Bacteria

Carbohydr Res, 1997 Oct 28, 304(1), 69 - 76
Novel alginate lyases from marine bacterium Alteromonas sp . strain H-4; Sawabe T et al.; A bacterium Alteromonas sp . strain H-4 isolated from Laminaria fronds produced extra- and intra-cellular alginate lyases and utilized alginate as its sole carbon source . An extracellular alginate lyase was purified from the culture supernatant of the strain and its substrate specificity was characterized . The estimated molecular mass of the enzyme was 32 kDa and the isoelectric point was 4.7 . Both polyM and polyG block degrading activities were observed using the substrate-containing gel overlay technique after isoelectric focusing of the enzyme . By analyzing the reaction products from the polyM block, polyG block, MG random block and intact alginate, three major peaks containing unsaturated tri-uronide through octa-uronide were detected for each substrate . The results indicate that the enzyme of Alteromonas sp . H-4 can degrade both polyM and polyG blocks with a K(m) in mg/mL 20-times higher for the polyM block.

J Bacteriol, 1997 Dec, 179(24), 7617 - 24
Characterization of genes encoding dimethyl sulfoxide reductase of Rhodobacter sphaeroides 2.4.1T: an essential metabolic gene function encoded on chromosome II; Mouncey NJ et al.; Rhodobacter sphaeroides 2.4.1T is a purple nonsulfur facultative phototrophic bacterium which exhibits remarkable metabolic diversity as well as genomic complexity . Under anoxic conditions, in the absence of light and the presence of dimethyl sulfoxide (DMSO) or trimethylamine N-oxide (TMAO), R . sphaeroides 2.4.1T utilizes DMSO or TMAO as the terminal electron acceptor for anaerobic respiration, which is mediated by the molybdoenzyme DMSO reductase . Sequencing of a 13-kb region of chromosome II revealed the presence of 10 putative open reading frames, of which 5 possess homology to genes encoding the TMAO reductase (the tor system) of Escherichia coli . The dorS and dorR genes encode a sensor-regulator pair of the two-component sensory transduction protein family, homologous to the torS and torR gene products . The dorC gene was shown to encode a 44-kDa DMSO-inducible c-type cytochrome . The dorB gene encodes a membrane protein of unknown function homologous to the torD gene product . The dorA gene encodes DMSO reductase, containing the molybdopterin active site . Mutations were constructed in each of these dor genes, and the resulting mutants were shown to be impaired for DMSO-dependent anaerobic growth in the dark . The mutant strains exhibited negligible levels of DMSO reductase activity compared to the wild-type strain under similar growth conditions . Further, no DorA protein was detected in DorS and DorR mutant strains with anti-DorA antisera, suggesting that the products of these genes are required for the positive regulation of dor expression in response to DMSO . This characterization of the dor gene cluster is the first evidence that genes of chromosome CII encode metabolic functions which are essential under particular growth conditions.

Helicobacter, 1996 Mar, 1(1), 62 - 4
Culture of Helicobacter pylori: effect of preimmersion of biopsy forceps in formalin; Yousfi MM et al.; BACKGROUND: Treatment of antibiotic-resistant Helicobacter pylori should be based on bacterial sensitivity testing that requires the ability to isolate the bacterium from gastric mucosal biopsies . The aim of this study was to determine whether the yield for detecting H . pylori infection by culture is reduced by immersion of biopsy forceps in formalin prior to obtaining the specimen . MATERIALS AND METHODS: Gastric antral mucosal biopsies (100 specimens) from 50 patients were obtained for culture of H . pylori . An antral biopsy was taken for culture, and with the same forceps a biopsy was taken for histological examination . The biopsy specimen was removed by shaking, whereas the forceps was immersed in 10% buffered formalin for the histological investigation . The forceps was then used without rinsing to obtain a second specimen for culture from an area adjacent to the first site . H . pylori status was determined by histological assessment with the Genta stain and a rapid urease test . RESULTS: Fifty patients with H . pylori infection documented by histological inquiry and positive rapid urease testing entered the study; 29 had duodenal ulcers, 5 had gastric ulcers, 1 had mucosal associated lymphoid tissue (MALT) lymphoma, and 15 were without ulcer disease . The results of culture both before and after immersion in formalin were identical . One patient had both cultures negative; the sensitivity of culture for detection of H . pylori infection was 98% (95% confidence interval = 93%-100%) . CONCLUSION: Preimmersion of biopsy forceps in formalin does not adversely affect the ability to culture H . pylori.

