention-to-treat analyses (P = 0.71) . OAC and RAC eradicated H . pylori in 67% and 70% of cases on per protocol (P = 0.68) and in 57% and 64% of cases on intention-to-treat analyses (P = 0.41) . In contrast, there was significant difference between OAM and OAC (P<0.01) and between RAM and RAC (P<0.05) . Side-effects occurred in 15%, 10%, 17% and 16% of patients with respect to the above four subgroups . CONCLUSIONS: Omeprazole and ranitidine combined with two antibiotics for 1 week are equally effective in the eradication of H . pylori infection, and these results question the role of profound acid suppression in the eradication of the bacterium.

Scand J Gastroenterol, 1999 Mar, 34(3), 238 - 43
The therapeutic effect of bovine lactoferrin in the host infected with Helicobacter pylori; Wada T et al.; BACKGROUND: It remains unclarified whether bovine lactoferrin (bLF) can exert a therapeutic effect on the host infected with Helicobacter pylori . METHODS: Germfree BALB/c mice were orally inoculated with H . pylori to induce infection . Three weeks after infection the mice were given bLF orally once daily for 2 or 4 weeks and were then killed to examine the bacterial number in the stomach and the serum antibody titer to H . pylori . To count the number of epithelium-bound H . pylori, the resected stomach was agitated in phosphate-buffered saline to remove non-bound H . pylori before bacterial enumeration . RESULTS: The administration of 10 mg bLF for 3 to 4 weeks decreased the number of H . pylori in the stomach to one-tenth and also exerted a significant inhibitory effect on the attachment of H . pylori to the stomach . As a result, the serum antibody titer to H . pylori, whose level is presumed to represent the size of the immune response by the host, thereby reflecting the degree of bacterial attack, decreased to an undetectable level . CONCLUSIONS: These findings suggest that bLF exerts an inhibitory effect on colonizing H . pylori by detaching the bacterium from the gastric epithelium and by exerting a direct anti-bacterial effect.

J Mol Evol, 1999 Jun, 48(6), 717 - 22
Intracellular bacterial symbionts of aphids possess many genomic copies per bacterium; Komaki K et al.; Although Buchnera, the endosymbiotic bacteria of aphids, are close relatives of Escherichia coli, their genome size is only a seventh that of E . coli . In this study, we estimated the genomic copy number of Buchnera by dot-blot hybridization and fluorimetry using a video-intensified microscope photon-counting system and obtained convincing evidence that each cell of these bacteria contains an average of 120 genomic copies . Thus, the Buchnera symbiont, with many copies of a small-sized genome, is reminiscent of cell organelles such as mitochondria and chloroplasts.

Hepatogastroenterology, 1999 Jan-Feb, 46(25), 308 - 11
The effect on survival of thoracic duct ligation in experimental peritonitis; Aydin M et al.; BACKGROUND/AIMS: It has been shown that systemic bacteremia and endotoxemia in peritonitis is mainly related to lymphatic transport via the thoracic duct . This study was performed to investigate the effect on mortality of thoracic duct ligation in experimental peritonitis . METHODOLOGY: Thirty dogs were divided into three groups . Groups I, II, and III were control, unligated, and ligated thoracic duct peritonitis groups, respectively . Liver biopsy, blood and peritoneal fluid cultures were taken and survival time was established . RESULTS: Bacteria were determined in peritoneal fluid in all animals in groups II and III . Growing bacteria numbers in group III were two times higher than in group II . While bacterium was grown on blood cultures in all group II animals, growing was determined on blood cultures in only 2 animals in group III . Diffuse necrosis was determined in the liver of 2 animals who died within 72 hours in group II . Another 8 animals had minimal focal necrosis in their livers . Diffuse and progressive necrosis was determined in the liver of all animals in group III . The difference between liver necrosis in group II and group III was found to be statistically significant (p = 0.002) . CONCLUSIONS: This experimental study demonstrates that thoracic duct ligation decreases bacteremia rates clearly but that mortality increases significantly.

Infect Immun, 1999 May, 67(5), 2258 - 65
Ehrlichia chaffeensis and E . sennetsu, but not the human granulocytic ehrlichiosis agent, colocalize with transferrin receptor and up-regulate transferrin receptor mRNA by activating iron-responsive protein 1; Barnewall RE et al.; Ehrlichia chaffeensis and E . sennetsu are genetically divergent obligatory intracellular bacteria of human monocytes and macrophages, and the human granulocytic ehrlichiosis (HGE) agent is an obligatory intracellular bacterium of granulocytes . Infection with both E . chaffeensis and E . sennetsu, but not HGE agent, in the acute monocytic leukemia cell line THP-1 almost completely inhibited by treatment with deferoxamine, a cell-permeable iron chelator . Transferrin receptors (TfRs) accumulated on both E . chaffeensis and E . sennetsu, but not HGE agent, inclusions in THP-1 cells or the cells of the promyelocytic leukemia cell line HL-60 . Reverse transcription-PCR showed an increase in the level of TfR mRNA 6 h postinfection which peaked at 24 h postinfection with both E . chaffeensis and E . sennetsu infection in THP-1 or HL-60 cells . In contrast, HGE agent in THP-1 or HL-60 cells induced no increase in TfR mRNA levels . Heat treatment of E . chaffeensis or the addition of monodansylcadaverine, a transglutaminase inhibitor, 3 h prior to infection inhibited the up-regulation of TfR mRNA . The addition of oxytetracycline 6 h after E . chaffeensis infection caused a decrease in TfR mRNA which returned to the basal level by 24 h postinfection . These results indicate that both internalization and continuous proliferation of ehrlichial organisms or the production of ehrlichial proteins are required for the up-regulation of TfR mRNA . Results of electrophoretic mobility shift assays showed that both E . chaffeensis and E . sennetsu infection increased the binding activity of iron-responsive protein 1 (IRP-1) to the iron-responsive element at 6 h postinfection and remained elevated at 24 h postinfection . However, HGE agent infection had no effect on IRP-1 binding activity . This result suggests that activation of IRP-1 and subsequent stabilization of TfR mRNA comprise the mechanism of TfR mRNA up-regulation by E . chaffeensis and E . sennetsu infection.

Appl Environ Microbiol, 1999 May, 65(5), 1941 - 8
Interactions between carbon and nitrogen metabolism in Fibrobacter succinogenes S85: a 1H and 13C nuclear magnetic resonance and enzymatic study; Matheron C et al.; The effect of the presence of ammonia on {1-13C}glucose metabolism in the rumen fibrolytic bacterium Fibrobacter succinogenes S85 was studied by 13C and 1H nuclear magnetic resonance (NMR) . Ammonia halved the level of glycogen storage and increased the rate of glucose conversion into acetate and succinate 2.2-fold and 1.4-fold, respectively, reducing the succinate-to-acetate ratio . The 13C enrichment of succinate and acetate was precisely quantified by 13C-filtered spin-echo difference 1H-NMR spectroscopy . The presence of ammonia did not modify the 13C enrichment of succinate C-2 (without ammonia, 20.8%, and with ammonia, 21.6%), indicating that the isotopic dilution of metabolites due to utilization of endogenous glycogen was not affected . In contrast, the presence of ammonia markedly decreased the 13C enrichment of acetate C-2 (from 40 to 31%), reflecting enhanced reversal of the succinate synthesis pathway . The reversal of glycolysis was unaffected by the presence of ammonia as shown by 13C-NMR analysis . Study of cell extracts showed that the main pathways of ammonia assimilation in F . succinogenes were glutamate dehydrogenase and alanine dehydrogenase . Glutamine synthetase activity was not detected . Glutamate dehydrogenase was active with both NAD and NADP as cofactors and was not repressed under ammonia limitation in the culture . Glutamate-pyruvate and glutamate-oxaloacetate transaminase activities were evidenced by spectrophotometry and 1H NMR . When cells were incubated in vivo with {1-13C}glucose, only 13C-labeled aspartate, glutamate, alanine, and valine were detected . Their labelings were consistent with the proposed amino acid synthesis pathway and with the reversal of the succinate synthesis pathway.

FEMS Microbiol Lett, 1999 Apr 1, 173(1), 1 - 8
Endosymbiosis in protozoa of the Trypanosomatidae family; de Souza W et al.; A small number of trypanosomatids present bacterium endosymbionts in the cytoplasm, which divide synchronously with the host cell . Crithidia oncopleti, Crithidia deanei . Crithidia desouzai, Blastocrithidia culicis and Herpetomonas roitmani are the best characterized species . The endosymbiont is surrounded by two membranes separated from each other by an electron-lucent space . The presence of the endosymbiont led to the appearance of morphological changes which include the lack of the paraflagellar rod associated to the axoneme, the morphology of the kinetoplast and the association of the sub-pellicular microtubules with portions of the protozoan plasma membrane . Aposymbiotic strains could be obtained by antibiotic treatment, opening the possibility to make comparative analysis of endosymbiont-containing an endosymbiont-free populations of the same species . It is clear that metabolic cycles are established between the prokaryiont and the host cell . The results obtained show that endosymbiont-containing species of trypanosomatids constitute an excellent model to study basic processes on the endosymbiont-host cell relationship and the origin of new organelles.

Proc Natl Acad Sci U S A, 1999 Apr 27, 96(9), 4924 - 9
Energy transduction in the sodium F-ATPase of Propionigenium modestum; Dimroth P et al.; The F-ATPase of the bacterium Propionigenium modestum is driven by an electrochemical sodium gradient between the cell interior and its environment . Here we present a mechanochemical model for the transduction of transmembrane sodium-motive force into rotary torque . The same mechanism is likely to operate in other F-ATPases, including the proton-driven F-ATPases of Escherichia coli.

Microbiology, 1999 Apr, 145 ( Pt 4), 925 - 34
Multiple genes involved in chitin degradation from the marine bacterium Pseudoalteromonas sp . strain S91; Techkarnjanaruk S et al.; A cluster of three closely linked chitinase genes organized in the order chiA, chiB and chiC, with the same transcriptional direction, and two unlinked genes, chiP and chiQ, involved in chitin degradation in Pseudoalteromnas sp . strain S91 were cloned, sequenced and characterized . The deduced amino acid sequences revealed that ChiA, ChiB and ChiC exhibited similarities to chitinases belonging to family 18 of the glycosyl hydrolases while ChiP and ChiQ belonged to family 20 . ChiP and ChiQ showed different enzymic activities against fluorescent chitin analogues, but neither was able to degrade colloidal chitin . ChiA possessed chitinase activity but did not bind chitin; ChiB bound chitin but had no chitinase activity; ChiC possessed strong chitinase activity and also bound chitin . Production of ChiC in S91 appeared to be controlled by chiA expression, since insertion of a transposon into the ORF of chiA resulted in the loss of chitinase activity as well as loss of ChiC proteins in a chitinase-negative mutant . In Escherichia coli, ChiC appeared to be expressed from its own promoter.

Microbiology, 1999 Apr, 145 ( Pt 4), 899 - 904
Genotypic analysis of Mycobacterium tuberculosis from medieval human remains; Taylor GM et al.; Three medieval bone samples with osteological evidence of tuberculosis infection were analysed for the presence of DNA sequences from Mycobacterium tuberculosis using a series of PCRs . In each case amplification of IS6110 and part of the beta-subunit of RNA polymerase identified infection with a bacterium belonging to the M . tuberculosis complex . Amplification of the mtp40 genome fragment and the presence of a guanine residue at position 285 in the oxyR pseudogene, demonstrated the infecting strain to be similar to present day M . tuberculosis isolates rather than to Mycobacterium bovis . Spoligotyping, based on amplification of the direct repeat (DR) region of the mycobacterial genome, provided further evidence of similarity to M . tuberculosis and indicated a close relationship between isolates associated with two separate medieval burials . The study demonstrates the feasibility of amplifying multiple M . tuberculosis loci in ancient human remains and suggests important applications in the study of the palaeoepidemiology and virulence of tuberculosis in past populations.

FEBS Lett, 1999 Mar 19, 447(1), 99 - 105
Re-evaluation of the primary structure of Ralstonia eutropha phasin and implications for polyhydroxyalkanoic acid granule binding; Hanley SZ et al.; Sequence analysis of several cDNAs encoding the phasin protein of Ralstonia eutropha indicated that the carboxyl terminus of the resulting derived protein sequence is different from that reported previously . This was confirmed by: (1) sequencing of the genomic DNA; (2) SDS-PAGE and peptide analysis of wild-type and recombinant phasin; and (3) mass spectrometry of wild-type phasin protein . The results have implications for the model proposed for the binding of this protein to polyhydroxyalkanoic acid granules in the bacterium.

Trends Microbiol, 1999 Apr, 7(4), 149 - 54
Developmental biology of Coxiella burnettii; Heinzen RA et al.; The obligate intracellular bacterial agent of human Q fever, Coxiella burnetii, has a remarkable ability to persist in the extracellular environment . It replicates only when phagocytosed and delivered to the phagolysosome, where it resists degradation . Different morphological forms of the bacterium have different resistance properties and appear to be stages of a developmental cycle . Despite the lack of genetic systems, the molecular events surrounding C . burnetii development are now being unraveled.

J Bacteriol, 1999 May, 181(9), 2689 - 96
Molecular and functional characterization of the Rhodopseudomonas palustris no . 7 phosphoenolpyruvate carboxykinase gene; Inui M et al.; The pckA gene, encoding the gluconeogenic enzyme phosphoenolpyruvate carboxykinase (PEPCK), was cloned by PCR amplification from the purple nonsulfur bacterium Rhodopseudomonas palustris No . 7 . Sequencing of a 2.5-kb chromosomal SmaI-PstI fragment containing the structural gene revealed an open reading frame encoding 537 amino acids, homologous to known pckA genes . Primer extension analysis identified a transcriptional start site 72 bp upstream of the pckA initiation codon and an upstream sequence similar to sigma70 promoters . Studies of a pckA-lacZ gene fusion indicated that when cells were grown in minimal media with various carbon sources, such as succinate, malate, pyruvate, lactate, or ethanol, under both anaerobic light and aerobic dark conditions, the pckA gene was induced in log phase, irrespective of the carbon source . A R . palustris No . 7 PEPCK-deficient strain showed growth characteristics identical to those of the wild-type strain either anaerobically in the light or aerobically in the dark when a C4-dicarboxylic acid, such as succinate or malate, was used as a carbon source . These results indicate that in R . palustris No . 7, an alternative gluconeogenic pathway may exist in addition to PEPCK.

J Nat Prod, 1999 Apr, 62(4), 608 - 10
Luisols A and B, new aromatic tetraols produced by an estuarine marine bacterium of the genus Streptomyces (Actinomycetales); Cheng XC et al.; Luisols A (1) and B (2), two new aromatic tetraols, have been isolated from the cultivation broth of an estuarine marine actinomycete of the genus Streptomyces (strain #CNH-370) . The structures of luisols A and B were assigned by combined spectroscopic methods, including extensive 2D NMR experiments . Luisol A appears related to the anthraquinone antibiotics of the granaticin class, while the structure of luisol B contains the rare epoxynaphtho{2,3c}furan, a structural feature found in only one natural product, the fungal metabolite anthrinone.

J Nat Prod, 1999 Apr, 62(4), 605 - 7
Arenaric acid, a new pentacyclic polyether produced by a marine bacterium (Actinomycetales); Cheng XC et al.; Arenaric acid (1a), a new pentacyclic polyether related to the antibiotics K-41A and oxolonomycin, was isolated as its sodium salt (1b) from the culture broth of an estuarine bacterial isolate of the genus Streptomyces . The structure of arenaric acid was established by spectroscopic methods involving comprehensive 2D NMR measurements.

Zentralbl Veterinarmed B, 1999 Mar, 46(2), 73 - 84
Isolation and purification of a protective protein antigen of Erysipelothrix rhusiopathiae; Sato H et al.; The rapid growth and high survival rate of Erysipelothrix rhusiopathiae was determined using a culture of the bacterium in tryptic soy broth supplemented with 0.3% Tris-hydroxymethyl aminomethane and 0.1% Tween 80 (TT-TS broth) . High concentrations of 64, 66 and 43 kDa proteins, which are associated with protection against E . rhusiopathiae infection in mice, were obtained by alkaline treatment of whole cells using 0.05-1 N NaOH . The supernatant of alkaline treated cells (alkaline extract; AE) was stable at alkaline or neutral pH . However, aggregates appeared at neutral pH in the absence of sodium dodecyl sulphate (SDS) . A high yield of 64, 66 and 43 kDa proteins was obtained from strain Agata (serovar 5) . The proteins were eluted from gel bands following SDS-polyacrylamide gel electrophoresis (SDS-PAGE) of the AE from strain Agata and designated P64 and P43 . The amounts of P64 and P43 isolated were 0.7 and 0.3 mg/16 g of wet bacteria, respectively . In a mouse protection test, 50% protective doses (PD50) of P64 and P43 were 0.58 and 0.63 microgram, respectively . Upon Western blotting of the AE, both anti-P64 and anti-P43 antibodies reacted with the 64 and 43 kDa proteins . From these results, it is suggested that P64 is the most effective protective antigen and that P43 (43 kDa protein) is a degradation product of P64 . Therefore, the 64 kDa structural proteins are associated with the induction of a protective activity against E . rhusiopathiae infection in mice.

Eur J Biochem, 1999 Apr, 261(2), 438 - 43
The superoxide dismutase activity of desulfoferrodoxin from Desulfovibrio desulfuricans ATCC 27774; Romao CV et al.; Desulfoferrodoxin (Dfx), a small iron protein containing two mononuclear iron centres (designated centre I and II), was shown to complement superoxide dismutase (SOD) deficient mutants of Escherichia coli {Pianzzola, M.J., Soubes M . & Touati, D . (1996) J . Bacteriol . 178, 6736-6742} . Furthermore, neelaredoxin, a protein from Desulfovibrio gigas containing an iron site similar to centre II of Dfx, was recently shown to have a significant SOD activity {Silva, G., Oliveira, S., Gomes, C.M., Pacheco, I., Liu, M.Y., Xavier, A.V., Teixeira, M., Le Gall, J . & Rodrigues-Pousada, C . (1999) Eur . J . Biochem . 259, 235-243} . Thus, the SOD activity of Dfx isolated from the sulphate-reducing bacterium Desulfovibrio desulfuricans ATCC 27774 was studied . The protein exhibits a SOD activity of 70 U x mg-1, which increases approximately 2.5-fold upon incubation with cyanide . Cyanide binds specifically to Dfx centre II, yielding a low-spin iron species with g-values at 2.27 (g perpendicular) and 1.96 (g parallel) . Upon reaction of fully oxidized Dfx with the superoxide generating system xanthine/xanthine oxidase, Dfx centres I and II become partially reduced, suggesting that Dfx operates by a redox cycling mechanism, similar to those proposed for other SODs . Evidence for another SOD in D . desulfuricans is also presented - this enzyme is inhibited by cyanide, and N-terminal sequence data strongly indicates that it is an analogue to Cu,Zn-SODs isolated from other sources . This is the first indication that a Cu-containing protein may be present in a sulphate-reducing bacterium.

Biochemistry, 1999 Apr 20, 38(16), 5185 - 90
Characterization of the different peripheral light-harvesting complexes from high- and low-light grown cells from Rhodopseudomonas palustris; Gall A et al.; In this paper we demonstrate that the spectroscopically different peripheral light-harvesting complexes from Rhodopseudomonas palustris, strain 2.6.1, isolated from high- and low-light grown cells have widely differing bacteriochlorophyll a (BChl a) resonance Raman spectra in the high-frequency carbonyl region (1550-1750 cm-1) . Complexes synthesized in low-light grown cells exhibit Raman spectra characteristic of B800-850 and B800-820 complexes, depending on the excitation conditions . The in vivo strategy for low-light adaptation in this bacterium is thus somewhat different from that generally encountered in the Rhodospirillaceae . In these bacteria, as typified by Rps . acidophila and Rps . cryptolactis, low-light conditions induce the synthesis of B800-820 only complexes in which the hydrogen bonds between the acetyl carbonyl and the B850 binding pocket are broken, inducing changes in the absorption properties of the monomeric bacteriochlorophylls . In the case of Rps . palustris, additional spectral effects occur due to the coupling of the electronic levels of the differently interacting dimers . The extensive use of differential alpha/beta-polypeptide expression {Tadros et al . (1993) Eur . J . Biochem . 217, 867-875} thus allows Rps . palustris to alter its BChl a binding site environments causing the observed spread of BChl a Qy transitions, ranging from 801 to 856 nm.

Lett Appl Microbiol, 1999 Apr, 28(4), 321 - 6
Streptomyces lividans as a host for the production and secretion of Escherichia coli TEM beta-lactamase; Isiegas C et al.; The regulatory region and the region coding for the signal peptide of an extracellular agarase have been used to synthesize and secrete the heterologous Escherichia coli TEM beta-lactamase in Streptomyces lividans . The transcriptional regulation of the chimeric gene, and the secretion pattern of the chimeric gene product, coincided with those of the agarase gene . The negative glucose effect on the secretion of the protein was reverted when the recombinant bacterium was grown in the chemostat under phosphate limiting conditions.

Lett Appl Microbiol, 1999 Apr, 28(4), 245 - 9
Influence of sample preparation technique on two-dimensional gel electrophoresis of proteins from Porphyromonas gingivalis; Pridmore AM et al.; Sample preparation methods were compared for two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) of cellular proteins from the proteolytic bacterium Porphyromonas gingivalis . Standard solubilization buffer yielded poorly resolved protein spots, but pre-treatment of cells with trichloroacetic acid or inclusion of the protease inhibitor TLCK during solubilization improved definition and separation . The latter approach allowed reliable detection of a 55 kDa immunodominant surface antigen by Western immunoblotting . Further improvements in resolution occurred when SDS was included in the sample preparation . Thus, controlling proteolysis and optimizing protein solubilization were essential for reproducible separations and maximal protein recovery during 2D-PAGE of P . gingivalis.

Jpn J Med Sci Biol, 1998, 51 Suppl, S91 - 100
Enteropathogenic E . coli interactions with host cells; Finlay BB et al.; Enteropathogenic E . coli (EPEC) interacts with intestinal epithelial cells, causing diarrhea and associated diseases . This pathogen binds to epithelial cells using sophisticated mechanisms that exploit existing epithelial signal transduction pathways and host cytoskeletal components, ultimately resulting in the bacterium resting upon a pedestal on host cell surfaces . Recent data indicates that similar mechanisms occur in vivo . EPEC interactions with host cells illustrate several principles of pathogenesis that are used by bacteria that interact with mammalian host cells.

Aliment Pharmacol Ther, 1999 Mar, 13 Suppl 1, 3 - 11
Review article: exploring the link between Helicobacter pylori and gastric cancer; Kuipers EJ; Cancer of the distal stomach, both of the intestinal and diffuse type, is strongly associated with Helicobacter pylori colonization . This bacterium causes chronic active inflammation of the gastric mucosa in the majority of colonized subjects . In a considerable number of them, this will eventually lead to a loss of gastric glands, and thus the establishment of atrophic gastritis, which is associated with the development of intestinal metaplasia and dysplasia . Development of atrophy and metaplasia of the gastric mucosa are thus strongly associated with H . pylori infection, instead of a direct and inevitable consequence of ageing . Approximately 40-50% of infected subjects develop these conditions, but they are rare in non-infected subjects . The presence of these consecutive disorders leads to a 5-90-fold increased risk for cancer of the distal stomach, in particular of the intestinal type . This sequence explains the increased risk for gastric cancer in H . pylori-infected subjects, as has been shown in various cross-sectional and longitudinal studies . In a combined analysis of three longitudinal studies, a significant trend was observed towards an increased odds ratio with longer intervals between (retrospective) serological diagnosis of H . pylori infection and observation of gastric cancer, this risk being more than eight-fold increased if the interval had been at least 15 years . This is thought to reflect development of atrophic gastritis and intestinal metaplasia with loss of H . pylori colonization in the years prior to development of cancer . Atrophic gastritis and gastric cancer thus appear closely associated with the presence of H . pylori, yet not all infected subjects will eventually develop atrophy and only a small minority develop gastric cancer . Factors that influence the risks for atrophy and cancer in the presence of infection may be related to the time that infection occurred and to characteristics of the bacterial strain and the host . Evidence for the role of these factors is now increasing . Recognition of the causal role of H . pylori in the induction of gastric cancer theoretically presents tools for cancer prevention . The efficacy of screening and bacterial eradication for prevention of distal gastric cancer is being studied in a number of large-scale intervention studies in different populations . It is hoped that these studies will also provide answers to the potential preventive role of H . pylori colonization in the development of gastro-oesophageal reflux disease and associated conditions, in particular development of cancer of the proximal stomach . Infection with H . pylori plays an important role in the aetiology of atrophic gastritis and gastric cancer . Studies suggest an eight-fold increased risk for both conditions in the presence of infection . Factors that influence the risk for both conditions in the presence of infection are the age at which infection occurred and the presence of cagA as a marker for more pathogenetic H . pylori strains . The efficacy and side-effects of intervention for the prevention of distal gastric cancer has yet to be established.

Biochim Biophys Acta, 1999 Apr 14, 1418(1), 245 - 50
Molecular cloning and sequencing of the cDNA for vacuolar H+-pyrophosphatase from Chara corallina1; Nakanishi Y et al.; We have cloned a cDNA for vacuolar proton-translocating pyrophosphatase of Chara corallina that is one of the closest green algae to the land plants . The deduced protein consists of 793 amino acid residues . Its sequence is 71% identical to the H+-pyrophosphatases of land plants, and is less than 46% identical to those of marine alga and phototrophic bacterium.

Acta Otorhinolaryngol Ital, 1998 Aug, 18(4 Suppl 59), 51 - 4
{Progressive sensorineural hearing loss from infectious agents}; Scasso CA et al.; The progressive sensorineural hearing loss due to infectious causes can involve different etiological agents like bacteria, viruses, protozoons or mycetes . These infectious agents can act in various ways: directly through a labyrinthitis that may destroy the neuroepithelium; through an ischaemic process secondary to a septic embolus; or through a thrombus . In some cases the damage can occur in a meningitis context, because of the passage of the germ in the inner through the nerves, the vases or the labyrinthine liquids . Bacterial meningitis is one of the causes of progressive sensorinueral hearing loss . Among bacteria, the Mycobacterium Tuberculosis has nowadays acquired a remarkable importance which is also due to its considerable diffusion, despite modern therapy, and to its association with HIV infection . Bacteria can also cause a labyrinthitis acting directly on the inner ear: among these, Treponemas Pallidum, a spirochaete which causes syphilis and Borrelia Burgdorferi, a spirochaete that causes Lyme Disease, must be mentioned . The viruses that are certainly involved in the etiology of progressive sensorineural hearing loss are Cytomegalovirus and Rubella virus . The virus usually causes a labyrinthitis after the viraemia, wich may be due to the passage of the virus from the blood to the endolymph, through the stria vascularis with the consequent infection of the sensorial cells of the organ of Corti . Less frequently the viral damage to the inner ear can occur after a vasculitis, a meningitis or an alteration of the cell-mediated immunity . Progressive sensorineural hearing loss can also occur because of some congenital viral infections such as those caused by Cytomegalovirus and Rubella virus . More recently even the Human Parvovirus B19 seems to have been involved . This virus seems to act through autoimmune and/or immunologic processes, like that causing sudden hearing loss in Lassa fever . Another viral infection which can nowadays more frequently be considered among the cause of progressive hearing loss is HIV . In the HIV infection the neurological toxic lesions due to the administered ototoxic drugs are added up to the damages caused by the opportunistic infectious agents (virus, bacterium, protozoon mycete) . However, in these patients HIV itself could be the cause of the auditory and vestibular lesions . More rarely, a progressive hearing loss may be due to the action of a protozoon or mycete only.

Nat Struct Biol, 1999 Apr, 6(4), 313 - 8
Structure of the cell-adhesion fragment of intimin from enteropathogenic Escherichia coli; Kelly G et al.; Enteropathogenic Escherichia coli (EPEC) induce gross cytoskeletal rearrangement within epithelial cells, immediately beneath the attached bacterium . The C-terminal 280 amino acid residues of intimin (Int280; 30.1 kDa), a bacterial cell-adhesion molecule, mediate the intimate bacterial host-cell interaction . Recently, interest in this process has been stimulated by the discovery that the bacterial intimin receptor protein (Tir) is translocated into the host cell membrane, phosphorylated, and after binding intimin triggers the intimate attachment . Using multidimensional nuclear magnetic resonance (NMR) and combining perdeuteration with site-specific protonation of methyl groups, we have determined the global fold of Int280 . This represents one of the largest, non-oligomeric protein structures to be determined by NMR that has not been previously resolved by X-ray crystallography . Int280 comprises three domains; two immunoglobulin-like domains and a C-type lectin-like module, which define a new family of bacterial adhesion molecules . These findings also imply that carbohydrate recognition may be important in intimin-mediated cell adhesion.

Mol Microbiol, 1999 Mar, 31(5), 1573 - 87
Inactivation of the antigen 85C gene profoundly affects the mycolate content and alters the permeability of the Mycobacterium tuberculosis cell envelope; Jackson M et al.; The antigen 85 complex of Mycobacterium tuberculosis consists of three abundantly secreted proteins . The recent characterization of a mycoloyltransferase activity associated in vitro with each of these antigens suggested that they are potentially important for the building of the unusual cell envelope of mycobacteria . To define the physiological role of these proteins, the gene coding for antigen 85C was inactivated by transposon mutagenesis . The resulting mutant was shown to transfer 40% fewer mycolates to the cell wall with no change in the types of mycolates esterifying arabinogalactan or in the composition of non-covalently linked mycolates . As a consequence, the diffusion of the hydrophobic chenodeoxycholate and the hydrophilic glycerol, but not that of isoniazid, was found to be much faster through the cell envelope of the mutant than that of the parent strain . Taken together, these data demonstrate that: (i) antigen 85C is involved directly or indirectly in the transfer of mycolates onto the cell wall of the whole bacterium; (ii) the enzyme is not specific for a given type of mycolate; and (iii) the cell wall-linked mycolate layer may represent a barrier for the diffusion of small hydrophobic and hydrophilic molecules.

J Bacteriol, 1999 Apr, 181(8), 2440 - 7
Azospirillum irakense produces a novel type of pectate lyase; Bekri MA et al.; The pelA gene from the N2-fixing plant-associated bacterium Azospirillum irakense, encoding a pectate lyase, was isolated by heterologous expression in Escherichia coli . Nucleotide sequence analysis of the region containing pelA indicated an open reading frame of 1,296 bp, coding for a preprotein of 432 amino acids with a typical amino-terminal signal peptide of 24 amino acids . N-terminal amino acid sequencing confirmed the processing of the protein in E . coli at the signal peptidase cleavage site predicted by nucleotide sequence analysis . Analysis of the amino acid sequence of PelA revealed no homology to other known pectinases, indicating that PelA belongs to a new pectate lyase family . PelA macerates potato tuber tissue, has an alkaline pH optimum, and requires Ca2+ for its activity . Of several divalent cations tested, none could substitute for Ca2+ . Methyl-esterified pectin (with a degree of esterification up to 93%) and polygalacturonate can be used as substrates . Characterization of the degradation products formed upon incubation with polygalacturonate indicated that PelA is an endo-pectate lyase generating unsaturated digalacturonide as the major end product . Regulation of pelA expression was studied by means of a translational pelA-gusA fusion . Transcription of this fusion is low under all growth conditions tested and is dependent on the growth phase . In addition, pelA expression was found to be induced by pectin . An A . irakense pelA::Tn5 mutant still displayed pectate lyase activity, suggesting the presence of multiple pectate lyase genes in A . irakense.

Lett Appl Microbiol, 1999 Mar, 28(3), 199 - 202
Actinorhodin production by Streptomyces coelicolor A3(2) in iron-restricted media; Coisne S et al.; Production of the polyketide antibiotic actinorhodin by Streptomyces coelicolor A3(2) was investigated using a defined medium with or without iron supplementation . Iron limitation was found to enhance the intracellular production and export of the pigmented antibiotic . The effect of iron deficiency was particularly pronounced when the bacterium was grown with nitrate instead of ammonium . Analysis of the excreted pigment led to the identification of the lactone form of actinorhodin, gamma-actinorhodin.

J Biol Chem, 1999 Apr 16, 274(16), 10795 - 801
A new cytochrome subunit bound to the photosynthetic reaction center in the purple bacterium, Rhodovulum sulfidophilum; Masuda S et al.; The nucleotide sequence of the puf operon, which contains the genes encoding the B870 light-harvesting protein and the reaction center complex of the purple photosynthetic bacterium, Rhodovulum sulfidophilum, was determined . The operon, which consisted of six genes, pufQ, pufB, pufA, pufL, pufM, and pufC, is a new variety in photosynthetic bacteria in the sense that pufQ and pufC coexist . The amino acid sequence of the cytochrome subunit of the reaction center deduced from the pufC sequence revealed that this cytochrome contains only three possible heme-binding motifs; the heme-1-binding motif of the corresponding tetraheme cytochrome subunits was not present . This is the first exception of the "tetraheme" cytochrome family in purple bacteria and green filamentous bacteria . The pufC sequence also revealed that the sixth axial ligands to heme-1 and heme-2 irons were not present in the cytochrome either . This cytochrome was actually detected in membrane preparation as a 43-kDa protein and shown to associate functionally with the photosynthetic reaction center as the immediate electron donor to the photo-oxidized special pair of bacteriochlorophyll . This new cytochrome should be useful for studies on the role of each heme in the cytochrome subunit of the bacterial reaction center and the evolution of proteins in photosynthetic electron transfer systems.

Biochemistry, 1999 Apr 6, 38(14), 4633 - 9
Thermodynamic study of phosphoglycerate kinase from Thermotoga maritima and its isolated domains: reversible thermal unfolding monitored by differential scanning calorimetry and circular dichroism spectroscopy; Zaiss K et al.; The folding of phosphoglycerate kinase (PGK) from the hyperthermophilic bacterium Thermotoga maritima and its isolated N- and C-terminal domains (N1/2 and C1/2) was characterized by differential scanning calorimetry (DSC) and circular dichroism (CD) spectroscopy . At pH 3.0-4.0, reversible thermal denaturation of TmPGK occurred below 90 degrees C . The corresponding peaks in the partial molar heat capacity function were fitted by a four-state model, describing three well-defined unfolding transitions . Using CD spectroscopy, these are ascribed to the disruption of the domain interactions and subsequent sequential unfolding of the two domains . The isolated N-terminal domain unfolds reversibly between pH 3.0 and pH 4.0 to >90% and at pH 7.0 to about 70% . In contrast, the isolated engineered C-terminal domain only shows reversible thermal denaturation between pH 3.0 and pH 3.5 . Neither N1/2 nor C1/2 obeys the simple two-state mechanism of unfolding . Instead, both unfold via a partially structured intermediate . In the case of N1/2, the intermediate exhibits native secondary structure and perturbed tertiary structure, whereas for C1/2 the intermediate could not be defined with certainty.

Int J Cancer, 1999 Apr 12, 81(2), 229 - 35
Mitogenic autoantibodies in Helicobacter pylori-associated stomach cancerogenesis; Hensel F et al.; Colonization of the bacterium Helicobacter pylori of gastric mucosa plays an important role in stomach carcinogenesis, while the gastric mucosa and nearby lymphoid tissue are active sites of humoral immunity against both bacteria and tumor . In a broad study on the humoral immunity of stomach-cancer patients (5 patients with diffuse- and intestinal-type stomach carcinoma), we immortalized spleen cells by using human hybridoma technology and isolated 11 hybrid clones (9 IgM, 1 IgG and 1 IgA) which react with defined proteins on different stomach-cancer cells and, interestingly, also with distinct proteins on H . pylori; 4 of these antibodies are mitogenic and stimulate the proliferation of stomach-cancer cells in vitro . Furthermore, immunohistochemical studies define these 4 clearly as autoantibodies, in view of their reactivity to normal epithelial cells . Sequence analysis of the genes for the immunoglobulin heavy (V(H)) and light (V(L)) chain variable regions revealed that most of the human antibodies belong to the V(H)3, Vlambda I and III gene families (DP-49, DPL-5 and DPL-23) and are germ-line configured.

Biochemistry (Mosc), 1999 Feb, 64(2), 189 - 93
Site-specific endonuclease NspLKI is an isoschizomer of endonuclease HaeIII; Zabaznaya EV et al.; Site-specific endonuclease NspLKI has been isolated and purified to functionally pure state from soil bacterium Nocardia species LK by successive chromatography on columns with phosphocellulose, HTP hydroxyapatite, and heparin-Sepharose . The isolated enzyme recognizes the 5'-GG downward arrowCC-3' sequence on DNA and cleaves it as indicated by the arrow, i.e., it is an isoschizomer of HaeIII . The final enzyme yield is 1.105 units per gram of wet biomass . The enzyme is active in the temperature range of 25-60 degrees C with an optimum at 48-55 degrees C; it does not lose activity on storage for three days at room temperature . An optimal buffer is HRB containing 10 mM Tris-HCl, pH 7.4, 200 microgram/ml albumin, 10 mM MgCl2, and 100 mM NaCl.

Eur J Oral Sci, 1999 Feb, 107(1), 14 - 20
LPS from Actinobacillus actinomycetemcomitans and the expression of beta2 integrins and L-selectin in an ex vivo human whole blood system; Blix IJ et al.; Actinobacillus actinomycetemcomitans is assumed to be an important etiological agent in localized juvenile periodontitis (LJP) and to have the ability to invade periodontal tissues . This bacterium has also been noted for its potential to cause serious extraoral infections . In this study, the effect of lipopolysaccharides (LPS) extracted from A . actinomycetemcomitans on the expression of the leukocyte adhesion molecules CD11a/CD18, CD11b/CD18, CD11c/CD18 and L-selectin (CD 62L) were measured in an ex vivo whole blood system by use of fluorescent antibodies followed by flow cytometry . LPS from Escherichia coli, which is known to elicit a strong inflammatory response was used as a reference . The expression of the beta2 integrins CD11a/CD18, CD11b/CD18, and CD11c/CD18 were significantly upregulated in granulocytes and monocytes . This expression was dose-dependent . The baseline levels of L-selectin was high on all three types of leukocytes, but on granulocytes and monocytes it decreased dramatically after stimulation with LPS . The LPS from A . actinomycetemcomitans was equally potent as LPS from E . coli in its ability to affect the expression of the leukocyte integrins and L-selectin.

J Biochem (Tokyo), 1999 Apr, 125(4), 649 - 57
Halobacterial rhodopsins; Mukohata Y et al.; Following the discovery of the bacteriorhodopsin proton pump in Halobacterium halobium (salinarum), not only the halorhodopsin halide pump and two photosensor rhodopsins (sensory rhodopsin and phoborhodopsin) in the same species, but also homologs of these four rhodopsins in strains of other genera of Halobacteriaceae have been reported . Twenty-eight full (and partial) sequences of the genomic DNA of these rhodopsins have been analyzed . The deduced amino acid sequences have led to new strategies and tactics for understanding bacterial rhodopsins on a comparative basis, as summarized briefly in this article . The data discussed include (i) alignment of the sequences to qualify/characterize the conserved residues; (ii) assignment of residues that cause differences in function(s)/properties; and (iii) phylogeny of the halobacterial rhodopsins to suggest their evolutionary paths . The four kinds of rhodopsin in each strain are assumed, on the basis of their genera-specific distributions, to have arisen by at least two gene-duplication processes during evolution prior to generic speciation . The first duplication of the rhodopsin ancestor gene yielded two genes, each of which was duplicated again to give four genes in the ancestor halobacterium . The bacterium carrying four rhodopsin genes, after accumulating mutations, became ready for generic speciation and the delivery of four rhodopsins to each species . The original rhodopsin ancestor is speculated to be closest to the proton pump (bacteriorhodopsin).

Trends Biotechnol, 1999 Jan, 17(1), 21 - 4
The metabolic effects of native and transgenic hemoglobins on plants; Bulow L et al.; The strictly aerobic bacterium Vitreoscilla expresses a hemoglobin-like protein, VHb, when subjected to oxygen stress . When expressed in plants, this has several intriguing physiological effects, such as improving the overall growth rate, speeding germination and flowering, and increasing the productivity of certain oxygen-requiring metabolic pathways . Although the mechanisms behind the effects of VHb in heterologous hosts are not yet fully characterized, it has been suggested that VHb facilitates oxygen transport and/or storage . This hypothesis is supported by the kinetic properties of VHb, which allow very rapid dissociation of oxygen from the protein.

J Bacteriol, 1999 Apr, 181(7), 2142 - 7
Iron reductase for magnetite synthesis in the magnetotactic bacterium Magnetospirillum magnetotacticum; Noguchi Y et al.; Ferric iron reductase was purified from magnetotactic bacterium Magnetospirillum (formerly Aquaspirillum) magnetotacticum (ATCC 31632) to an electrophoretically homogeneous state . The enzyme was loosely bound on the cytoplasmic face of the cytoplasmic membrane and was found more frequently in magnetic cells than in nonmagnetic cells . The molecular mass of the purified enzyme was calculated upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis to be about 36 kDa, almost the same as that calibrated by gel filtration analysis . The enzyme required NADH and flavin mononucleotide (FMN) as optimal electron donor and cofactor, respectively, and the activity was strongly inhibited by Zn2+ acting as a partial mixed-type inhibitor . The Km values for NADH and FMN were 4.3 and 0 . 035 microM, respectively, and the Ki values for Zn2+ were 19.2 and 23.9 microM for NADH and FMN, respectively . When the bacterium was grown in the presence of ZnSO4, the magnetosome number in the cells and the ferric iron reductase activity declined in parallel with an increase in the ZnSO4 concentration of the medium, suggesting that the ferric iron reductase purified in the present study may participate in magnetite synthesis.

J Bacteriol, 1999 Apr, 181(7), 2102 - 9
BadR, a new MarR family member, regulates anaerobic benzoate degradation by Rhodopseudomonas palustris in concert with AadR, an Fnr family member; Egland PG et al.; A cluster of genes for the anaerobic degradation of benzoate has been described for the phototrophic bacterium Rhodopseudomonas palustris . Here we provide an initial analysis of the regulation of anaerobic benzoate degradation by examining the contributions of two regulators: a new regulator, BadR, encoded by the benzoate degradation gene cluster, and a previously described regulator, AadR, whose gene lies outside the cluster . Strains with single mutations in either badR or aadR grew slowly on benzoate but were relatively unimpaired in growth on succinate and several intermediates of benzoate degradation . A badR aadR double mutant was completely defective in anaerobic growth on benzoate . Effects of the regulators on transcriptional activation were monitored with an R . palustris strain carrying a chromosomal fusion of 'lacZ to the badE gene of the badDEFG operon . This operon encodes benzoyl-coenzyme A (benzoyl-CoA) reductase, an unusual oxygen-sensitive enzyme that catalyzes the benzene ring reduction reaction that is the rate-limiting step in anaerobic benzoate degradation . Expression of badE::'lacZ was induced 100-fold when cells grown aerobically on succinate were shifted to anaerobic growth on succinate plus benzoate . The aadR gene was required for a 20-fold increase in expression that occurred in response to anaerobiosis, and badR was responsible for a further 5-fold increase in expression that occurred in response to benzoate . Further studies with the badE::'lacZ fusion strain grown with various kinds of aromatic acids indicated that BadR probably responds to benzoyl-CoA acting as an effector molecule . Sequence information indicates that BadR is a member of the MarR family of transcriptional regulators . These studies expand the range of functions regulated by MarR family members to include anaerobic aromatic acid degradation and provide an example of a MarR-type protein that acts as a positive regulator rather than as a negative regulator, as do most MarR family members . AadR resembles the Escherichia coli Fnr regulator in sequence and contains cysteine residues that are spaced appropriately to serve in the capacity of a redox-sensing protein.

J Bacteriol, 1999 Apr, 181(7), 1994 - 2000
The presence of ADP-ribosylated Fe protein of nitrogenase in Rhodobacter capsulatus is correlated with cellular nitrogen status; Yakunin AF et al.; The photosynthetic bacterium Rhodobacter capsulatus has been shown to regulate its nitrogenase by covalent modification via the reversible ADP-ribosylation of Fe protein in response to darkness or the addition of external NH4+ . Here we demonstrate the presence of ADP-ribosylated Fe protein under a variety of steady-state growth conditions . We examined the modification of Fe protein and nitrogenase activity under three different growth conditions that establish different levels of cellular nitrogen: batch growth with limiting NH4+, where the nitrogen status is externally controlled; batch growth on relatively poor nitrogen sources, where the nitrogen status is internally controlled by assimilatory processes; and continuous culture . When cultures were grown to stationary phase with different limiting concentrations of NH4+, the ADP-ribosylation state of Fe protein was found to correlate with cellular nitrogen status . Additionally, actively growing cultures (grown with N2 or glutamate), which had an intermediate cellular nitrogen status, contained a portion of their Fe protein in the modified state . The correlation between cellular nitrogen status and ADP-ribosylation state was corroborated with continuous cultures grown under various degrees of nitrogen limitation . These results show that in R . capsulatus the modification system that ADP-ribosylates nitrogenase in the short term in response to abrupt changes in the environment is also capable of modifying nitrogenase in accordance with long-term cellular conditions.

Mol Microbiol, 1998 Nov, 30(4), 883 - 93
The structure of an ECF-sigma-dependent, light-inducible promoter from the bacterium Myxococcus xanthus; Martinez-Argudo I et al.; Expression of the Myxococcus xanthus gene crtl is controlled by a light-inducible promoter . The activity of this promoter depends on CarQ, a sigma factor of the extracytoplasmic function (ECF) subfamily . Here, we show thatthe minimum DNA stretch reproducing normal expression of crtl extends from a few bases upstream of the -35 position to a site well downstream of the transcriptional start . The downstream DNA contains an enhancer-like element that remains active when displaced upstream of the promoter . Experimental evidence is provided for the activity of the crtl promoter being critically dependent on a pentanucleotide sequence centred at the -31 position . The similarity of this sequence with the consensus for ECF-sigma-dependent promoters from other bacteria is discussed . The activity of the crtl promoter also depends on certain basepairs at the -10 region . Hence, the operation of ECF-sigma-factors seems to require binding to two different DNA sites, although the -10 sequences of different ECF-sigma-dependent promoters are unrelated to one another, and the ECF-sigma-factors themselves lack the conserved domain known to mediate binding of other sigma-factors to the -10 DNA site.

Mol Microbiol, 1998 Nov, 30(4), 761 - 5
Protein-mediated DNA transfer into liposomes; Lambert O et al.; The transfer of a foreign genome into a bacterium by means of phage infection is a very efficient but poorly understood process . To analyse the mechanism of phage DNA transfer at a molecular level, we have reconstituted FhuA, the receptor for phage T5 in the outer membrane of Escherichia coli, into unilamellar vesicles made of natural phospholipids . Cryoelectron microscopy studies showed that the binding of the phage to FhuA triggered the transfer of its double-stranded DNA (121000 bp) into the proteoliposomes . DNA was entrapped within vesicles with a diameter ranging from 70 to 150 nm . The DNA appeared to be densely packed, but its presence did not alter the morphology of the liposomes, suggesting no DNA-lipid interactions . These liposomes represent an attractive model system for studying the mechanisms of DNA transport and condensation . They may also serve as alternative vehicles for the transfer of foreign genes into eukaryotic cells.

FEBS Lett, 1999 Feb 26, 445(2-3), 409 - 14
ATP-synthase of Rhodobacter capsulatus: coupling of proton flow through F0 to reactions in F1 under the ATP synthesis and slip conditions; Feniouk BA et al.; A stepwise increasing membrane potential was generated in chromatophores of the phototrophic bacterium Rhodobacter capsulatus by illumination with short flashes of light . Proton transfer through ATP-synthase (measured by electrochromic carotenoid bandshift and by pH-indicators) and ATP release (measured by luminescence of luciferin-luciferase) were monitored . The ratio between the amount of protons translocated by F0F1 and the ATP yield decreased with the flash number from an apparent value of 13 after the first flash to about 5 when averaged over three flashes . In the absence of ADP, protons slipped through F0F1 . The proton transfer through F0F1 after the first flash contained two kinetic components, of about 6 ms and 20 ms both under the ATP synthesis conditions and under slip . The slower component of proton transfer was substantially suppressed in the absence of ADP . We attribute our observations to the mechanism of energy storage in the ATP-synthase needed to couple the transfer of four protons with the synthesis of one molecule of ATP . Most probably, the transfer of initial protons of each tetrad creates a strain in the enzyme that slows the translocation of the following protons.

Eur J Biochem, 1999 Feb, 259(3), 709 - 18
The cytochrome bc1 complex from Rhodovulum sulfidophilum is a dimer with six quinones per monomer and an additional 6-kDa component; Montoya G et al.; A highly active, large-scale preparation of cytochrome bc1 complex has been obtained from the photosynthetic purple bacterium Rhodovulum (Rhv.) sulfidophilum . It has been characterized using mass spectrometry, quinone and lipid analysis as well as inhibitor binding . About 35 mg of pure complex can be obtained from 1 g of membrane protein . EPR spectroscopy and optical titrations have been used to obtain the redox midpoint potentials of the cofactors . The Em-value of 310 mV for the Rieske protein is the most positive midpoint potential for this protein in a bc1 complex so far . The bc1 complex from Rhv . sulfidophilum is very stable and consists of three subunits and a 6-kDa polypeptide . The complex appears as a dimer in solution and contains six quinone molecules per monomer which are tightly bound . EPR spectroscopy shows that the Q(o) site is highly occupied . High detergent concentrations convert the complex into an inactive, monomeric form that has lost the Rieske protein as well as the quinones and the 6-kDa component.

Eur J Biochem, 1999 Feb, 260(1), 258 - 67
The tricarboxylic acid cycle of Helicobacter pylori; Pitson SM et al.; The composition and properties of the tricarboxylic acid cycle of the microaerophilic human pathogen Helicobacter pylori were investigated in situ and in cell extracts using {1H}- and {13C}-NMR spectroscopy and spectrophotometry . NMR spectroscopy assays enabled highly specific measurements of some enzyme activities, previously not possible using spectrophotometry, in in situ studies with H . pylori, thus providing the first accurate picture of the complete tricarboxylic acid cycle of the bacterium . The presence, cellular location and kinetic parameters of citrate synthase, aconitase, isocitrate dehydrogenase, alpha-ketoglutarate oxidase, fumarate reductase, fumarase, malate dehydrogenase, and malate synthase activities in H . pylori are described . The absence of other enzyme activities of the cycle, including alpha-ketoglutarate dehydrogenase, succinyl-CoA synthetase, and succinate dehydrogenase also are shown . The H . pylori tricarboxylic acid cycle appears to be a noncyclic, branched pathway, characteristic of anaerobic metabolism, directed towards the production of succinate in the reductive dicarboxylic acid branch and alpha-ketoglutarate in the oxidative tricarboxylic acid branch . Both branches were metabolically linked by the presence of alpha-ketoglutarate oxidase activity . Under the growth conditions employed, H . pylori did not possess an operational glyoxylate bypass, owing to the absence of isocitrate lyase activity; nor a gamma-aminobutyrate shunt, owing to the absence of both gamma-aminobutyrate transaminase and succinic semialdehyde dehydrogenase activities . The catalytic and regulatory properties of the H . pylori tricarboxylic acid cycle enzymes are discussed by comparing their amino acid sequences with those of other, more extensively studied enzymes.

Ital J Gastroenterol Hepatol, 1999 Jan-Feb, 31(1), 70 - 2
Detection of Tropheryma whippelii DNA (Whipple's disease) in faeces; Gross M et al.; To date the diagnosis of Whipple's disease is based mainly on the histopathological analysis of duodenal biopsies since Tropheryma whippelii cannot be cultured in vitro . We investigated the possibility to diagnose Whipple's disease by detection of bacterial DNA in faces . Nested polymerase chain reaction with amplification of part of the 16S rRNA gene of this bacterium in DNA extracted from faeces of a patient with Whipple's disease was performed . Sequencing of the polymerase chain reaction product revealed the sequence of Tropheryma whippelii . We conclude that Whipple's disease will be able to be diagnosed non-invasively by DNA analysis from the faeces as soon as more specific sequences of this bacteria are known.

Acta Crystallogr D Biol Crystallogr, 1999 Jan, 55 ( Pt 1), 329 - 31 Epub 1999 Jan 01.
Crystallization and preliminary crystallographic analysis of the pyruvate-ferredoxin oxidoreductase from Desulfovibrio africanus; Pieulle L et al.; For the first time, crystals of a pyruvate-ferredoxin oxidoreductase (PFOR) suitable for X-ray analysis have been obtained . This enzyme catalyzes, in anaerobic organisms, the crucial energy-yielding reaction of pyruvate decarboxylation to acetylCoA . Polyethylene glycol and divalent metal cations have been used to crystallize the PFOR from the sulfate-reducing bacterium Desulfovibrio africanus . Two different orthorhombic (P212121 ) crystal forms have been grown with unit-cell dimensions a = 86.1, b = 146.7, c = 212.5 A and a = 84.8, b = 144.9, c = 203.0 A . Both crystals diffract to 2.3 A resolution using synchrotron radiation.

Acta Crystallogr D Biol Crystallogr, 1999 Apr, 55 ( Pt 4), 927 - 8
Crystallization and preliminary x-ray analysis of glutamate racemase from Aquifex pyrophilus, a hyperthermophilic bacterium; Hwang KY et al.; Glutamate racemase catalyzes the reversible reaction of L-glutamate to D-glutamate, an essential component of the bacterial cell wall . Glutamate racemase from Aquifex pyrophilus has been crystallized by the hanging-drop vapor-diffusion method using polyethylene glycol 6000 as a precipitant . The crystals belong to space group P6122 or P6522 with unit-cell parameters a = b = 72.1, c = 185.02 A . The asymmetric unit contains one molecule, corresponding to a Vm value of 2.35 A3 Da-1 . Complete data sets from a native and a mercury-derivative crystal have been collected at 2.0 and 2.3 A resolution, respectively, using a synchrotron-radiation source.

Acta Crystallogr D Biol Crystallogr, 1999 Apr, 55 ( Pt 4), 877 - 9
Crystallization and preliminary x-ray analysis of a nitrate reductase from Desulfovibrio desulfuricans ATCC 27774; Dias JM et al.; Periplasmic nitrate reductase from the sulfate-reducing bacterium Desulfovibrio desulfuricans ATCC 27774 contains two molybdopterin guanine dinucleotide cofactors and one {4Fe-4S} cluster as prosthetic groups and catalyzes the conversion of nitrate to nitrite . Crystals of the oxidized form of this enzyme were obtained using PEG as precipitant and belong to space group P3121 or P3221, with unit-cell dimensions a = b = 106.3, c = 135.1 A . There is one monomer of 80 kDa in the asymmetric unit, which corresponds to a Matthews ratio of 2.75 A3 Da-1 . Using cryo-cooling procedures and X-rays from a rotating-anode generator, diffraction was observed to beyond 3.0 A resolution.

Trends Cell Biol, 1999 Jan, 9(1), 11 - 4
Putting E . coli on a pedestal: a unique system to study signal transduction and the actin cytoskeleton; Goosney DL et al.; Enteropathogenic Escherichia coli (EPEC) subverts host signalling pathways and the cytoskeleton during infection, resulting in disease characterized by diarrhoea . Recent studies have revolutionized our understanding of the infection process by showing that this bacterium inserts its own receptor into the plasma membrane overlying the host actin cytoskeleton . The reorganized actin forms a pedestal-like structure with the bacterium at the tip . This review discusses the mechanism of infection and pedestal formation and how this system might be a powerful tool for studying actin dynamics at the plasma membrane.

Extremophiles, 1999 Jan, 3(1), 35 - 43
The mer operon of the acidophilic bacterium Thiobacillus T3.2 diverges from its Thiobacillus ferrooxidans counterpart; Velasco A et al.; The chromosomal mercury resistance (mer) region of the acidophilic bacterium Thiobacillus T3.2 was cloned, characterized, and compared to reported homologous sequences . The Thiobacillus T3.2 mer resistance system is organized as an operon that transcribes into a polycistronic mRNA encoding the Hg2+ ion transport MerT and MerP proteins and the mercuric reductase MerA . In contrast to the Thiobacillus ferrooxidans mer determinant, no merC gene was detected . Transcription of structural genes is regulated by the product of the regulatory merR gene . On the basis of sequence data and expression experiments in E . coli, both merTPA and merR transcription units could be located close to each other and in different strands, with their promoters (PTPA and PR, respectively) overlapping the putative MerR binding site in the intergenic operator/promoter (O/P) region . Amino acid sequences of mer gene products were compared to their homologs . Some sequence features, such as the number and position of cysteine residues, are unique for the Mer proteins of this bacterium . Similarities (-10 and -35 boxes are 19bp apart in both PR and PTPA promoters) and differences (inverted repeats in the Thiobacillus T3.2 MerR-binding site are 2bp shorter than in Thiobacillus ferrooxidans) exist between the O/P intergenic regions of both Thiobacilli . In vivo experiments showed inducible expression of mercury resistance in E . coli cells transformed with the entire Thiobacillus T3.2 mer genetic determinant (structural plus regulatory genes), and little or no expression in clones containing only the structural merT, merP, and merA genes.

Infect Immun, 1999 Apr, 67(4), 1798 - 805
Human embryonic gastric xenografts in nude mice: a new model of Helicobacter pylori infection; Lozniewski A et al.; In vitro or animal models have been used to investigate the pathogenesis of Helicobacter pylori infection . However, extrapolation to humans of results obtained with these heterologous models remains difficult . We have developed a new model for the study of H . pylori infection that uses human entire embryonic stomachs engrafted in nude mice . At 80 days after implantation, 22 of these xenografts, which exhibited a mature gastric epithelium, were inoculated with 10(7) to 10(8) CFU of either H . pylori LB1, a freshly isolated H . pylori strain (n = 12), or H . pylori ATCC 49503 (n = 10) . After 12-week examination, H . pylori LB1 persistently colonized the antrum of all inoculated grafts, as assessed by culture (mucus and mucosa), immunohistochemistry (mucosa), and a rapid urease test (mucus) . H . pylori ATCC 49503, either before or after in vivo passage, permitted only a transient 2-week colonization in one of the five inoculated grafts in both groups . Colonization was always associated with an increase of gastric juice pH . A mild neutrophil infiltration of the gastric mucosa was noted solely in infected grafts . Transmission electron microscopy showed adherence of H . pylori organisms to epithelial cell surface . In six animals, intracytoplasmic location of this bacterium was observed in the antrum or the fundus . These results allow us to propose this model as a new ex vivo model for the study of specific H . pylori-gastric cell interactions.

Tijdschr Diergeneeskd, 1999 Feb 15, 124(4), 108 - 10
{Capnocytophaga canimorsus infections: a possibly fatal complication of bite wounds}; Kramer AM et al.; Practising veterinarians and their assistants run the risk of being bitten by their patients, mostly cats and dogs, and many have experienced that bites and bite-wound infections can have unpleasant consequences . In recent years, more insight has been gained into a 'new' bacterial infection of bite wounds that not only has severe local effects but also potentially fatal systemic consequences . The bacterium involved is Capnocytophaga canimorsus . All bite wounds should be treated adequately, but this is especially so when wounds are infected with C . canimorsus . In this article, dog and cat bites are briefly described and then an overview is given of current knowledge of C . canimorsus and appropriate prophylactic measures.

Emerg Infect Dis, 1999 Jan-Feb, 5(1), 164 - 7
Mycoplasma penetrans bacteremia and primary antiphospholipid syndrome; Yanez A et al.; Mycoplasma penetrans, a rare bacterium so far only found in HIV-infected persons, was isolated in the blood and throat of a non-HIV-infected patient with primary antiphospholipid syndrome (whose etiology and pathogenesis are unknown).

Ital J Gastroenterol Hepatol, 1998 Oct, 30 Suppl 3, S310 - 2
Helicobacter pylori infection and autoimmune processes: an emerging field of study; Sorrentino D et al.; Evidence is accumulating that Helicobacter pylori infection may be closely associated with autoimmunity . However, whether autoimmunity plays a causal role in the pathogenesis of some of the diseases attributed to this bacterium or whether it is rather an epiphenomenon remains to be determined . In this brief review, a summary is made of current knowledge regarding the potential general mechanisms by which Helicobacter pylori causes mucosal damage . A review is then made of the evidence linking this bacterium to the production of different gastric autoantibodies . Finally, the reported association between Helicobacter pylori infection and some known autoimmune diseases is discussed . Although the data are still not sufficiently complete to draw definite conclusions, autoimmunity appears to be an important aspect of this infection and will certainly become a major field of study in the next few years.

Ital J Gastroenterol Hepatol, 1998 Oct, 30 Suppl 3, S307 - 9
Helicobacter pylori infection and vascular diseases; Gasbarrini A et al.; Helicobacter pylori infection has recently been associated to some organic functional vascular disorders and, both observational and interventional studies have been carried out . A correlation between Helicobacter pylori infection and ischaemic heart disease was first described in 1994 . Recent data, moreover, indicate a role of the bacterium in some functional vascular disorders such as primary headache and primary Raynaud phenomenon; indeed, some patients following eradication of Helicobacter pylori showed a significant improvement in the manifestations of these diseases . The host immune response against the bacterium may play an important role in the pathogenesis of vascular disorders, probably through a chronic stimulation of the release of vasoactive substances, such as cytokines, prostaglandins and others . However, various confounding factors such as co-infections, genetic and immunological host-factors, different strains of Helicobacter pylori or other may influence the data . Well designed case/control and randomized interventional studies are still needed to assess the real role of Helicobacter pylori in vascular disorders.

Ital J Gastroenterol Hepatol, 1998 Oct, 30 Suppl 3, S289 - 93
Gastro-oesophageal reflux and Helicobacter pylori; Pace F et al.; The nature of the relationship between Helicobacter pylori infection and reflux oesophagitis is still not completely understood . To review the available evidence from the literature concerning the relationship between Helicobacter pylori infection and reflux oesophagitis with or without Barrett's metaplasia, we performed a Medline search to discover all published reports available in this field since the first description of Helicobacter pylori (1984) to April 1998 . A total of 37 papers were found addressing the issue . From the available data, it can be concluded that Hp infection has probably only a minor protective role against the development of reflux oesophagitis . The related mechanisms are, however, still to be clarified . The presence of Helicobacter pylori infection is also likely to increase the efficacy of PPI drugs and, conversely, the eradication of the bacterium decreases the drug effect . The bacterial production of ammonia is the most likely factor explaining this observation . Furthermore, it is now sufficiently clear that patients placed on long-term PPI therapy develop atrophic gastritis only if Helicobacter pylori infection is not eradicated . Finally, both the development of metaplastic changes and the progression to severe dysplasia and adenocarcinoma in Barrett's oesophagus patients are phenomena not related to the presence of Helicobacter pylori antral or oesophageal colonization.

Ital J Gastroenterol Hepatol, 1998 Oct, 30 Suppl 3, S286 - 8
Helicobacter pylori infection and dyspepsia; Quina MG; Dyspepsia is defined as a persistent or recurrent pain or discomfort, localised in the upper abdomen, which may or may not be related to meals . Its prevalence in the general population is extraordinarily high (20-40%) . Several pathological conditions can provoke dyspepsia, although non ulcer dyspepsia is the main cause . The relationships between Helicobacter pylori and non ulcer dyspepsia are discussed, namely the prevalence of Helicobacter pylori infection and the efficacy of its eradication in non ulcer dyspepsia . The management of dyspeptic patients in the community is analysed according to the Maastricht Consensus of 1996 . Our opinion is that, in Helicobacter pylori-positive dyspeptic patients, after a careful investigation with exclusion of other organic diseases, the bacterium should be eradicated.

J Clin Microbiol, 1999 Apr, 37(4), 1077 - 83
Evaluation of modified BACTEC 12B radiometric medium and solid media for culture of Mycobacterium avium subsp . paratuberculosis from sheep; Whittington RJ et al.; Definitive diagnosis of Johne's disease in ruminants depends on confirming the presence of the causative bacterium, Mycobacterium avium subsp . paratuberculosis, in tissues of the host . This is readily achieved in most ruminant species by culture . However, culture of clinical specimens from sheep in many countries has been unrewarding . Such a culture from sheep was achieved recently in Australia by using a radiometric culture medium . The aims of the present study were to evaluate the culture of M . avium subsp . paratuberculosis from sheep by using modified BACTEC 12B radiometric medium, to determine the sensitivity of culture in relation to histopathology, and to evaluate a range of solid media . Culture of M . avium subsp . paratuberculosis from sheep with Johne's disease is a sensitive method of diagnosis: intestinal tissues from all 43 animals with multibacillary disease and all 22 animals with paucibacillary disease were culture positive, while 98% of feces from 53 animals with multibacillary disease and 48% of feces from 31 animals with paucibacillary disease were culture positive . Of sheep without histological evidence of Johne's disease from infected flocks, intestinal tissue from 32% of 41 were culture positive, while feces from 17% of 41 were culture positive . Consequently, culture is recommended as the "gold standard" test for detection of ovine Johne's disease . Of the wide range of solid media that were evaluated, only modified Middlebrook 7H10 and 7H11 agars, which were very similar in composition to modified BACTEC 12B medium, yielded growth of ovine strains of M . avium subsp . paratuberculosis . The sensitivity of detection of M . avium subsp . paratuberculosis on solid media was slightly lower than that in modified BACTEC 12B radiometric medium . Both egg yolk and mycobactin J were essential additives for growth of ovine strains of M . avium subsp . paratuberculosis in both liquid and solid media.

J Clin Microbiol, 1999 Apr, 37(4), 987 - 92
Rapid and sensitive quantification of Borrelia burgdorferi-infected mouse tissues by continuous fluorescent monitoring of PCR; Morrison TB et al.; The quantity of Borrelia burgdorferi organisms in tissue samples is an important determinant for infection studies in the mouse model of Lyme disease . This report presents the development of a rapid and sensitive external-standard-based PCR assay for the absolute quantification of B . burgdorferi in mouse tissue samples . The assay uses a double-stranded DNA dye to continuously monitor product formation and in less than an hour was able to quantify samples ranging up to 6 log units in concentration . The PCR efficiencies of the sample and the standard were matched by using a standard composed of purified B . burgdorferi chromosome mixed with tissue-matched mouse genome lacking bacterial DNA . Normalization of B . burgdorferi quantities to the mouse nidogen gene allowed comparison of B . burgdorferi numbers in samples isolated from different tissues and strains . PCR analysis of the chromosomal gene recA in cultured B . burgdorferi was consistent with a single recA per bacterium . The parameters defined in this assay should be applicable to quantification of other organisms, even infectious agents for which no ready source of DNA standard is available . In summary, this report presents a rapid external-standard-based PCR method for the quantification of B . burgdorferi in mouse DNA samples.

Biochemistry, 1999 Mar 9, 38(10), 2861 - 5
Interaction site for high-potential iron-sulfur protein on the tetraheme cytochrome subunit bound to the photosynthetic reaction center of Rubrivivax gelatinosus; Osyczka A et al.; We have recently demonstrated, using site-directed mutagenesis, that soluble cytochromes interact with the Rubrivivax gelatinosus photosynthetic reaction center (RC) in the vicinity of the low-potential heme 1 (c-551, Em = 70 mV) of the tetraheme cytochrome subunit, the fourth heme from the special pair of bacteriochlorophyll {Osyczka, A., et al . (1998) Biochemistry 37, 11732-11744} . Although the mutations generated in that study did not show clear effects on the electron transfer from high-potential iron-sulfur protein (HiPIP), which is the major physiological electron donor to the RC in this bacterium, we report here that other site-directed mutations near the solvent-exposed edge of the same low-potential heme 1, V67K (valine-67 substituted by lysine) and E79K/E85K/E93K (glutamates-79, -85, and -93, all replaced by lysines), considerably inhibit the electron transfer from HiPIP to the RC . Thus, it is concluded that HiPIP, like soluble cytochromes, binds to the RC in the vicinity of the exposed part of the low-potential heme 1 of the cytochrome subunit, although some differences in the configurations of the HiPIP-RC and cytochrome c-RC transient complexes may be postulated.

J Virol, 1999 Apr, 73(4), 2994 - 3003
HMG protein family members stimulate human immunodeficiency virus type 1 and avian sarcoma virus concerted DNA integration in vitro; Hindmarsh P et al.; We have reconstituted concerted human immunodeficiency virus type 1 (HIV-1) integration in vitro with specially designed mini-donor HIV-1 DNA, a supercoiled plasmid acceptor, purified bacterium-derived HIV-1 integrase (IN), and host HMG protein family members . This system is comparable to one previously described for avian sarcoma virus (ASV) (A . Aiyar et al., J . Virol . 70:3571-3580, 1996) that was stimulated by the presence of HMG-1 . Sequence analyses of individual HIV-1 integrants showed loss of 2 bp from the ends of the donor DNA and almost exclusive 5-bp duplications of the acceptor DNA at the site of integration . All of the integrants sequenced were inserted into different sites in the acceptor . These are the features associated with integration of viral DNA in vivo . We have used the ASV and HIV-1 reconstituted systems to compare the mechanism of concerted DNA integration and examine the role of different HMG proteins in the reaction . Of the three HMG proteins examined, HMG-1, HMG-2, and HMG-I(Y), the products formed in the presence of HMG-I(Y) for both systems most closely match those observed in vivo . Further analysis of HMG-I(Y) mutants demonstrates that the stimulation of integration requires an HMG-I(Y) domain involved in DNA binding . While complexes containing HMG-I(Y), ASV IN, and donor DNA can be detected in gel shift experiments, coprecipitation experiments failed to demonstrate stable interactions between HMG-I(Y) and ASV IN or between HMG-I(Y) and HIV-1 IN.

Plant Physiol, 1999 Mar, 119(3), 1057 - 64
Further studies of the role of cyclic beta-glucans in symbiosis . An NdvC mutant of Bradyrhizobium japonicum synthesizes cyclodecakis-(1-->3)-beta-glucosyl; Bhagwat AA et al.; The cyclic beta-(1-->3),beta-(1-->6)-D-glucan synthesis locus of Bradyrhizobium japonicum is composed of at least two genes, ndvB and ndvC . Mutation in either gene affects glucan synthesis, as well as the ability of the bacterium to establish a successful symbiotic interaction with the legume host soybean (Glycine max) . B . japonicum strain AB-14 (ndvB::Tn5) does not synthesize beta-glucans, and strain AB-1 (ndvC::Tn5) synthesizes a cyclic beta-glucan lacking beta-(1-->6)-glycosidic bonds . We determined that the structure of the glucan synthesized by strain AB-1 is cyclodecakis-(1-->3)-beta-D-glucosyl, a cyclic beta-(1-->3)-linked decasaccharide in which one of the residues is substituted in the 6 position with beta-laminaribiose . Cyclodecakis-(1-->3)-beta-D-glucosyl did not suppress the fungal beta-glucan-induced plant defense response in soybean cotyledons and had much lower affinity for the putative membrane receptor protein than cyclic beta-(1-->3),beta-(1-->6)-glucans produced by wild-type B . japonicum . This is consistent with the hypothesis presented previously that the wild-type cyclic beta-glucans may function as suppressors of a host defense response.

J Inorg Biochem, 1998 Dec, 72(3-4), 179 - 85
Concentration of Cu, EPR-detectable Cu, and formation of cupric-ferrocyanide in membranes with pMMO; Yuan H et al.; EPR spectra were obtained for the type 2 Cu(2+) site in particulate methane monooxygenase, pMMO, from membrane fractions of Methylomicrobium album BG8 . In addition to the EPR signal with g parallel = 2.24 and A parallel = 185 G found in both cells and membrane fractions, a second EPR signal with g parallel = 2.29 and A parallel = 146 G was found in membrane fractions and attributed to oxidation of cuprous sites . Comparison of EPR-detectable Cu(2+) with total copper determined by atomic absorption suggests that there are two or three EPR-silent coppers for every EPR-detectable copper and that there are approximately four coppers per enzyme composed of the 47, 27, and 25 kDa subunits . Treatment of membrane fractions loaded with pMMO with Fe(CN)6(3-) results in a new EPR signal that is attributed to CuFe(CN)6(2-), not to an intrinsic trimeric copper cluster as previously reported in studies with a related bacterium.

Biol Chem, 1999 Jan, 380(1), 75 - 80
Activation of protein C by arginine-specific cysteine proteinases (gingipains-R) from Porphyromonas gingivalis; Hosotaki K et al.; In order to determine the effect of bacterial proteinases on activation of the protein C system, a negative regulator of blood coagulation, two arginine-specific cysteine proteinases (gingipains R) from Porphyromonas gingivalis, a causative bacterium of adult periodontitis, were examined . Each enzyme activated human protein C in a dose- and incubation time-dependent manner . Interestingly, the form of enzyme being composed of a non-covalent complex containing both catalytic and adhesion domains (RgpA) produced activated protein C 14-fold more efficiently than RgpB which contained the catalytic domain alone . The kcat/Km value of RgpA was 18-fold higher than that of RgpB and comparable to that of the thrombin-thrombomodulin complex, the physiological activator of protein C . RgpA catalyzed protein C activation was augmented 1.4-fold by phospholipids, ubiquitous cell membrane components . Furthermore, RgpA, but not RgpB, could activate protein C in plasma and this resulted in a decrease of the protein C concentration in plasma, which is often observed in patients with sepsis during the development of disseminated intravascular coagulation (DIC) . These data indicate that RgpA is a more potent activator of protein C than RgpB and suggest that only the former enzyme can cause protein C activation in vivo . The present study further suggests that bacterial proteinases may possibly contribute to the consumption of plasma protein C which predisposes to DIC and/or promotes a thrombotic tendency towards DIC in sepsis.

Proc Natl Acad Sci U S A, 1999 Mar 2, 96(5), 2054 - 9
Multiple pathways for ultrafast transduction of light energy in the photosynthetic reaction center of Rhodobacter sphaeroides; van Brederode ME et al.; A pathway of electron transfer is described that operates in the wild-type reaction center (RC) of the photosynthetic bacterium Rhodobacter sphaeroides . The pathway does not involve the excited state of the special pair dimer of bacteriochlorophylls (P*), but instead is driven by the excited state of the monomeric bacteriochlorophyll (BA*) present in the active branch of pigments along which electron transfer occurs . Pump-probe experiments were performed at 77 K on membrane-bound RCs by using different excitation wavelengths, to investigate the formation of the charge separated state P+HA- . In experiments in which P or BA was selectively excited at 880 nm or 796 nm, respectively, the formation of P+HA- was associated with similar time constants of 1.5 ps and 1 . 7 ps . However, the spectral changes associated with the two time constants are very different . Global analysis of the transient spectra shows that a mixture of P+BA- and P* is formed in parallel from BA* on a subpicosecond time scale . In contrast, excitation of the inactive branch monomeric bacteriochlorophyll (BB) and the high exciton component of P (P+) resulted in electron transfer only after relaxation to P* . The multiple pathways for primary electron transfer in the bacterial RC are discussed with regard to the mechanism of charge separation in the RC of photosystem II from higher plants.