Helicobacter, 1996 Sep, 1(3), 159 - 64
Gastric juice polymerase chain reaction: an alternative to histology in the diagnosis of Helicobacter pylori infection; Basso D et al.; BACKGROUND: Infection from Helicobacter pylori plays a role in several gastroduodenal diseases . The recent availability of molecular techniques, particularly the polymerase chain reaction (PCR), allows us to detect small amounts of this bacterium . The aims of this study were to compare PCR and histological findings and to ascertain the clinical usefulness of H . pylori PCR identification in different biological samples . MATERIALS AND METHODS: We studied 94 consecutive patients . Saliva, gastric juice, and four antral and four body biopsies were obtained from each patient . H . pylori was evaluated histologically in two antral and two body biopsies (Giemsa or Warthin-Starry stain) . After extraction, DNA was submitted for PCR amplification using the two primers HPU1 and HPU2, which amplified a 411-bp product from the urease gene A . RESULTS: Forty-nine patients were H . pylori-positive at histological workup . The sensitivity of PCR was 92% for gastric juice, 73% for antral biopsies, 61% for body biopsies, and 13% for saliva . Of the 45 H . pylori-negative patients at histological assessment, 7 (16%) had positive findings on PCR, mainly when gastric juice was examined . CONCLUSIONS: These results indicate that PCR is as sensitive as histological assessment . We suggest that PCR H . pylori detection in gastric juice is a sensitive method for diagnosing this infection.

Biochem Biophys Res Commun, 1997 Nov 26, 240(3), 787 - 92
A pluripotent polyphenol oxidase from the melanogenic marine Alteromonas sp shares catalytic capabilities of tyrosinases and laccases; Sanchez-Amat A et al.; The recently characterized marine melanogenic bacterium MMB-1 contains a pluripotent polyphenol oxidase (PPO) which catalyzes the oxidation of a very wide range of substrates considered specific for tyrosinase or laccase . This range includes monophenols such as L-tyrosine, o-diphenols such as L-dopa, p-diphenols such as hydroquinone, o-aminophenols such as 3-hydroxyanthranilic acid, activated monophenols such as 2,6-dimethoxyphenol and syringaldazine, and chromophores such as ABTS . This is the first report of an enzyme that is able to catalyze the oxidation of compounds so far considered specific for tyrosinases (L-tyrosine) or laccase (syringaldazine), showing cresolase, catechol oxidase and laccase activities . Such PPO could be a very useful model to study the structural requirements, catalytic mechanisms and involvement of the copper sites existing in non-blue and blue copper-oxidases.

Eur J Biochem, 1997 Nov 1, 249(3), 820 - 5
Characterization, gene cloning and expression of isocitrate lyase involved in the assimilation of one-carbon compounds in Hyphomicrobium methylovorum GM2; Tanaka Y et al.; Isocitrate lyase of an obligate methylotrophic bacterium, Hyphomicrobium methylovorum GM2, was purified to homogeneity and characterized . The enzyme is a homotetramer of identical 62-kDa subunits . After the enzyme had been incubated at temperatures up to 25 degrees C for 30 min, no loss of activity was observed . The enzyme was stable in the pH range of 7.5-9.0 . Maximum activity was observed at pH 7.5 and around 45 degrees C . The Km value for Ds-isocitrate was 0.51 mM . The activity required Mg2+ and was inhibited by oxalate, succinate and glycolate . The gene encoding the isocitrate lyase and its flanking regions were isolated from H . methylovorum GM2 . Nucleotide sequencing of recombinant plasmids revealed that the isocitrate lyase gene codes for a 540-amino-acid protein . The amino acid sequence of the enzyme is similar to those of the enzymes from Escherichia coli (40% identity) and cucumber (37% identity) . The recombinant plasmid, which was constructed by ligation of the cloned gene and an expression vector, pKK223-3, was introduced into E . coli HB101 . The transformed E . coli cells expressed isocitrate lyase, which was indistinguishable from the purified H . methylovorum GM2 isocitrate lyase on analysis by SDS/PAGE.

Gastroenterology, 1997 Dec, 113(6 Suppl), S31 - 4; discussion S50
How does Helicobacter pylori cause mucosal damage? Direct mechanisms; Smoot DT; Helicobacter pylori-associated gastritis is characterized by an abundant inflammatory response and gastric epithelial cell injury . Adherence of H . pylori to gastric epithelial cells seems to be required for bacterial colonization of the gastric mucosa . Attachment of the bacterium to polarized gastric epithelial cells causes damage to microvilli and stimulates actin polymerization, which is associated with adherence pedestal formation . Studies suggest that H . pylori directly contributes to the injury of gastric epithelial cells by the elaboration of cytotoxic factors . The first toxin identified from H . pylori strains, known as vacuolating cytotoxin, induces vacuole formation in eukaryotic cells . Elaborated enzymes by H . pylori may also contribute directly to epithelial cell injury . Ammonia produced through urease activity may be toxic to gastric epithelial cells . H . pylori protease and lipase degrade gastric mucus and disrupt the phospholipid-rich layer at the apical epithelial cell surface, allowing for cell injury from back diffusion of gastric acid . This cell injury may lead to cell death, believed to result from induction of apoptosis . There are sufficient data to suggest that H . pylori, through direct pathogenic mechanisms, contributes significantly to the gastric mucosal injury associated with this infection, and may enhance the susceptibility of gastric epithelial cells to carcinogenic conversion.