Genetics, 1999 Mar, 151(3), 1165 - 72
Temporal and multiple quantitative trait loci analyses of resistance to bacterial wilt in tomato permit the resolution of linked loci; Mangin B et al.; Ralstonia solanacearum is a soil-borne bacterium that causes the serious disease known as bacterial wilt in many plant species . In tomato, several QTL controlling resistance have been found, but in different studies, markers spanning a large region of chromosome 6 showed strong association with the resistance . By using two different approaches to analyze the data from a field test F3 population, we show that at least two separate loci approximately 30 cM apart on this chromosome are most likely involved in the resistance . First, a temporal analysis of the progression of symptoms reveals a distal locus early in the development of the disease . As the disease progresses, the maximum LOD peak observed shifts toward the proximal end of the chromosome, obscuring the distal locus . Second, although classical interval mapping could only detect the presence of one locus, a statistical "two-QTL model" test, specifically adapted for the resolution of linked QTL, strongly supported the hypothesis for the presence of two loci . These results are discussed in the context of current molecular knowledge about disease resistance genes on chromosome 6 and observations made by tomato breeders during the production of bacterial wilt-resistant varieties.

Appl Environ Microbiol, 1999 Mar, 65(3), 1015 - 9
Involvement of two plasmids in the degradation of carbaryl by Arthrobacter sp . strain RC100; Hayatsu M et al.; A bacterium capable of utilizing carbaryl (1-naphthyl N-methylcarbamate) as the sole carbon source was isolated from carbaryl-treated soil . This bacterium was characterized taxonomically as Arthrobacter and was designated strain RC100 . RC100 hydrolyzes the N-methylcarbamate linkage to 1-naphthol, which was further metabolized via salicylate and gentisate . Strain RC100 harbored three plasmids (designated pRC1, pRC2, and pRC3) . Mutants unable to degrade carbaryl arose at a high frequency after treating the culture with mitomycin C . All carbaryl-hydrolysis-deficient mutants (Cah-) lacked pRC1, and all 1-naphthol-utilization-deficient mutants (Nat-) lacked pRC2 . The plasmid-free strain RC107 grew on gentisate as a carbon source . These two plasmids could be transferred to Cah- mutants or Nat- mutants by conjugation, resulting in the restoration of the Cah and Nah phenotypes.

J Theor Biol, 1999 Jan 21, 196(2), 251 - 61
Colicin diversity: a result of eco-evolutionary dynamics; Pagie L et al.; Colicins are plasmids that are carried in Escherichia coli . They code for a toxic protein and for proteins that confer on the host immunity against this toxin . When bacteria carry plasmids their growth rate is reduced . At the same time, the production of toxins makes it possible for colicinogenic bacteria to invade bacterium strains that are not immune . In natural bacterium populations there is a high diversity of colicin types . The reason for the maintenance of this diversity has been the subject of much recent debate . We have studied a simple eco-evolutionary model of the interaction of bacteria with colicins and show that high diversity of colicins is to be expected . We find two different dynamical modes each with a high diversity: a hyperimmunity mode and a multitoxicity mode . Bacteria are immune to most toxins in the first mode but in fact produce very few toxins . In the second mode bacteria are immune only to those toxins that they actually produce . In the second mode toxin levels per bacterium are much higher, whereas immunity levels per bacterium are lower.

J Bacteriol, 1999 Mar, 181(5), 1684 - 8
Genomic complexity among strains of the facultative photoheterotrophic bacterium Rhodobacter sphaeroides; Nereng KS et al.; Pulsed-field gel electrophoresis following the use of rare cutting restriction endonucleases together with Southern hybridization, using markers distributed on chromosomes I and II of Rhodobacter sphaeroides 2.4.1, has been used to examine approximately 25 strains of R . sphaeroides in an effort to assess the occurrence of genome complexity in these strains . The results suggest that genome complexity is widespread and is accompanied by substantial genomic heterogeneity.

J Bacteriol, 1999 Mar, 181(5), 1677 - 83
Model for bacteriophage T4 development in Escherichia coli; Rabinovitch A et al.; Mathematical relations for the number of mature T4 bacteriophages, both inside and after lysis of an Escherichia coli cell, as a function of time after infection by a single phage were obtained, with the following five parameters: delay time until the first T4 is completed inside the bacterium (eclipse period, nu) and its standard deviation (sigma), the rate at which the number of ripe T4 increases inside the bacterium during the rise period (alpha), and the time when the bacterium bursts (mu) and its standard deviation (beta) . Burst size {B = alpha(mu - nu)}, the number of phages released from an infected bacterium, is thus a dependent parameter . A least-squares program was used to derive the values of the parameters for a variety of experimental results obtained with wild-type T4 in E . coli B/r under different growth conditions and manipulations (H . Hadas, M . Einav, I . Fishov, and A . Zaritsky, Microbiology 143:179-185, 1997) . A "destruction parameter" (zeta) was added to take care of the adverse effect of chloroform on phage survival . The overall agreement between the model and the experiment is quite good . The dependence of the derived parameters on growth conditions can be used to predict phage development under other experimental manipulations.

Biophys J, 1999 Mar, 76(3), 1401 - 9
Helicobacter pylori vacuolating toxin forms anion-selective channels in planar lipid bilayers: possible implications for the mechanism of cellular vacuolation; Tombola F et al.; The Helicobacter pylori VacA toxin plays a major role in the gastric pathologies associated with this bacterium . When added to cultured cells, VacA induces vacuolation, an effect potentiated by preexposure of the toxin to low pH . Its mechanism of action is unknown . We report here that VacA forms anion-selective, voltage-dependent pores in artificial membranes . Channel formation was greatly potentiated by acidic conditions or by pretreatment of VacA at low pH . No requirement for particular lipid(s) was identified . Selectivity studies showed that anion selectivity was maintained over the pH range 4.8-12, with the following permeability sequence: Cl- approximately HCO3- > pyruvate > gluconate > K+ approximately Li+ approximately Ba2+ > NH4+ . Membrane permeabilization was due to the incorporation of channels with a voltage-dependent conductance in the 10-30 pS range (2 M KCl), displaying a voltage-independent high open probability . Deletion of the NH2 terminus domain (p37) or chemical modification of VacA by diethylpyrocarbonate inhibited both channel activity and vacuolation of HeLa cells without affecting toxin internalization by the cells . Collectively, these observations strongly suggest that VacA channel formation is needed to induce cellular vacuolation, possibly by inducing an osmotic imbalance of intracellular acidic compartments.

J Antibiot (Tokyo), 1998 Dec, 51(12), 1093 - 8
Structure of actinotetraose hexatiglate, a unique glucotetraose from an actinomycete bacterium; Rickards RW et al.; An Actinomycete strain A499 belonging to the genera Amycolatopsis or Amycolata isolated from a Western Australian soil sample produced the cyclic decapeptide antibiotic quinaldopeptin (1), together with the actinotetraose hexatiglate (2), the hexa-ester of a novel non-reducing glucotetraose.

Wien Klin Wochenschr, 1998 Dec 23, 110(24), 874 - 81
Clinical manifestations, pathogenesis, and effect of antibiotic treatment on Lyme borreliosis in dogs; Straubinger RK et al.; BACKGROUND: Borrelia burgdorferi, the causative agent of Lyme disease, infects humans and animals . In humans, the disease primarily affects the skin, large joints, and the nervous system days to months after infection . Data generated with appropriate animal model help to understand the fundamental mechanisms of the disease . OBJECTIVE: 1) More clearly define the clinical manifestation and pathogenetic mechanisms of Lyme disease in dogs; 2) evaluate the effect of antibiotics in dogs infected with B . burgdorferi; 3) describe the effects of corticosteroids on dogs persistently infected with B . burgdorferi . DESIGN: Specific-pathogen-free beagles were infected with B . burgdorferi using ticks collected in an endemic Lyme disease area . Clinical signs were recorded daily . Antibody titers were measured by ELISA at two-week intervals . B . burgdorferi organisms were detected in tissues by culture and PCR . Synovial fluids were evaluated microscopically and with a chemotaxis cell migration assay . Histological sections were examined for pathological lesions . Specific cytokine up-regulation in tissues was detected by RT-PCR . INTERVENTIONS: In three separate experiments, B . burgdorferi-infected dogs received antibiotic treatment (amoxicillin; azithromycin; ceftriaxone; doxycycline) for 30 consecutive days . Two subclinical persistently infected dogs received oral prednisone for 14 consecutive days starting at day 420 post-infection . RESULTS: Dogs developed acute arthritis in the joints closest to the tick bites after a median incubation period of 68 days . Synovial membranes of lame and non-lame dogs produced the chemokine IL-8 in response to B . burgdorferi . Antibiotic treatment prevented or resolved episodes of acute arthritis, but failed to eliminate the bacterium from infected dogs . Corticosteroid treatment reactivated Lyme disease in persistently infected dogs, which had not received antibiotics previously . CONCLUSIONS: B . burgdorferi disseminates through tissue by migration following tick inoculation, produces episodes of acute arthritis, and establishes persistent infection . The spirochete survives antibiotic treatment and disease can be reactivated in immunosuppressed animals.

Curr Opin Microbiol, 1999 Feb, 2(1), 94 - 8
Gene-for-gene interactions: bacterial avirulence proteins specify plant disease resistance; Bonas U et al.; Resistance of plants to bacterial pathogens is often controlled by corresponding genes for resistance and avirulence in host and pathogen, respectively . Fifty years after discovery of the genetic basis of gene-for-gene interactions, several avirulence and plant resistance genes have been isolated and are being studied on the molecular level . Tremendous progress has been made due to a better understanding of type III secretion systems that are required for bacterial pathogenicity . We are beginning to grasp how the plant actually recognizes bacterial avirulence determinants . The current view is that the bacterium translocates avirulence proteins into the host cell by the Hrp type III secretion system and that recognition occurs in the plant cell.

Curr Opin Microbiol, 1999 Feb, 2(1), 78 - 82
Interactions of Bartonella henselae with vascular endothelial cells; Dehio C; The emerging human pathogen Bartonella henselae has the remarkable capacity to colonise vascular tissues and to stimulate vasoproliferative tumour growth . Although the molecular principle of bacterium-induced neovascularisation (angiogenesis) is still unclear, recent studies have indicated a novel mechanism of endothelial colonisation that involves the formation, engulfment and uptake of a large bacterial aggregate.

J Mol Biol, 1999 Mar 5, 286(4), 1059 - 74
A free-energy-based stochastic simulation of the Tar receptor complex; Morton-Firth CJ et al.; We recently developed a stochastic-based program that allows individual molecules in a cell signalling pathway to be simulated . This program has now been used to model the Tar complex, a multimeric signalling complex employed by coliform bacteria . This complex acts as a solid-state computational cassette, integrating and disseminating information on the presence of attractants and repellents in the environment of the bacterium . In our model, the Tar complex exists in one of two conformations which differ in the rate at which they generate labile phosphate groups and hence signal to the flagellar motor . Individual inputs to the complex (aspartate binding, methylation at different sites, binding of CheB, CheR and CheY) are represented as binary flags, and each combination of flags confers a different free energy to the two conformations . Binding and catalysis by the complex are performed stochastically according to the complete set of known reactions allowing the swimming performance of the bacterium to be predicted.The assumption of two conformational states together with the use of free energy values allows us to bring together seemingly unrelated experimental parameters . Because of thermodynamic constraints, we find that the binding affinity for aspartate is linked to changes in phosphorylation activity . We estimate the pattern of Tar methylation and effective affinity constant of receptors over a range of aspartate levels . We also obtain evidence that both the methylating and demethylating enzymes must operate exclusively on one or other of the two conformations, and that sites of methylation of the complex are occupied in sequential order rather than independently . Detailed analysis of the response to aspartate reveals several quantitative discrepancies between simulated and experimental data which indicate areas for future research .

Nippon Rinsho, 1999 Jan, 57(1), 23 - 31
{Immune response to an H . pylori infection in the stomach}; Wakatsuki Y; Helicobacter pylori infection associates with chronic infiltration by various cell types including T cells whose cytokine production may regulate local immune response to the bacterium . Indeed in the antral biopsy tissue taken from both H . pylori infected and uninfected stomach, CD3 positive T cells predominate over neutrophils, monocytes and plasma cells . Depending on the rout of antigen priming, these tissue infiltrating T cells play roles either as effector cells of tissue damage or as protecting cells to H . pylori challenge . Emerging evidences suggests that H . pylori infection appear to direct regional immune response to a Th1 type . However, even in the absence of H . pylori infection, almost ninety percent of gastric CD4 T cells are comprised of IFN-gamma producing cells . Therefore activation of tissue infiltrating T cells, either by antigen specific and non-specific manner, would lead to a perpetual inflammation in the H . pylori infected stomach.

FEMS Immunol Med Microbiol, 1999 Jan, 23(1), 57 - 66
Transmission electron microscopy studies of Moraxella (Branhamella) catarrhalis; Fitzgerald M et al.; A trypsin-sensitive 200-kDa protein has been reported to be exclusively associated with haemagglutinating isolates of Moraxella (Branhamella) catarrhalis . Transmission electron microscopy studies revealed that haemagglutination by M . catarrhalis to both human and rabbit erythrocytes was mediated by a trypsin-sensitive outer fibrillar coat . This fibrillar layer was absent on non-haemagglutinating isolates examined . Immuno-electron microscopy, using a polyclonal antiserum containing antibodies to the 200-kDa protein as a probe, showed that the 200-kDa protein is present on the outer fibrillar layer of the bacterium . These findings suggest that the haemagglutinin of M . catarrhalis is a 200-kDa protein present on the outer fibrillar coat.

Berl Munch Tierarztl Wochenschr, 1999 Jan, 112(1), 5 - 9
{Multiplex PCR for the diagnostic detection of Coxiella burnetii in cow's milk}; Edingloh M et al.; A multiplex PCR based assay was developed for the highly sensitive and specific detection of Coxiella (C.) burnetii in cow's milk . The assay simultaneously amplifies a diagnostic target within the C . burnetii IS1111 sequence and a control target within the bovine CD18 gene . The internal PCR amplification control allows the discrimination of false negative results (single tube reaction failures) from negative results due to true absence of target sequences . In order to maximize the sensitivity of the assay, a sample preparation method including a centrifugation step to concentrate the bacterium was developed . In milk samples artificially contaminated with serial dilutions of C . burnetii, about four particles per ml could reproducibly be detected . The sensitivities of both assays, multiplex PCR and PCR with only a single pair of primers ('simplex' PCR), were observed to be similar.

Int J Syst Bacteriol, 1999 Jan, 49 Pt 1, 267 - 75
Sodalis gen . nov . and Sodalis glossinidius sp . nov., a microaerophilic secondary endosymbiont of the tsetse fly Glossina morsitans morsitans; Dale C et al.; A secondary intracellular symbiotic bacterium was isolated from the haemolymph of the tsetse fly Glossina morsitans morsitans and cultured in Aedes albopictus cell line C6/36 . Pure-culture isolation of this bacterium was achieved through the use of solid-phase culture under a microaerobic atmosphere . After isolation of strain M1T, a range of tests was performed to determine the phenotypic properties of this bacterium . Considering the results of these tests, along with the phylogenetic position of this micro-organism, it is proposed that this intracellular symbiont from G . m . morsitans should be classified in a new genus Sodalis gen . nov., as Sodalis glossinidius gen . nov., sp . nov . Strain M1T is the type strain for this new species.

Infect Immun, 1999 Mar, 67(3), 1157 - 71
A 55-kilodalton immunodominant antigen of Porphyromonas gingivalis W50 has arisen via horizontal gene transfer; Hanley SA et al.; A 55-kDa outer membrane protein of Porphyromonas gingivalis W50 is a significant target of the serum immunoglobulin G antibody response of periodontal disease patients and hence may play an important role in host-bacterium interactions in periodontal disease . The gene encoding the 55-kDa antigen (ragB, for receptor antigen B) was isolated on a 9.5-kb partial Sau3AI fragment of P . gingivalis W50 chromosomal DNA in pUC18 by immunoscreening with a monoclonal antibody to this antigen . The 1.6-kb open reading frame (ORF) encoding RagB was located via subcloning and nested-deletion analysis . Sequence analysis demonstrated the presence of an upstream 3.1-kb ORF (ragA) which is cotranscribed with ragB . A number of genetic characteristics suggest that the ragAB locus was acquired by a horizontal gene transfer event . These include a significantly reduced G+C content relative to that of the P . gingivalis chromosome (42 versus 48%) and the presence of mobility elements flanking this locus in P . gingivalis W50 . Furthermore, Southern blotting and PCR analyses showed a restricted distribution of this locus in laboratory and clinical isolates of this bacterium . The association of ragAB+ P . gingivalis with clinical status was examined by PCR analysis of subgingival samples . ragAB+ was not detected in P . gingivalis-positive shallow pockets from periodontal disease patients but was present in 36% of the P . gingivalis-positive samples from deep pockets . These data suggest that the ragAB locus was acquired by certain P . gingivalis strains via horizontal gene transfer and that the acquisition of this locus may facilitate the survival of these strains at sites of periodontal destruction.

Infect Immun, 1999 Mar, 67(3), 1056 - 62
Role of Bordetella pertussis virulence factors in adherence to epithelial cell lines derived from the human respiratory tract; van den Berg BM et al.; During colonization of the respiratory tract by Bordetella pertussis, virulence factors contribute to adherence of the bacterium to the respiratory tract epithelium . In the present study, we examined the roles of the virulence factors filamentous hemagglutinin (FHA), fimbriae, pertactin (Prn), and pertussis toxin (PT) in the adherence of B . pertussis to cells of the human bronchial epithelial cell line NCI-H292 and of the laryngeal epithelial cell line HEp-2 . Using B . pertussis mutant strains and purified FHA, fimbriae, Prn, and PT, we demonstrated that both fimbriae and FHA are involved in the adhesion of B . pertussis to laryngeal epithelial cells, whereas only FHA is involved in the adherence to bronchial epithelial cells . For PT and Prn, no role as adhesion factor was found . However, purified PT bound to both bronchial and laryngeal cells and as such reduced the adherence of B . pertussis to these cells . These data may imply that fimbriae play a role in infection of only the laryngeal mucosa, while FHA is the major factor in colonization of the entire respiratory tract.

Bioorg Med Chem Lett, 1999 Jan 4, 9(1), 49 - 54
Toxicity and osmoprotective activities of analogues of glycine betaine obtained by solid phase organic synthesis towards Sinorhizobium meliloti; Cosquer A et al.; Seven analogues of the bacterial osmoprotectant glycine betaine (GB, trimethylammonioacetate), in which the methyl groups of the Me3N+ moiety are replaced by various substituents, were obtained by SPOS using Wang resin . Their biological activities (osmoprotection vs toxicity), appeared closely related to their uptake efficiency and their catabolism in the betaine-demethylating model bacterium Sinorhizobium meliloti.

Eur J Biochem, 1998 Dec 15, 258(3), 1050 - 8
Thermotoga maritima maltosyltransferase, a novel type of maltodextrin glycosyltransferase acting on starch and malto-oligosaccharides; Meissner H et al.; A novel enzyme acting on starch and malto-oligosaccharides was identified and characterised . The non-hydrolytic enzyme, designated maltosyltransferase (MTase), of the hyperthermophilic bacterium Thermotoga maritima MSB8 disproportionates malto-oligosaccharides via glycosyl transfer reactions . The enzyme has a unique transfer specificity strictly confined to the transfer of maltosyl units . Incubation of MTase with starch or its constituents . i.e . amylose and amylopectin, led to the formation of a set of multiples of maltose (i.e . maltose, maltotetraose, maltohexaose etc.) . Malto-oligosaccharides with a degree of polymerization (DP) X were disproportionated to products with a DP of X +/- 2n (with X > or = 3 and n = 0,1,2,...) . Maximum activity in a 10-min assay was recorded at pH 6.5 and 85-90 degrees C . The enzyme displayed extraordinary resistance to thermal inactivation . For example, at 90, 85, and 70 degrees C (pH 6.5, 0.34 mg ml-1 protein), MTase half-lives of about 2.5 h, 17 h, and 21 days, respectively, were recorded . The gene for MTase, designated mmtA, was isolated from a gene library of T . maritima strain MSB8 . Analysis of the MTase primary structure as deduced from the nucleotide sequence of mmtA revealed that the enzyme is not closely related to known protein sequences . However, low-level local similarity between MTase and the alpha-amylase enzyme family (glycosyl hydrolase family 13) was detected, including conserved acidic residues essential for catalysis . Therefore, MTase should be assigned to this family . Based on detailed sequence analyses and comparison with amylolytic enzymes of known crystal structure we propose that MTase contains a (beta/alpha)8-fold as the core supersecondary structure which is typical for the alpha-amylase family . On the other hand, MTase is unique in that it lacks several residues highly conserved throughout this family . Also, MTase possesses an extraordinarily large domain B (a domain typical for the alpha-amylase family, inserted between beta-strand 3 and alpha-helix 3 of the (beta/alpha)8-barrel fold).

J Exp Med, 1999 Feb 15, 189(4), 647 - 56
Treponema pallidum major sheath protein homologue Tpr K is a target of opsonic antibody and the protective immune response; Centurion-Lara A et al.; We have identified a family of genes that code for targets for opsonic antibody and protective immunity in T . pallidum subspecies pallidum using two different approaches, subtraction hybridization and differential immunologic screening of a T . pallidum genomic library . Both approaches led to the identification of a polymorphic multicopy gene family with predicted amino acid homology to the major sheath protein of Treponema denticola . One of the members of this gene family, tpr K, codes for a protein that is predicted to have a cleavable signal peptide and be located in the outer membrane of the bacterium . Reverse transcription polymerase chain reaction analysis of T . pallidum reveals that Tpr K is preferentially transcribed in the Nichols strain of T . pallidum . Antibodies directed to purified recombinant variable domain of Tpr K can opsonize T . pallidum, Nichols strain, for phagocytosis, supporting the hypothesis that this portion of the protein is exposed at the surface of the treponeme . Immunization of rabbits with the purified recombinant variable domain of Tpr K provides significant protection against infection with the Nichols strain of T . pallidum . This gene family is hypothesized to be central to pathogenesis and immunity during syphilis infection.

J Med Microbiol, 1999 Feb, 48(2), 173 - 80
Saccharopolyspora rectivirgula from Quebec dairy barns: application of simplified criteria for the identification of an agent responsible for farmer's lung disease; Duchaine C et al.; Saccharopolyspora rectivirgula (Micropolyspora faeni) is one of the major agents responsible for farmer's lung disease, a form of hypersensitivity pneumonitis . It is frequently isolated from the air of contaminated barns . The identification of this actinomycete is difficult because most of its phenotypic characteristics are variable and classical tests are not easy to perform on actinomycetes . Fatty acid analysis is very useful for the identification of these strains, but is not available except in some research or reference laboratories . Morphological (microscopic and macroscopic observations), physiological and biochemical tests (growth properties; macromolecules degraded; citrate utilisation and acid production from carbohydrates; resistance to antibiotics, lysozyme and heat), cell wall and fatty acid analyses and IgG analyses with serum from patients with farmer's lung were performed on 12 environmental isolates presumed to be S . rectivirgula and two control strains of S . rectivirgula . From this, a simple and rapid scheme for the identification of this actinomycete is proposed: optimal growth temperature (55 degrees C); colony appearance based on morphology (filamentous) and colour (beige to orange-brown); microscopic morphology (chains of spores on both aerial and substrate mycelium); growth on NaCl 10%; cell-wall analysis (type IV); and the verification of antibody response with serum from a patient with farmer's lung . This last criterion is important to confirm the immunogenicity of the strains identified as S . rectivirgula . This scheme provides an accurate and efficient way of identifying S . rectivirgula strains and evaluating exposure to this bacterium . The study shows the limited value and the lack of reproducibility of some classical biochemical tests.

Nucleic Acids Res, 1999 Mar 1, 27(5), 1223 - 42
Conserved domains in DNA repair proteins and evolution of repair systems; Aravind L et al.; A detailed analysis of protein domains involved in DNA repair was performed by comparing the sequences of the repair proteins from two well-studied model organisms, the bacterium Escherichia coli and yeast Saccharomyces cerevisiae, to the entire sets of protein sequences encoded in completely sequenced genomes of bacteria, archaea and eukaryotes . Previously uncharacterized conserved domains involved in repair were identified, namely four families of nucleases and a family of eukaryotic repair proteins related to the proliferating cell nuclear antigen . In addition, a number of previously undetected occurrences of known conserved domains were detected; for example, a modified helix-hairpin-helix nucleic acid-binding domain in archaeal and eukaryotic RecA homologs . There is a limited repertoire of conserved domains, primarily ATPases and nucleases, nucleic acid-binding domains and adaptor (protein-protein interaction) domains that comprise the repair machinery in all cells, but very few of the repair proteins are represented by orthologs with conserved domain architecture across the three superkingdoms of life . Both the external environment of an organism and the internal environment of the cell, such as the chromatin superstructure in eukaryotes, seem to have a profound effect on the layout of the repair systems . Another factor that apparently has made a major contribution to the composition of the repair machinery is horizontal gene transfer, particularly the invasion of eukaryotic genomes by organellar genes, but also a number of likely transfer events between bacteria and archaea . Several additional general trends in the evolution of repair proteins were noticed; in particular, multiple, independent fusions of helicase and nuclease domains, and independent inactivation of enzymatic domains that apparently retain adaptor or regulatory functions.

J Mol Biol, 1999 Feb 19, 286(2), 365 - 74
ErmE methyltransferase recognizes features of the primary and secondary structure in a motif within domain V of 23 S rRNA; Villsen ID et al.; The Erm methyltransferases confer resistance to macrolide, lincosamide and streptogramin B (MLS) antibiotics by methylation of a single adenosine base within bacterial 23 S ribosomal RNA . The ErmE methyltransferase, from the macrolide-producing bacterium Saccharopolyspora erythraea, recognizes a motif within domain V of the rRNA that specifically targets adenosine 2058 (A2058) for methylation . Here, we define the structure of the RNA motif by a combination of molecular genetics and biochemical probing . The core of the motif has the primary sequence 2056-GGAHA-2060, where H is any nucleotide except guanosine, and ErmE methylates at the adenosine in bold . For efficient recognition by ErmE, this sequence must be displayed within a particular secondary structure . An irregular stem (helix 73) is required immediately 5' to A2058, with an unpaired nucleotide, preferably a cytidine residue, at position 2055 . Nucleotides 2611 to 2616 are collectively required to form part of the 3'-side of helix 73, but there is little or no restriction on the identities of individual nucleotides here . There are minor preferences in the identities of nucleotides 2051 to 2055 that are adjacent to the motif core, although their main role is in maintaining the irregular secondary structure . The essential elements of the ErmE motif are conserved in bacterial 23 S rRNAs, and thus presumably also form the recognition motif for other Erm methyltransferases .

J Immunol, 1999 Feb 15, 162(4), 2217 - 26
IL-12 administered during Chlamydia psittaci lung infection in mice confers immediate and long-term protection and reduces macrophage inflammatory protein-2 level and neutrophil infiltration in lung tissue; Huang J et al.; Protection against infections with the intracellular bacterium Chlamydia spp . requires Th1-polarized CD4+ T cell immunity . In BALB/c mouse lung infections, immediate innate and nascent Chlamydia-specific immune responses following intranasal inoculation of Chlamydia psittaci strain B577 were modulated by 7-day i.p . administration of murine rIL-12, the initiation cytokine for Th1 immunity . Treatment with IL-12 reduced the severity of chlamydial pneumonia, abolished mortality (37.5% in untreated mice), and significantly reduced numbers of chlamydial organisms in lungs . On day 4 after inoculation, the neutrophil:macrophage ratio in bronchointerstitial pneumonias was 1.96 in untreated mice and 0.51 in IL-12-treated mice . This immediate, IL-12-mediated shift in innate inflammatory phenotype was correlated with a significant reduction of lung concentrations of the neutrophil chemoattractant macrophage inflammatory protein (MIP)-2 (putative murine homologue of human IL-8), monocyte chemotactic protein-1, and TNF-alpha; and a reduction in MIP-1alpha and IFN-gamma, at high-dose infection only, and IL-12-independent IL-10 levels . Chlamydia-specific Ab titers and Ig isotype ratios indicated an IL-12-dependent Th1 shift . Recall responses of IL-12-primed mice to secondary chlamydial lung infection eliminated chlamydiae more effectively and generated a lung cytokine profile conducive to perpetuation of the Th1 memory population . These data support the hypothesis that genetic differences in endogenous IL-12 production and response pathways could determine disease outcomes characterized by poor chlamydial clearance and a purulent inflammatory infiltrate vs effective elimination of chlamydiae in a macrophage-dominated response.

J Bacteriol, 1999 Feb, 181(4), 1196 - 202
Two nucleotide transport proteins in Chlamydia trachomatis, one for net nucleoside triphosphate uptake and the other for transport of energy; Tjaden J et al.; The genome of Chlamydia trachomatis, one of the most prominent human pathogens, contains two structural genes coding for proteins, herein called Npt1Ct and Npt2Ct (nucleoside phosphate transporters 1 and 2 of C . trachomatis), exhibiting 68 and 61% similarity, respectively, to the ATP/ADP transporter from the intracellular bacterium Rickettsia prowazekii at the deduced amino acid level . Hydropathy analysis and sequence alignments suggested that both proteins have 12 transmembrane domains . The putative transporters were expressed as histidine-tagged proteins in Escherichia coli to study their biochemical properties . His10-Npt1Ct catalyzed ATP and ADP transport in an exchange mode . The apparent Km values were 48 (ATP) and 39 (ADP) microM . ATP and ADP transport was specific since AMP, GTP, CTP, UTP, dATP, dCTP, dGTP, and dTTP did not inhibit uptake . In contrast, His10-Npt2Ct transported all four ribonucleoside triphosphates with apparent Km values of 31 microM (GTP), 302 microM (UTP), 528 microM (CTP), and 1,158 microM (ATP) . Ribonucleoside di- and monophosphates and deoxyribonucleotides were not substrates . The protonophore m-chlorocarbonylcyanide phenylhydrazone abolished uptake of all nucleoside triphosphates by Npt2Ct . This observation indicated that His10-Npt2Ct acts as a nucleosidetriphosphate/H+ symporter energized by the proton motive force across the Escherichia coli cytoplasmic membrane . We conclude that Npt1Ct provides chlamydiae with energy whereas Npt2Ct catalyzes the net uptake of ribonucleoside triphosphates required for anabolic reactions.

J Bacteriol, 1999 Feb, 181(4), 1118 - 25
Cell cycle control of a holdfast attachment gene in Caulobacter crescentus; Janakiraman RS et al.; Attachment to surfaces by the prosthecate bacterium Caulobacter crescentus is mediated by an adhesive organelle, the holdfast, found at the tip of the stalk . Indirect evidence suggested that the holdfast first appears at the swarmer pole of the predivisional cell . We used fluorescently labeled lectin and transmission electron microscopy to detect the holdfast in different cell types . While the holdfast was readily detectable in stalked cells and at the stalked poles of predivisional cells, we were unable to detect the holdfast in swarmer cells or at the flagellated poles of predivisional cells . This suggests that exposure of the holdfast to the outside of the cell occurs during the differentiation of swarmer to stalked cells . To investigate the timing of holdfast synthesis and exposure to the outside of the cell, we have examined the regulation of a holdfast attachment gene, hfaA . The hfaA gene is part of a cluster of four genes (hfaABDC), identified in strain CB2A and involved in attachment of the holdfast to the polar region of the cell . We have identified the hfaA gene in the synchronizable C . crescentus strain CB15 . The sequence of the CB2A hfaA promoter suggested that it was regulated by sigma54 . We show that the transcription of hfaA from either strain is not dependent on sigma54 . Using a hfaA-lacZ fusion, we show that the transcription of hfaA is temporally regulated during the cell cycle, with maximal expression in late-predivisional cells . This increase in expression is largely due to the preferential transcription of hfaA in the swarmer pole of the predivisional cell.

J Bacteriol, 1999 Feb, 181(4), 1088 - 98
Presence of acetyl coenzyme A (CoA) carboxylase and propionyl-CoA carboxylase in autotrophic Crenarchaeota and indication for operation of a 3-hydroxypropionate cycle in autotrophic carbon fixation; Menendez C et al.; The pathway of autotrophic CO2 fixation was studied in the phototrophic bacterium Chloroflexus aurantiacus and in the aerobic thermoacidophilic archaeon Metallosphaera sedula . In both organisms, none of the key enzymes of the reductive pentose phosphate cycle, the reductive citric acid cycle, and the reductive acetyl coenzyme A (acetyl-CoA) pathway were detectable . However, cells contained the biotin-dependent acetyl-CoA carboxylase and propionyl-CoA carboxylase as well as phosphoenolpyruvate carboxylase . The specific enzyme activities of the carboxylases were high enough to explain the autotrophic growth rate via the 3-hydroxypropionate cycle . Extracts catalyzed the CO2-, MgATP-, and NADPH-dependent conversion of acetyl-CoA to 3-hydroxypropionate via malonyl-CoA and the conversion of this intermediate to succinate via propionyl-CoA . The labelled intermediates were detected in vitro with either 14CO2 or {14C}acetyl-CoA as precursor . These reactions are part of the 3-hydroxypropionate cycle, the autotrophic pathway proposed for C . aurantiacus . The investigation was extended to the autotrophic archaea Sulfolobus metallicus and Acidianus infernus, which showed acetyl-CoA and propionyl-CoA carboxylase activities in extracts of autotrophically grown cells . Acetyl-CoA carboxylase activity is unexpected in archaea since they do not contain fatty acids in their membranes . These aerobic archaea, as well as C . aurantiacus, were screened for biotin-containing proteins by the avidin-peroxidase test . They contained large amounts of a small biotin-carrying protein, which is most likely part of the acetyl-CoA and propionyl-CoA carboxylases . Other archaea reported to use one of the other known autotrophic pathways lacked such small biotin-containing proteins . These findings suggest that the aerobic autotrophic archaea M . sedula, S . metallicus, and A . infernus use a yet-to-be-defined 3-hydroxypropionate cycle for their autotrophic growth . Acetyl-CoA carboxylase and propionyl-CoA carboxylase are proposed to be the main CO2 fixation enzymes, and phosphoenolpyruvate carboxylase may have an anaplerotic function . The results also provide further support for the occurrence of the 3-hydroxypropionate cycle in C . aurantiacus.

Biosci Biotechnol Biochem, 1998 Dec, 62(12), 2446 - 9
Cloning and nucleotide sequence of the gene encoding a serine proteinase inhibitor named marinostatin from a marine bacterium, Alteromonas sp . strain B-10-31; Miyamoto K et al.; The gene (mstI) encoding a serine proteinase inhibitor named marinostatin from marine Alteromonas sp . strain B-10-31 was cloned and its nucleotide sequence was analyzed . A short open reading frame of 192 bp encoded 63 amino acids with a molecular weight of 6,985 . Furthermore, the initial product of marinostatin (marinostatin L) was purified and its amino acid sequence was analyzed . These results indicate that marinostatin is produced as a unique precursor consisting of the mature peptide and the leader peptide for an ATP-binding cassette (ABC) transporter, and furthermore the initial product of marinostatin is dehydrated and processed by proteolysis to give homologous forms of marinostatin.

Ter Arkh, 1998, 70(11), 28 - 31
{Isoenzymes of transferases in serum from diphtheria patients}; Fedianin IuP; AIM: Investigation of serum spectrum of isoenzymes AlAT, AsAT, GGT in diphtheria in adults . MATERIALS AND METHODS: The serum spectrum of cytoplasmic and mitochondrial isoenzymes AlAT, AsAT, GGT were studied in 12 bacterium carriers and 65 patients with diphtheria . Quite original evidence has been obtained on mechanism of hyperenzymemia . RESULTS: Diphtheria is associated with enhanced activity of cytoplasmic isoenzymes in correlation with intoxication severity . Spectrum of isoenzymes AlAT and especially GGT shifted to the side of mitochondrial fractions . There were no statistically significant differences in the activity and spectrum of isoenzymes in myocarditis and in its absence . In clinical recovery the spectrum of isoenzyme activity remained changed . CONCLUSION: It is suggested that dysisoenzymemia is related to active role of hepatocytes in utilization of diphtheria toxin, that carriers are not healthy people as hyperenzymemia is associated with morphological changes in the myocardium.

Can J Microbiol, 1998 Oct, 44(10), 920 - 8
Monoclonal antibodies to 2,4-dichlorophenol hydroxylase as probes for the 2,4-D-degradative phenotype; Farhana L et al.; Two different monoclonal antibodies (MAb) were raised against 2,4-dichlorophenol hydroxylase (DCP-hydroxylase) of Ralstonia eutropha JMP134 (pJP4), the second enzyme in the 2,4-D-degradative pathway of this bacterium . The utility of these antibodies in detecting and characterizing 2,4-D-degrading soil bacteria was investigated . One MAb (F6) reacted with DCP-hydroxylase from 27 out of 36 strains tested, while the other (MAb C3) reacted with only 17 isolates . When used with the colony blot technique, MAb F6 was useful for detecting cross-reacting strains on plates of pure cultures or of mixtures containing nondegraders even when 2,4-D degraders were outnumbered 60 to 1 . 2,4-D-degrading strains could also be detected from plates spread with enrichment cultures but not from primary isolation plates spread from soil dilutions, presumably because the ratio of degraders to nondegraders was too low . Colonies of some strains that were very distantly related genetically, but produced functionally similar DCP-hydroxlase enzymes, were detected by MAb F6 . This result suggests that MAbs could be useful for detecting functionally similar proteins expressed from tfdB analogs, even in the absence of detectable DNA homology between the genes encoding them.