Gastroenterology, 1997 Dec, 113(6 Suppl), S21 - 8
Helicobacter pylori factors associated with disease development; Mobley HL; Although certain factors appear to predispose the host to infection by Helicobacter pylori, clearly the bacterium possesses a well-defined battery of virulence factors that allow the organism to: (1) colonize the gastric mucosa (urease, flagella, adhesins, acid-inhibitory protein, iron acquisition proteins, and heat shock proteins); (2) evade host defense (shedding of surface proteins, catalase, superoxide dismutase, and poorly reactive lipopolysaccharide); and (3) damage host tissue (vacuolating cytotoxin, protease, CagA-related factors, inducers of cytokines, and chemotaxins) . Together these factors allow H . pylori to persist in the host, establishing a chronic infection . Although many of these virulence factors are produced by all strains of H . pylori, there are also well-defined pathogenicity islands (contiguous stretches of chromosomal DNA) present in some strains that encode additional proteins including CagA that potentiate virulence . Strains possessing these "virulence cassettes" are isolated more frequently from patients with the more serious clinical manifestations associated with duodenal ulcer than from patients with gastritis alone or nonulcer dyspepsia.

Infect Immun, 1997 Dec, 65(12), 5222 - 30
Identification and characterization of a K88- and CS31A-like operon of a rabbit enteropathogenic Escherichia coli strain which encodes fimbriae involved in the colonization of rabbit intestine; Adams LM et al.; Initiation of attaching-effacing lesions, which characterize infections with rabbit enteropathogenic Escherichia coli (REPEC), requires bacteria to adhere to the intestinal epithelium . This adherence is reflected in vitro by the affinity of these E . coli strains for various types of eukaryotic cells . TnphoA mutants of REPEC 83/39 (O15:H-) which had lost the ability to adhere to HEp-2 epithelial cells, guinea pig ileal brush borders, and mouse erythrocytes were generated . DNA sequencing of the region surrounding the inactivating transposon insertions within a 95-kb plasmid, designated pRAP for REPEC adherence plasmid, revealed extensive homology between that region and the structural genes of enterotoxigenic E . coli operons encoding the K88 and CS31A fimbrial adhesins and the genes for the afr2 adhesin from REPEC B10 (O103:H2) . Seven genes of the ral operon (for REPEC adherence locus), including three putative minor fimbrial subunit genes (ralC, ralF, and ralH), a major fimbrial subunit gene (ralG), a gene of unknown function (ralI), and genes for two fimbrial subunit chaperones (ralD and ralE), were sequenced . When inoculated perorally into weanling rabbits, a mutant with a TnphoA insertion in the ralE gene showed a 10-fold reduction in colonizing ability, with only 1 of 10 rabbits excreting bacteria compared to all 5 of those infected with the wild-type parent strain (P = 0.002) . The severity of the diarrheal illness caused by the mutant strain was also reduced . Western blotting of surface protein extracts of strain 83/39 with hyperimmune anti-83/39 antiserum, adsorbed with the ralE mutant, revealed a 32-kDa protein which was absent from protein extracts of two nonadherent mutants . The adsorbed antiserum also bound to the surface of strain 83/39 but not to nonadherent mutants, as detected by immunogold labeling . These results indicate that the ral operon of REPEC 83/39 contains genes necessary for the biosynthesis of fine fimbriae which are responsible for in vitro adherence of the bacteria and play a role in their colonization of, and hence virulence for, rabbits . The putative major fimbrial subunit is a protein with an observed molecular size of approximately 32 kDa which, when assembled, appears to form a capsule of fimbriae surrounding the bacterium similar to that described for CS31A.

J Bacteriol, 1997 Dec, 179(23), 7435 - 45
Genetic requirements and mutational specificity of the Escherichia coli SOS mutator activity; Fijalkowska IJ et al.; To better understand the mechanisms of SOS mutagenesis in the bacterium Escherichia coli, we have undertaken a genetic analysis of the SOS mutator activity . The SOS mutator activity results from constitutive expression of the SOS system in strains carrying a constitutively activated RecA protein (RecA730) . We show that the SOS mutator activity is not enhanced in strains containing deficiencies in the uvrABC nucleotide excision-repair system or the xth and nfo base excision-repair systems . Further, recA730-induced errors are shown to be corrected by the MutHLS-dependent mismatch-repair system as efficiently as the corresponding errors in the rec+ background . These results suggest that the SOS mutator activity does not reflect mutagenesis at so-called cryptic lesions but instead represents an amplification of normally occurring DNA polymerase errors . Analysis of the base-pair-substitution mutations induced by recA730 in a mismatch repair-deficient background shows that both transition and transversion errors are amplified, although the effect is much larger for transversions than for transitions . Analysis of the mutator effect in various dnaE strains, including dnaE antimutators, as well as in proofreading-deficient dnaQ (mutD) strains suggests that in recA730 strains, two types of replication errors occur in parallel: (i) normal replication errors that are subject to both exonucleolytic proofreading and dnaE antimutator effects and (ii) recA730-specific errors that are not susceptible to either proofreading or dnaE antimutator effects . The combined data are consistent with a model suggesting that in recA730 cells error-prone replication complexes are assembled at sites where DNA polymerization is temporarily stalled, most likely when a normal polymerase insertion error has created a poorly extendable terminal mismatch . The modified complex forces extension of the mismatch largely at the exclusion of proofreading and polymerase dissociation pathways . SOS mutagenesis targeted at replication-blocking DNA lesions likely proceeds in the same manner.

J Bacteriol, 1997 Dec, 179(23), 7264 - 73
Analysis of the fnrL gene and its function in Rhodobacter capsulatus; Zeilstra-Ryalls JH et al.; The fnr gene encodes a regulatory protein involved in the response to oxygen in a variety of bacterial genera . For example, it was previously shown that the anoxygenic, photosynthetic bacterium Rhodobacter sphaeroides requires the fnrL gene for growth under anaerobic, photosynthetic conditions . Additionally, the FnrL protein in R . sphaeroides is required for anaerobic growth in the dark with an alternative electron acceptor, but it is not essential for aerobic growth . In this study, the fnrL locus from Rhodobacter capsulatus was cloned and sequenced . Surprisingly, an R . capsulatus strain with the fnrL gene deleted grows like the wild type under either photosynthetic or aerobic conditions but does not grow anaerobically with alternative electron acceptors such as dimethyl sulfoxide (DMSO) or trimethylamine oxide . It is demonstrated that the c-type cytochrome induced upon anaerobic growth on DMSO is not synthesized in the R . capsulatus fnrL mutant . In contrast to wild-type strains, R . sphaeroides and R . capsulatus fnrL mutants do not synthesize the anaerobically, DMSO-induced reductase . Mechanisms that explain the basis for FnrL function in both organisms are discussed.

Scand J Immunol, 1997 Nov, 46(5), 500 - 5
Down-regulatory effect of Mycobacterium leprae cell wall lipids on phagocytosis, oxidative respiratory burst and tumour cell killing by mouse bone marrow derived macrophages; Moura AC et al.; The authors have previously demonstrated that lipids from Mycobacterium leprae cell walls inhibit macrophage functions and are endowed with anti-inflammatory properties in vivo . To investigate these observations further, the authors describe here the influence of dead M . leprae or of the lipids extracted from the cell wall of the mycobacterium, enclosed in liposomes, on the phagocytic, oxidative respiratory burst and tumouricidal ability of bone marrow derived macrophages in vitro . Dead M . leprae or its cell wall lipids abrogated the oxidative respiratory burst and phagocytic ability of mouse bone marrow derived macrophages . A dose-dependent inhibitory effect of the bacterial lipid extract on tumour cell killing by lipopolysaccharide (LPS)-activated bone marrow derived macrophages was demonstrated . However, when delipidated M . leprae was added to cultures of bone marrow derived macrophages, immune phagocytosis and superoxide production was up-regulated . Mycobacterium leprae or its lipids did not appear to be toxic to those cells assayed by the MTT (methyl thiazol tetrazolium) test . These data, added to our preceding observations, support the hypothesis that the down-regulatory activity of M . leprae wall lipids on macrophage function might be one of the evasive mechanisms of the bacterium to enable it to perpetuate itself in the host tissues.

Mol Gen Genet, 1997 Oct, 256(3), 211 - 22
Differential activation of two ACC oxidase gene promoters from melon during plant development and in response to pathogen attack; Lasserre E et al.; ACC (1-aminocyclopropane-1-carboxylate) oxidase genes are differentially expressed in melon during development and in response to various stresses . We investigated the molecular basis of their transcription by analyzing the 5' untranslated regions of the ACC oxidase genes CM-ACO1 and CM-ACO3 . In order to determine how their temporal and spatial expression patterns were established, we fused the promoter regions of CM-ACO1 (726 bp) and CM-ACO3 (2260 bp) to the beta-glucuronidase (GUS) reporter gene and examined their regulation in transgenic tobacco plants . The CM-ACO1 promoter was able to drive GUS expression in response to wounding, and to treatment with ethylene or copper sulfate . It was also rapidly induced (8-12 h postinoculation) in tobacco leaves inoculated with the hypersensitive response (HR)-inducing bacterium Ralstonia solanacearum . Expression was also observed during compatible interactions but was delayed . In contrast, the CM-ACO3 promoter was not expressed in response to infection, but was up-regulated during flower development . Both promoters were regulated during leaf senescence but in different patterns . The CM-ACO1-driven GUS activity increased sharply concomitantly with the onset of chlorophyll breakdown, while the CM-ACO3 promoter drove strong GUS expression in green, fully expanded leaves and this declined at the onset of senescence . This result is consistent with the expression patterns of these two genes in senescent melon leaves . These data suggest that the regulation of expression of CM-ACO1 is related preferentially to stress responses, whereas CM-ACO3 seems to be associated with developmental processes . The possible role of ethylene is discussed, particularly in the regulation of the CM-ACO1 gene in response to stress and during senescence.