Biochemistry, 1999 Jan 26, 38(4), 1276 - 83
Membrane-bound electron transfer chain of the thermohalophilic bacterium Rhodothermus marinus: characterization of the iron-sulfur centers from the dehydrogenases and investigation of the high-potential iron-sulfur protein function by in vitro reconstitution of the respiratory chain; Pereira MM et al.; Rhodothermus marinus, a thermohalophilic bacterium, has a unique electron-transfer chain, containing, besides a cbb3 and a caa3 terminal oxidases, a novel cytochrome bc complex {Pereira, M . M., Carita, J . N., and Teixeira, M . (1999) Biochemistry 38, 1268-1275} . The membrane-bound iron-sulfur centers of this bacterium were studied by electron paramagnetic resonance (EPR) spectroscopy, leading to the identification of its main electron-transfer complexes . The resonances typical for the Rieske-type centers are not detected . Clusters S1 and S3 from succinate dehydrogenase were identified; interestingly, center S3 is shown to be present in two different conformations, with g values at 2.035, 2.009, and 2.001 and at 2.025, 2.002, and 2.000 . Upon addition of NADH and dithionite, EPR signals assigned to resonances characteristic of binuclear and tetranuclear clusters develop and are attributed to the iron-sulfur centers of complexes I and II . A high-potential iron-sulfur protein- (HiPIP-) type center previously detected in the membranes of this bacterium {Pereira et al . (1994) FEBS Lett . 352, 327-330} is shown to belong indeed to a canonical HiPIP . This protein was purified and extensively characterized . It is a small water-soluble protein of approximately 10 kDa, containing a single {4Fe-4S}3+/2+ cluster . The reduction potential, determined by EPR redox titrations in intact and detergent-solubilized membranes as well as by cyclic voltammetry in solution, has a pH-independent value of 260 +/- 20 mV, in the range 6-9 . In vitro reconstitution of the R . marinus electron-transfer chain shows that the HiPIP plays a fundamental role in the chain, as the electron shuttle between R . marinus cytochrome bc complex and the caa3 terminal oxidase, being thus simultaneously identified a HiPIP reductase and a HiPIP oxidase.

Biochemistry, 1999 Jan 26, 38(4), 1268 - 75
Membrane-bound electron transfer chain of the thermohalophilic bacterium Rhodothermus marinus: a novel multihemic cytochrome bc, a new complex III; Pereira MM et al.; A novel multihemic cytochrome bc complex was isolated from the membranes of Rhodothermus marinus . It is a complex with a minimum of three subunits (43, 27, and 18 kDa), containing five low-spin heme centers of the B and C types, in a 1:4 ratio . All the C-type hemes are in the same subunit (27 kDa) . Three distinct redox transitions, at 235, 80, and -45 mV, were observed by visible redox titrations . The first involves one B- and one C-type hemes, and in the other two transitions one and two C-type hemes are involved, respectively . Spectroscopic data strongly suggest that the two hemes intervening in the last transition are in van der Waals contact, yielding a split Soret band . Electron paramagnetic resonance spectra of the oxidized complex show resonances of five low-spin ferric heme centers . Upon reduction with ascorbate, all these resonances vanish and a new one attributed to the last pair of hemes appears . A {3Fe-4S}1+/0 center copurifies with this complex, having a high reduction potential of +140 mV . No Rieske-type centers are detected in R . marinus and no effect is observed in the respiratory rates when the typical bc1 complex inhibitors are present, suggesting that such a complex is absent in R . marinus {Pereira et al . (1994) FEBS Lett . 352, 327-330} . The newly isolated cytochrome bc complex has quinol:cytochrome c or high-potential iron-sulfur protein (HiPIP) oxidoreductase activity, being a functional analogue of the canonical bc1 complexes; i.e., it is the complex III in R . marinus . This complex plays a central role in this bacterium's electron-transfer chain, coupling the electron transfer between the quinols reduced by the dehydrogenases and the HiPIP, the final electron donor to the terminal oxidases {Pereira, M . M., Carita, J . N., and Teixeira, M . (1999) Biochemistry 38, 1276-1283}.

J Clin Invest, 1999 Feb, 103(3), 407 - 12
Leukocyte infection by the granulocytic ehrlichiosis agent is linked to expression of a selectin ligand; Goodman JL et al.; Human granulocytic ehrlichiosis (HGE) is an emerging tickborne illness caused by an intracellular bacterium that infects neutrophils . Cells susceptible to HGE express sialylated Lewis x (CD15s), a ligand for cell selectins . We demonstrate that adhesion of HGE to both HL60 cells and normal bone marrow cells directly correlates with their CD15s expression . HGE infection of HL60 cells, bone marrow progenitors, granulocytes, and monocytes was blocked by monoclonal antibodies against CD15s . However, these antibodies did not inhibit HGE binding, and anti-CD15s was capable of inhibiting the growth of HGE after its entry into the target cell . In contrast, neuraminidase treatment of HL60 cells prevented both HGE binding and infection . A cloned cell line (HL60-A2), derived from HL60 cells and resistant to HGE, was deficient in the expression of alpha-(1, 3)fucosyltransferase (Fuc-TVII), an enzyme known to be required for CD15s biosynthesis . Less than 1% of HL60-A2 cells expressed CD15s, and only these rare CD15s-expressing cells bound HGE and became infected . After transfection with Fuc-TVII, cells regained CD15s expression, as well as their ability to bind HGE and become infected . Thus, CD15s expression is highly correlated with susceptibility to HGE, and it, and/or a closely related sialylated and alpha-(1,3) fucosylated molecule, plays a key role in HGE infection, an observation that may help explain the organism's tropism for leukocytes.

Genetics, 1999 Feb, 151(2), 569 - 84
Extracellular complementation and the identification of additional genes involved in aerial mycelium formation in Streptomyces coelicolor; Nodwell JR et al.; Morphogenesis in the bacterium Streptomyces coelicolor involves the formation of a lawn of hair-like aerial hyphae on the colony surface that stands up in the air and differentiates into chains of spores . bld mutants are defective in the formation of this aerial mycelium and grow as smooth, hairless colonies . When certain pairs of bld mutants are grown close to one another on rich sporulation medium, they exhibit extracellular complementation such that one mutant restores aerial mycelium formation to the other . The extracellular complementation relationships of most of the previously isolated bld mutants placed them in a hierarchy of extracellular complementation groups . We have screened for further bld mutants with precautions intended to maximize the discovery of additional genes . Most of the 50 newly isolated mutant strains occupy one of three of the previously described positions in the hierarchy, behaving like bldK, bldC, or bldD mutants . We show that the mutations in some of the strains that behave like bldK are bldK alleles but that others fall in a cluster at a position on the chromosome distinct from that of any known bld gene . We name this locus bldL . By introducing cloned genes into the strains that exhibit bldC or bldD-like extracellular complementation phenotypes, we show that most of these strains are likely to contain mutations in genes other than bldC or bldD . These results indicate that the genetic control of aerial mycelium formation is more complex than previously recognized and support the idea that a high proportion of bld genes are directly or indirectly involved in the production of substances that are exchanged between cells during morphological differentiation.

EMBO J, 1999 Feb 1, 18(3), 534 - 42
Supramolecular organization of the photosynthetic apparatus of Rhodobacter sphaeroides; Jungas C et al.; Native tubular membranes were purified from the purple non-sulfur bacterium Rhodobacter sphaeroides . These tubular structures contain all the membrane components of the photosynthetic apparatus, in the relative ratio of one cytochrome bc1 complex to two reaction centers, and approximately 24 bacteriochlorophyll molecules per reaction center . Electron micrographs of negative-stained membranes diffract up to 25 A and allow the calculation of a projection map at 20 A . The unit cell (a = 198 A, b = 120 A and gamma = 103 degrees) contains an elongated S-shaped supercomplex presenting a pseudo-2-fold symmetry . Comparison with density maps of isolated reaction center and light-harvesting complexes allowed interpretation of the projection map . Each supercomplex is composed of light-harvesting 1 complexes that take the form of two C-shaped structures of approximately 112 A in external diameter, facing each other on the open side and enclosing the two reaction centers . The remaining positive density is tentatively attributed to one cytochrome bc1 complex . These features shed new light on the association of the reaction center and the light-harvesting complexes . In particular, the organization of the light-harvesting complexes in C-shaped structures ensures an efficient exchange of ubihydroquinone/ubiquinone between the reaction center and the cytochrome bc1 complex.

Acta Gastroenterol Latinoam, 1998, 28(5), 335 - 6
{Helicobacter pylori detection by polymerase chain reaction in gastric juice and its correlation with the histology (Giemsa)}; Monti J et al.; HP infection is involved in the pathogenesis of several gastroduodenal diseases, as type B chronic gastritis, duodenal and gastric ulcer, MALT lymphoma and gastric cancer . The recent availability of molecular techniques, specifically the PCR, allow us to detect very low amounts of the bacterium . The aim of the study is to evaluate the presence of HP in gastric juice by PCR technique and to correlate this findings with histology (Giemsa) of gastric mucosa . Gastric juice PCR positive findings were found in 10/31 (32.3%) HP positive patients at histology . We concluded that HP in gastric juice is possible to detect by molecular techniques . In our study 32.3% of the patients showed the presence of HP in gastric juice.

Appl Environ Microbiol, 1999 Feb, 65(2), 611 - 7
Cold-active serine alkaline protease from the psychrotrophic bacterium Shewanella strain ac10: gene cloning and enzyme purification and characterization; Kulakova L et al.; The gene encoding serine alkaline protease (SapSh) of the psychrotrophic bacterium Shewanella strain Ac10 was cloned in Escherichia coli . The amino acid sequence deduced from the 2,442-bp nucleotide sequence revealed that the protein was 814 amino acids long and had an estimated molecular weight of 85,113 . SapSh exhibited sequence similarities with members of the subtilisin family of proteases, and there was a high level of conservation in the regions around a putative catalytic triad consisting of Asp-30, His-65, and Ser-369 . The amino acid sequence contained the following regions which were assigned on the basis of homology to previously described sequences: a signal peptide (26 residues), a propeptide (117 residues), and an extension up to the C terminus (about 250 residues) . Another feature of SapSh is the fact that the space between His-65 and Ser-369 is approximately 150 residues longer than the corresponding spaces in other proteases belonging to the subtilisin family . SapSh was purified to homogeneity from the culture supernatant of E . coli recombinant cells by affinity chromatography with a bacitracin-Sepharose column . The recombinant SapSh (rSapSh) was found to have a molecular weight of about 44,000 and to be highly active in the alkaline region (optimum pH, around 9.0) when azocasein and synthetic peptides were used as substrates . rSapSh was characterized by its high levels of activity at low temperatures; it was five times more active than subtilisin Carlsberg at temperatures ranging from 5 to 15 degreesC . The activation energy for hydrolysis of azocasein by rSapSh was much lower than the activation energy for hydrolysis of azocasein by the subtilisin . However, rSapSh was far less stable than the subtilisin.

Dis Aquat Organ, 1998 Nov 30, 34(3), 223 - 9
Development of a nested polymerase chain reaction for amplification of a sequence of the p57 gene of Renibacterium salmoninarum that provides a highly sensitive method for detection of the bacterium in salmonid kidney; Chase DM et al.; Nucleic acid-based assays have shown promise for diagnosing Renibacterium salmoninarum in tissues and body fluids of salmonids . Development of a nested polymerase chain reaction (PCR) method to detect a 320 bp DNA segment of the gene encoding the p57 protein of R . salmoninarum is described . Whereas a conventional PCR for a 383 bp segment of the p57 gene reliably detected 1000 R . salmoninarum cells per reaction in kidney tissue, the nested PCR detected as few as 10 R . salmoninarum per reaction in kidney tissue . Two DNA extraction methods for the nested PCR were compared and the correlation between replicate samples was generally higher in samples extracted by the QIAamp system compared with those extracted by the phenol/chloroform method . The specificity of the nested PCR was confirmed by testing DNA extracts of common bacterial fish pathogens and a panel of bacterial species reported to cause false-positive reactions in the enzyme-linked immunosorbent assay (ELISA) and the fluorescent antibody test (FAT) for R . salmoninarum . Kidney samples from 74 naturally infected chinook salmon were examined by the nested PCR, the ELISA, and the FAT, and the detected prevalences of R . salmoninarum were 61, 47, and 43%, respectively.

Curr Microbiol, 1999 Mar, 38(3), 176 - 82
Isolation of DNA from the uncultured "Candidatus Liberobacter" species associated with citrus Huanglongbing by RAPD; Hocquellet A et al.; "Candidatus Liberobacter," the uncultured bacterium associated with citrus Huanglongbing (HLB) disease, is an alpha-Proteobacteria, and two species, "Candidatus L . africanum" and "Candidatus L . asiaticum, " have been characterized by sequence analysis of the 16S rDNA and beta operon (rplKAJL-rpoBC) genes . These genes were isolated by PCR and random cloning of DNA from infected plants . However, this strategy is laborious and allowed selection of only three Liberobacter DNA fragments . In this paper, we described isolation of additional genes using Random Amplified Polymorphic DNA (RAPD) . In total, 102 random 10-mer primers were used in PCR reactions on healthy and Liberobacter-infected plant DNA . Eight DNA bands amplified from infected plant DNA were cloned and analyzed . Six of them were found to be part of the Liberobacter genome by sequence and hybridization experiments . On these DNA fragments, four genes were identified: nusG, pgm, omp, and a hypothetical protein gene . These results indicate that RAPD can be used to clone DNA of uncultured organisms.

Proc R Soc Lond B Biol Sci, 1998 Dec 22, 265(1413), 2407 - 13
Phylogeny of Wolbachia in filarial nematodes; Bandi C et al.; Intracellular bacteria have been observed in various species of filarial nematodes (family Onchocercidae) . The intracellular bacterium of the canine filaria Dirofilaria immitis has been shown to be closely related to Wolbachia, a rickettsia-like micro-organism that is widespread among arthropods . However, the relationships between endosymbionts of different filariae, and between these and the arthropod wolbachiae, appear not to have been studied . To address these issues we have examined ten species of filarial nematodes for the presence of Wolbachia . For nine species, all samples examined were PCR positive using primers specific for the ftsZ gene of Wolbachia . For one species, the examined samples were PCR negative . Sequences of the amplified ftsZ gene fragments of filarial wolbachiae fall into two clusters (C and D), which are distinct from the A and B clusters recognized for arthropod wolbachiae . These four lineages (A-D) are related in a star-like phylogeny, with higher nucleotide divergence observed between C and D wolbachiae than that observed between A and B wolbachiae . In addition, within each of the two lineages of filarial wolbachiae, the phylogeny of the symbionts is consistent with the host phylogeny . Thus, there is no evidence for recent Wolbachia transmission between arthropods and nematodes . Endosymbiont 16S ribosomal DNA sequences from a subset of filarial species support these findings.

Ann Biol Clin (Paris), 1999 Jan-Feb, 57(1), 29 - 36
{Bartonellosis . II . Other Bartonella responsible for human diseases}; Piemont Y et al.; In addition to Bartonella henselae, five other Bartonella species were involved in human pathology . As for B . henselae, ectoparasites seem to be responsible for the transmission of most or all these bacterial species . B . bacilliformis is responsible for Carrion's disease that occurs in some valleys of Colombia, Ecuador and Peru . This disease is transmitted by biting of infected sandflies . The bacterial reservoir is constituted by humans only . That disease occurs either as an acute form with severe infectious hemolytic anemia (or Oroya fever), or as benign cutaneous tumors, also called verruga peruana . Healthy blood carriers of the bacterium exist . Trench fever was described during the First World War . This non-lethal disease is constituted of recurrent febrile attacks associated particularly with osseous pains . The causative agent of the disease is B . quintana, transmitted by the body louse . Humans seem to be the reservoir of that bacterium . In some patients, B . quintana can be responsible for endocarditis, bacillary angiomatosis and chronic or recurrent bacteremia . Other human infections due to Bartonella sp . have been described: B . vinsonii, isolated from blood of small rodents, and B . elizabethae, the reservoir of which is currently unknown, can be responsible for endocardites . B . clarridgeiae (isolated from blood of 5% of pet cats and 17% of stray cats) may be responsible for human cat scratch disease . All these bartonelloses are diagnosed by non-standard blood culture or by in vitro DNA amplification or by serological testing . Their treatment requires tetracyclines or chloramphenicol or macrolides.

Curr Opin Rheumatol, 1999 Jan, 11(1), 17 - 23
Animal models of infection-mediated vasculitis; Dal Canto AJ et al.; The human vasculitides are idiopathic syndromes for which both autoimmune and infectious causes have been proposed . Although proof of a correlation between infection and human vasculitis would aid in patient management, it is difficult to confirm causality . To study infection-mediated vascular disease, different animal models have been developed . Infections with the bacterium Chlamydia pneumoniae, an RNA virus, and herpesviruses all cause vascular pathology and are reviewed here . Many aspects of the human diseases are recapitulated in these models, so that further animal studies may help elucidate mechanisms of infection-mediated vasculitis . Such results may improve management, and potentially prevention, of these important human diseases.

Infect Immun, 1999 Feb, 67(2), 862 - 70
Apoptosis in macrophages and alveolar epithelial cells during early stages of infection by Legionella pneumophila and its role in cytopathogenicity; Gao LY et al.; The hallmark of Legionnaires' disease is intracellular replication of Legionella pneumophila within cells in the alveolar spaces . Cytopathogenicity of this bacterium to the host cell has been well demonstrated, but the mechanisms of host cell death due to infection by L . pneumophila are not well understood . In this study, induction of apoptosis in macrophages and alveolar epithelial cells by L . pneumophila during early stages of infection was confirmed by using multiple criteria, including DNA fragmentation by agarose gel electrophoresis, terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling, surface exposure of phosphatidylserine, and cellular morphology by transmission electron microscopy . Induction of nuclear apoptosis in L . pneumophila-infected macrophages is mediated by activation of the caspase cascade death machinery . We provide genetic and biochemical evidence that L . pneumophila-induced apoptosis in macrophages and alveolar epithelial cells does not require intracellular bacterial replication or new protein synthesis . In addition, extracellular L . pneumophila is capable of inducing apoptosis . Furthermore, induction of apoptosis by L . pneumophila correlates with cytopathogenicity . We conclude that L . pneumophila-induced apoptosis in macrophages and alveolar epithelial cells plays an important role in cytopathogenicity to the host cell during early stages of infection.

N Engl J Med, 1999 Jan 21, 340(3), 184 - 9
Chronic Bartonella quintana bacteremia in homeless patients; Brouqui P et al.; BACKGROUND: Infection with Bartonella quintana can cause trench fever, endocarditis, bacillary angiomatosis, and peliosis . An outbreak of bacteremia due to B . quintana has been reported among homeless people in Seattle, and the seroprevalence is high among homeless people in both the United States and Europe . Body lice are known to be the vectors of B . quintana . METHODS: We studied all the homeless people who presented in 1997 to the emergency departments of the University Hospital, Marseilles, France . Blood was collected for microimmunofluorescence testing for antibodies against B . quintana and for culture of the bacterium . Body lice were collected and analyzed by the polymerase chain reaction and sequencing of a portion of the citrate synthase gene of B . quintana . RESULTS: In 10 of 71 homeless patients (14 percent), blood cultures were positive for B . quintana, and 21 of the patients (30 percent) had high titers of antibody against the organism . A total of 17 patients (24 percent) had evidence of recent infection (bacteremia or seroconversion) . Tests of lice from 3 of the 15 patients from whom they were collected were positive for B . quintana . The homeless people with B . quintana bacteremia were more likely to have been exposed to lice (P=0.002), were more likely to have headaches (P=0.03) and severe leg pain (P<0.001), and had lower platelet counts (P=0.006) than the homeless people who were seronegative for B . quintana and did not have bacteremia; 8 of the 10 patients with bacteremia were afebrile . Five patients had chronic bacteremia, as indicated by positive blood cultures over a period of several weeks . CONCLUSIONS: In an outbreak of urban trench fever among homeless people in Marseilles, B . quintana infections were associated with body lice in patients with nonspecific symptoms or no symptoms.

Aliment Pharmacol Ther, 1999 Jan, 13(1), 43 - 7
Optimal duration of therapy combining ranitidine bismuth citrate with clarithromycin and metronidazole in the eradication of Helicobacter pylori infection; Savarino V et al.; BACKGROUND: Ranitidine bismuth citrate (RBC) co-prescribed with clarithromycin and metronidazole for 1 week has been shown to be an effective eradicating regimen for Helicobacter pylori . AIM: To determine the optimal duration of this regimen . METHODS: A series of 165 dyspeptic patients were recruited for this randomized, open, parallel-group study . They were subdivided into three groups receiving RBC 400 mg b.d . plus clarithromycin 250 mg b.d . and metronidazole 500 mg b.d . for three different periods (4, 7 and 10 days) . H . pylori infection was assessed by the concomitant positivity of CLO-test and histology performed at the pre-entry endoscopy . The bacterium was considered eradicated on the basis of a negative 13C-urea breath test performed at least 28 days after the completion of treatment . RESULTS: The three subgroups were well matched and 16 patients dropped out of the study for many reasons (six in the 4-day, five in the 7-day and five in the 10-day treatment regimens) . Intention-to-treat cure rates were 60%, 84% and 85%, and the per-protocol rates 67%, 92% and 94% in the 4-day, 7-day and 10-day treatment regimens, respectively . There was a significant difference, P = 0.003-0.006 on intention-to-treat and P = 0.001-0 . 002 on per protocol analysis between the 4-day and the 7-day and the 4-day and the 10-day periods, respectively . The 7-day and 10-day periods did not differ from each other . Side-effects were reported in 9%, 14% and 20% of the 4-, 7- and 10-day regimens . They led to stopping treatment in four cases (one in the 7-day and three in the 10-day period) . There was no statistical difference among them . CONCLUSIONS: Reducing the duration of RBC-based triple therapy to 4 days provides a low and unacceptable rate of H . pylori eradication . As there is no difference between 7 and 10 days of treatment, 1 week represents the optimal time period for this kind of treatment, based on RBC plus two antibiotics.

J Bacteriol, 1999 Jan, 181(2), 666 - 9
Targeted mutagenesis by duplication insertion in the radioresistant bacterium Deinococcus radiodurans: radiation sensitivities of catalase (katA) and superoxide dismutase (sodA) mutants; Markillie LM et al.; Deinococcus radiodurans R1 is extremely resistant to both oxidative stress and ionizing radiation . A simple and general targeted mutagenesis method was developed to generate catalase (katA) and superoxide dismutase (sodA) mutants . Both mutants were shown to be more sensitive to ionizing radiation than the wild type.

Biochem J, 1999 Jan 15, 337 ( Pt 2), 243 - 51
Magnesium chelatase from Rhodobacter sphaeroides: initial characterization of the enzyme using purified subunits and evidence for a BchI-BchD complex; Gibson LC et al.; The enzyme magnesium-protoporphyrin IX chelatase (Mg chelatase) catalyses the insertion of Mg into protoporphyrin IX, the first committed step in (bacterio)chlorophyll biosynthesis . In the photosynthetic bacterium Rhodobacter sphaeroides, this reaction is catalysed by the products of the bchI, bchD and bchH genes . These genes have been expressed in Escherichia coli so that the BchI, BchD and BchH proteins are produced with N-terminal His6 affinity tags, which has led to the production of large amounts of highly purified, highly active Mg chelatase subunits from a single chromatography step . Furthermore, BchD has been purifed free of contamination with the chaperone GroEL, which had proven to be a problem in the past . BchD, present largely as an insoluble protein in E . coli, was purified in 6 M urea and refolded by addition of BchI, MgCl2 and ATP, yielding highly active protein . BchI/BchD mixtures prepared in this way were used in conjunction with BchH to determine the kinetic parameters of R . sphaeroides Mg chelatase for its natural substrates . We have been able to demonstrate for the first time that BchI and BchD form a complex, and that Mg2+ and ATP are required to establish and maintain this complex . Gel filtration data suggest that BchI and BchD form a complex of molecular mass 200 kDa in the presence of Mg2+ and ATP . Our data suggest that, in vivo, BchD is only folded correctly and maintained in its correct conformation in the presence of BchI, Mg2+ and ATP.

J Med Microbiol, 1998 Feb, 47(2), 159 - 68
Outer-membrane antigen expression by Moraxella (Branhamella) catarrhalis influences pulmonary clearance; Kyd JM et al.; Moraxella (Branhamella) catarrhalis is a common respiratory tract pathogen in man . The bacterium shows a strong tendency to form aggregates in vitro . A variant strain of M . catarrhalis that showed a reduced tendency to form aggregates was selected by successive in-vitro passage in broth culture from which aggregates had settled . The non-clumping variant strain showed alteration in expression of outer-membrane antigens, including the HMW-OMP, an outer-membrane protein of c . 200 kDa, outer-membrane protein CD and lipo-oligosaccharide . A mouse model for pulmonary challenge with M . catarrhalis revealed significant differences in the rate of clearance of the isogenic variant strains from the lung . The parent strain caused enhanced recruitment of neutrophils to the lung and more rapid clearance of bacteria from the lungs in comparison to the non-clumping variant . It is concluded that alteration of expression of surface molecules by M . catarrhalis has a significant impact in an in-vivo model of pulmonary clearance.

J Periodontal Res, 1998 Nov, 33(8), 509 - 16
Involvement of prostaglandin E2 and interleukin-1 alpha in the differentiation and survival of osteoclasts induced by lipopolysaccharide from Actinobacillus actinomycetemcomitans Y4; Ueda N et al.; Lipopolysaccharide (LPS) is a bacterial cell component that plays multifunctional roles in inflammatory reactions . LPS from various periodontal pathogens is supposed to be a major virulence factor of periodontal diseases . In the present study, we demonstrated that LPS from periodontopathic bacterium Actinobacillus actinomycetemcomitans Y4 (Y4 LPS) stimulated osteoclast formation in mouse bone marrow culture systems . Addition of anti-interleukin-1 alpha (IL-1 alpha) antibody or indomethacin in the marrow cultures resulted in the suppression of osteoclast differentiation . Quantitative analyses revealed that Y4 LPS stimulated the production of IL-1 alpha and prostaglandin E2 (PGE2) by bone marrow cells . Furthermore, an immunoblot analysis showed that Y4 LPS stimulated bone marrow cells to upregulate the expression of cyclooxygenase-2, a rate-limiting enzyme for the conversion of arachidonic acid to prostanoids . These findings suggest that both IL-1 alpha and PGE2 are involved in the LPS-mediated osteoclast differentiation . In addition, we found that Y4 LPS supported the survival of osteoclasts . Addition of anti-IL-1 alpha antibody in the osteoclast culture resulted in a reduction of osteoclast survival . Indomethacin, however, showed no effect on osteoclast survival . These findings suggest that the increased PGE2 and IL-1 alpha synthesis by bone marrow cells may play an important role in the differentiation and survival of osteoclasts induced by A . actinomycetemcomitans LPS.

Biophys J, 1999 Jan, 76(1 Pt 1), 580 - 7
Torque generated by the flagellar motor of Escherichia coli while driven backward; Berry RM et al.; The technique of electrorotation was used to apply torque to cells of the bacterium Escherichia coli tethered to glass coverslips by single flagella . Cells were made to rotate backward, that is, in the direction opposite to the rotation driven by the flagellar motor itself . The torque generated by the motor under these conditions was estimated using an analysis that explicitly considers the angular dependence of both the viscous drag coefficient of the cell and the torque produced by electrorotation . Motor torque varied approximately linearly with speed up to over 100 Hz in either direction, placing constraints on mechanisms for torque generation in which rates of proton transfer for backward rotation are limiting . These results, interpreted in the context of a simple three-state kinetic model, suggest that the rate-limiting step in the torque-generating cycle is a powerstroke in which motor rotation and dissipation of the energy available from proton transit occur synchronously.

Vet Microbiol, 1998 Nov, 64(1), 1 - 5
Survey of seroprevalence of Q fever in dogs in the southeast of France, French Guyana, Martinique, Senegal and the Ivory Coast; Boni M et al.; A serological survey was carried out on 429 dogs belonging to the French military in France, French Guyana, Martinique, Senegal and the Ivory Coast . Serology against phase I and II antigens of Coxiella burnetii, the intracellular zoonotic bacterium was performed using indirect immunofluorescence techniques . Specific antibodies were found in dogs from France (9.8%), Senegal (11.6%), Ivory Coast (8.3%), French Guyana (5.2%) but not in those from Martinique . The seroprevalence among 77 dogs who had contact with sheep compared with 352 dogs who had had no contact, demonstrated a significantly higher seroprevalence in the former . Our results indicate that dogs, living close to sheep, may be infected by Coxiella burnetii and should be considered as possible sources of infection for humans.

Appl Environ Microbiol, 1999 Jan, 65(1), 1 - 5
Fate of Escherichia coli O157:H7 on fresh-cut apple tissue and its potential for transmission by fruit flies; Janisiewicz WJ et al.; Pathogenic Escherichia coli O157:H7, as well as nonpathogenic strains ATCC 11775 and ATCC 23716, grew exponentially in wounds on Golden Delicious apple fruit . The exponential growth occurred over a longer time period on fruit inoculated with a lower concentration of the bacterium than on fruit inoculated with a higher concentration . The bacterium reached the maximum population supported in the wounds regardless of the initial inoculum concentrations . Populations of E . coli O157:H7 in various concentrations of sterilized apple juice and unsterilized cider declined over time and declined more quickly in diluted juice and cider . The decline was greater in the unsterilized cider than in juice, which may have resulted from the interaction of E . coli O157:H7 with natural populations of yeasts that increased with time . Experiments on the transmission of E . coli by fruit flies, collected from a compost pile of decaying apples and peaches, were conducted with strain F-11775, a fluorescent transformant of nonpathogenic E . coli ATCC 11775 . Fruit flies were easily contaminated externally and internally with E . coli F-11775 after contact with the bacterium source . The flies transmitted this bacterium to uncontaminated apple wounds, resulting in a high incidence of contaminated wounds . Populations of the bacterium in apple wounds increased significantly during the first 48 h after transmission . Further studies under commercial conditions are necessary to confirm these findings.

J Clin Gastroenterol, 1998, 27 Suppl 1, S159 - 62
Relationship between the eradication of Helicobacter pylori and the healing pattern of peptic ulcer; Suzuki J et al.; We investigated the relationship between the eradication of Helicobacter pylori and the stage after the eradication of this bacterium . Eighty-six patients with H . pylori who had gastric ulcer (n=45) or duodenal ulcer (n=41) were enrolled in the study . As eradication therapy, patients received 1,500 mg amoxicillin, 400 mg clarithromycin, and 30 mg lansoprazole for 2 weeks, followed by 30 mg lansoprazole for 6 weeks in patients with gastric ulcer or for 4 weeks in those with duodenal ulcer . The ulcer stages were evaluated using the indigocarmine method at the third and sixth month after entry . The overall eradication rate of H . pylori was 85% in this study . In the gastric ulcer group, 62% of H . pylori-eradicated patients and 37.5% of H . pylori-uneradicated patients showed the S2 stage (white scar) at the sixth month . In the duodenal ulcer group, 61% of H . pylori-eradicated patients showed the S2 stage and none of the H . pylori-uneradicated patients showed the S2 stage (p < 0.05) . We concluded that H . pylori eradication might promote not ulcer healing but maturity of the ulcer scar, especially in duodenal ulcer patients, and that this might also be related to the reduction of ulcer relapse.

J Appl Microbiol, 1998 Dec, 85(6), 941 - 8
Specific PCR primers from the 16S-23S rRNA spacer region for the rapid detection and identification of Actinobacillus seminis; Appuhamy S et al.; Actinobacillus seminis is a common cause of ovine epididymitis and ram infertility . The ability to detect and identify this organism promptly is important commercially for the quality control of ram semen samples . Actinobacillus seminis is a fastidious and slow-growing bacterium and primary isolation and presumptive identification can be difficult and time-consuming . In this study, two ribosomal operons, termed rrnA and rrnB, have been characterized in the A . seminis genome, and these contain one and two tRNAs, respectively, in the spacer region between the 16S and 23S rRNA genes . Species-specific primers for A . seminis were developed from the sequence of the spacer region of rrnB for the identification and detection of A . seminis by PCR . The PCR assay was specific for A . seminis and gave no amplification products with phenotypically similar organisms such as Histophilus ovis . Storage solution used to preserve semen for long-term storage was found to inhibit the PCR . Therefore, for diagnostic purposes, the assay would best be performed after primary isolation or perhaps on fresh semen prior to storage if obvious contamination is indicated.

Arch Microbiol, 1998 Dec, 171(1), 13 - 8
Effect of high sugar concentration on nitrogenase activity of Acetobacter diazotrophicus; Reis VM et al.; Acetobacter diazotrophicus is a nitrogen-fixing bacterium that growth inside sugar cane plant tissue where the sucrose concentration is approximately 10% . The influence of high sugar content on nitrogenase was measured in the presence of oxygen and of nitrogen added in the form of ammonium and amino acids . In all parameters analyzed, 10% sucrose protected nitrogenase against inhibition by oxygen, ammonium, some amino acids, and also to some extent by salt stress . The oxygen concentration at which inhibition occurred increased from 2 kPa in 1% glucose or gluconic acid, to 4 kPa (0.4 atm) in 10% sucrose . Nitrogenase activity was partially inhibited by increased ammonium levels (2.0, 5.0, and 10.0 mM) in the presence of 1% sucrose, but the cells maintained their nitrogenase activity at 10% sucrose . This could be explained by the slow ammonium assimilation by the cells in the presence of high sucrose concentrations, i.e., independent of its concentration between 2 and 10 mM, the assimilation of ammonium was reduced to one-third in cells grown with 10% sucrose . Some amino acids were also tested in the presence of 1 and 10% sucrose . Cells grown in 1% sucrose had their nitrogenase activity reduced by 50-98% in the presence of glutamic acid, glutamine, alanine, asparagine, or threonine, whereas with 10% sucrose, nitrogenase activity was increased by glutamic acid and was reduced by only 61-73% by the other amino acids . The effect of NaCl concentrations (0.0, 0.25, 0.5, 0.75, or 1.0%) was also studied at the two concentrations of sucrose . Nitrogenase activity and growth of A . diazotrophicus, which was visualized by the pellicle formation in semi-solid medium, showed sensitivity even to low NaCl concentrations, which was somewhat relieved at the higher sucrose level . These observations indicate different osmotolerance mechanisms for sucrose and salt.

Gastroenterology, 1999 Jan, 116(1), 90 - 6
Host specificity of Helicobacter pylori strains and host responses in experimentally challenged nonhuman primates; Dubois A et al.; BACKGROUND & AIMS: The specificity of colonization by Helicobacter pylori and complex host-bacterium interactions cannot be readily examined in humans . The aim of this study was to perform such analyses in rhesus monkeys . METHODS: Four animals that had been cured of natural H . pylori colonization were challenged with a mixture of 7 strains of human origin, and bacteria recovered during periodic videogastroscopy were DNA fingerprinted . RESULTS: Three animals carried mixtures of several strains for 4 months, after which strain J166 predominated . In the fourth animal, only strain J238 was isolated from the earliest phase of colonization through 7 months, but strain J166 again became predominant by 10 months after the challenge . Gastritis scores and plasma gastrin and anti-H . pylori immunoglobulin G titers reached levels observed in naturally colonized animals by 4 months after the challenge; however, no plasma immunoglobulin A response was observed up to 10 months . CONCLUSIONS: These results show that (1) natural colonization does not elicit protective immunity against subsequent H . pylori challenge; (2) individuals differ in susceptibility to different H . pylori strains during initial stages of colonization; and (3) certain strains are better suited than others for long-term survival in different hosts . These observations show the complexity of H . pylori-host interactions.

Plant Mol Biol, 1998 Dec, 38(6), 1011 - 9
Metabolic engineering of rice leading to biosynthesis of glycinebetaine and tolerance to salt and cold; Sakamoto A et al.; Genetically engineered rice (Oryza sativa L.) with the ability to synthesize glycinebetaine was established by introducing the codA gene for choline oxidase from the soil bacterium Arthrobacter globiformis . Levels of glycinebetaine were as high as 1 and 5 micromol per gram fresh weight of leaves in two types of transgenic plant in which choline oxidase was targeted to the chloroplasts (ChlCOD plants) and to the cytosol (CytCOD plants), respectively . Although treatment with 0.15 M NaCl {corrected} inhibited the growth of both wild-type and transgenic plants, the transgenic plants began to grow again at the normal rate after a significantly less time than the wild-type plants after elimination of the salt stress . Inactivation of photosynthesis, used as a measure of cellular damage, indicated that ChlCOD plants were more tolerant than CytCOD plants to photoinhibition under salt stress and low-temperature stress . These results indicated that the subcellular compartmentalization of the biosynthesis of glycinebetaine was a critical element in the efficient enhancement of tolerance to stress in the engineered plants.

Biochemistry (Mosc), 1998 Oct, 63(10), 1205 - 8
Effect of modification of histidine residues on organization of the pigment--protein complexes of chromatium minutissimum
Prokhorenko IR, Makhneva ZK, Erokhin YE.
The effect of diethyl pyrocarbonate on chromatophores and isolated pigment--protein complexes of Chromatium minutissimum was studied . It is shown that modification of histidine residues results in the destruction of the core antenna LHI (B880) and in a spectral shift from 850 to 830 nm in the peripheral antenna LHII (B800-850) . In the purple sulfur bacterium Chromatium minutissimum the pigment--protein complexes B800-B850 (peripheral antenna, LHII) and B880 (core antenna, LHI) collect and transmit the absorbed light energy to the reaction centers . The composition of pigments and proteins as well as primary structure of the majority of polypeptides in both types of complexes from various photosynthetic bacteria have been determined.

Nippon Jibiinkoka Gakkai Kaiho, 1998 Nov, 101(11), 1316 - 20
{Epidemiologic study of Chlamydia pneumoniae with ELISA}; Dake Y et al.; Chlamydia Pneumoniae (C . pneumoniae) is a bacterium involved in infections of the upper airway as well as in lower respiratory diseases . It has been reported that infections from this bacterium are prevalent worldwide and that the proportion of the population having the antibody is high . It is, however, difficult to identify the pathogen by routine bacterial examination because it is an obligate cytozoic bacterium . In addition, the examination has been feasible in only a limited number of laboratories because the determination of the serum antibody titer has required preparation of the antigen as well as sophisticated skill . We conducted an epidemiological survey of 320 healthy males and females in their 20s living in the three cities of Osaka, Kobe and Oita using "HITAZYME C . Pneumoniae", which is a recently developed kit for determination of anti-C . pneumoniae specific antibody . The statistical method used was the chi square test for the comparison of proportions . Mean proportion of the population showing positive antibody test was 58.1% . IgA antibody was positive in 42.8% and IgG in 46.5% of the above population . There were no statistically significant differences between districts of between genders in the percentages of cases positive for these antibodies . The results were comparable to those previously reported suggesting that C . pneumoniae is prevalent all over Japan . This kit was found to be an easy way to use the ELISA method and therefore to be clinically useful.

Biochemistry (Mosc), 1998 Oct, 63(10), 1209 - 15
Alpha-galactosidase of the marine bacterium Pseudoalteromonas sp . KMM 701; Bakunina IY et al.; An alpha-galactosidase that inactivates the group specificity of B erythrocytes (group III) of human blood and does not affect A erythrocytes (group II) was isolated from the marine bacterium Pseudoalteromonas sp . KMM 701 . The enzyme preparation did not contain lectin, hemolytic, sialidase, endoglycanase, or glycosidase activities . The enzyme is stable at 20 degreesC for 24 h, has pH optimum for catalysis within the range of 6.7-7.7, and is stable to high concentrations of NaCl . It is 4-fold more efficient than the alpha-galactosidase from green coffee beans . At pH 7.0 the Km for p-nitrophenyl-alpha-D-galactopyranoside is 0.29 mM . The molecular weight of the enzyme determined by gel-filtration is 195 +/- 5 kD . The alpha-galactosidase is denatured by urea and guanidine hydrochloride . Its activity does not depend on the presence of metal ions . It contains a sulfhydryl group essential for its catalytic activity.

J Bacteriol, 1999 Jan, 181(1), 262 - 9
Repair of oxidized bases in the extremely radiation-resistant bacterium Deinococcus radiodurans; Bauche C et al.; Deinococcus radiodurans is able to resist and survive extreme DNA damage induced by ionizing radiation and many other DNA-damaging agents . It is believed that it possesses highly efficient DNA repair mechanisms . To characterize the repair pathway of oxidized purines in this bacteria, we have purified, from crude extracts, proteins that recognize these oxidized bases . We report here that D . radiodurans possesses two proteins excising the oxidized purines (formamidopyrimidine and 8-oxoguanine) by a DNA glycosylase-a purinic/apyrimidine lyase mechanism . Moreover, one of those proteins is endowed with a thymine glycol DNA glycosylase activity . One of these proteins could be the homolog of the Escherichia coli Fpg enzyme, which confirms the existence of a base excision repair system in this bacteria.

J Bacteriol, 1999 Jan, 181(1), 100 - 6
Thioredoxin is involved in oxygen-regulated formation of the photosynthetic apparatus of Rhodobacter sphaeroides; Pasternak C et al.; Thioredoxin, a redox active protein, has been previously demonstrated to be essential for growth of the anoxygenic photosynthetic bacterium Rhodobacter sphaeroides . In the present study, the involvement of thioredoxin in the formation of the photosynthetic apparatus of R . sphaeroides WS8 was investigated by construction and analysis of a mutant strain disrupted for the chromosomal trxA copy and carrying a plasmid-borne copy of trxA under the control of the hybrid ptrc promoter inducible by IPTG (isopropyl-beta-D-thiogalactopyranoside) . This strain was viable in the absence of IPTG but was affected in pigmentation . When shifted from high to low oxygen tension conditions, the trxA mutant showed a reduced bacteriochlorophyll content in comparison to that of the wild type . Although thioredoxin is able to regulate aminolevulinic acid (ALA) synthase (the first enzyme of the tetrapyrrole biosynthetic pathway) activity by a dithiol-disulfide exchange, our mutant strain exhibited a level of ALA synthase activity identical to that of the wild type, suggesting that thioredoxin is involved in other steps to regulate the synthesis of the photosynthetic apparatus . Accordingly, we showed that the trxA mutation affects the oxygen-regulated expression of the puf operon encoding the pigment-binding proteins of the light-harvesting and reaction center complexes . Upon transition from aerobic to semiaerobic growth conditions, the maximal puf mRNA level was found to be 40 to 50% lower in the mutant strain than in the wild type . The stability of the puf transcripts was identical in both strains grown under low oxygen tension, indicating that the role of thioredoxin in regulating puf expression occurs at the transcriptional level.

Infect Immun, 1999 Jan, 67(1), 384 - 94
Interleukin-6 (IL-6) and IL-8 are induced in human oral epithelial cells in response to exposure to periodontopathic Eikenella corrodens; Yumoto H et al.; Periodontitis is the inflammatory response in periodontal tissues elicited by bacterial colonization in periodontal pockets . In this response, pocket epithelial cells are the first cells to come into contact with bacteria . To elucidate this mechanism, we determined the adherence of the periodontopathic bacterium Eikenella corrodens 1073, which has a GalNAc-sensitive lectin-like adhesin (EcLS), to a human oral epithelial carcinoma cell line (KB) and the induction of proinflammatory cytokine production in the cells following exposure to this bacterium in vitro . In the adherence assay, EcLS played a role as the adhesin of this bacterium in adherence to KB cells . In a reverse transcriptase PCR, significant interleukin-8 (IL-8) and IL-6 mRNA levels were induced in response to exposure to this bacterium . In an enzyme-linked immunosorbent assay after an 8-h bacterial exposure, the IL-8 and IL-6 protein levels were 13.5- and 8.3-fold higher than those in the nonexposed controls, respectively . These protein responses were time dependent . Interestingly, when E . corrodens was separated from KB cells by cell culture inserts, a slight stimulation of the IL-6 and IL-8 mRNA and secreted protein levels was seen . These results imply that the direct contact of E . corrodens 1073 with oral epithelial cells is not necessarily required for the stimulation of IL-6 and IL-8 secretion . We suggest that E . corrodens induces the epithelial cells to secrete proinflammatory cytokines which serve as an early signaling system to host immune and inflammatory cells in underlying connective tissues.

Infect Immun, 1999 Jan, 67(1), 286 - 93
Cloning and characterization of a novel membrane-associated antigenic protein of Helicobacter pylori; Yoshida M et al.; Infection by Helicobacter pylori, a noninvasive bacterium, induces chronic leukocyte infiltration in the stomach by still largely unknown molecular mechanisms . We investigated the possibility that a membrane protein of H . pylori induces an inflammatory reaction in the subepithelial tissue of the stomach . By generating an expression library of H . pylori chromosomal DNA and screening with rabbit antiserum raised to a membrane fraction of H . pylori and sera of infected patients, we cloned a 16.0-kDa protein (HP-MP1) which appeared to attach to the inner membrane of the H . pylori in a homodimeric form . Anti-HP-MP1 antibodies were detected in the sera of infected patients but not in those of uninfected controls . Coincubation of monocytes with recombinant HP-MP1 led to cell activation and production of interleukin-1alpha (IL-1alpha), tumor necrosis factor alpha, IL-8, and macrophage inflammatory protein 1alpha . The results indicate that HP-MP1 is an antigenic membrane-associated protein of H . pylori which potentially activates monocytes . This suggests that HP-MP1 may play roles in the pathogenesis of perpetual tissue inflammation associated with H . pylori infection.

Cas Lek Cesk, 1998 Oct 19, 137(20), 619 - 23
{Chlamydia trachomatis--a risk for reproductive capacity in women}; Rezacova J et al.; BACKGROUND: Chlamydia trachomatis is a bacterium which causes in man among others urogenital infections . So far no data were published pertaining to the herd immunity of the population to this infection in the Czech Republic . The objective of the present investigation was to summarize clinical and laboratory results of examined women and to evaluate the assembled data by statistical methods . Special attention was paid to patients examined on account of infertility . METHODS AND RESULTS: 506 women aged 17-50 years (mean age 28 years) were examined . They were divided into five groups (1-cervicitis, 2-pelvic pain, 3-pregnant, 4-sterile, 5-before UPT) . Group 4 (sterile) was further subdivided into three categories (4a IVF on account of tubal sterility, 4b IVF on account of non-tubal sterility and 4c others) . With regard to the domicile the women were divided into those residing in Prague and those from other towns and rural areas . In all antigenic examinations of smears from the endocervix were made and serological examinations of specific IgA, IgG antichlamydial antibodies . During collection of the smears the appearance of the cervix; discharge, bleeding and pain on examination were evaluated . For statistical evaluation the chi 2 test was used . In 57% of the women from the whole group positive IgG antibodies were detected suggesting a past or present infection . Differences of antigenic or serological positivity between the five different groups were not statistically significant . In category 4a a significantly higher positivity of IgG (2 alpha < 0.001) of antichlamydial antibodies was found (92% patients) than in categories 4b and 4c . 71% women had complaints during examination . The authors did not find a correlation between antigen- and serum positive women indifferent groups and their domicile . CONCLUSIONS: The results provide evidence of a high herd immunity of the population of the Czech Republic to Chlamyata infection . At the same time they prove that post-inflammatory changes caused by Chlamydia trachomatis are the most frequent cause of occlusion of the oviducts and tubal sterility.

FEMS Microbiol Rev, 1998 Oct, 22(4), 323 - 32
The genome of Treponema pallidum: new light on the agent of syphilis; Weinstock GM et al.; Treponema pallidum subsp, pallidum, the causative agent of the sexually transmitted disease syphilis, is a fastidious, microaerophilic obligate parasite of humans . This bacterium is one of the few prominent infectious agents that has not been cultured continuously in vitro and consequently relatively little is known about its virulence mechanisms at the molecular level . T . pallidum therefore represented an attractive candidate for genomic sequencing . The complete genome sequence of T . pallidum has now been completed and comprises 1,138,006 base pairs containing 1041 predicted protein coding sequences . An important goal of this project is to identify possible virulence factors . Analysis of the genome indicates a number of potential virulence factors including a family of 12 proteins related to the Msp protein of Treponema denticola, a number of putative hemolysins, as well as several other classes of proteins of interest . The results of this analysis are reviewed in this article and indicate the value of whole genome sequences for rapidly advancing knowledge of infectious agents.

Am J Chin Med, 1998, 26(3-4), 353 - 64
Effects of garlic compounds diallyl sulfide and diallyl disulfide on arylamine N-acetyltransferase activity in strains of Helicobacter pylori from peptic ulcer patients; Chung JG et al.; Arylamine N-acctyltransferase (NAT) activities with p-aminobenzoic acid (PABA) and 2-aminofluorene (2-AF) were determined in the bacterium Helicobacter pylori collected from peptic ulcer patients . Two assay systems were performed, one with cellular cytosols, the other with intact cell suspensions . Cytosols or suspensions of H . pylori with or without specific concentrations of diallyl sulfide (DAS) or diallyl disulfide (DADS) co-treatment showed different percentages of 2-AF and PABA acetylation . The data indicated that there was decreased NAT activity associated with increased levels of DAS or DADS in H . pylori cytosols and suspensions . Viability studies on H . pylori demonstrated that DAS or DADS elicited dose-dependent bactericide affects on H . pylori cultures . The data also indicated that DAS and DADS decreased the apparent values of K(m) and Vmax of NAT enzyme from H . pylori in both systems examined . This report is the first demonstration that garlic components can affect H . pylori growth and NAT activity.

J Biomater Sci Polym Ed, 1998, 9(12), 1361 - 73
Evaluating the interaction of bacteria with biomaterials using atomic force microscopy; Razatos A et al.; Bacterial infection of biomaterials represents one of the most important reasons for the failure of transdermal or implanted medical devices . The first and least understood step in biomaterial-associated infections is the initial interaction between bacteria and a surface . This initial interaction can be either attractive or repulsive depending on the physiochemical nature of the biological and synthetic surfaces, as well as the properties of the interstitial fluid . We have shown that atomic force microscopy (AFM) can be employed as an exquisitely sensitive and versatile tool for quantifying the interaction between bacteria and surfaces in physiological solutions . The forces of interaction between an AFM cantilever tip and a uniform lawn of bacteria immobilized on glass were determined . By comparing the interactions of cantilever tips with lawns of isogenic E . coli strains carrying genetic lesions that alter their cell surface composition, it was possible to evaluate the effect of macromolecules such as lipopolysaccharide and capsular polysaccharide on the adhesion process . Mutations that result in the synthesis of truncated lipopolysaccharide or in the overproduction of the negatively charged capsular polysaccharide colanic acid render the interaction of the bacteria with the AFM tip unfavorable due to increased electrostatic repulsion . Furthermore, AFM could be used to evaluate the adhesion of bacteria onto commercially relevant biomaterials . In one approach, micron-size polystyrene beads were attached to AFM tips which were then used to measure forces . Unfortunately, this approach is limited by the meager number of materials manufactured as beads of a size suitable for AFM measurements . As an alternative approach, AFM cantilever tips were coated with a confluent layer of bacteria and used to probe planar surfaces . In this configuration, AFM could be employed to measure the force of interaction between virtually any bacterium and surface of interest.

Psychother Psychosom Med Psychol, 1998 Nov, 48(11), 425 - 9
{Reductionism once again . The example of Helicobacter pylori}; Weiner H; The contrasting evidence for a multifactorial pathogenesis of gastroduodenal disease rather than the reductionistic, monocausal role of Helicobacter pylori is presented . Evidence for the former is derived from epidemiological, physiological, immunological and experimental behavioral studies in animals . The high prevalence of the bacterium in popularies and the low incidence of peptic ulcer strongly suggests that it alone cannot play the only pathogenetic role . The evidence that peptic ulcer is not one disease raises the problem of identifying the contributions of psychosocial factors in combination with infection by H . pylori in the various forms of the human disease . The most likely role it plays is that of an "opportunist", when the gastroduodenal mucosal "defense" is compromised by many possible factors.

J Biol Chem, 1998 Dec 25, 273(52), 35319 - 25
AppA, a redox regulator of photosystem formation in Rhodobacter sphaeroides 2.4.1, is a flavoprotein . Identification of a novel fad binding domain; Gomelsky M et al.; The AppA protein is required for increased photosystem gene expression upon transition of the facultatively photoheterotrophic bacterium Rhodobacter sphaeroides 2.4.1 from aerobic to anaerobic photosynthetic conditions . AppA shows no obvious similarity to proteins with established function . Genetic evidence suggests that its effect is exerted through modulation of the activity of the repressor PpsR, which controls expression of multiple photosystem genes . To gain insight into the nature of AppA involvement in redox-dependent photosystem gene expression, the appA gene was overexpressed in Escherichia coli . AppA was produced as insoluble inclusion bodies . The purified inclusion bodies were found to contain FAD . By overexpressing various deletion derivatives, we were able to localize the region of AppA sufficient for FAD binding to approximately 120 amino-terminal residues . To assess the role of FAD binding in AppA function, we constructed an AppA derivative lacking the entire FAD binding domain . Surprisingly, this derivative complemented the AppA null mutant undergoing transition from aerobic to anaerobic photosynthetic growth conditions almost to the same extent as the full-length AppA protein . When the sequence of the amino-terminal portion of AppA was examined, it was shown not to contain any known flavin binding motifs . However, two open reading frames of unknown function, showing significant similarity to the amino terminus of AppA, were identified, i.e . Synechocystis sp . Srl1694 and E . coli F403 . The latter gene was amplified and overexpressed in E . coli, and the partially purified F403 protein was found to contain FAD as a cofactor . We have therefore concluded that the amino terminus of AppA represents a novel FAD binding domain present in a small group of bacterial proteins . The binding of FAD by AppA may be the first clue as to how this regulatory protein is involved in redox-regulated reactions.

Mol Cells, 1998 Oct 31, 8(5), 524 - 9
Cloning and characterization of ribulose bisphosphate carboxylase gene of a carboxydobacterium, hydrogenophagea pseudoflava DSM 1084; Lee SN et al.; The ribulose bisphosphate carboxylase/oxygenase rbcL and rbcS genes of a carbon monoxide-oxidizing bacterium, Hydrogenophaga pseudoflava DSM 1084, were cloned and sequenced . The cloned rbcL and rbcS genes had open reading frames of 1422 and 351 nucleotides encoding RbcL and RbcS with calculated molecular masses of 52,689 and 13,541, respectively . The known active site residues in other RbcL proteins were conserved in the H . pseudoflava proteins . The H . pseudoflava RbcS protein lacked the 12-residue internal sequence found in the plant enzymes . The 2 genes were separated by a 134 bp intergenic region and cotranscribed as a 2.0 kb rbcLS mRNA . Novel two perfect 9 bp direct repeats overlapping with two dyad symmetries were found in the rbcLS promoter region.

Int J Mol Med, 1998 May, 1(5), 817 - 22
Inhibition in a microgravity environment of the recovery of Escherichia coli cells damaged by heavy ion beams during the NASDA ISS phase I program of NASA Shuttle/Mir mission no . 6; Harada K et al.; We participated in a space experiment, part of the National Space Development Agency of Japan (NASDA) Phase I Space Radiation Environment Measurement Program, conducted during the National Aeronautics and Space Administration (NASA) Shuttle/Mir Mission No . 6 (S/MM-6) project . The aim of our study was to investigate the effects of microgravity on the DNA repair processes of living organisms in the <Realtime Radiation Monitoring Device III (RRMD III)> in orbit . Heavy ion beam radiation- or c-irradiation-damaged biological samples of Escherichia coli and the radioresistant bacterium Deinococcus radiodurans were prepared and placed in a biospecimen box, which was loaded into the RRMD III sensor unit of the Space Shuttle . Two identical sets of samples were left in the Spacehab's Payload Processing Facility (SPPF) in Florida, USA, as a control . (flight No . STS-84) was launched from NASA John F . Kennedy Space Center (KSC) in Florida, USA, on May 15, 1997 . The mission duration was 9.22 days . An astronaut activated the biological samples in the biospecimen box in the Spacehab during orbit in order to start repair of the DNA damaged by heavy ion beams or c-irradiation and the samples were incubated for 19 h 35 min at about 22uC, the cabin temperature . The control specimens in the SPPF were subjected to the same treatment under terrestrial gravity . After returned to earth, we investigated cell recovery by comparing the repair of the radiation-damaged DNA of E . coli and D . radiodurans in the microgravity environment in space with that on Earth . The results indicated that the DNA repair process of E . coli, but not of D . radiodurans, cells was inhibited in a microgravity environment.

J Bacteriol, 1998 Dec, 180(24), 6544 - 50
Inhibition of development of Myxococcus xanthus by eukaryotic protein kinase inhibitors; Jain R et al.; Myxococcus xanthus is a social bacterium that lives in the soil and undergoes spectacular development to form multicellular fruiting bodies . It contains a large family of eukaryote-like serine/threonine protein kinases . We found that a number of inhibitors for eukaryotic protein serine, threonine, and tyrosine kinases could inhibit the development and sporulation of M . xanthus to various degrees . These results suggest that serine/threonine and tyrosine phosphorylation may be involved in development of M . xanthus . None of the inhibitors tested had any effect on vegetative growth of M . xanthus . Most of them seemed to act during the early stages of development . However, the expression of a very early development-specific gene, Omega4521, was not significantly affected by the inhibitors . The patterns of protein phosphorylation during development were also not significantly altered by the inhibitors, suggesting that the targets of the inhibitors are minor or unstable phosphoproteins but play key roles in fruiting-body formation in M . xanthus.

Antonie Van Leeuwenhoek, 1998 May, 73(4), 299 - 313
The strategy of Myxococcus xanthus for group cooperative behavior; Koch AL; New evidence has been presented from our laboratory that the gliding bacterium, Myxococcus xanthus, does not home by chemotaxis toward a nutrient source . Our experiments, those of others, and the theory presented here combine to suggest a model, called the 'Pied Piper' model . It hypothesizes a gene that has a high mutation rate forward and back (say something in excess 10(-4) mutations per cell generation) which leads to switching between two motility states . Occasionally rare organisms become genetically, but reversibly, changed so that they move unidirectionally instead of mostly forward and back as do the bulk of the cells . When such a 'leader' cell arises, it continues to move in its original orientation, and causes a cohort of cells to move together away from the bulk of the cells . That is, in the less common mutational state it counteracts the usual tendency to just move forward and backward achieving little net movement . The assumption of a genetic element that mutates in a reversible way is suggested by numerous cases of reversible switches now known in a wide range of bacteria serving a variety of functions . A second aspect of the model is that mechanisms exist that cause cells to move in the same direction as their nearby neighbors . This process results in a regular spacing of bands of cells to form mounds in the absence of a leader . The action of C-factor, a factor-secreted by the cells which has been largely studied in the laboratory of Dale Kaiser, and extracellular fibrils, (rod-shaped protein and carbohydrate bodies) largely studied in the laboratory of Martin Dworkin, may be key elements in coordinating (or linking) the movements of neighboring cells . Based on the assumption of the absence of chemotaxis, computer simulations of pattern formation for gliding bacterial swarms and flares are consistent with observed behaviors and thus are additional evidence that chemotactic motility of the type exhibited by Escherichia coli, is not necessary for the group movements of M . xanthus . Some tests for this model are suggested.

Int, J . Angiol. . 1998 Aug, 7(4), 310 - 2
Primary Headache and Helicobacter Pylori; Gasbarrini A et al.; H . pylori infection has recently been associated with various vascular disorders . The aim of this study was to investigate its role in primary headache, a pathology strictly associated with vascular alterations . A total of 200 subjects affected by primary headache were evaluated . H . pylori infection was diagnosed by the 13C urea breath test . Headache was classified in tension-type headache, cluster headache, and migraine with or without aura . Prevalence of H . pylori infection and gastrointestinal (GI) symptoms were evaluated . H . pylori infection was found in 40% of the patients; prevalence of migraine without aura was found to be significantly greater in infected patients . The positive group showed no significant differences in the prevalence of the GI symptoms evaluated . In 30 infected patients, it was assessed whether the eradication of the bacterium was able to reduce frequency, intensity, and duration of clinical attacks of headache . After eradication, clinical attacks of headache completely disappeared in 17% of patients . Moreover, intensity, duration, and frequency of headache attacks were reduced in 69% of the remaining subjects . In conclusion, H . pylori infection is common in primary headache; bacterium eradication appears to be related to a significant reduction in clinical attacks of the disease.

Int, J . Angiol. . 1998 Aug, 7(4), 307 - 9
Raynaud's Phenomenon and Helicobacter Pylori Infection; Gasbarrini A et al.; Raynaud's phenomenon is an intermittent vasospasm of the arterioles of the distal limbs which occurs mostly following exposure to cold or emotional stimuli . Helicobacter pylori infection, the most relevant cause of gastritis, has been associated with some vascular diseases . The aim of this study was to assess the prevalence of H . pylori infection and effects of eradication on Raynaud attacks . Twenty-five patients affected by primary Raynaud's phenomenon were evaluated . H . pylori infection was assessed by the 13C urea breath test and 20 subjects were found to be infected by the bacterium . Triple therapy consisting of amoxicillin, clarithromycin, and lansoprazole was given to the patients for 7 days at the time of diagnosis . Duration and frequency of clinical attacks of Raynaud's phenomenon per week were assessed for 12 weeks after eradication . In 80 percent of the patients Raynaud's was eradicated after therapy . Clinical attacks of Raynaud's phenomenon completely disappeared in 18% of the eradicated patients . Duration and frequency of attacks were significantly reduced in 68% of the remaining patients . The preliminary findings from this study show that H . pylori eradication significantly ameliorates primary Raynaud's disease.

J Colloid Interface Sci, 1998 Sep 1, 205(1), 89 - 96
Influence of Divalent Cations and pH on Adsorption of a Bacterial Polysaccharide Adhesin; Bhosle N et al.; Hyphomonas MHS-3 (MHS-3) elaborates a diffuse capsular material, primarily composed of polysaccharide, which has been implicated to serve as the holdfast of this prosthecate marine bacterium . A purified polysaccharide (fr2ps) from this capsular material exhibits a relatively large affinity for (Ge), or more precisely for the Ge oxide surface film . In its natural habitat MHS-3 attaches to marine sediments . This suggests that molecular properties of fr2ps have evolved to render it adhesive toward mineral oxides . In order to characterize these molecular interactions, the effect of divalent cations and pH on the adsorption of fr2ps to Ge has been measured using attenuated total internal reflection Fourier transform infrared (ATR/FT-IR) spectroscopy . The effect of adsorption of fr2ps on the Ge oxide film has been investigated using X-ray photoelectron spectroscopy (XPS) . The results indicate that divalent cations participate in binding of fr2ps to Ge oxide and that atomic size of the cation is important . Evidence for significant participation of hydrogen bonding to the oxide surface is lacking .

Nucleic Acids Res, 1999 Jan 1, 27(1), 61 - 2
RsGDB, the Rhodobacter sphaeroides Genome Database; Choudhary M et al.; This report provides a summary of the sequencing project of the small chromosome (CII) of Rhodobacter sphaeroides 2.4.1(T),and introduces the first version of the genome database of this bacterium . The database organizes and describes diverse sets of biological information . The main role of the R.sphaeroides genome database (RsGDB) is to provide public access to the collected genomic information for R.sphaeroides via the World-Wide Web at The database allows the user access to hundreds of low redundancy R.sphaeroides sequences for further database searching, a summary of our current search results, and other allied information pertaining to this bacterium.

Microbiology, 1998 Nov, 144 ( Pt 11), 3019 - 26
A lipid A-associated protein of Porphyromonas gingivalis, derived from the haemagglutinating domain of the RI protease gene family, is a potent stimulator of interleukin 6 synthesis; Sharp L et al.; There is evidence that the lipid A-associated proteins (LAPs) of enteric bacteria can induce the synthesis of interleukin 1 (IL-1) and therefore may be important virulence factors . Porphyromonas gingivalis is now recognized as a major pathogen in the chronic inflammatory periodontal diseases and it has previously been reported that a crude LAP fraction from this organism could induce IL-1 and interleukin 6 (IL-6) synthesis . In the present study the chemical and biological properties of the LAPs of this bacterium have been further characterized . Analysis by SDS-PAGE has shown that the LAPs comprise nine proteins of molecular masses 81, 68, 48, 47, 28, 25, 20, 17 and 16 kDa . These LAPs, at concentrations as low as 100 ng ml(-1), were shown to stimulate human gingival fibroblasts, human peripheral blood mononuclear cells and whole human blood to produce the pro-inflammatory cytokine IL-6 . The cytokine-inducing activity of the LAPs was reduced after heat-inactivation and trypsinization, suggesting that the activity was not due to contaminating LPS . To establish which proteins in this mixture were the active cytokine inducers, the LAPs were separated by electrophoresis on polyacrylamide gels . The majority of the activity was associated with the 17 kDa LAP . N-terminal sequence analysis demonstrated that this protein was homologous to an internal region of a conserved adhesin domain contained within a family of P . gingivalis extracellular proteins including the RI protease, Lys-gingipain, porphypain and haemagglutinin A . In addition to a role in adherence, the adhesin domain(s) of these proteins may also have cytokine-inducing properties . Furthermore, it has also been shown that the previously observed degradation of cytokines by P . gingivalis may be attributable to the catalytic domain of the RI protease . Thus, different domains within the same molecule appear to have opposing actions on pro-inflammatory cytokine levels and the balance between these two activities may influence the cytokine status of the periodontium in patients with the common chronic inflammatory conditions known as the periodontal diseases.

Biochem Mol Biol Int, 1998 Nov, 46(4), 807 - 19
Sequence of electron carriers in the process of methanol oxidation by a new obligate methylotrophic bacterium; Yang SS et al.; From pink soluble fractions prepared from cells cultured in a copper-free medium, active methanol dehydrogenase (MDH) and two soluble c-type cytochromes (c-I and c-II) were purified homogeneously . The green fractions from cells grown on a medium containing 1.0 mg/l of copper had inactive MDH, cytochrome c-II, and blue copper protein . The amount of copper retained in the blue copper protein increased with cultivation time . The oxidized blue copper protein was similar to the classical type I blue copper proteins since it had the novel absorption peak at 625 nm . However, when the blue protein was reduced with MDH or dithionite, it showed the same spectrum as ferrocytochrome c-I . The isoelectric points of cytochrome c-I, blue copper protein and cytochrome c-II were 9.08, 9.08 and 6.52, respectively . These results suggest that the identity of the purified blue copper protein is cytochrome c-I, and copper ions bind to the cytochrome as methanol is depleted in the culture medium . In addition, MDH activity was not detected at all in the methanol-depleted condition . The data suggest that blue copper protein acts as a negative regulator for MDH . The electrons were transferred as follows: MDH-->cytochrome c-II-->cytochrome c-I (blue copper protein) . It was also revealed that the initial 'docking' of MDH and cytochrome c-II is accompanied by electrostatic interactions between lysine or arginine residues on the alpha-subunit of MDH and carboxyl groups on the cytochrome c-II.

Proc Natl Acad Sci U S A, 1998 Dec 8, 95(25), 14658 - 63
Structural assignments to the Mycoplasma genitalium proteins show extensive gene duplications and domain rearrangements; Teichmann SA et al.; The parasitic bacterium Mycoplasma genitalium has a small, reduced genome with close to a basic set of genes . As a first step toward determining the families of protein domains that form the products of these genes, we have used the multiple sequence programs PSI-BLAST and GEANFAMMER to match the sequences of the 467 gene products of M . genitalium to the sequences of the domains that form proteins of known structure {Protein Data Bank (PDB) sequences} . PDB sequences (274) match all of 106 M . genitalium sequences and some parts of another 85; thus, 41% of its total sequences are matched in all or part . The evolutionary relationships of the PDB domains that match M . genitalium are described in the structural classification of proteins (SCOP) database . Using this information, we show that the domains in the matched M . genitalium sequences come from 114 superfamilies and that 58% of them have arisen by gene duplication . This level of duplication is more than twice that found by using pairwise sequence comparisons . The PDB domain matches also describe the domain structure of the matched sequences: just over a quarter contain one domain and the rest have combinations of two or more domains.

Curr Microbiol, 1999 Jan, 38(1), 51 - 6
Metabolism of L-phenylalanine and L-tyrosine by the phototrophic bacterium Rhodobacter capsulatus; Saez LP et al.; The phototrophic bacterium Rhodobacter capsulatus utilizes the aromatic amino acids L-phenylalanine and L-tyrosine as nitrogen source . L-Phenylalanine is hydroxylated to L-tyrosine, which is further converted into p-hydroxyphenyl pyruvate (pHPP) by a transamination reaction . The bacterium is unable to grow at the expense of these amino acids as the sole carbon source, although it is able to degrade them to homogentisate, probably by unspecific hydroxylation reactions . Metabolization of L-phenylalanine or L-tyrosine as nitrogen source requires phototrophic growth conditions and does not produce free ammonium inside the cells . A low aminotransferase activity with 2-oxoglutarate and L-tyrosine as substrates can be detected in crude extracts of R . capsulatus . Uptake of both amino acids by R . capsulatus was completely inhibited by ammonium addition, which also prevents aminotransferase induction.

J Gastroenterol, 1998, 33 Suppl 10, 48 - 52
Are there more clinically important complications of Helicobacter pylori infection than peptic ulcer disease? A review of current literature; Unge P; Helicobacter pylori gastritis (i.e . H . pylori infection and complications) is a focus of tremendous research activity today . Besides peptic ulcer disease, a large number of reports suggest that other diseases are associated with H . pylori . The International Agency for Research on Cancer sponsored by the World Health organization classified the bacterium as a group I carcinogen in 1994 . Population-based studies of H . pylori and gastric cancer in 1991 showed an increased odds ratio, of 3-6, in infected patients, and a calculation of odds ratios in different age groups showed a markedly increased odds ratio, to about 20, in younger ages . Studies of non-ulcer dyspepsia and the effect of cure of H . pylori show either none, small, or significant symptom relief, suggesting a positive effect in a subgroup of non-ulcer dyspepsia patients . Mucosa-associated lymphoid tissue-lymphoma caused by H . pylori could be eradicated, at least in its mild forms . Barrett's ulcer is a possible H . pylori-associated disease as well as gastroesophageal reflux disease . Normal feedback in the acid regulation system is changed in infected patients, which may facilitate an increased gastroesophageal acidic reflux . Gastropathy and/or peptic ulcer due to use of nonsteroidal antiinflammatory drugs is probably aggravated by the infection . The infectious disease H . pylori gastritis is associated with a large number of complications, some of which are serious . There are no data showing any advantages of the infection . Giving anti-H . pylori therapy to infected patients should be regarded as essential.

Ital J Gastroenterol Hepatol, 1998 Oct, 30(5), 484 - 9
Polyamine profile in human gastric mucosa infected by Helicobacter pylori; Linsalata M et al.; BACKGROUND: Several studies have demonstrated increased gastric epithelial cell proliferation associated with Helicobacter pylori infection, which is reversed after bacterium eradication . Among the substances involved in cell proliferation and differentiation, polyamines are a group of polycations found in high concentrations both in normal and neoplastic cells . AIMS: Of the study were: a) to examine the influence of Helicobacter pylori infection on the polyamine profile in the gastric antrum and body, by comparing infected, to uninfected, patients, b) to evaluate the effect of successful and unsuccessful bacterium eradication on polyamine levels . PATIENTS AND METHODS: Twenty-six consecutive dyspeptic patients (20 Helicobacter pylori positive and 6 Helicobacter pylori negative) undergoing gastroscopy were enrolled . Polyamines were evaluated in antral and body biopsies by High Performance Liquid Chromatography . RESULTS: Antral and body biopsies from Helicobacter pylori positive patients contained higher polyamine levels than those from Helicobacter pylori negative subjects . In Helicobacter pylori positive patients, the baseline polyamine levels were higher in the antrum than in the body . In Helicobacter pylori negative subjects, levels in the two stomach regions were similar . After therapy, polyamine levels decreased in patients with successful eradication, whereas these levels remained unchanged in patients in whom infection persisted . CONCLUSIONS: These findings indicate enhanced antral cellular proliferation linked to the presence of Helicobacter pylori and add weight to the postulation of an association between Helicobacter pylori infection and increased risk of neoplastic changes in gastric antral mucosa . Differences in antral and body levels of polyamines may also be considered as a further indication of the different mucosal reactivity between the two regions of the stomach towards bacterial invasion.

Ital J Gastroenterol Hepatol, 1998 Oct, 30(5), 478 - 80
Autoantibodies reacting with gastric antigens in Helicobacter pylori associated body gastritis of dyspeptic children; Ierardi E et al.; BACKGROUND: Helicobacter pylori induced antibodies reacting with fundal mucosa have been shown to be involved in the pathogenesis of chronic atrophic gastritis in adults . Furthermore, previous reports have indicated that Helicobacter pylori increases the risk of gastric carcinoma, suggesting that the bacterium plays a role in mucosal changes representing especially in adulthood important steps in the progression from gastritis to cancer . PATIENTS AND METHODS: We investigated 16 Helicobacter pylori+ children from a series of 53 dyspeptic patients . Diagnosis of Helicobacter pylori infection was based on the positivity of at least 3 of the following tests: serology, 13C-urea breath test, rapid urease test and histology . Autoreaction was detected by incubation of gastric body sections with autologous sera and revealed by immunohistochemistry . Positive sera were tested with samples of unaffected gastric body to exclude a link with residual bacterial antigens . Proliferating cell nuclear antigen immunohistostain was also performed to evaluate the epithelial proliferative state . RESULTS: Histologically 6 out of 16 Helicobacter pylori+ patients showed chronic pangastritis . In the remaining 10 Helicobacter pylori-related mucosal inflammation was confined to the antrum . All 6 subjects with pangastritis and 3 out of 10 with antral gastritis were anti-CagA+ . The autoreaction was found in a 10-year old male child with Helicobacter pylori+ pangastritis and a clinical history of ulcer-like dyspepsia . Parietal cells, in particular, were involved and showed diffuse cytoplasm staining . Proliferating cell nuclear antigen expression demonstrated, only in this case, a zone of regeneration extending from the normal site in the neck towards the base of the glands . CONCLUSIONS: Our finding demonstrates that an autoreaction of gastric mucosa may be found in Helicobacter pylori gastritis of childhood . Its association with some known risk factors (i.e., cytotoxic strains and increased proliferation of gastric epithelium with a changed pattern) may play a role in the progression from gastritis to atrophy and account for the increased risk of late gastric cancer when Helicobacter pylori infection occurs in paediatric age.

Acta Derm Venereol, 1998 Nov, 78(6), 440 - 2
Chronic urticaria and Helicobacter pylori; Valsecchi R et al.; Chronic urticaria can result from multiple causes . A number of factors have been identified that can appear to be important in the pathogenesis of individual cases, including intolerance to food, drugs, some internal diseases and some infections . Recently a possible relationship between chronic urticaria and Helicobacter pylori has been suggested . One hundred and twenty-five patients were investigated for Helicobacter pylori infection by means of ELISA assay and 13C urea-breath tests . When the two tests were positive, gastric biopsy was performed after informed consent . Patients with Helicobacter pylori infection were randomly assigned to receive triple therapy for the eradication of bacterium for one week, or no treatment . As controls, 25 patients with chronic urticaria and with negative results on ELISA and urea-breath tests were treated with the same triple therapy course . Forty-six unrelated blood donors of both sexes were examined for the presence of anti-Helicobacter pylori antibodies in the normal population . Seventy-eight patients had circulating specific IgG antibodies against the bacterium and positive urea-breath tests . Among these patients, 31 received eradication therapy, 34 were enrolled in the control group, and 13 patients neglected the study . Three patients in the eradication therapy group showed complete remission of urticaria after 12 months of follow-up as compared with 1 patient in the control group . Twenty blood donors out of 46 were IgG anti-Helicobacter pylori positive . In conclusion, our data show that the prevalence of Helicobacter pylori infection is high in chronic urticaria patients, but eradication of the bacterium does not appear to influence the skin disorders nor the symptoms.