J Wildl Dis, 1997 Oct, 33(4), 733 - 7
Humoral immune response of cottontail rabbits naturally infected with Francisella tularensis in southern Illinois; Shoemaker D et al.; Cottontail rabbits (Sylvilagus floridanus) usually are thought to succumb to infection with Francisella tularensis . Reports of a rabbit population from southern Illinois (USA) with a high prevalence of F . tularensis antibodies suggested that some cottontails survived infection with this typically fatal bacterium . Our goal was to examine the humoral response of cottontails from a study area in southern Illinois for which multiple serum samples existed . Multiple sera were collected from 79 cottontails from 1986 to 1990 and 63% gained, lost, or maintained ELISA titers of IgM and IgG isotype antibodies . The typical pattern of antibody response appeared to be IgM isotype antibodies first, followed by IgG isotype antibodies, with both generally increasing to high titers . Negative culture attempts of liver tissue from 51 cottontails with varying antibody responses suggested that chronic infection did not occur in rabbits that developed antibody . The significance of the cottontail antibody response in resolution or prevention of tularemia infection remains unclear.

Eur J Gastroenterol Hepatol, 1997 Oct, 9(10), 935 - 7
Helicobacter pylori and non-steroidal anti-inflammatory drugs: lone agents or partners in crime?
Roseveare CD, Colin-Jones DG.
A variety of mechanisms are responsible for the gastric and duodenal mucosal injury known to result from the consumption of non-steroidal anti-inflammatory drugs (NSAIDs) . Many of these mechanisms may be influenced by coexistent infection with Helicobacter pylori . However, evidence of increased risk from NSAIDs in patients with this bacterium is contradictory . While some authors have reported that symptoms, severity and prevalence of mucosal damage are higher in H . pylori-positive individuals taking NSAIDs than in those who are H . pylori negative, others have noted no significant difference . Reasons for this conflict may include the age of the subjects studied, duration of treatment, toxicity of the NSAID employed and pathogenicity factors related to different strains of H . pylori.

J Bacteriol, 1997 Nov, 179(22), 7161 - 4
Plasmid pRQ7 from the hyperthermophilic bacterium Thermotoga species strain RQ7 replicates by the rolling-circle mechanism; Yu JS et al.; The hyperthermophilic bacterium Thermotoga species strain RQ7 harbors an 846-bp plasmid, pRQ7, with a single open reading frame . Previously published analyses of the DNA sequence of pRQ7 suggested that it may replicate by a rolling-circle (RC) replication mechanism, and this report provides experimental evidence supporting this hypothesis . Single-stranded pRQ7 DNA accumulates in strain RQ7, as evidenced by the facts that this DNA bound to nitrocellulose membranes under nondenaturing conditions, was sensitive to S1 nuclease digestion, and hybridized to only one of two homologous DNA probes specific for each strand of the plasmid . The DNA encoding the open reading frame was cloned and expressed in Escherichia coli and gave a protein with a molecular mass of 26 kDa, similar to that deduced by sequence analysis . This protein bound to a fragment of pRQ7 that contains a putative double-stranded replication region in a magnesium-dependent reaction and made this fragment sensitive to S1 nuclease activity . It did not cause this same S1 nuclease sensitivity in the remainder of pRQ7 . This activity on pRQ7 DNA suggests that this protein plays a role in plasmid replication.

Eur J Biochem, 1997 Oct 15, 249(2), 630 - 6
Electron transfer towards the RCI-type photosystem in the green sulphur bacterium Chlorobium limicola forma thiosulphatophilum studied by time-resolved optical spectroscopy in vivo; Albouy D et al.; Flash-induced spectral changes in the wavelength region of the alpha-peaks of heme proteins and in the time domain from microseconds to seconds have been recorded on whole cells of the green sulphur bacterium Chlorobium limicola forma thiosulfatophilum . Extensive flash-excitation by trains of flashes resulted in oxidation of 7-8 c-type heme molecules/photosynthetic reaction centre . The complement of heme species was found to be spectrally heterogeneous allowing the study of electron transfer events induced by an isolated single-turnover flash . Under single-flash conditions, a c553 heme was seen to become oxidised with tau = 30 micros, concommitant with the reduction of the primary donor of the reaction centre . Subsequently, the alpha-peak of the photooxidised heme broadened and shifted towards longer wavelengths (tau = 70 micros) indicating equilibration of the positive charge over two differing heme species . In the time domain t > 1 ms, rereduction of c-type hemes was seen to be paralleled by a blue shift and further broadening of the alpha-peak . Concommitantly, b-type hemes were observed to first become reduced (within a few milliseconds), then over-oxidised (t > 200 ms) and eventually rereduced to their redox state prior to the flash . The results obtained are discussed with respect to the question of the identity of the immediate electron donor to the photosynthetic reaction centre and with respect to the involvement of a cytochrome bc complex in photo-induced electron transport of green sulphur bacteria.