Med Microbiol Immunol (Berl), 1998 Oct, 187(2), 115 - 20
Viability and gene expression in Chlamydia trachomatis during persistent infection of cultured human monocytes; Gerard HC et al.; The principal host cell for persistently infecting synovial Chlamydia trachomatis is the macrophage . During infection of human monocytes/macrophages in culture this bacterium displays aberrant morphology and produces no new elementary bodies, reflecting the situation in synovium . Here we investigate the metabolic status of C . trachomatis (serovar K) during an extended infection of human peripheral monocytes in vitro . Using reverse transcription-polymerase chain reaction assays, we have shown that primary transcripts from the chlamydial rRNA operons are present throughout a 10-day course of infection . Other assays targeting mRNAs from chlamydial genes encoding r-proteins S5 and L5, the glycyl-tRNA synthetase, the 60-kDa cysteine-rich outer membrane protein, and the KDO transferase indicate that these messengers are also present throughout the entire 10-day period . The gene encoding the 57-kDa heat-shock protein (hsp60) is expressed by the bacterium throughout the 10-day infection of cultured monocytes, but transcript levels from the gene encoding the major outer membrane protein (omp1) appear to be attenuated . Western analyses targeting these latter proteins confirm the presence of the hsp60 gene product, and the virtual absence of major outer membrane protein, in chlamydia-infected cultured human monocytes . Thus, during extended infection of human monocytes in vitro, chlamydia are non-productive but transcriptionally active; the pattern of transcriptional activity reflects that known for persistent C . trachomatis infection in vivo in synovial tissue.

Eur J Gastroenterol Hepatol, 1998 Oct, 10(10), 893 - 5
Abdominal lymphomas, convulsive seizure and coma: a case of successfully treated, advanced Whipple's disease with cerebral involvement; Mohm J et al.; Whipple's disease is a rare, generalized inflammatory disorder due to the recently described bacterium Tropheryma whippelii . We report an unusual, successfully treated case of a 32-year-old woman, who presented with a 25 month history of large abdominal lymphomas, polyserositis and cachexia . The diagnosis of Whipple's disease was confirmed by duodenoscopy, lymph node and duodenal histology and polymerase chain reaction analysis of biopsy material and cerebrospinal fluid . A prolonged convulsive seizure with a subsequent 5 day period of coma were interpreted as signs of cerebral involvement . Under antibiotic treatment with trimethoprim-sulfamethoxazole (co-trimoxazole) the patient recovered completely, CT scans showed a complete regression of abdominal lymphomas . The therapy was continued over 18 months without the occurrence of a relapse.

J Pathol, 1998 Aug, 185(4), 409 - 12
High frequency of CagA+ Helicobacter pylori infection in high-grade gastric MALT B-cell lymphomas; Peng H et al.; A high incidence of Helicobacter pylori infection has been found in patients with gastric MALT (mucosa-associated lymphoid tissue) B-cell lymphoma . Recent studies have indicated that the aggressive strains of the bacterium containing the CagA gene may have direct effects on tumourigenesis . To investigate the involvement of CagA+ strains in MALT lymphomagenesis, a sensitive polymerase chain reaction (PCR)-based detection assay for the gene was developed . DNA extracts from paraffin sections of 123 H . pylori-related gastric biopsies from Italy were analysed, including 56 cases of chronic gastritis, 37 low-grade, and 30 high-grade MALT lymphomas: 30.3 per cent (17/56) of the gastritis cases, 37.8 per cent (14/37) of the low-grade, and 76.7 per cent (23/30) of the high-grade MALT lymphomas were found to contain the CagA gene . The frequency of CagA+ strain infection was significantly higher (P < 0.05) in high-grade than in low-grade MALT lymphoma or gastritis . These results suggest that high-grade gastric MALT lymphoma transformation may be more likely to occur following infection by CagA+ strains of H . pylori.

J Bacteriol, 1998 Dec, 180(23), 6392 - 5
Short-term regulation of nitrogenase activity by NH4+ in Rhodobacter capsulatus: multiple in vivo nitrogenase responses to NH4+ addition; Yakunin AF et al.; The photosynthetic bacterium Rhodobacter capsulatus has been shown to carry out nitrogenase "switch-off," a rapid, reversible inhibition of in vivo activity . Here, we demonstrate that highly nitrogen-limited cultures of both the wild-type strain and a draT draG mutant are capable of nitrogenase switch-off while moderately nitrogen-limited cultures show instead a "magnitude" response, with a decrease in in vivo nitrogenase activity that is proportional to the amount of added NH4+.

Membr Cell Biol, 1998, 12(1), 9 - 26
The structural organization of the antenna chromophore protein complexes in membranes of the photosynthetic bacterium Rhodopseudomonas viridis; Klevanik AV; A possible structural and functional organization of the antenna chromophore protein complexes (CPC) in the Rhodopseudomonas viridis membranes was considered in terms of structural models proposed by Zuber and Brunisholz (in Chlorophylls, ed . H . Scheer (Boca Raton: CRC Press, 1991):626-703) . Analysis of the absorption spectra led to the conclusion that the number of the antenna bacteriochlorophyll molecules per reaction center (RC) is 30 +/- 3 both for chromophores and quantasomes of Rps . viridis . It implies a multicentral organization of the CPCs around RCs, when the CPC of cyclic structure is formed by (alpha beta gamma)4 polypeptides . A multicentral model predicts an almost linear dependence of the antenna fluorescence yield on the oxidized primary donor concentration if the antenna fluorescence lifetimes are assumed to be 60-70 and 110-120 ps for the open and closed RCs, respectively, which is in agreement with the experimental observations . We conclude that the Rps . viridis membrane domain consists of 4-6 RCs surrounded by 6-22 CPCs, and both of these protein subsystems are packed into a hexagonal array.

Biosens Bioelectron, 1998 Sep 15, 13(6), 665 - 73
Binding characteristics of anti-atrazine monoclonal antibodies and their fragments synthesised in bacteria and plants; Strachan G et al.; Single-chain antibody fragments (scAb), specific for the herbicide atrazine, have been expressed in the bacterium Escherichia coli and in transgenic tobacco plants . The scAb could be purified as a monomer (monovalent) via a hexa-histidine tail or as a dimer (divalent) by antibody affinity chromatography . In competition ELISA, the bacterial scAb showed the same specificity for atrazine and related triazine herbicides as the parental mAb cell line, but both plant and bacterial monomeric scAbs showed increased sensitivity to free atrazine . Surface plasmon resonance (BIAcore 2000) analysis confirmed that purified scAb, derived from plant or bacteria, retained similar association rates as the mAb . However, the monomeric plant and bacterial scAbs showed a lower affinity for immobilised antigen, than the equivalent dimeric scAbs or mAb . This decrease in affinity was due to a 10 fold slower dissociation rate and is likely due to loss of the avidity contribution of dimeric molecules.

Biophys J, 1998 Dec, 75(6), 2877 - 87
Total lipids with short and long acyl chains from Acholeplasma form nonlamellar phases; Andersson AS et al.; The cell-wall-less bacterium Acholeplasma laidlawii A-EF22 synthesizes eight glycerolipids . Some of them form lamellar phases, whereas others are able to form normal or reversed nonlamellar phases . In this study we examined the phase properties of total lipid extracts with limiting average acyl chain lengths of 15 and 19 carbon atoms . The temperature at which these extracts formed reversed hexagonal (HII) phases differed by 5-10 degreesC when the water contents were 20-30 wt% . Thus the cells adjust the ratio between lamellar-forming and nonlamellar-forming lipids to the acyl chain lengths . Because short acyl chains generally increase the potential of lipids to form bilayers, it was judged interesting to determine which of the A . laidlawii A lipids are able to form reversed nonlamellar phases with short acyl chains . The two candidates with this ability are monoacyldiglucosyldiacylglycerol (MADGlcDAG) and monoglucosyldiacylglycerol . The average acyl chain lengths were 14.7 and 15.1 carbon atoms, and the degrees of acyl chain unsaturation were 32 and 46 mol%, respectively . The only liquid crystalline phase formed by MADGlcDAG is an HII phase . Monoglucosyldiacylglycerol forms reversed cubic (Ia3d) and HII phases at high temperatures . Thus, even when the organism is grown with short fatty acids, it synthesizes two lipids that have the capacity to maintain the nonlamellar tendency of the lipid bilayer . MADGlcDAG in particular contributes very powerfully to this tendency.

Infect Immun, 1998 Dec, 66(12), 5980 - 7
Actinobacillus actinomycetemcomitans toxin induces both cell cycle arrest in the G2/M phase and apoptosis; Ohguchi M et al.; We found that the culture supernatant of the periodontopathic bacterium Actinobacillus actinomycetemcomitans had a cytotoxic effect on several cell lines . In this study, we purified the toxin from the culture supernatant of A . actinomycetemcomitans Y4 by a four-step procedure: ammonium sulfate precipitation, POROS HQ/M column chromatography, polymyxin B matrix column chromatography, and Mono-Q column chromatography . The purified toxin gave two major bands of protein with molecular masses of 80 and 85 kDa upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis . The mechanism of cell death of the B-cell hybridoma cell line HS-72 was examined by observing changes in nuclear morphology, an increase in the proportion of fragmented DNA, and the typical ladder pattern of degraded chromosomal DNA, indicating the induction of apoptosis . Overexpression of human Bcl-2 suppressed apoptosis in HS-72 cells, indicating that the toxin from A . actinomycetemcomitans induces apoptosis by a Bcl-2-inhibitable mechanism . Flow cytometric analysis revealed that the toxin caused cell cycle arrest in the G2/M phase and apoptosis in HS-72 cells . In addition, aurintricarboxylic acid, a DNA endonuclease inhibitor, markedly decreased the percentage of apoptotic cells but had no effect on cell cycle arrest in the G2/M phase . Taken together, these findings suggest that the toxin from A . actinomycetemcomitans could mediate the development of periodontal diseases through cell cycle arrest in the G2/M phase and apoptosis in B lymphocytes of periodontal tissue.

Infect Immun, 1998 Dec, 66(12), 5763 - 70
Function and protective capacity of Treponema pallidum subsp . pallidum glycerophosphodiester phosphodiesterase; Cameron CE et al.; Infectious syphilis, caused by the spirochete bacterium Treponema pallidum subsp . pallidum, remains a public health concern worldwide . The immune-response evasion mechanisms employed by T . pallidum are poorly understood, and prior attempts to identify immunoprotective antigens for subsequent vaccine design have been unsuccessful . Previous investigations conducted in our laboratory identified the T . pallidum glycerophosphodiester phosphodiesterase as a potential immunoprotective antigen by using a differential immunologic expression library screen . In studies reported here, heterologous expression of the T . pallidum glycerophosphodiester phosphodiesterase in Escherichia coli yielded a full-length, enzymatically active protein . Characterization of the recombinant molecule showed it to be bifunctional, in that it exhibited specific binding to human immunoglobulin A (IgA), IgD, and IgG in addition to possessing enzymatic activity . IgG fractionation studies revealed specific binding of the recombinant enzyme to the Fc fragment of human IgG, a characteristic that may play a role in enabling the syphilis spirochete to evade the host immune response . In further investigations, immunization with the recombinant enzyme significantly protected rabbits from subsequent T . pallidum challenge, altering lesion development at the sites of challenge . In all cases, animals immunized with the recombinant molecule developed atypical pale, flat, slightly indurated, and nonulcerative reactions at the challenge sites that resolved before lesions appeared in the control animals . Although protection in the immunized rabbits was incomplete, as demonstrated by the presence of T . pallidum in the rabbit infectivity test, glycerophosphodiester phosphodiesterase nevertheless represents a significantly immunoprotective T . pallidum antigen and thus may be useful for inclusion in an antigen cocktail vaccine for syphilis.

Gut, 1998 Oct, 43(4), 456 - 7
The medium is the messenger; Phillips AD; Enteropathogenic Escherichia coli (EPEC), like many bacterial pathogens, employ a type III secretion system to deliver effector proteins across the bacterial cell . In EPEC, four proteins are known to be exported by a type III secretion system--EspA, EspB and EspD required for subversion of host cell signal transduction pathways and a translocated intimin receptor (Tir) protein (formerly Hp90) which is tyrosine-phosphorylated following transfer to the host cell to become a receptor for intimin-mediated intimate attachment and "attaching and effacing" (A/E) lesion formation . The structural basis for protein translocation has yet to be fully elucidated for any type III secretion system . Here, we describe a novel EspA-containing filamentous organelle that is present on the bacterial surface during the early stage of A/E lesion formation, forms a physical bridge between the bacterium and the infected eukaryotic cell surface and is required for the translocation of EspB into infected epithelial cells.

Mol Microbiol, 1998 Nov, 30(3), 595 - 602
A surface active protein involved in aerial hyphae formation in the filamentous fungus Schizophillum commune restores the capacity of a bald mutant of the filamentous bacterium Streptomyces coelicolor to erect aerial structures; Tillotson RD et al.; The filamentous bacterium Streptomyces coelicolor undergoes a complex process of morphological differentiation involving the formation of a dense lawn of aerial hyphae that grow away from the colony surface into the air to form an aerial mycelium . Bald mutants of S . coelicolor, which are blocked in aerial mycelium formation, regain the capacity to erect aerial structures when exposed to a small hydrophobic protein called SapB, whose synthesis is temporally and spatially correlated with morphological differentiation . We now report that SapB is a surfactant that is capable of reducing the surface tension of water from 72 mJ m-2 to 30 mJ m-2 at a concentration of 50 microgram ml-1 . We also report that SapB, like the surface-active peptide streptofactin produced by the species S . tendae, was capable of restoring the capacity of bald mutants of S . tendae to erect aerial structures . Strikingly, a member (SC3) of the hydrophobin family of fungal proteins involved in the erection of aerial hyphae in the filamentous fungus Schizophyllum commune was also capable of restoring the capacity of S . coelicolor and S . tendae bald mutants to erect aerial structures . SC3 is unrelated in structure to SapB and streptofactin but, like the streptomycetes proteins, the fungal protein is a surface active agent . Scanning electron microscopy revealed that aerial structures produced in response to both the bacterial or the fungal proteins were undifferentiated vegetative hyphae that had grown away from the colony surface but had not commenced the process of spore formation . We conclude that the production of SapB and streptofactin at the start of morphological differentiation contributes to the erection of aerial hyphae by decreasing the surface tension at the colony surface but that subsequent morphogenesis requires additional developmentally regulated events under the control of bald genes.

J Antibiot (Tokyo), 1998 Sep, 51(9), 805 - 15
B-90063, a novel endothelin converting enzyme inhibitor isolated from a new marine bacterium, Blastobacter sp . SANK 71894; Takaishi S et al.; A novel endothelin-converting enzyme (ECE) inhibitor, B-90063, was isolated from the culture supernatant of the newly discovered marine bacterium Blastobacter sp . SANK 71894 . Based on spectral analyses and chemical reactions, the structure of B-90063 was determined to be bis{6-formyl-4-hydroxy-2-(2'-n-pentyloxazol-4'-yl)-4-pyridon -3-yl}-disulfide (1a) . Human and rat ECEs were inhibited more potently by B-90063, with respective IC50 values of 1.0 and 3.2 microM, than were other neutral endopeptidases such as NEP and type-I and -IV collagenases . B-90063 also inhibited the binding of ET-1 to rat ET(A) and bovine ET(B) receptors, though its antagonistic activities were weak . B-90063, thus, may abolish the physiological actions of endothelins through the ECE inhibitory and receptor antagonistic mechanisms.

Zentralbl Veterinarmed B, 1998 Oct, 45(8), 473 - 9
Detection of Mycobacterium bovis in milk by polymerase chain reaction; Zanini MS et al.; A simple method was developed for DNA extraction of Mycobacterium bovis in milk and further detection of the bacterium by polymerase chain reaction (PCR) . Milk previously seeded with M . bovis was used as the starting material . The procedure involved overnight digestion of a milk sample with proteinase K at 56 degrees C and phenol extraction, followed by ethanol precipitation and PCR . The amplification pattern obtained was analyzed with primers BW8-BW9 which amplify a 248 bp in strains of M . bovis . By using the BW8-BW9 primers, 10(3) CFU were detected on silver-stained PAGE gels . The procedure was validated by PCR analysis of milk in tuberculin-positive animals . It is anticipated that this method can be used for routine diagnosis of M . bovis in milk samples.

Biochemistry, 1998 Nov 17, 37(46), 16280 - 8
Steps toward constructing a cytochrome b6 f complex in the purple bacterium Rhodobacter sphaeroides: an example of the structural plasticity of a membrane cytochrome; Kuras R et al.; We have modified the cytochrome b subunit of the cytochrome bc1 complex from the purple bacterium Rhodobacter sphaeroides to introduce two distinctive features of cytochrome b6 f complexes . In the first one, we have split cyt b into two polypeptides thus mimicking the organization of cyt b6 and subunit IV in the b6 f complexes . In the second, an extra residue was added between His198 and Phe199, thus extending the span between the histidine ligands for the two b-hemes in helix D . The properties of the mutant strains were determined using thermodynamic and kinetic analysis . The two mutant enzymes were assembled and functioned so as to allow the photosynthetic growth of the mutant strains . For the split enzyme, we show that two independently translated fragments of cyt b are inserted in the membrane . Our results indicate a decrease in the stability of the semiquinone formed at the quinone reduction (Qi) site in this mutant . This property, characteristic for b6 f complexes, indicates the functional importance of the connecting span between helices D and E . The presence of the inserted threonine in helix D modified the spectrum and redox potential of the bL-heme, shifting the potential difference between the two b-hemes from 140 mV in the wild-type to 55 mV in the mutant strain . This change in the driving force of electron transfer through the membrane was reflected in an inability of the mutant strain to accumulate a large transmembrane electrical potential on successive flashes.

Berl Munch Tierarztl Wochenschr, 1998 Oct, 111(10), 368 - 73
{Bovine paratuberuclosis: history and resulrs of new efforts to control an old disease}; Gerlach GF et al.; The problem of bovine paratuberculosis is being reviewed . The historic development as well as the cultivation and characterization of the infectious agent are described . The current knowledge of the epidemiology is being discussed with particular emphasis on excretion and resistance of the bacterium, potential hosts, and transmission pathways . Subsequently, the economic importance of the disease is described from an international and a German point of view . International, European and German regulations on para tuberculosis are discussed with respect to their possible influence on future development of animal trade politics.

Scand J Infect Dis, 1998, 30(4), 371 - 6
Risk factors for infection with Helicobacter pylori--a study of children in rural Ethiopia; Lindkvist P et al.; The public health impact of Helicobacter pylori (HP) infection is gradually becoming obvious, the bacterium now being implicated as an aetiologic agent in a variety of gastric diseases . Transmission routes still remain unknown, although single risk factors, such as domestic crowding (especially bed-sharing) in childhood and low parental socioeconomic status, have been pointed out in studies from developed countries . In an attempt to study the risk factors in a developing country, we performed a case control study of 242 randomly selected children aged 2-4 y in Butajira rural area in Ethiopia . Blood samples were drawn and a questionnaire administered . The total prevalence of IgG antibody to HP among the children in the region was 48% (116/242) . Several risk factors such as: crowding, water, animals, sanitation, etc . correlated strongly to seropositivity in a univariate analysis . After controlling for possible confounding, independent predictors of seropositivity were: living in town (OR = 2.15, p = 0.001), increasing age (OR = 1.71, p = 0.060), and being a Muslim (OR = 1.54, p = 0.005) . It could not be excluded that a bad water supply in town could explain the difference in seroprevalence between town and village . These results indicate that, in developing countries, factors relating to community and religion might be as important risk factors for infection with HP in children as characteristics of the family or the home.

Digestion, 1998 Nov-Dec, 59(6), 638 - 45
Decreased Helicobacter pylori-specific gastric secretory IgA antibodies in infected patients; Birkholz S et al.; Despite an increase in local Helicobacter pylori-specific IgA production in H . pylori infection, the bacterium is able to persist over decades . We focused on IgA and secretory IgA (sIgA) in gastric juice because sIgA is more relevant in local protection and more resistant to degradation than nonsecretory IgA . H . pylori-specific IgA and sIgA in gastric juice, saliva, and serum of H . pylori-infected patients were compared . Samples from 28 H . pylori-positive and 16 negative patients were tested by means of immunoblotting for the presence of H . pylori-specific IgA and sIgA . In gastric juice the majority of H . pylori-specific IgA was not of the secretory type, whereas total IgA was bound mainly to the secretory component as shown by immunoblot and slot blot . In contrast H . pylori-specific IgA antibodies in saliva of infected patients were of the secretory type as shown by immunoblot . The presence of specific, nonsecretory IgA may be a consequence of the damaged mucosal epithelium at the site of H . pylori infection allowing IgA to bypass the secretory transport system . Considering the resistance of secretory IgA against hydrolysis and proteolysis, these data suggest that the predominantly nonsecretory IgA specific for H . pylori may lead to a decreased protection against H . pylori.

Biochim Biophys Acta, 1998 Sep 7, 1366(3), 301 - 16
The LH1-RC core complex of Rhodobacter sphaeroides: interaction between components, time-dependent assembly, and topology of the PufX protein; Pugh RJ et al.; Mutant strains of the photosynthetic bacterium Rhodobacter sphaeroides, lacking either LH1, the RC or PufX, were analysed by mild detergent fractionation of the cores . This reveals a hierarchy of binding of PufX in the order RC:LH1 > LH1 > RC . The assembly of photosynthetic membranes was studied by switching highly aerated cells to conditions of low aeration in the dark . The RC-H subunit appears before other components, followed by the pufBALMX then pufBA transcripts . Synthesis of the PufX polypeptide precedes that of LH1alpha and beta, which suggests that PufX associates with a limited amount of LH1alpha, beta and the RC, and prior to the encirclement of the RC by the rest of the LH1 complex . The topology of PufX within the intracytoplasmic membrane was determined by proteolytic treatment of membrane vesicles followed by protein sequencing; PufX is N-terminally exposed on the cytoplasmic surface of the photosynthetic membrane.

J Mol Biol, 1998 Nov 27, 284(2), 421 - 33
Molecular analysis of the trimethylamine N-oxide (TMAO) reductase respiratory system from a Shewanella species; Dos Santos JP et al.; Trimethylamine N-oxide (TMAO) is an abundant compound of tissues of marine fish and invertebrates . During fish spoilage, certain marine bacteria can reduce TMAO to nauseous trimethylamine (TMA) . One such bacterium has been isolated and identified as a new Shewanella species, and called Shewanella massilia . The anaerobic growth of S . massilia is greatly increased when TMAO is added, indicating that TMAO reduction involves a respiratory pathway . The TorA enzyme responsible for TMAO reduction is a molybdenum cofactor-containing protein of 90 kDa located in the periplasm . Whereas TorA is induced by both TMAO and dimethylsulfoxide (DMSO), this enzyme has a high substrate specificity and appears to only efficiently reduce TMAO as a natural compound . The structural torA gene encoding the TMAO reductase (TorA) and its flanking regions were amplified using PCR techniques . The torA gene is the third gene of a TMAO-inducible operon (torECAD) encoding the TMAO respiratory components . The torC gene, located upstream from torA encodes a pentahemic c-type cytochrome, likely to be involved in electron transfer to the TorA terminal reductase . TorC was shown to be anchored to the membrane and, like TorA, is induced by TMAO . Except for the TorE protein, which is encoded by the first gene of the torECAD operon, all the tor gene products are homologous to proteins found in the TMAO/DMSO reductase systems from Escherichia coli and Rhodobacter species . In addition, the genetic organization of these systems is similar . Although these bacteria are found in different ecological niches, their respiratory systems appear to be phylogenetically related, suggesting that they come from a common ancestor .

J Biol Chem, 1998 Nov 20, 273(47), 30888 - 96
Molecular cloning, overexpression, and characterization of steroid-inducible 3alpha-hydroxysteroid dehydrogenase/carbonyl reductase from Comamonas testosteroni . A novel member of the short-chain dehydrogenase/reductase superfamily; Mobus E et al.; 3alpha-Hydroxysteroid dehydrogenase/carbonyl reductase (3alpha-HSD/CR) from Comamonas testosteroni, a bacterium that is able to grow on steroids as the sole carbon source, catalyzes the oxidoreduction at position 3 of a variety of C19-27 steroids and the carbonyl reduction of a variety of nonsteroidal aldehydes and ketones . The gene of this steroid-inducible 3alpha-HSD/CR was cloned by screening a C . testosteroni gene bank with a homologous DNA probe that was obtained by polymerase chain reaction with two degenerative primers based on the N-terminal sequence of the purified enzyme . The 3alpha-HSD/CR gene is 774 base pairs long, and the deduced amino acid sequence comprises 258 residues with a calculated molecular mass of 26.4 kDa . A homology search revealed that amino acid sequences highly conserved in the short-chain dehydrogenase/reductase (SDR) superfamily are present in 3alpha-HSD/CR . Two consensus sequences of the SDR superfamily were found, an N-terminal Gly-X-X-X-Gly-X-Gly cofactor-binding motif and a Tyr-X-X-X-Lys segment (residues 155-159 in the 3alpha-HSD/CR sequence) essential for catalytic activity of SDR proteins . 3alpha-HSD/CR was overexpressed and purified to homogeneity, and its activity was determined for steroid and nonsteroidal carbonyl substrates . These results suggest that inducible 3alpha-HSD/CR from C . testosteroni is a novel member of the SDR superfamily.

C R Acad Sci III, 1998 Sep, 321(9), 777 - 87
Modelling Escherichia coli . The concept of competitive coherence; Norris V; Many of the characteristics of life are plainly evident . Others are not . In principle, computer models might be devised to test whether a hypothetical characteristic is likely to be a characteristic of real life . Here, I propose that the bacterium Escherichia coli can be considered as passing through a series of states in which a distinct set of its constituent molecules or 'elements' are active . The activity of these elements is determined by a competition between two processes . One of these processes depends on the previous cell state whilst the other depends on the internal coherence of the developing state . The simultaneous operation of these two processes I term 'competitive coherence' . To clarify and eventually test these related hypotheses, a possible computer model is outlined.

Curr Microbiol, 1998 Dec, 37(6), 431 - 2
News & notes: paper digestion by the cellulolytic ruminal bacterium Fibrobacter succinogenes; Martin SA et al.; The objective of this study was to evaluate the ability of the predominant cellulolytic ruminal bacterium Fibrobacter succinogenes S85 to digest filter paper, office paper, newspaper, and magazine paper . When F . succinogenes S85 was incubated with all four paper sources, little digestion of any paper occurred between 0 and 48 h . However, digestion of filter paper increased from 12% at 48 h to 80% at 120 h . No significant digestion of office paper, newspaper, or magazine paper occurred between 48 and 120 h . These results suggest that F . succinogenes S85 is unable to digest office paper, newspaper, or magazine paper.

Fertil Steril, 1998 Nov, 70(5), 945 - 8
Viability of Chlamydia trachomatis in fallopian tubes of patients with ectopic pregnancy; Gerard HC et al.; OBJECTIVE: To use standard molecular methods to define the prevalence and metabolic characteristics of Chlamydia trachomatis during infection of fallopian tubes in women with ectopic pregnancies . DESIGN: Polymerase chain reaction (PCR)- and reverse transcription-PCR (RT-PCR)-based assessment of presence of chlamydial DNA and various RNA species in fallopian tube biopsy samples . SETTING: Hospital and molecular genetics laboratory . PATIENTS: Ten women of varying ages, each presenting with ectopic pregnancy . MAIN OUTCOME MEASURE(S): Positive signal in specific chlamydia-directed PCR and RT-PCR assays . RESULT(S): Nucleic acid preparations from 7 of the 10 fallopian tube patient samples were PCR-positive for C . trachomatis DNA . Each of the 7 PCR-positive samples also showed the presence of several transcripts from the bacterium, including primary transcripts from the ribosomal RNA operons . CONCLUSION(S): A higher proportion of ectopic pregnancies than was believed previously may be attributable to infection of the fallopian tubes by C . trachomatis . The presence of various chlamydial RNA molecules suggests that viable, metabolically active bacteria were present in fallopian tubes of the patients studied.

Biochim Biophys Acta, 1998 Nov 2, 1409(1), 39 - 49
Purification and characterization of pyruvate oxidoreductase from the photosynthetic bacterium Rhodobacter capsulatus; Yakunin AF et al.; Pyruvate:ferredoxin (flavodoxin) oxidoreductase (POR) was purified 3050-fold to apparent homogeneity from the photosynthetic bacterium Rhodobacter capsulatus using ion-exchange, Reactive Red, and gel filtration chromatography . The isolated enzyme was sensitive to dilution and oxygen (especially when in dilute solution) . The molecular mass of the native enzyme was determined by high performance liquid chromatography gel filtration to be 270+/-20 kDa . Since a subunit molecular mass of 130+/-5 kDa was found by denaturing gel electrophoresis, POR from R . capsulatus thus appears to be a homodimer . Electron paramagnetic resonance analysis showed that a free radical was formed upon the addition of pyruvate . This POR is shown to be an indiscriminate electron donor causing the full reduction of R . capsulatus flavodoxin (Fld), R . capsulatus ferredoxin I (FdI), R . capsulatus ferredoxin II (FdII), as well as the major plant-type ferredoxin (FdI) from Anabaena variabilis . The purified enzyme can couple the oxidation of pyruvate to the reduction of nitrogenase in a coupled system with either R . capsulatus ferredoxins or nif-specific flavodoxin, NifF; (Fld>FdI>FdII) . Immunoblot analysis shows that R . capsulatus POR is constitutively synthesized, with synthesis augmented under nitrogen-fixing conditions (34+/-13%) and decreased in acetate and aerobically grown cells.

Microbiology, 1998 Oct, 144 ( Pt 10), 2837 - 44
Divercin V41, a new bacteriocin with two disulphide bonds produced by Carnobacterium divergens V41: primary structure and genomic organization; Metivier A et al.; Divercin V41 is a new bacteriocin produced by Carnobacterium divergens V41, a lactic acid bacterium isolated from fish viscera . The amino acid sequence of divercin V41 showed high homologies with pediocin PA-1 and enterocin A . Two disulphide bonds were present in the hydrophilic N-terminal domain and in the highly variable hydrophobic C-terminal domain, respectively . A DNA probe designed from the N-terminal sequence of the purified peptide was used to locate the structural gene of divercin V41 . A 6 kb chromosomal fragment containing the divercin V41 structural gene (dvnA) was cloned and sequenced . The results indicate that divercin V41 is synthesized as a pre-bacteriocin of 66 amino acids . The 23-residue N-terminal extension is cleaved off to yield the mature 43-amino-acid divercin V41 . In addition, the fragment encodes putative proteins commonly found within bacteriocin operons, including an ATP-dependent transporter, two immunity-like proteins and the two components of a lantibiotic-type signal-transducing system . The genetic organization of the fragment suggested important gene rearrangements.

Antonie Van Leeuwenhoek, 1998 Apr, 73(3), 279 - 88
The interaction between saliva and Actinobacillus actinomycetemcomitans influenced by the zeta potential; Groenink J et al.; The adhesion of Actinobacillus actinomycetemcomitans is a virulence factor in the aetiology of periodontitis and is determined by physico-chemical properties, e.g . surface charge and hydrophobicity, of the bacterial cell surface . Although oral surfaces are constantly coated with saliva, few studies have dealt with the binding of A . actinomycetemcomitans with saliva . In this report, the charge properties of A . actinomycetemcomitans have been studied through measurement of the zeta potential and the saliva-bacteria interaction investigated at different pH-values . At physiological conditions the zeta potential was negative, varying from -11 to -26 mV, for two laboratory and two fresh isolates of A . actinomycetemcomitans . Under these conditions, binding of the low-molecular-weight salivary mucin, lactoferrin, and S-IgA was confirmed using salivary samples and purified salivary fractions in liquid-phase and in ELISA . The iso-electric points of the laboratory and fresh clinical isolates of A . actinomycetemcomitans were determined at pH 4.6 and 3.8, respectively . At pH below the iso-electric point, giving positive values of the zeta potential, additional salivary protein species bound to A . actinomycetemcomitans, including the high-molecular-weight salivary mucin (MG1) and agglutinin . Binding of the low-molecular-weight salivary mucin (MG2), lactoferrin, and S-IgA, was hardly affected by this change in zeta potential . A salivary coating formed on the bacterium at pH 7 reduced the zeta potential of the laboratory strain Y4 greatly and an iso-electric point for the bacterium could not be determined . Overall, the study suggests that upon changes in environmental pH additional salivary attachment sites on the micro-organism are exposed.

Biochemistry, 1998 Nov 3, 37(44), 15320 - 6
Electrostatic interactions at the donor side of the photosynthetic reaction center of Rhodopseudomonas viridis; Baymann F et al.; The photosynthetic reaction center of Rhodopseudomonas viridis, a purple bacterium, contains a tetraheme cytochrome subunit . After its photoinduced oxidation, the primary donor, P, is reduced by the nearby heme (c559) of the tetraheme subunit in about 200 ns . This heme, in turn, is reduced by another heme (c556) of the subunit in about 2 micro(s) . The midpoint potentials of P, c559, and c556 are known to be +500, +380, and +320 mV, respectively . The reduction kinetics of P+ are strongly biphasic in living cells, membrane fragments, and isolated reaction centers . We show here that this biphasicity reflects a small equilibrium constant (lower than 10) for the electron-transfer reaction between P and c559, which arises from a significant difference between the operating redox potentials of the P+/P and c559+/c559 couples and their equilibrium midpoint potentials . This difference is partly due to the effect of the permanent transmembrane potential, which arises from the cell metabolism, and to significant electrostatic interactions which develop between the electron carriers of the reaction center . Interestingly, the kinetic parameters of P+ reduction in decoupled cells or membrane fragments are identical to those reported for isolated reaction centers . We estimate an interaction of about 20 mV between c556 and c559 and about 90 mV between c559 and P . Consequently, the operating redox potential of the P+/P couple is 410 mV.

Eur J Biochem, 1998 Oct 1, 257(1), 249 - 54
The influence of 5' codon context on translation termination in Saccharomyces cerevisiae; Mottagui-Tabar S et al.; Translation termination in vivo was studied in the yeast Saccharomyces cerevisiae using a translation-assay system . Codon changes that were made at position -2 relative to the stop codon, gave a 3.5-fold effect on termination in a release-factor-defective (sup45) mutant strain, in line with the effect observed in a wild-type strain . The influence of the -2 codon could be correlated to the charge of the corresponding amino acid residue in the nascent peptide; an acidic residue favoring efficient termination . Thus, the C-terminal end of the nascent peptide influences translation termination both in the bacterium Escherichia coli and to a lesser extent in the yeast S . cerevisiae . However, the sensitivity to the charge of the penultimate amino acid is reversed when the E . coli and S . cerevisiae are compared . Changing - 1 (P-site) codons in yeast gave a 10-fold difference in effect on the efficiency of termination . This effect could not be related to any property of the encoded last amino acid in the nascent peptide . Iso-codons read by the same tRNA (AAA/G, GAA/G) gave similar readthrough values . Codons for glutamine (CAA/G), glutamic acid (GAA/G) and isoleucine (AUA/C) that are read by different isoaccepting tRNAs are associated with an approximately twofold difference in each case in termination efficiency . This suggests that the P-site tRNA is able to influence termination at UGAC in yeast.

Eur J Biochem, 1998 Oct 1, 257(1), 185 - 91
Biosynthesis of the 3-acetyl and 13(1)-oxo groups of bacteriochlorophyll a in the facultative aerobic bacterium, Rhodovulum sulfidophilum--the presence of both oxygenase and hydratase pathways for isocyclic ring formation; Porra RJ et al.; Using (18)O-labelling and mass spectrometry, we have examined bacteriochlorophyll a formation in Rhodovulum sulfidophilum, formerly known as Rhodobacter sulfidophilus, which forms large amounts of BCh1 a both aerobically in the dark and anaerobically in the light . R . sulfidophilum, growing under strict anaerobiosis in the light, possesses hydratases which incorporate (18)O label from H2(18)O into both the 13(1)-oxo and 3-acetyl oxygens; in addition, the four carboxyl oxygens at C13(3) and C17(3) were labelled by H2(18)O . Under aerobic conditions in the dark, the labelling of the 13(1)-oxo group by H2(18)O was reduced indicating that (16)O was being incorporated into this group from air . R . sulfidophilum, grown in the dark under an atmosphere initially containing 50% (18)O2 in Ar, possessed an oxygenase which incorporated (18)O label from (18)O2 specifically into the 13(1)-oxo group; under these conditions the acetyl and carboxyl groups remained unlabelled . Thus, both an oxygenase and hydratase operate in R . sulfidophilum to form the 13(1)-oxo group of ring E of BCh1 a; the 3-acetyl group oxygen, however, arises only from water via a hydratase.

J Food Prot, 1998 Oct, 61(10), 1265 - 8
Removal of Escherichia coli O157:H7 from surface tissues of beef carcasses inoculated with wet and dry manure; Delazari I et al.; Beef tissues were contaminated with wet and dry manure . The manure was previously inoculated with Escherichia coli O157:H7 GFP, genetically modified with a plasmid encoding a protein that fluoresces green when exposed to long-wave ultraviolet light . After incubation at 37 degrees C for 5 days, the wet manure was spread on the surface of beef tissues at an average E . coli O157:H7 GFP level of 6.62 log CFU/cm2 . Dry manure was obtained by subjecting wet manure to natural drying (simulating dry manure adhering to the hides of cattle) and was also applied to the surfaces of beef tissues . The degree of removal of E . coli O157:H7 GFP by washing was compared to the removal of cells of the same strain that had been inoculated as a suspension . The E . coli O157:H7 mixed into feces of cattle adhered more strongly to meat surfaces than that applied as a suspension, complicating the removal by conventional washing procedures . The fate of the bacterium mixed into wet or dry manure was evaluated . An initial decrease of the inoculated population was observed; this was probably an effect of the changed environment represented by the manure . After adaptation, the inoculated bacteria grew in the wet manure; a maximum population was reached in 5 days at 37 degrees C; levels declined with drying . The use of the GFP marker was of great value, since it allowed enumeration of E . coli O157:H7 in the presence of the natural flora of manure.

Appl Environ Microbiol, 1998 Nov, 64(11), 4581 - 7
Quantification of 16S rRNAs in complex bacterial communities by multiple competitive reverse transcription-PCR in temperature gradient gel electrophoresis fingerprints; Felske A et al.; A novel approach was developed to quantify rRNA sequences in complex bacterial communities . The main bacterial 16S rRNAs in Drentse A grassland soils (The Netherlands) were amplified by reverse transcription (RT)-PCR with bacterium-specific primers and were separated by temperature gradient gel electrophoresis (TGGE) . The primer pair used (primers U968-GC and L1401) was found to amplify with the same efficiency 16S rRNAs from bacterial cultures containing different taxa and cloned 16S ribosomal DNA amplicons from uncultured soil bacteria . The sequence-specific efficiency of amplification was determined by monitoring the amplification kinetics by kinetic PCR . The primer-specific amplification efficiency was assessed by competitive PCR and RT-PCR, and identical input amounts of different 16S rRNAs resulted in identical amplicon yields . The sequence-specific detection system used for competitive amplifications was TGGE, which also has been found to be suitable for simultaneous quantification of more than one sequence . We demonstrate that this approach can be applied to TGGE fingerprints of soil bacteria to estimate the ratios of the bacterial 16S rRNAs.

Appl Environ Microbiol, 1998 Nov, 64(11), 4467 - 76
A small, dilute-cytoplasm, high-affinity, novel bacterium isolated by extinction culture and having kinetic constants compatible with growth at ambient concentrations of dissolved nutrients in seawater; Button DK et al.; Dilutions of raw seawater produced a bacterial isolate capable of extended growth in unamended seawater . Its 2.9-Mb genome size and 40-fg dry mass were similar to values for many naturally occurring aquatic organotrophs, but water and DNA comprised a large portion of this small chemoheterotroph, as compared to Escherichia coli . The isolate used only a few aromatic hydrocarbons and acetate, and glucose and amino acid incorporation were entirely absent, although many membrane and cytoplasmic proteins were inducible; it was named Cycloclasticus oligotrophus . A general rate equation that incorporates saturation phenomena into specific affinity theory is derived . It is used to relate the kinetic constants for substrate uptake by the isolate to its cellular proteins . The affinity constant KA for toluene was low at 1.3 microg/liter under optimal conditions, similar to those measured in seawater, and the low value was ascribed to an unknown slow step such as limitation by a cytoplasmic enzyme; KA increased with increasing specific affinities . Specific affinities, a degreess, were protocol sensitive, but under optimal conditions were 47.4 liters/mg of cells/h, the highest reported in the literature and a value sufficient for growth in seawater at concentrations sometimes found . Few rRNA operons, few cytoplasmic proteins, a small genome size, and a small cell size, coupled with a high a degreess and a low solids content and the ability to grow without intentionally added substrate, are consistent with the isolation of a marine bacterium with properties typical of the bulk of those present.

J Immunol, 1998 Nov 1, 161(9), 4834 - 41
Mycobacterium avium-intracellulare complex activates nuclear transcription factor-kappaB in different cell types through reactive oxygen intermediates; Giri DK et al.; Mycobacterium avium-intracellulare complex (MAC) is one of the most common opportunistic pathogens in HIV-infected patients . Their synergistic interaction leads to a rapid deterioration of the host defense . In vivo, MAC manifests as a disseminated granulomatous disease that produces a massive inflammatory tissue response perhaps through its activation of inflammatory cytokines . The intracellular signaling following interaction of the mycobacterium with host cells is incompletely understood . Because the response is dependent, in part, on the activation of NF-kappaB, we investigated the effect of MAC on this nuclear transcription factor in cells of macrophage and nonmacrophage lineage . We demonstrate that both high and low virulence strains of MAC potently and rapidly activated NF-kappaB . In supershift assays, using specific Abs against the NF-kappaB subunits, we identified a p50/p65 heterodimer that was formed within 5 min after incubation with the bacterium too rapidly for cytokines to be involved in the activation . This activation was instead mediated through the generation of reactive oxygen intermediates, inasmuch as preincubation of cells with a variety of antioxidants inhibited NF-kappaB activation . Likewise, the transfection of cells with Mn-superoxide dismutase blocked the NF-kappaB activation induced by the bacterium . These data suggest that NF-kappaB activation is a consequence of interaction of host cells with the bacterium and that the interaction may play a pivotal role in the pathogenesis of the disease.

J Biol Chem, 1998 Nov 6, 273(45), 29545 - 53
Accumulation in marine sponge grafts of the mRNA encoding the main proteins of the cell adhesion system; Fernandez-Busquets X et al.; Specific cell adhesion in the marine sponge Microciona prolifera is mediated by an extracellular aggregation factor complex, whose main protein component, termed MAFp3, is highly polymorphic . We have now identified MAFp4, an approximately 400-kDa protein, from the aggregation factor that is translated from the same mRNA as MAFp3 . The existence of multiple potential sites for N-glycosylation and calcium binding suggests a direct involvement of MAFp4 in the species-specific aggregation of sponge cells . The deduced partial polypeptide consists of a 16-fold reiterated motif that shows significant similarity to a repeat in an endoglucanase from the symbiontic bacterium Azorhizobium caulinodans and to the intracellular loop of mammalian Na+-Ca2+ exchangers . Restriction fragment length polymorphism analysis indicated that the genomic variability of MAFp4 is high and comparable to that of MAFp3 . Their combined polymorphism correlates with allogeneic responses studied in a population of 23 sponge individuals . Peptide mass fingerprinting of tryptic digests of the polymorphic MAFp3 bands observed on polyacrylamide gels after chemical deglycosylation of the Microciona aggregation factor revealed that the variability detected on Southern blots at least partially reflects the individual variability of aggregation factor protein components . Polyclonal antibodies raised against MAFp3 strongly cross-reacted with a 68-kDa protein localized in sponge cell membranes . Immunohistochemical use of the anti-MAFp3 antibodies strongly stained a cell layer along the line of contact in allogeneic grafts . We show that the transcription level of the MAFp3/MAFp4 mRNA in sponge allo- and isografts is clearly increased in comparison with non-grafted tissue . These data are discussed with respect to a possible evolutionary relationship between cell adhesion and histocompatibility systems.

Eur Biophys J, 1998, 27(6), 582 - 9
Confocal Raman microspectroscopy of the activation of single neutrophilic granulocytes; Otto C et al.; Confocal Raman micro-spectroscopy has been applied to investigate the activation process of single, living neutrophilic granulocytes . Both resting cells as well as activated cells were measured . The activation of cells was performed with phorbol-12-myristate-13-acetate activator and Escherichia Coli bacteria . Raman microspectroscopy combines a high spatial resolution inside a single, living cell with detailed material information . Using this approach it can be concluded that activation of the cells with phorbol-12-myristate-13-acetate causes a change in the redox state of cytochrome b558 . This protein is a part of the NADPH-oxidase complex that neutrophilic granulocytes employ to generate O-2, superoxide anion . Additionally a change in the redox state of myeloperoxidase can be observed . Myeloperoxidase is known to react with O-2 . Activation of the cells with bacteria gives rise to corresponding changes in the Raman spectra . From this single cell study it can be concluded that the enzymes cytochrome b558 and myeloperoxidase are present inside the cytoplasm of the living cell, while participating in the redox processes . Activation causes an intra-cellular release of oxygen metabolites . Activation with bacteria of neutrophilic granulocytes from a patient with chronic granulomatous disease, that contain no cytochrome b558, led to typical changes in the redox state of myeloperoxidase . This indicates that in the bacterium/neutrophilic granulocyte system oxygen metabolites are generated that are capable of reacting with MPO.

Mol Microbiol, 1998 Oct, 30(2), 275 - 84
The Myxococcus xanthus lipopolysaccharide O-antigen is required for social motility and multicellular development; Bowden MG et al.; The gliding bacterium Myxococcus xanthus aggregates to form spore-filled fruiting bodies when nutrients are limiting . Defective fruiting-body formation and sporulation result from mutations in the sasA locus, which encodes the wzm wzt wbgA (formerly rfbABC) lipopolysaccharide (LPS) O-antigen biosynthesis genes . Mutants carrying these same sasA mutations are defective in social motility and form small glossy colonies . We report here that the developmental and motility phenotypes of four mutants each containing different Tn5 insertions in LPS O-antigen biosynthesis genes are similar to those of the original sasA locus mutants . All of the LPS O-antigen mutants tested exhibited defective developmental aggregation and sporulated at only 0.02-15% of the wild-type level . In addition, all of the LPS O-antigen mutants were determined by genetic analyses to be wild type for adventurous motility and defective in social motility, indicating that the LPS O-antigen is necessary for normal development and social motility . The two previously identified cell-surface components required for social motility, type IV pili and the protein-associated polysaccharide material termed fibrils, were detected on the surfaces of all of the LPS O-antigen mutants . This indicates that LPS O-antigen is a third cell-surface component required for social motility.

J Bacteriol, 1998 Nov, 180(21), 5668 - 75
Regulation of motility behavior in Myxococcus xanthus may require an extracytoplasmic-function sigma factor; Ward MJ et al.; Using interaction trap technology, we identified a putative extracytoplasmic-function (ECF) sigma factor (RpoE1) in Myxococcus xanthus, a bacterium which has a complex life cycle that includes fruiting body formation . The first domain of the response regulator protein FrzZ, a component of the Frz signal transduction system, was used as bait . Although the RpoE1 protein displayed no interactions with control proteins presented as bait, a weak interaction with a second M . xanthus response regulator (AsgA) was observed . While the specificity of the FrzZ-RpoE1 interaction therefore remains speculative, cloning and sequencing of the region surrounding rpoE1 localized it to a position downstream of the frzZ gene . A potential promoter site for binding of an ECF sigma factor was identified upstream of rpoE1, suggesting the gene may be autoregulated . However, primer extension studies suggested that transcription of rpoE1 occurs under both vegetative and developmental conditions from a sigma70-like promoter . Dot blot analysis of RNA preparations confirmed the low-level, constitutive expression of rpoE1 during both stages of the life cycle . Analysis of an insertion mutant also indicated a role for RpoE1 under both vegetative and developmental conditions, since swarming was reduced on nutrient-rich agar and developmental aggregation was effected under starvation conditions, especially at high cell densities . An insertion mutation introduced into the gene directly downstream of rpoE1 (orf5) did not result in either swarming or developmental aggregation defects, even though the gene is transcribed as part of the same operon . Therefore, we propose that this new ECF sigma factor could play a role in the transcriptional regulation of genes involved in motility behavior during both stages of the complex M . xanthus life cycle.

J Bacteriol, 1998 Nov, 180(21), 5574 - 9
A novel mechanism for resistance to the antimetabolite N-phosphonoacetyl-L-aspartate by Helicobacter pylori; Burns BP et al.; The mechanism of resistance to N-phosphonoacetyl-L-aspartate (PALA), a potent inhibitor of aspartate carbamoyltransferase (which catalyzes the first committed step of de novo pyrimidine biosynthesis), in Helicobacter pylori was investigated . At a 1 mM concentration, PALA had no effects on the growth and viability of H . pylori . The inhibitor was taken up by H . pylori cells and the transport was saturable, with a Km of 14.8 mM and a Vmax of 19.1 nmol min-1 microliters of cell water-1 . By 31P nuclear magnetic resonance (NMR) spectroscopy, both PALA and phosphonoacetate were shown to have been metabolized in all isolates of H . pylori studied . A main metabolic end product was identified as inorganic phosphate, suggesting the presence of an enzyme activity which cleaved the carbon-phosphorus (C-P) bonds . The kinetics of phosphonate group cleavage was saturable, and there was no evidence for substrate inhibition at higher concentrations of either compound . C-P bond cleavage activity was temperature dependent, and the activity was lost in the presence of the metal chelator EDTA . Other cleavages of PALA were observed by 1H NMR spectroscopy, with succinate and malate released as main products . These metabolic products were also formed when N-acetyl-L-aspartate was incubated with H . pylori lysates, suggesting the action of an aspartase . Studies of the cellular location of these enzymes revealed that the C-P bond cleavage activity was localized in the soluble fraction and that the aspartase activity appeared in the membrane-associated fraction . The results suggested that the two H . pylori enzymes transformed the inhibitor into noncytotoxic products, thus providing the bacterium with a mechanism of resistance to PALA toxicity which appears to be unique.

J Bacteriol, 1998 Nov, 180(21), 5515 - 9
Functional consequences of changing proline residues in the phenylalanine-specific permease of Escherichia coli; Pi J et al.; The PheP protein is a high-affinity phenylalanine-specific permease of the bacterium Escherichia coli . A topological model based on genetic analysis involving the construction of protein fusions with alkaline phosphatase has previously been proposed in which PheP has 12 transmembrane segments with both N and C termini located in the cytoplasm (J . Pi and A . J . Pittard, J . Bacteriol . 178:2650-2655, 1996) . Site-directed mutagenesis has been used to investigate the functional importance of each of the 16 proline residues of the PheP protein . Replacement of alanine at only three positions, P54, P341, and P442, resulted in the loss of 50% or more activity . Substitutions at P341 had the most dramatic effects . None of these changes in transport activity were, however, associated with any defect of the mutant protein in inserting into the membrane, as indicated by {35S}methionine labelling and immunoprecipitation using anti-PheP serum . A possible role for each of these three prolines is discussed . Inserting a single alanine residue at different sites within span IX and the loop immediately preceding it also had major effects on transport activity, suggesting an important role for a highly organized structure in this region of the protein.

Protein Expr Purif, 1998 Nov, 14(2), 275 - 82
Purification and characterization of Chromatium vinosum GroEL and GroES proteins overexpressed in Escherichia coli cells lacking the endogenous groESL operon; Dionisi HM et al.; Using an Escherichia coli strain (RF101) in which the endogenous chromosomal groESL operon was removed, we overexpressed the GroEL and GroES chaperonins cloned from the photosynthetic bacterium Chromatium vinosum . The identities of these proteins were confirmed by immunological and N-terminal sequence analyses . The native molecular masses of GroEL and GroES, as determined by size-exclusion chromatography, were 830 and 74 kDa, respectively . This suggests a tetradecameric structure for GroEL and a heptameric structure for GroES . C . vinosum GroEL catalyzed a K+-stimulated ATP hydrolysis with a specific activity at 25 degreesC of 50.2 +/- 3.8 nmol Pi released min-1 mg protein-1 . GroEL ATPase was inhibited by GroES, reaching about 50% inhibition at a ratio GroES-7mer/GroEL-14mer of 1 in the presence of 10 mM KCl . The ATPase Vmax increased almost fivefold in the 25 to 65 degreesC temperature range; higher temperatures led to a rapid inactivation of this activity . The chaperone activity of the C . vinosum GroEL/GroES system was characterized by its effect on the refolding of guanidinium chloride-unfolded rhodanese . In the presence of ATP and GroES, C . vinosum GroEL assisted rhodanese refolding . The heterologous combination C . vinosum GroEL/E . coli GroES or E . coli GroEL/C . vinosum GroES was as effective as the homologous complexes . In summary, this strategy allowed the purification at high yields of fully functional, homogenous C . vinosum GroEL and GroES chaperonins from E . coli .

Mol Gen Genet, 1998 Sep, 259(4), 379 - 82
Ribonuclease-charged vector for facile direct cloning with positive selection; Deyev SM et al.; Plasmid vectors for positive selection of cloned inserts in Escherichia coli were devised, based on an expression plasmid (pMT416) for the bacterial ribonuclease barnase . In addition to the barnase gene under control of a synthetic tac promoter, these plasmids carry the gene for the barnase inhibitor, barstar, the constitutive expression of which protects the bacterium from the detrimental effects of moderate barnase production . Full expression of the barnase gene overcomes protection by barstar and becomes lethal . Having a unique SmaI/XmaI site in the barnase structural gene, pMT416 itself can be used as a selective vector: uncut or religated pMT416 will preclude growth while plasmids with inserts in the barnase gene will allow the cells to survive . The entire pUC polylinker was inserted into the barnase gene in place of the Val-36 codon . This insert of nineteen largely hydrophilic amino acids does not prevent the lethal effect of full expression of the gene . The resulting plasmid, pMT440, is a generally useful selective cloning vector representing the "kill-the-rest" approach.

FEMS Microbiol Lett, 1998 Oct 1, 167(1), 41 - 9
Concomitant expression of E . coli cytosine deaminase and uracil phosphoribosyltransferase improves the cytotoxicity of 5-fluorocytosine; Tiraby M et al.; The prodrug activation system formed by the E . coli codA gene encoding cytosine deaminase (CD) and 5-fluorocytosine (5-FC) developed for selective cancer chemotherapy suffers from a sensitivity limitation in many tumour cells . In an attempt to improve the CD/5-FC suicide association, we combined the E . coli upp gene encoding uracil phosphoribosyltransferase (UPRT) with codA gene to create the situation prevailing in E . coli, a bacterium very efficient in metabolising 5-FC . The constitutive expression of the two genes cloned on an E . coli-animal cell shuttle plasmid either in a linked or in a fused configuration was evaluated in E . coli strains selected and engineered to mimic the 5-FC metabolism encountered in mammalian cells . The simultaneous expression of codA and upp genes generated a cooperative effect resulting in a dramatic increase in 5-FC sensitivity of cells compared to the expression of codA alone . Furthermore, it was shown that the association of UPRT with CD facilitated the uptake of 5-FC, in the situation where the drug penetrates cells by passive diffusion as in mammalian cells, by directly channeling 5-fluorouracil, the product of CD, to 5-fluoroUMP, the product of UPRT.

Infect Immun, 1998 Nov, 66(11), 5580 - 6
Translocated intimin receptors (Tir) of Shiga-toxigenic Escherichia coli isolates belonging to serogroups O26, O111, and O157 react with sera from patients with hemolytic-uremic syndrome and exhibit marked sequence heterogeneity; Paton AW et al.; The capacity to form attaching and effacing (A/E) lesions on the surfaces of enterocytes is an important virulence trait of several enteric pathogens, including enteropathogenic Escherichia coli (EPEC) and Shiga-toxigenic E . coli (STEC) . Formation of such lesions depends upon an interaction between a bacterial outer membrane protein (intimin) and a bacterially encoded receptor protein (Tir) which is exported from the bacterium and translocated into the host cell membrane . Intimin, Tir, and several other proteins necessary for generation of A/E lesions are encoded on a chromosomal pathogenicity island termed the locus for enterocyte effacement (LEE) . Reports of sequence heterogeneity and antigenic variation in the region of intimin believed to be responsible for receptor binding raise the possibility that the receptor itself is also heterogeneous . We have examined this by cloning and sequencing tir genes from three different STEC strains belonging to serogroups O26, O111, and O157 . The deduced amino acid sequences for the Tir homologues from these strains varied markedly, exhibiting only 65.4, 80.2, and 56.7% identity, respectively, to that recently reported for EPEC Tir . STEC Tir is also highly immunogenic in humans . Western blots of E . coli DH5alpha expressing the various STEC tir genes cloned in pBluescript {but not E . coli DH5alpha(pBluescript)} reacted strongly with convalescent sera from patients with hemolytic-uremic syndrome (HUS) caused by known LEE-positive STEC . Moreover, no reaction was seen when the various clone lysates were probed with serum from a patient with HUS caused by a LEE-negative STEC or with serum from a healthy individual . Covariation of exposed epitopes on both intimin and Tir may be a means whereby STEC avoid host immune responses without compromising adhesin-receptor interaction.

Infect Immun, 1998 Nov, 66(11), 5527 - 33
Coxiella burnetii induces reorganization of the actin cytoskeleton in human monocytes; Meconi S et al.; Coxiella burnetii, an obligate intracellular bacterium which survives in myeloid cells, causes Q fever in humans . We previously demonstrated that virulent C . burnetii organisms are poorly internalized by monocytes compared to avirulent variants . We hypothesized that a differential mobilization of the actin cytoskeleton may account for this distinct phagocytic behavior . Scanning electron microscopy demonstrated that virulent C . burnetii stimulated profound and polymorphic changes in the morphology of THP-1 monocytes, consisting of membrane protrusions and polarized projections . These changes were transient, requiring 5 min to reach their maximum extent and vanishing after 60 min of incubation . In contrast, avirulent variants of C . burnetii did not induce any significant changes in cell morphology . The distribution of filamentous actin (F-actin) was then studied with a specific probe, bodipy phallacidin . Virulent C . burnetii induced a profound and transient reorganization of F-actin, accompanied by an increase in the F-actin content of THP-1 cells . F-actin was colocalized with myosin in cell protrusions, suggesting that actin polymerization and the tension of actin-myosin filaments play a role in C . burnetii-induced morphological changes . In addition, contact between the cell and the bacterium seems to be necessary to induce cytoskeleton reorganization . Bacterial supernatants did not stimulate actin remodeling, and virulent C . burnetii organisms were found in close apposition with F-actin protrusions . The manipulation of the actin cytoskeleton by C . burnetii may therefore play a critical role in the internalization strategy of this bacterium.

Infect Immun, 1998 Nov, 66(11), 5364 - 71
Fusion of Chlamydia trachomatis-containing inclusions is inhibited at low temperatures and requires bacterial protein synthesis; Van Ooij C et al.; The human pathogen Chlamydia trachomatis is an obligate intracellular bacterium with a unique developmental cycle . Within the host cell cytoplasm, it resides within a membrane-bound compartment, the inclusion . A distinguishing characteristic of the C . trachomatis life cycle is the fusion of the chlamydia-containing inclusions with each other in the host cell cytoplasm . We report that fusion of inclusions does not occur at 32 degreesC in multiple mammalian cell lines and with three different serovars of C . trachomatis . The inhibition of fusion was inclusion specific; the fusion with sphingolipid-containing secretory vesicles and the interaction with early endosomes were unaffected by incubation at 32 degreesC . The inhibition of fusion of the inclusions was not primarily the result of delayed maturation of the inclusion, as infectious progeny was produced in host cells incubated at 32 degreesC, and the unfused inclusions remained competent to fuse up to 48 h postinfection . The ability to reverse the inhibition of fusion by shifting the infected cells from 32 to 37 degreesC allowed the measurement of the rate and the time of fusion of the inclusions after entry of the bacteria . Most significantly, we demonstrate that fusion of inclusions with each other requires bacterial protein synthesis and that the required bacterial protein(s) is present, but inactive or not secreted, at 32 degreesC.

J Infect Dis, 1998 Nov, 178(5), 1379 - 90
Type I Helicobacter pylori shows Lewis(b)-independent adherence to gastric cells requiring de novo protein synthesis in both host and bacteria; Su B et al.; Type I Helicobacter pylori strains frequently recognize the Lewisb (Leb) blood group antigen . This binding property and expression of the Leb oligosaccharide were required for adherence to fixed normal or pathologic gastric tissue . In contrast, both type I and type II strains adhered to cultured cells in the absence of the Leb epitope . For the gastric cell line AGS, adherence was significantly higher when viable type I strains were allowed to interact with viable AGS cells compared with fixed cells . The observation that chloramphenicol and cycloheximide, inhibitors of bacterial and eukaryotic protein synthesis, respectively, significantly reduced adherence of type I but not type II isolates suggests that in type I strains, adherence depends on the up-regulation of one or more host cell receptors triggered by the bacterium.

J Immunol, 1998 Oct 15, 161(8), 4220 - 6
Apoptosis of epithelial cells and macrophages due to infection with the obligate intracellular pathogen Chlamydia psittaci; Ojcius DM et al.; We have characterized the cytotoxic activity of the obligate intracellular bacterium Chlamydia psittaci, which resides within a membrane-bound vacuole during the 2-day infection cycle . We have established that infected epithelial cells and macrophages die through apoptosis, which is measurable within 1 day of infection and requires productive infection by the bacteria . Inhibition of host cell protein synthesis has no effect on cell death, but blocking bacterial entry or bacterial protein synthesis prevents apoptosis, implying that bacterial growth is required for death of the host cell . Apoptosis was confirmed through the use of electron microscopy, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling, gel agarose electrophoresis of fragmented DNA, and propidium-iodide labeling of host cell nuclei . Although infected cells died preferentially, both infected and uninfected cells became apoptotic, suggesting that the infected cells may secrete proapoptotic factors . Inhibition of either of two proapoptotic enzymes, caspase-1 or caspase-3, did not significantly affect Chlamydia-induced apoptosis . These results suggest that, as in the case of apoptosis due to Bax expression or oncogene dysregulation, which initiate the apoptotic program within the cell interior, the Chlamydia infection may trigger an apoptotic pathway that is independent of known caspases . As apoptotic cells secrete proinflammatory cytokines, Chlamydia-induced apoptosis may contribute to the inflammatory response of the host.

Biochemistry, 1998 Oct 20, 37(42), 14875 - 80
The effect of pressure on the bacteriochlorophyll a binding sites of the core antenna complex from Rhodospirillum rubrum; Sturgis JN et al.; In this paper we examine the effect of pressure on the absorption spectrum and binding site of the core antenna complex from the photosynthetic bacterium Rhodospirillum rubrum . Absorption spectra and Raman spectra in preresonance with the Qy transition of the bacteriochlorophyll a were studied at pressures up to 625 MPa . In agreement with previous work we observe a pressure-induced red shift and broadening of the absorption spectrum . We show that at these pressures the pigments within the protein matrix at room temperature experience little if any distortion, and the hydrogen-bonding network involving the C2 and C9 carbonyl groups of the pigment molecules are undisturbed . Having shown the lack of sensitivity to pressure of the binding site interactions, which are known to modulate the absorption spectrum, we feel that it is relatively safe to attribute the pressure-induced red shift broadly to solvatochromic effects and, in particular, to the modulation of the pigment-pigment interactions by the pressure . This paper represents the first vibrational study of photosynthetic complexes at high pressure and the first application of FT Raman spectroscopy to biological molecules at high pressure.

J Med Entomol, 1998 Sep, 35(5), 653 - 9
Minimum infection rate of Ambylomma americanum (Acari: Ixodidae) by Ehrlichia chaffeensis (Rickettsiales: Ehrlichieae) in southern Indiana; Burket CT et al.; In 1994 and 1995, 8 cases of human monocytic ehrlichiosis were confirmed . These cases originated from southern counties where the putative tick vector Ambylomma americanum (L.) is well established . To confirm the presence of Ehrlichia chaffeensis in ticks in southern Indiana and to determine the minimum infection rate, specimens of A . americanum were collected from 5 counties (7 sites) . Nucleic acid was isolated from 88 pools of ticks (430 individuals) using an optimized phenol/CTAB (cetyltrimethylammonium bromide) extraction procedure and subjected to polymerase chain reaction analysis using species-specific 16S rRNA gene bacterial primers . Twenty-one of 88 pools (a minimum of 21 of 430 individuals) were positive for the presence of E . Chaffeensis, yielding an average minimum infection rate of 4.9% . Minimum infection rates at individual sites ranged from 0 to 9.4% . These data extend the known distribution of the bacterium to 3 southern counties of Indiana and suggest a higher prevalence of E . chaffeensis than previously reported for Missouri, North Carolina, or Kentucky.

J Periodontol, 1998 Sep, 69(9), 998 - 1007
Relationship between conversion of localized juvenile periodontitis-susceptible children from health to disease and Actinobacillus actinomycetemcomitans leukotoxin promoter structure; Bueno LC et al.; The periodontal pathogen Actinobacillus actinomycetemcomitans produces a leukotoxin that is considered a primary virulence factor in localized juvenile periodontitis (LJP) . Select strains of the bacterium contain a 530-bp deletion in the promoter region of the leukotoxin gene operon which results in enhanced transcription of the leukotoxin . DNA hybridization and polymerase chain reaction (PCR) were used to examine genetic variants of A . actinomycetemcomitans in 24 LJP-susceptible children from 21 families having a history of the disease and 34 control children from non-LJP families . A significant association was found between the detection of variants that had a deletion in the leukotoxin promoter region, indicative of a high level expression leukotoxin genotype, and conversion from a healthy periodontal status to disease . Subjects harboring A . actinomycetemcomitans of this genotype were more likely to convert to LJP than those subjects who had variants containing the full length leukotoxin promoter region (odds ratio = 22.5; 95% C.I., 2.84 < 206.66) {corrected} . These findings support the concept that highly virulent strains or clonal types of periodontal pathogens play a major role in the initiation of periodontal disease in susceptible hosts.

J Clin Microbiol, 1998 Nov, 36(11), 3420 - 2
Effect of oxygen on growth of Mycobacterium ulcerans in the BACTEC system; Palomino JC et al.; The effect of low oxygen concentration on the growth of 15 strains of Mycobacterium ulcerans was evaluated in the BACTEC system . Reduced oxygen tension enhanced the growth of M . ulcerans, suggesting that this organism has a preference for microaerobic environments . Application of this observation may improve rates of isolation of M . ulcerans in primary culture from clinical samples and promote isolation of the bacterium from environmental sources.

J Clin Microbiol, 1998 Nov, 36(11), 3285 - 90
Detection of point mutations associated with resistance of Helicobacter pylori to clarithromycin by hybridization in liquid phase; Pina M et al.; When the standard procedure for determining antibiotic susceptibility of bacteria is used, the results are delayed, especially for bacteria that grow slowly, such as Helicobacter pylori . Treatment for this bacterium may involve clarithromycin, a compound for which resistance has been associated with point mutations on the 23S rRNA gene . This resistance is currently found in organisms isolated from 0 to 15% of patients and jeopardizes the success of the treatment . We have designed a test involving amplification and colorimetric hybridization in the liquid phase to detect the mutation at the molecular level . First, four reference strains, including the wild type and three strains with the mutations A2143C, A2143G, and A2144G, were used to optimize the method . Amplification was carried out with primers previously published . The amplified products were added to probe-coated microtiter wells . A DNA enzyme immunoassay was used to detect the hybrids . The optimal conditions of the hybridization were defined for each probe . Nineteen H . pylori strains resistant to clarithromycin and 22 susceptible according to phenotypic data were submitted to restriction with BsaI and BbsI, and part of the 23S rRNA gene was sequenced in order to determine the mutation involved for the resistant strains . The new assay showed a complete correlation with the reference methods, except for one strain . Cross-hybridizations as well as application of the reaction to other bacteria did not lead to optical densities higher than the cutoff values chosen with the receiving operating characteristic curve . This method can be easily standardized and gives a result within a day . Its application directly to the biopsy specimens or infected gastric juice is planned in the future.

Proc Natl Acad Sci U S A, 1998 Oct 13, 95(21), 12376 - 80
Loss of social behaviors by myxococcus xanthus during evolution in an unstructured habitat
Velicer GJ, Kroos L, Lenski RE.
Social behaviors are often targets of natural selection among higher organisms, but quantifying the effects of such selection is difficult . We have used the bacterium Myxococcus xanthus as a model system for studying the evolution of social interactions . Changes in the social behaviors of 12 M . xanthus populations were quantified after 1,000 generations of evolution in a liquid habitat, in which interactions among individuals were continually hindered by shaking and low cell densities . Derived lineages were compared with their ancestors with respect to maximum growth rate, motility rates on hard and soft agar, fruiting body formation ability, and sporulation frequency during starvation . Improved performance in the liquid selective regime among evolved lines was usually associated with significant reductions in all of the major social behaviors of M . xanthus . Maintenance of functional social behaviors is apparently detrimental to fitness under asocial growth conditions.

Acta Virol, 1998 Apr, 42(2), 95 - 101
Isolation and characterization of the dnaA gene of Rickettsia prowazekii; Waite RT et al.; The dnaA gene encoding the initiator protein of DNA replication was isolated from the obligate intracellular bacterium, Rickettsia prowazekii . Comparison of the deduced amino acid sequence of R . prowazekii DnaA with other bacterial DnaA proteins revealed extensive similarity . However, the rickettsial sequence is unique in the number of basic lysine residues found within a highly conserved portion of the putative DNA binding region, suggesting that the rickettsial protein may recognize a DNA sequence that differs from the consensus DnaA box sequence identified in other bacteria . Consensus DnaA box sequences, found upstream of many bacterial dnaA genes, were not identified upstream of rickettsial dnaA gene . In addition, gene organization within this region differed from that of other bacteria . The putative start of transcription of the rickettsial dnaA gene was localized to a site 522 nucleotides (nt) upstream of the DnaA start codon.

Aliment Pharmacol Ther, 1998 Sep, 12(9), 857 - 64
Helicobacter pylori-positive peptic ulcer patients do not adapt to aspirin; Konturek JW et al.; BACKGROUND: Recent studies indicate that eradication of Helicobacter pylori might prevent peptic ulcer formation in patients treated with non-steroidal anti-inflammatory drugs (NSAIDs) . On the other hand, gastric adaptation after repeated exposures to aspirin (ASA) is well documented but the influence of H . pylori on this process remains to be elucidated . AIM: To compare gastric damage and adaptation following repeated exposures to ASA in a group of patients with H . pylori infection, before and after eradication of the bacterium, and in H . pylori-negative controls . METHODS: Eight healthy volunteers without H . pylori infection and eight patients with duodenal ulcer (DU) history and H . pylori infection before and after H . pylori eradication were given ASA 2 g/day for a period of 14 days . Mucosal damage was evaluated by endoscopy and histology of biopsy samples . Gastric microbleeding, DNA synthesis in the gastric mucosa and mucosal expression, as well as luminal content of transforming growth factor-alpha (TGFalpha) were determined on days 0, 3, 7 and 14 of the ASA course . RESULTS: In all patients aspirin-induced gastric damage reached a maximum on day 3 . In H . pylori-positive patients, this damage was maintained at a similar level up to day 14, whereas in H . pylori-negative controls and H . pylori-eradicated patients this damage significantly lessened on day 14 and was accompanied by elevated DNA synthesis as well as increased mucosal expression and luminal release of TGFalpha.

Curr Microbiol, 1998 Nov, 37(5), 324 - 32
Protection of mice against challenge with homologous and heterologous serovars of Actinobacillus pleuropneumoniae after live vaccination; Prideaux CT et al.; Protective immune responses and the virulence of Actinobacillus pleuropneumoniae (APP) have been attributed, in part, to toxins (Apx) produced by the bacterium . A mutant of the serovar 7 strain HS93 (HS93Tox-), lacking the genes encoding the structural toxin ApxA and the post-translational activating protein ApxC, but retaining the genes required for secretion ApxB and ApxD, was isolated and shown to be attenuated in a mouse model . A plasmid vector system was developed and used to express the ApxA gene from within the HS93Tox- strain . The resulting strain, HS93Tox-/pIG-T1K, expresses the Apx structural protein in a non-activated form . HS93Tox-/pIG-T1K was shown to be attenuated in a mouse model and to be capable of inducing Apx-specific antibodies, which were boosted on re-inoculation . Live vaccination of mice with HS93Tox-/pIG-T1K offered protection against homologous wild-type serovar 7 challenge, and also heterologous challenge with a serovar 1 strain . This is in contrast to vaccination with the HS93Tox- strain, which failed to protect mice against a heterologous challenge.

J Membr Biol, 1998 Oct 1, 165(3), 213 - 25
Charge displacements in interfacial layers containing reaction centers; Brzezinski P et al.; Reaction centers from the photosynthetic bacterium Rhodobacter sphaeroides were oriented in phospholipid interfacial layers adsorbed to a Teflon film separating two electrolyte-filled compartments of a Teflon cell . Light-induced voltage changes were measured as a function of time across electrodes immersed in the cell compartments . The experimental system is characterized both experimentally and theoretically to relate the measured signals to the light-induced displacement currents in the reaction centers . Mathematical relations between the measured signals and the distances and geometries of the charge-transfer reactions are derived . At pH 8.0 the reaction centers were found to be oriented with approximately 60% of the population oriented with the donor facing the aqueous phase . The density of the reaction centers in the layer was approximately 10(11) cm-2, which is close to that found in the native system . Reconstitution of the secondary quinone, QB, in 90% of the RCs was achieved with an approximately 100-fold excess of ubiquinone in the vesicle preparation.

Mol Microbiol, 1998 Aug, 29(4), 963 - 73
Morphological adaptation and inhibition of cell division during stationary phase in Caulobacter crescentus; Wortinger MA et al.; During exponential growth, each cell cycle of the alpha-purple bacterium Caulobacter crescentus gives rise to two different cell types: a motile swarmer cell and a sessile stalked cell . When cultures of C . crescentus are grown for extended periods in complex (PYE) medium, cells undergo dramatic morphological changes and display increased resistance to stress . After cultures enter stationary phase, most cells are arrested at the predivisional stage . For the first 6-8 days after inoculation, the colony-forming units (cfu) steadily decrease from 10(9) cfu ml(-1) to a minimum of 3x10(7) cfu ml(-1) after which cells gradually adopt an elongated helical morphology . For days 9-12, the cfu of the culture increase and stabilize around 2 x 10(8) cfu ml(-1) . The viable cells have an elongated helical morphology with no constrictions and an average length of 20 microm, which is 15-20 times longer than exponentially growing cells . The level of the cell division initiation protein FtsZ decreases during the first week in stationary phase and remains at a low constant level consistent with the lack of cell division . When resuspended in fresh medium, the elongated cells return to normal size and morphology within 12 h . Cells that have returned from stationary phase proceed through the same developmental changes when they are again grown for an extended period and have not acquired a heritable growth advantage in stationary phase (GASP) compared with overnight cultures . We conclude that the changes observed in prolonged cultures are the result of entry into a new developmental pathway and are not due to mutation.