Eur J Biochem, 1997 Oct 15, 249(2), 564 - 75
Molecular characterisation of the pifC gene encoding translation initiation factor 3, which is required for normal photosynthetic complex formation in Rhodobacter sphaeroides NCIB 8253; Babic S et al.; In order to determine whether translation initiation events play a selective role in regulating the expression of photosynthetic complexes in the photosynthetic bacterium Rhodobacter sphaeroides, we have undertaken an initial study to investigate the potential role of translation initiation factor IF3, which also behaves as a pleiotropic regulatory factor in some bacteria . Following the isolation and purification of a 24-kDa IF3-like protein (PifC) from R . sphaeroides, we used nested PCR to clone and characterise the encoding gene, pifC (photosynthesis-affecting initiation factor) . The 545-bp pifC encodes a protein exhibiting 60% identity (78.6% similarity) with the Escherichia coli IF3 (InfC) protein and, in common with all other IF3 genes identified to date, pifC possesses a rare initiation codon (AUA) . Furthermore, in common with IF3, PifC was shown here to perform a discriminatory function towards CUG start codons, confirming its role and function as an IF3 in R . sphaeroides . Insertion of a kanamycin resistance cassette into the 5' end of pifC resulted in a viable phenotype which exhibits growth rates similar to wild type but which possesses reduced bacteriochlorophyll and photosynthetic complexes in semi-aerobic cultures . It is shown here that the mutant is still able to produce a PifC protein but that it possesses reduced IF3 activity . This may account for the viable nature of the mutant strain, and may indicate that the effect of the mutation on photosynthesis can be more severe than shown in the present study . The mechanisms by which PifC may exert its selective regulatory effect on photosynthesis expression are discussed.

Gene, 1997 Oct 1, 198(1-2), 171 - 80
Codon usage and nucleotide composition in Coxiella burnetii; Lin Z et al.; Coxiella burnetii, the causative agent of Q fever, is an obligate intracellular bacterium . With the development of molecular biology techniques, there have been increasing efforts on gene cloning and other genetic analyses of this organism . In this report, we tabulate the codon usage (CU) and nucleotide (nt) co-occurrence in C . burnetii, based on available nt sequence data . The average G+C content of the C . burnetii genome is 42.4%, where the G+C content is 42.7% for the chromosome and 38.7% for the plasmid . In comparison to Escherichia coli, there is biased CU . Some codons are frequently used in C . burnetii, but rarely used in E . coli and vice versa . Plasmid genes prefer A or T at the first or third position of a codon . However, TAA remains the most used stop codon . In the AT-rich DNA of C . burnetii, A or T tend to occur together, forming A or T tracks.

Mol Microbiol, 1997 Sep, 25(5), 847 - 58
Assembly of the cell division protein FtsZ into ladder-like structures in the aerial hyphae of Streptomyces coelicolor; Schwedock J et al.; In the filamentous bacterium Streptomyces coelicolor, the cell division protein FtsZ is required for the conversion of multinucleoidal aerial hyphae into chains of uninucleoidal spores, although it is not essential for viability . Using immunofluorescence microscopy, we have shown that FtsZ assembles into long, regularly spaced, ladder-like arrays in developing aerial hyphae, with an average spacing of about 1.3 microm . Within individual hyphae, ladder formation was relatively synchronous and extended for distances over 100 microm . These ladders were present only transiently, decreasing in intensity as chromosomes separated into distinct nucleoids and disappearing upon the completion of septum formation . Evidence from the overall intensity of immunofluorescence staining suggested that ladder formation was regulated in part at the level of the accumulation and degradation of FtsZ within individual aerial hyphae . Finally, FtsZ ladder formation was under developmental control in that long arrays of FtsZ rings could not be detected in certain so-called white mutants (whiG, whiH and whiB), which are blocked in spore formation . The assembly of FtsZ into ladders represents the earliest known molecular manifestation of the process of spore formation, and its discovery provides insight into the role of whi genes in the conversion of aerial hyphae into chains of spores . We have also described a novel use of a cell wall-staining technique to visualize apical tip growth in vegetatively growing hyphae.

Vaccine, 1997 Nov, 15(16), 1779 - 83
Comparative efficacy of a Coxiella burnetii chloroform:methanol residue (CMR) vaccine and a licensed cellular vaccine (Q-Vax) in rodents challenged by aerosol; Waag DM et al.; Q fever is an acute and self-limited febrile illness caused by the obligate intracellular bacterium Coxiella burnetii . While phase I cellular Q fever vaccines are efficacious in humans, vaccination of immune individuals may result in sterile abscesses and granulomas . The chloroform:methanol residue vaccine (CMR) was developed as a safer alternative . The efficacy of a licensed phase I cellular vaccine (Q-Vax) was compared with that of CMR vaccine in A/J mice and Hartley guinea pigs challenged with virulent phase I C . burnetii by aerosol . Both vaccines were efficacious . The CMR vaccine dose required to protect 50% of mice (PD50) against lethal aerosol challenge (11 LD50) was one-third of the Q-Vax dose . However, the PD50 for CMR was four times the Q-Vax dose in guinea pigs challenged by aerosol (60 LD50) . It was concluded that CMR is an efficacious alternative to cellular Q fever vaccines for the prevention of Q fever.