Res Microbiol, 1998 May, 149(5), 361 - 72
Stable electrotransformation of symbiont candidate diazotrophic bacterium with plasmids carrying selectable and screenable marker genes; Koranyi P et al.; Nitrogen-fixing symbioses had been established between the originally asymbiotic soil bacterium Azotobacter vinelandii CCM289 and different lower and higher plant species . Better characterization and further development of such artificial systems require a reliable genetic transformation method for the introduction of marker genes into symbiont candidates . The performance of electroporation was evaluated using pJB3 (4.8 kb), pBI121 (12.8 kb) and pFAJ31.2 (24 kb) plasmid DNAs containing selectable (Ap, Km, Tc) and screenable (gusA, lacZ) marker genes . The adapted methods for the preparation of transformation-competent azotobacters and their electroporation (18 kV/cm electric field strength, 5 ms time constant, 0 degree C) provided up to 6.8 x 10(5) transformants per microgram plasmid DNA, which is about 10(3) times the transformation efficiency achieved in control experiments . No electrotransformants were obtained with the 24-kb pFAJ31.2 . The size of plasmid DNA did not significantly affect the efficiency of transformation . Transformants were able to grow at antibiotic concentrations that were 100-200 times greater than the lowest amounts that completely inhibited the growth of wild-type bacteria . A constitutive expression of gusA gene was observed in transformants with the CaMV 35S promoter-gusA fusion containing pBI121, while lacZ expression was not detected under the control of the lac promoter in pJB3 transformants . Electroporated plasmids were reisolated from transformants in their original form, while non-transformed bacteria did not contain indigenous plasmids . PCR amplification and Southern DNA blot hybridization showed the integration of plasmid DNA into the host genome as well . Transformants retained their nitrogen-fixing ability and had normal morphological and growth characteristics . Experimental findings proved the stable maintenance of plasmid DNA in azotobacters, making possible the routine transformation and detection of these symbiont candidates.

AIDS Res Hum Retroviruses, 1998 Sep 20, 14(14), 1263 - 9
HIV peptide conjugated to heat-killed bacteria promotes antiviral responses in immunodeficient mice; Scott DE et al.; Enhancement of immunity in the setting of HIV infection is difficult owing to loss of functional CD4+ T cells . The MHC class II-deficient mouse (II-/-) environment simulates that of the immunocompromised HIV-infected individual, since these mice have low CD4+ T cell numbers, defective CD4-dependent responses, and are susceptible to opportunistic infection . This strain was used to test whether heat-killed Brucella abortus (BA), covalently conjugated to the V3 peptide of HIV-1 (MN), could elicit anti-HIV responses . V3-BA, but not the T-dependent antigen V3-KLH, induced high levels of IL-12, IFN-gamma, and IL-10 mRNA in both wild-type (WT) and II-/- mice within 24 hr of injection . V3-BA-treated, but not V3-KLH-treated, II-/- mice developed serum IgG and IgA anti-V3 antibodies, with IgG2b and IgG3 as the predominant isotype . Viral neutralization studies, using a syncytium inhibition assay, demonstrated that the antibodies generated by V3-BA in II-/- mice were capable of neutralizing HIV . These experiments demonstrate that a heat-inactivated bacterium such as BA, when used as a carrier, can generate a cytokine environment that results in the production of neutralizing antiviral antibodies in an immunodeficient host . Such strategies could be important in the development of immunotherapies and vaccines for HIV-1 patients.

Eur J Clin Microbiol Infect Dis, 1998 Jul, 17(7), 512 - 5
Five cases of Kingella kingae skeletal infection in a French hospital; La Scola B et al.; Five cases of Kingella kingae skeletal infections were diagnosed in children admitted to La Timone Hospital between 1992 and 1997 . Patients were between 6 and 31 months old and presented with septic spondylodiskitis, calcaneus osteomyelitis, and hip-joint arthritis . All displayed either an upper respiratory tract infection or eczema during the month prior to their admission . Laboratory findings included an elevated leukocyte count and an elevated erythrocyte sedimentation rate . Standard radiography was unrevealing, but 99mTc bone scans and magnetic resonance imaging showed significant abnormalities . Isolation of Kingella kingae was achieved in all cases by culture of fluid aspirates using the Bactec blood culture system . This bacterium was sensitive to the most common antibiotics tested, and the outcome was favourable in all cases.

Acta Crystallogr D Biol Crystallogr, 1998 Mar 1, 54 ( Pt 2), 284 - 7
Crystallization and preliminary X-ray diffraction analysis of cytochrome c' from Rubrivivax gelatinosus at 1.3 A resolution; Benini S et al.; Cytochrome c' from the purple non-sulfur phototrophic bacterium Rubrivivax gelatinosus has been crystallized by vapour diffusion at pH 5, 6.3 and 8, in sodium acetate, sodium citrate, and Tris-HCl buffers, respectively . Crystals grown at pH 5 and 6.3 diffract, respectively, to 2.0 A (298 K) and 1.4 A (100 K) using synchrotron radiation . Data up to 1.3 A resolution with 99.8% completeness were collected at 100 K on a crystal grown at pH 8 . The space group is P3121 or P3221, and the unit-cell parameters are a = b = 69.63, c = 123.63 A.

Anal Biochem, 1978 Dec, 91(2), 421 - 31
A new, fast, and very sensitive bioluminescence assay for phospholipases A and C; Ulitzur S et al.; A new, simple, and very sensitive assay for phospholipase A and C is described . The assay is based on the bioluminescence developed by the mutant of the bacterium Beneckea harveyi as a response to myristic acid released from dimyristoyl phosphatidylcholine by either phospholipase A or by a phospholipase C-lipase coupled system . It is possible to assay these enzymes at a rate corresponding to a release of as little as 1 to 2 pmol of myristic acid per minute.

Microbiol Res, 1998 Aug, 153(2), 163 - 71
Purification and characterization of four catechol 1,2-dioxygenase isozymes from the benzamide-assimilating bacterium Arthrobacter species BA-5-17; Murakami S et al.; When Arthrobacter sp . BA-5-17 was grown on benzamide, the bacterium synthesized four different catechol 1,2-dioxygenase (CD, EC 1.13.11.1) isozymes (CD-I, II, III-1, and III-2) . We purified each CD to homogeneity by a series of column chromatography . The molecular masses of the four CDs were between 68 and 72 kDa . The enzymes were made up of two identical subunits each with the molecular mass of 33 kDa . CD-I and II were indistinguishable in enzymatic properties tested . Most properties of CD-III-1 were similar to those of CD-III-2 . However, CD-III-1 had a marked adsorption peak at 325 nm, which disappeared in CD-III-2 as well as in CD-I and II . CD-III-1 and III-2 were much more resistant to heating and inhibitors than CD-I and II.

Appl Environ Microbiol, 1998 Oct, 64(10), 3663 - 8
A plant growth-promoting bacterium that decreases nickel toxicity in seedlings
Burd GI, Dixon DG, Glick BR.
A plant growth-promoting bacterium, Kluyvera ascorbata SUD165, that contained high levels of heavy metals was isolated from soil collected near Sudbury, Ontario, Canada . The bacterium was resistant to the toxic effects of Ni2+, Pb2+, Zn2+, and CrO4-, produced a siderophore(s), and displayed 1-aminocyclopropane-1-carboxylic acid deaminase activity . Canola seeds inoculated with this bacterium and then grown under gnotobiotic conditions in the presence of high concentrations of nickel chloride were partially protected against nickel toxicity . In addition, protection by the bacterium against nickel toxicity was evident in pot experiments with canola and tomato seeds . The presence of K . ascorbata SUD165 had no measurable influence on the amount of nickel accumulated per milligram (dry weight) of either roots or shoots of canola plants . Therefore, the bacterial plant growth-promoting effect in the presence of nickel was probably not attributable to the reduction of nickel uptake by seedlings . Rather, it may reflect the ability of the bacterium to lower the level of stress ethylene induced by the nickel.

Scand J Gastroenterol, 1998 Sep, 33(9), 916 - 22
Effect of sucralfate on gastric mucosal inflammatory responses induced by Helicobacter pylori lipopolysaccharide; Slomiany BL et al.; BACKGROUND: Helicobacter pylori lipopolysaccharide is emerging as a primary factor in the bacterium virulence, and its involvement in causing gastric mucosal responses typical of gastritis has recently been shown . In this study we investigated the effect of the antiulcer agent sucralfate on the expression of regulatory cytokines, tumor necrosis factor-alpha (TNF-alpha) and interleukin-4 (IL-4), and epithelial cell apoptosis during H . pylori lipopolysaccharide-induced acute gastritis . METHODS: The experiments were conducted with rats pretreated intragastrically twice daily for 3 days with sucralfate at 100 mg/kg or the vehicle . The rats were then subjected to intragastric surface epithelial application of H . pylori lipopolysaccharide at 50 microg per animal and maintained on the sucralfate or vehicle regimen for an additional 4 days . The animals were killed 16 h after the last dose, and their gastric mucosal tissue used for histologic assessment, quantitation of TNF-alpha and IL-4 expression, and the assay of epithelial cell apoptosis . RESULTS: In the absence of sucralfate, H . pylori lipopolysaccharide induced acute mucosal responses characterized by the inflammatory infiltration of the lamina propria, hyperemia, and epithelial hemorrhage . This was accompanied by an 11-fold increase in gastric epithelial cell apoptosis and a 9-fold enhancement of the mucosal expression of TNF-alpha, but the level of IL-4 fell by 15% . Intragastric administration of sucralfate produced a 62% reduction in the extent of mucosal damage caused by H . pylori lipopolysaccharide, a 51% decrease in the mucosal expression of TNF-alpha, and a 7-fold reduction in the extent of epithelial cell apoptosis, whereas the expression of IL-4 increased by 52% . CONCLUSIONS: Gastric mucosal inflammatory responses to H . pylori lipopolysaccharide are characterized by a massive enhancement of the proinflammatory cytokine TNF-alpha and epithelial cell apoptosis and repression of IL-4 . Our data also show that sucralfate is capable of inducing expression of the regulatory cytokine IL-4 and the suppression of apoptotic events triggered in gastric mucosa by the increase in TNF-alpha that is elicited by H . pylori lipopolysaccharide.

Acta Crystallogr D Biol Crystallogr, 1998 Sep 1, 54 ( Pt 5), 1012 - 3
Preliminary crystallographic studies of citrate synthase from an Antarctic psychrotolerant bacterium; Gerike U et al.; Recombinant citrate synthase from a psychrotolerant bacterium, DS2-3R, recently isolated in Antarctica, has been crystallized . The crystals belong to space group P6122 or P6522, with cell dimensions a = b = 70.8, c = 307.8 A . Diffraction data collected on a synchrotron from a cryoprotected crystal extends to at least 2.0 A . Knowledge of the structure of this enzyme will add to the understanding of cold activity and thermolability, and will be of biotechnological interest . Previously, the structure of citrate synthase from Archaea inhabiting environments at 328 and 373 K, has been reported . This present study will extend our understanding of the structural integrity and activity of proteins at the temperature extremes of life.

Biochim Biophys Acta, 1998 Jul 20, 1365(3), 373 - 384
The triplet state of the FMO complex of the green sulfur bacterium Prosthecochloris aestuarii studied with single-crystal EPR; Louwe RJW et al.; Triplet-electron paramagnetic resonance (EPR) spectra were obtained of single crystals of the FMO complex of the green sulfur bacterium Prostecochloris aestuarii . The experiments support the results presented in a previous paper (Louwe et al., J . Phys . Chem . 101 (1997) 11280), which showed that the experimental optical spectra of this pigment-protein complex are best reproduced by assuming that one bacteriochlorophyll (BChl 3) is energetically isolated and that this BChl is the triplet-carrying BChl of the FMO complex at cryogenic temperatures for low excitation density . When comparing the experimental and simulated data sets of the triplet-EPR spectra in single crystals, the best fit is obtained for two triplet states, one localized at BChl 3 and the other at BChl 1 . The existence of two different triplet states is traced to the relatively high excitation power necessary to observe the small triplet-EPR signal of the FMO single crystals.

Proc Natl Acad Sci U S A, 1998 Sep 29, 95(20), 11957 - 62
Chemotaxis in a gliding bacterium
Kearns DB, Shimkets LJ.
Myxococcus xanthus cells exhibit directed motility up phosphatidylethanolamine (PE) gradients, and we suggest that PE behaves as a chemoattractant . Computer-assisted stop-motion digital microscopy was used to record cell movements in slide culture . PE decreased cellular reversal frequency with molecular specificity that was correlated with the fatty acid composition . Synthetic dilauroyl (di C12:0) PE and dioleoyl (di C18:1 omega9c) PE suppressed direction reversals and stimulated movement up the gradient . Sensory adaptation occurred about 1 hr after the onset of stimulation . Null mutants in a methylated chemotaxis protein homolog (FrzCD) and a CheA/CheY homolog (FrzE) moved up a PE gradient at a reduced rate . The mutants displayed normal excitation but were defective in adaptation . A dominant, hyper-reversal mutant in the M . xanthus methyl accepting chemotaxis protein homolog, frzCD224, failed to respond to PE stimulation, which argued that PE was a transduced stimulus . Neither dilauroyl PE nor dioleoyl PE is present at high enough concentrations in vegetative or developmental PE to account for all of the chemotactic activity . It appears then that there are additional, as yet unknown, PE species that serve as autoattractants . We report on a discrete phospholipid chemoattractant in a gliding bacterium

Biochemistry, 1998 Sep 29, 37(39), 13674 - 80
Synthesis and biochemical characterization of an analogue of CheY-phosphate, a signal transduction protein in bacterial chemotaxis; Halkides CJ et al.; CheY is a signal transduction protein of the bacterial chemotaxis system that acts as a molecular switch to alter the swimming behavior of the bacterium . When CheY becomes phosphorylated at Asp57, CheY-Pi interacts with flagellar motor proteins, including FliM, to increase the likelihood that the flagellar motor will change its sense of rotation, increasing the frequency of tumbling . The structure of CheY in its dephosphorylated (inactive) state has been intensively investigated . The short lifetime ( approximately 20 s) of the aspartyl phosphate has precluded the complete structural determination of CheY-Pi . We have synthesized an analogue of CheY-Pi by alkylating an aspartate-to-cysteine mutant at position 57 of CheY to add a phosphonomethyl group at Cys57 . This analogue, phosphono-CheY, is stable for months . Phosphono-CheY binds to two of the targets of CheY-Pi, FliM and CheZ, in a manner similar to that of CheY-Pi and much better than either unphosphorylated CheY or the unmodified form of D57C CheY . Phosphono-CheY also binds Mg(II) with a dissociation constant of approximately 6 mM at neutral pH and moderate salt level . These observations indicate that phosphono-CheY is a good biochemical analogue of CheY-Pi.

Proc Natl Acad Sci U S A, 1998 Sep 29, 95(20), 11679 - 84
Conformational gating of the electron transfer reaction QA-.QB --> QAQB- . in bacterial reaction centers of Rhodobacter sphaeroides determined by a driving force assay; Graige MS et al.; The mechanism of the electron transfer reaction, QA-.QB --> QAQB-., was studied in isolated reaction centers from the photosynthetic bacterium Rhodobacter sphaeroides by replacing the native Q10 in the QA binding site with quinones having different redox potentials . These substitutions are expected to change the intrinsic electron transfer rate by changing the redox free energy (i.e., driving force) for electron transfer without affecting other events that may be associated with the electron transfer (e.g., protein dynamics or protonation) . The electron transfer from QA- . to QB was measured by three independent methods: a functional assay involving cytochrome c2 to measure the rate of QA- . oxidation, optical kinetic spectroscopy to measure changes in semiquinone absorption, and kinetic near-IR spectroscopy to measure electrochromic shifts that occur in response to electron transfer . The results show that the rate of the observed electron transfer from QA- . to QB does not change as the redox free energy for electron transfer is varied over a range of 150 meV . The strong temperature dependence of the observed rate rules out the possibility that the reaction is activationless . We conclude, therefore, that the independence of the observed rate on the driving force for electron transfer is due to conformational gating, that is, the rate limiting step is a conformational change required before electron transfer . This change is proposed to be the movement, controlled kinetically either by protein dynamics or intermolecular interactions, of QB by approximately 5 A as observed in the x-ray studies of Stowell et al . {Stowell, M . H . B., McPhillips, T . M., Rees, D . C., Soltis, S . M., Abresch, E . & Feher, G . (1997) Science 276, 812-816}.

Lett Appl Microbiol, 1998 Aug, 27(2), 83 - 5
Two restriction endonucleases in Selenomonas ruminantium subsp . lactilytica; Pristas P et al.; Crude protein extract from a recently isolated ruminal bacterium identified as Selenomonas ruminantium subsp . lactilytica specifically cleaved DNA . This ability was due to the presence of two site-specific restriction endonucleases . Sr/I, a NaeI schizomer, recognizes the 5'-GCCGGC-3' sequence . Sr/II, a NsiI schizomer, recognizes 5'-ATGCAT-3'.

Anal Biochem, 1998 Oct 1, 263(1), 102 - 12
Electrocatalytic reduction of S-nitrosoglutathione at electrodes modified with an electropolymerized film of a pyrrole-derived viologen system and their application to cellular S-nitrosoglutathione determinations; Wu Q et al.; The preparation, electrochemical characterization, and analytical applications of glassy carbon (GC) electrodes modified with electropolymerized films of the cation N,N'-di(3-pyrrol-1-yl-propyl)-4,4'-bipyridine (DPPB) are described . Electropolymerized films of DPPB on GC electrodes exhibit two one-electron redox processes centered at -0.45 and -0.85 V, respectively . S-Nitrosoglutathione (GSNO) can be electrocatalytically reduced at electrodes modified with electropolymerized films of DPPB at approximately -0.4 V vs sodium-saturated calomel electrode, which represents a dramatic diminution of about 600 mV in the overpotential in comparison with the reaction carried out at a bare GC electrode . The kinetics of the catalytic reaction have been characterized using cyclic voltammetry and rotated disk electrode techniques from which a value of (1.3 +/- 0.2) x 10(3)M-1 s-1 was obtained . Using electrodes modified with an electropolymerized film of DPPB we have carried out preliminary studies of the determination of intracellular GSNO concentrations in two strains of the bacterium Rhodobacter sphaeroides .

Med Microbiol Immunol (Berl), 1998 Jun, 187(1), 23 - 42
Identification and localization of Chlamydia pneumoniae in the Alzheimer's brain; Balin BJ et al.; We assessed whether the intracellular bacterium Chlamydia pneumoniae was present in post-mortem brain samples from patients with and without late-onset Alzheimer's disease (AD), since some indirect evidence seems to suggest that infection with the organism might be associated with the disease . Nucleic acids prepared from those samples were screened by polymerase chain reaction (PCR) assay for DNA sequences from the bacterium, and such analyses showed that brain areas with typical AD-related neuropathology were positive for the organism in 17/19 AD patients . Similar analyses of identical brain areas of 18/19 control patients were PCR-negative . Electron- and immunoelectron-microscopic studies of tissues from affected AD brain regions identified chlamydial elementary and reticulate bodies, but similar examinations of non-AD brains were negative for the bacterium . Culture studies of a subset of affected AD brain tissues for C . pneumoniae were strongly positive, while identically performed analyses of non-AD brain tissues were negative . Reverse transcription (RT)-PCR assays using RNA from affected areas of AD brains confirmed that transcripts from two important C . pneumoniae genes were present in those samples but not in controls . Immunohistochemical examination of AD brains, but not those of controls, identified C . pneumoniae within pericytes, microglia, and astroglia . Further immunolabelling studies confirmed the organisms' intracellular presence primarily in areas of neuropathology in the AD brain . Thus, C . pneumoniae is present, viable, and transcriptionally active in areas of neuropathology in the AD brain, possibly suggesting that infection with the organism is a risk factor for late-onset AD.

Mutat Res, 1998 Oct 12, 421(1), 73 - 82
Generation of a genetic polymorphism in clonal populations of the bacterium Streptomyces ambofaciens: characterization of different mutator states; Martin P et al.; In Streptomyces ambofaciens, colony pigmentation is an unstable character . Very unstable mutants selected from twelve wild type (WT) subclones of S . ambofaciens ATCC23877 were investigated . This research showed that the polymorphism in colony pigmentation had distinct features . The first aspect is the coexistence of four types of colonies: pigmented colonies (Pig+), pigment-defective colonies (Pigcol-), pigmented colonies harboring pigment-defective sectors (Pigsec+) or pigment-defective papillae (Pigpap+) . The second feature was revealed by the study on Pigpap+ colonies . We showed that WT progeny after 14 days of growth consisted almost totally of Pigpap+ colonies . Pigpap+ colonies were also found to be genetically different from each other . Characterization of twelve colonies presenting more than 20 papillae (Hyperpap colonies) led to the isolation of twelve mutator strains which produced at high frequency Pigcol- and Hyperpap colonies . Each exhibited a specific mutator phenotype and were distinct from each other . Such strains constituted a part of the polymorphism observed in each of the WT progeny and also generated a high variability . Finally, we showed that pigment-defective papillae were mutants and constituted a new form of genetic instability .

Infect Immun, 1998 Oct, 66(10), 4832 - 7
Urease plays an important role in the chemotactic motility of Helicobacter pylori in a viscous environment; Nakamura H et al.; Helicobacter pylori exhibits chemotactic responses to urea, flurofamide, acetohydroxamic acid, and sodium bicarbonate . In buffer, the chemotactic activities of a urease-positive strain were higher than those of the isogenic urease-negative strain . Moreover, the chemotactic activities of the urease-positive strain were increased in a viscous solution containing 3% polyvinylpyrrolidone, whereas those of the urease-negative mutant were not . These results are in accordance with the fact that the mutant strain did not show swarming in motility agar regardless of having flagella . Incubation of the wild-type strain with flurofamide resulted in partial inhibition of the chemotactic activities in the viscous solution . In addition, incubation with acetohydroxamic acid, a low-molecular-weight, diffusible urease inhibitor, resulted in complete loss of chemotactic activity in the viscous solution . The inhibition of the chemotactic activity by urease inhibitors paralleled the inhibition of urease . The chemotactic activity of H . pylori was also inhibited by the proton carrier carbonyl cyanide m-chlorophenylhydrazone, showing that H . pylori utilizes proton motive force for motility . These results indicate that cytoplasmic urease plays an important role in the chemotactic motility of H . pylori under a condition that mimics the ecological niche of the bacterium, the gastric mucous layer.

Infect Immun, 1998 Oct, 66(10), 4640 - 50
A second two-component regulatory system of Bordetella bronchiseptica required for bacterial resistance to oxidative stress, production of acid phosphatase, and in vivo persistence; Jungnitz H et al.; Random minitransposon mutagenesis was used to identify genes involved in the survival of Bordetella bronchiseptica within eukaryotic cells . One of the mutants which exhibited a reduced ability to survive intracellularly harbored a minitransposon insertion in a locus (ris) which displays a high degree of homology to two-component regulatory systems . This system exhibited less than 25% amino acid sequence homology to the only other two-component regulatory system described in Bordetella spp., the bvg locus . A risA'-'lacZ translational fusion was constructed and integrated into the chromosome of B . bronchiseptica . Determination of beta-galactosidase activity under different environmental conditions suggested that ris is regulated independently of bvg and is optimally expressed at 37 degrees C, in the absence of Mg2+, and when bacteria are in the intracellular niche . This novel regulatory locus, present in all Bordetella spp., is required for the expression of acid phosphatase by B . bronchiseptica . Although catalase and superoxide dismutase production were unaffected, the ris mutant was more sensitive to oxidative stress than the wild-type strain . Complementation of bvg-positive and bvg-negative ris mutants with the intact ris operon incorporated as a single copy into the chromosome resulted in the reestablishment of the ability of the bacterium to produce acid phosphatase and to resist oxidative stress . Mouse colonization studies demonstrated that the ris mutant is cleared by the host much earlier than the wild-type strain, suggesting that ris-regulated products play a significant role in natural infections . The identification of a second two-component system in B . bronchiseptica highlights the complexity of the regulatory network needed for organisms with a life cycle requiring adaptation to both the external environment and a mammalian host.

Rev Argent Microbiol, 1998 Apr-Jun, 30(2), 93 - 5
{Isolation of Helicobacter pylori from dental plaque}; Mattana CM et al.; It has been suggested that oral dissemination might be the major transmission vehicle for Helicobacter pylori, and that dental plaque might act as its reservoir . The presence of H . pylori was investigated in 62 odontological male and female patients (average age: 35 years old) . Samples were taken from supragingival plaque, placed in 0.3 ml of thioglycolate broth, cultured within 12 h in Mueller-Hinton agar with the addition of 5-7% of sheep blood and antibiotic supplement, and incubated at 37 degrees C in microaerophilia for 5-7 days . Typical colonies were identified by gram, urease, oxidase and catalase . H . pylori was detected in a 15 year-old patient suffering from gastric acidity (1.61% positivity index) . The medium used facilitated recovery of the agent from a sample abundant in germs . H . pylori was not recovered from the same patient 12 months later, suggesting that there might have been a transitory passage by gastric reflux or that the bacterium was acquired from an exogenous source.

J Mol Biol, 1998 Oct 2, 282(4), 833 - 45
Projection structures of three photosynthetic complexes from Rhodobacter sphaeroides: LH2 at 6 A, LH1 and RC-LH1 at 25 A; Walz T et al.; Three photosynthetic complexes, light-harvesting complex 2 (LH2), light-harvesting complex 1 (LH1), and the reaction centre-light-harvesting complex 1 photounit (RC-LH1), were purified from a single species of a purple bacterium, Rhodobacter sphaeroides, and reconstituted into two-dimensional (2-D) crystals . Vesicular 2-D crystals of LH1 and RC-LH1 were imaged in negative stain and projection maps at 25 A resolution were produced . The rings formed by LH1 have approximately the same mean diameter as the LH1 rings from Rhodospirillum rubrum ( approximately 90 A) and therefore are likely to be composed of 15 to 17 alphabeta subunits . In the projection map calculated from the RC-LH1 2-D crystals, the reaction centre is represented by an additional density in the centre of the ring formed by the LH1 subunits . The marked improvement of shape and fine structure after a rotational pre-alignment of the RC-LH1 unit cells before averaging strongly suggests that the RC is not in a unique orientation within the LH1 rings . Tubular crystals of LH2 showed a high degree of order and allowed calculation of a projection map at 6 A resolution from glucose-embedded specimens . The projection structure shows a ring of nine alphabeta subunits . Variation of the alpha-helical projection densities suggests that the 9-fold symmetry axis is tilted with respect to the membrane normal .

J Mol Biol, 1998 Oct 2, 282(4), 819 - 31
Are the light-harvesting I complexes from Rhodospirillum rubrum arranged around the reaction centre in a square geometry?
Stahlberg H, Dubochet J, Vogel H, Ghosh R.
The basic photosynthetic unit containing the reaction centre and the light-harvesting I complex (RC-LHI) of the purple non-sulphur bacterium Rhodospirillum rubrum was purified and reconstituted into two-dimensional (2D) membrane crystals . Transmission electron microscopy using conventional techniques and cryoelectron microscopy of the purified single particles and of 2D crystals yielded a projection of the RC-LHI complex at a resolution of at least 1.6 nm . In this projection the LHI ring appears to have a square symmetry and packs in a square crystal lattice . The square geometry of the LHI ring was observed also in images of single isolated particles of the RC-LHI complex . However, although the LHI units are packed identically within the crystal lattice, a new rotational analysis developed here showed that the reaction centres take up one of four possible orientations within the ring . This fourfold disorder supports our interpretation of a square ring symmetry and suggests that a hitherto undetected component may be present within the photosynthetic unit .

Syst Appl Microbiol, 1998 Mar, 21(1), 1 - 11
Properties of an alpha-galactosidase, and structure of its gene galA, within an alpha-and beta-galactoside utilization gene cluster of the hyperthermophilic bacterium Thermotoga maritima; Liebl W et al.; Thermotoga maritima represents one of the few hyperthermophilic bacteria currently known . The chromosomal alpha-galactosidase gene of T . maritima strain MSB8 has been cloned and its nucleotide sequence was determined . The gene, designated galA, has coding capacity for a 552 residue polypeptide with a calculated molecular mass of 63,653 Da . GalA was found to be flanked by other genes probably involved in galactoside breakdown and utilization . The previously sequenced beta-galactosidase gene, lacZ, is localized immediately upstream of galA while two open reading frames that putatively encode enzymes of galactose catabolism, i.e . galactose-1-phosphate uridylytransferase (galT) and galactokinase (galK), were found downstream of galA . The identified genes are extremely close together or even overlap and have the same orientation, so they could all be part of one galactoside utilization operon of T . maritima MSB8 . GalA displayed low-level amino acid sequence similarity with alpha-galactoside of glycosyl hydrolase family 36 . However, GalA is smaller than the other members of this enzyme family . The galA gene was expressed in Escherichia coli and the recombinant alpha-galactosidase was purified and characterized . The molecular mass of the recombinant enzyme was estimated at about 62 kDa by denaturting gel electrophoresis . Maximal hydrolysis of the chromogenic substrate p-nitrophenyl-alpha-D-galactopyranoside was measured at pH 5.0-5.5 and 90-95 degrees C (5 min assay) . Divalent cations were not required for activity . The enzyme released galactose from raffinose, melibiose and the synthetic substrates p-nitrophenyl-and omicron-nitrophenyl-alpha-D-galactopyranoside . The T . maritima alpha-galactosidase thus was highly specific for the galactose moiety and the alpha-anomeric configuration of the glycosidic linkage . Its extreme thermal stability (t 1/2 = 6.5 h at 85 degrees C) makes this enzyme an interesting candidate for biotechnological applications.

Eur J Biochem, 1998 Aug 1, 255(3), 604 - 10
Myxococcus xanthus spore coat protein S, a stress-induced member of the betagamma-crystallin superfamily, gains stability from binding of calcium ions; Wenk M et al.; Protein S, a calcium-binding spore coat protein from the soil bacterium Myxococcus xanthus, belongs to a group of structurally related proteins, the betagamma-crystallin superfamily . Common features of this protein family are the Greek-key structural motif or crystallin fold, and the fact that all members are extremely stable long term . To investigate the correlation between the stability and Greek-key topology, protein S was cloned, expressed in Escherichia coli and purified to homogeneity . Ca2+ binding influences the native tertiary structure of protein S, whereas the secondary structure remains unaffected as shown by spectroscopic methods . Ca2+ ions enhance the conformational stability of protein S significantly . The midpoints of urea and guanidinium chloride-induced transitions show a difference of 1.4 M and 0.5 M denaturant, respectively, in the absence and in the presence of calcium . An equilibrium intermediate indicating independent domain folding can be detected at pH 2 . In addition, thermal denaturation shows a clear deviation from the two-state model of folding, again with a strong stabilisation by Ca2+ ions . Temperature and denaturant-induced equilibrium transitions are fully reversible . Our data implicate a different strategy for achieving the high stability required for the biological function compared with the structurally related lens crystallins.

J Gastroenterol Hepatol, 1998 Jan, 13(1), 95 - 103
Gastrin release and gastric acid secretion in the rat infected with either Helicobacter felis or Helicobacter heilmannii; Danon SJ et al.; Helicobacter pylori infection in humans has been shown to be associated with changes in gastric physiology, including exaggerated basal and meal-stimulated gastrin levels . This has been suggested to be due to the direct effects of the bacterium through inflammation and its urease enzyme . The gastric bacteria Helicobacter felis and Helicobacter heilmannii colonize the antrum of rats in large numbers and induce no significant inflammatory response . Thus, the direct effect of Helicobacter infection on gastric physiology, independent of gastritis, could be studied . Basal, freely fed and stimulated acid and gastrin levels were recorded from animals infected with H . felis, H . heilmannii or uninfected controls over a 30 week period . No significant difference was found between freely fed gastrin over 7 weeks or fasting gastrin over 24 weeks or basal and stimulated acid over 30 weeks between all three groups . Triple therapy did not alter gastrin or acid output . The antrum of all Helicobacter-infected rats was well colonized; triple therapy cleared H . felis but not H . heilmannii . Very little inflammation was seen in control or Helicobacter-infected animals . In conclusion, Helicobacter-induced effects on gastric physiology are unlikely to be due to direct bacterial effects, but are best explained by other factors (i.e . inflammatory damage).

Int J Syst Bacteriol, 1998 Jul, 48 Pt 3, 1043 - 7
Proposal of Craurococcus roseus gen . nov., sp . nov . and Paracraurococcus ruber gen . nov., sp . nov., novel aerobic bacteriochlorophyll a-containing bacteria from soil; Saitoh S et al.; Sequences of the 16S rRNA gene were determined for three strains of aerobic bacteriochlorophyll a-containing bacteria isolated from soil . The sequence of two strains (NS89(T) and NS102) were identical for approximately 1500 nucleotides . Phylogenetic analysis revealed that the three strains belonged to the alpha-1 subclass of the Proteobacteria, constituting one line of descent . The three strains are comparatively related to Roseoccus thiosulfatophilus, which is an aerobic bacteriochlorophyll a-containing bacterium . The 16S rRNA gene sequence similarity and the DNA-DNA relatedness allow the proposal of two new genera, Craurococcus gen . nov . and Paracraurococcus gen . nov . The type species are Craurococus roseus sp . nov . and Paracraurococcus ruber sp . nov., and their type strains are NS130(T) (=JCM 9933(T)) and NS89(T) (=JCM 9931(T)), respectively.

Int J Syst Bacteriol, 1998 Jul, 48 Pt 3, 1037 - 41
Pseudoalteromonas prydzensis sp . nov., a psychrotrophic, halotolerant bacterium form Antarctic sea ice; Bowman JP; Species of the genus Pseudoalteromonas are frequently isolated from marine ecosystems and appear to be particularly abundant in Antarctic coastal waters . Most Pseudoalteromonas strains isolated form sea ice and underlying seawater samples are phenotypically similar to the species Pseudoalteromonas antarctica and Pseudoalteromonas nigrifaciens . However, a minority of isolates were recognized by phenotypic, DNA-DNA hybridization and 16S rRNA-based phylogenetic studies to represent a distinct genospecies clustering at the periphery of the non-pigmented, Pseudoalteromonas species clade . These strains are non-pigmented, halotolerant psychrotrophs that are capable of hydrolysing starch and chitin, and possess a DNA G+C content of 38-39 mol% . It is proposed that this group represents a novel species, Pseudoalteromonas prydzensis sp . nov., for which the type strain is ACAM 620(T).

J Biol Chem, 1998 Sep 18, 273(38), 24792 - 6
The subunit b of the F0F1-type ATPase of the bacterium Mycoplasma pneumoniae is a lipoprotein; Pyrowolakis G et al.; The DNA sequence analysis of the F0F1-ATPase operon of the bacterium Mycoplasma pneumoniae predicted that the subunit b, encoded by the gene atpF, is a lipoprotein of the murein lipoprotein type of Escherichia coli . Here we experimentally verify this prediction by metabolic labeling of subunit b with {14C}palmitic acid and by in vivo interfering with the processing of the prolipoprotein form of subunit b by the antibiotic globomycin, a specific inhibitor of the signal peptidase II . Our results suggest that the subunit b of the F0F1-ATPase of M . pneumoniae is anchored at the cytoplasmic membrane by an N-terminal lipid modification in addition to its transmembrane domain . The lipoprotein nature of subunit b and its proposed membrane topology seems to be characteristic for mycoplasmas, since among all sequenced bacterial atpF genes, only those from Mycoplasma gallisepticum and Mycoplasma genitalium code for a conserved lipoprotein consensus sequence.

Arch Microbiol, 1998 Oct, 170(4), 252 - 8
Environmental and physiological factors affecting the uptake of phosphate by chlorobium limicola
Baneras L, Garcia-Gil LJ.
The uptake of soluble phosphate by the green sulfur bacterium Chlorobium limicola UdG6040 was studied in batch culture and in continuous cultures operating at dilution rates of 0.042 or 0.064 h-1 . At higher dilution rates, washout occurred at phosphate concentrations below 7.1 &mgr;M . This concentration was reduced to 5 . 1 &mgr;M when lower dilution rates were used . The saturation constant for growth on phosphate (K&mgr;) was between 2.8 and 3.7 &mgr;M . The specific rates of phosphate uptake in continuous culture were fitted to a hyperbolic saturation model and yielded a maximum rate (Vamax) of 66 nmol P (mg protein)-1 h-1 and a saturation constant for transport (Kt) of 1.6 &mgr;M . In batch cultures specific rates of phosphate uptake up to 144 nmol P (mg protein)-1 h-1 were measured . This indicates a difference between the potential transport of cells and the utilization of soluble phosphate for growth, which results in a significant change in the specific phosphorus content . The phosphorus accumulated within the cells ranged from 0.4 to 1.1 &mgr;mol P (mg protein)-1 depending on the growth conditions and the availability of external phosphate . Transport rates of phosphate increased in response to sudden increases in soluble phosphate, even in exponentially growing cultures . This is interpreted as an advantage that enables Chl . limicola to thrive in changing environments.

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