Appl Environ Microbiol, 1997 Nov, 63(11), 4355 - 9
Glycogen biosynthesis via UDP-glucose in the ruminal bacterium Prevotella bryantii B1(4); Lou J et al.; Prevotella bryantii is an important amylolytic bacterium in the rumen that produces considerable amounts of glycogen when it is grown on maltose . Radiolabel studies indicated that glucose-1-phosphate was converted to UDP-glucose, and this latter intermediate served as the immediate precursor for glycogen synthesis . High levels of UDP-glucose pyrophosphorylase activities (> 1,492 nmol/min/mg of protein) were detected in cells grown on maltose, cellobiose, glucose, or sucrose, and activity was greatly stimulated (by approximately 60-fold) by the addition of fructose-1,6-bis phosphate (half-maximal activation concentration was 240 microM) . However, ADP-glucose pyrophosphorylase activity was not detected in any of the cultures . Glycogen synthase activity in maltose-grown cultures (48 nmol/min/mg of protein) was higher than that in cellobiose-, sucrose-, and glucose-grown cultures (< 26 nmol/min/mg of protein) . This is the first report of a bacterium that exclusively uses UDP-glucose to synthesize glycogen . The elucidation of this unique glycogen biosynthesis pathway provides information necessary to further investigate the role of bacterial glycogen accumulation in rumen metabolism.

J Wildl Dis, 1996 Oct, 32(4), 691 - 4
Isolation of Mycoplasma felis from a serval (Felis serval) with severe respiratory disease; Johnsrude JD et al.; We report cytologic observations and isolation of Mycoplasma felis in September 1992 from the lower respiratory tract of a 3-week-old captive serval (Felis serval) cub with pneumonia, in Florida (USA) . Septic, neutrophilic inflammation with a large, monomorphic population of unique, pleomorphic, intracellular and extracellular rods was diagnosed from a transtracheal wash . Mycoplasma felis was the only bacterium isolated in significant numbers from the transtracheal wash.

Microbiology, 1997 Oct, 143 ( Pt 10), 3085 - 99
Low-resolution sequencing of Rhodobacter sphaeroides 2.4.1T: chromosome II is a true chromosome; Choudhary M et al.; The photosynthetic bacterium Rhodobacter sphaeroides 2.4.1T has two chromosomes, CI (approximately 3.0 Mb) and CII (approximately 0.9 Mb) . In this study a low-redundancy sequencing strategy was adopted to analyse 23 out of 47 cosmids from an ordered CII library . The sum of the lengths of these 23 cosmid inserts was approximately 495 kb, which comprised approximately 417 kb of unique DNA . A total of 1145 sequencing runs was carried out, with each run generating 559 +/- 268 bases of sequence to give approximately 640 kb of total sequence . After editing, approximately 2.8% bases per run were estimated to be ambiguous . After the removal of vector and Escherichia coli sequences, the remaining approximately 565 kb of R . sphaeroides sequences were assembled, generating approximately 291 kb of unique sequences . BLASTX analysis of these unique sequences suggested that approximately 131 kb (45% of the unique sequence) had matches to either known genes, or database ORFs of hypothetical or unknown function (dORFs) . A total of 144 strong matches to the database was found; 101 of these matches represented genes encoding a wide variety of functions, e.g . amino acid biosynthesis, photosynthesis, nutrient transport, and various regulatory functions . Two rRNA operons (rrnB and rrnC) and five tRNAs were also identified . The remaining 160 kb of DNA sequence which did not yield database matches was then analysed using CODONPREFERENCE from the GCG package . This analysis suggested that 122 kb (42% of the total unique DNA sequence) could encode putative ORFs (pORFs), with the remaining 38 kb (13%) possibly representing non-coding intergenic DNA . From the data so far obtained, CII does not appear to be specialized for encoding any particular metabolic function, physiological state or growth condition . These data suggest that CII contains genes which are functionally as diverse as those found on any other bacterial chromosome and also contains sequences (pORFs), which may prove to be unique to this organism.

IARC Sci Publ, 1997, (138), 325 - 9
Infection with Helicobacter pylori and parasites, social class and cancer; Boffetta P; Three genera of parasites are known or suspected risk factors for cancer in humans: Schistosoma, Opisthorchis and Clonorchis . No adequate information is available on the determinants of infections related to social class . Infection with the bacterium Helicobacter pylori is an important cause of stomach cancer . Studies, in particular from the United Kingdom and the United States of America, strongly suggest that social class factors, especially those acting during childhood, are determinants of the infection, with odds ratios of seroprevalence of the order of 1.5-5 for lower social class as compared with higher social class . A conservative estimate of the contribution of social class, acting through an increased prevalence of H . pylori infection, to the burden of stomach cancer gives a figure of over 50,000 stomach cancers per year worldwide, or 8% of all stomach cancers . In countries with both high and low prevalence of infection with H . pylori, it is likely that a sizeable proportion of this difference is due to social-class-related risk factors of infection.

Infect Immun, 1997 Nov, 65(11), 4738 - 46
Utilization of similar mechanisms by Legionella pneumophila to parasitize two evolutionarily distant host cells, mammalian macrophages and protozoa; Gao LY et al.; The Legionnaires' disease bacterium, Legionella pneumophila, is an intracellular pathogen of humans that is amplified in the environment by intracellular multiplication within protozoa . Within both evolutionarily distant hosts, the bacterium multiplies in a rough endoplasmic reticulum-surrounded phagosome that is retarded from maturation through the endosomal-lysosomal degradation pathway . To gain an understanding of the mechanisms utilized by L . pneumophila to invade and replicate within two evolutionarily distant hosts, we isolated a collection of 89 mini-Tn10::kan insertion mutants that exhibited defects in cytotoxicity, intracellular survival, and replication within both U937 macrophage-like cells and Acanthamoeba polyphaga . Interestingly, the patterns of defects in intracellular survival and replication of the mutants within both host cells were highly similar, and thus we designated the defective loci in these mutants pmi (for protozoan and macrophage infectivity loci) . On the basis of their ability to attach to host cells and their growth kinetics during the intracellular infection, the mutants were grouped into five groups . Groups 1 and 2 included 41 mutants that were severely defective in intracellular survival and were completely or substantially killed during the first 4 h of infection in both host cells . Three members of group 1 were severely defective in attachment to both U937 cells and A . polyphaga, and another four mutants of group 1 exhibited severe defects in attachment to A . polyphaga but only a mild reduction in their attachment to U937 cells . Four members of groups 1 and 2 were serum sensitive . Intracellular replication of mutants of the other three groups was less defective than that of mutants of groups 1 and 2, and their growth kinetics within both host cells were similar . The mutants were tested for several other phenotypes in vitro, revealing that 14 of the pmi mutants were resistant to NaCl, 3 had insertions in dot or icm, 3 were aflagellar, 12 were highly intolerant to a hyperosmotic medium, and one failed to grow in a minimal medium . Our data indicated that similar mechanisms are utilized by L . pneumophila to replicate within two evolutionarily distant hosts . Although some mechanisms of attachment to both host cells were similar, other distinct mechanisms were utilized by L . pneumophila to attach to A . polyphaga . Our data supported the hypothesis that preadaptation of L . pneumophila to infection of protozoa may play a major role in its ability to replicate within mammalian cells and cause Legionnaires' disease.

Infect Immun, 1997 Nov, 65(11), 4531 - 8
A novel component different from endotoxin extracted from Prevotella intermedia ATCC 25611 activates lymphoid cells from C3H/HeJ mice and gingival fibroblasts from humans; Iki K et al.; A novel immunobiologically active fraction was prepared from a phenol-water extract of Prevotella intermedia ATCC 25611 by Sephadex G-100 column chromatography . The fraction consisted mainly of carbohydrate and protein and was devoid of fatty acid . The fraction showed high-molecular-weight bands (10,000 to 12,000) on deoxycholate polyacrylamide gel electrophoresis (DOC-PAGE) and was scarcely active in a Limulus test . We designated the fraction Prevotella glycoprotein (PGP) . The PGP fraction showed strong mitogenicity on splenocytes and cytokine-inducing activities on peritoneal macrophages from both C3H/HeJ and C3H/HeN mice, and it stimulated human gingival fibroblasts to produce cytokines . The activities of the PGP fraction were resistant to heat inactivation (100 degrees C for 1 h) and protease treatments and were scarcely inhibited by polymyxin B . In contrast, the purified lipopolysaccharide fraction (LPS-PCP) extracted from the same bacterium with a phenol-chloroform-petroleum ether mixture, which showed strong Limulus activity and a single low-molecular-weight band (approximately 3,000) on DOC-PAGE, lacked the activities on splenocytes and macrophages from C3H/HeJ mice and human gingival fibroblasts . The activities of the LPS-PCP fraction on cells from C3H/HeN mice were completely inhibited by polymyxin B . The LPS extracted from the same bacterium with hot phenol-water (LPS-PW) exhibited the properties of both the PGP fraction and the LPS-PCP fraction . These findings suggest that the unique bioactivities of the LPS-PW fraction of oral black-pigmented bacteria reported to date, which differed from those of the classical endotoxin, were derived from the PGP fraction and not from the LPS itself.

Appl Microbiol Biotechnol, 1997 Sep, 48(3), 367 - 72
Extracellular reduction of selenite by a novel marine photosynthetic bacterium; Yamada A et al.; A novel purple nonsulfur bacterium strain NKPB030619, which has resistance to over 5 mM selenite, was isolated from a marine environment . An initial concentration of 1.1 mM selenite, added to the medium, was decreased to under 0.05 mM within 5 days . The color of the cell suspension turned red within 2 days . The red coloration gradually decreased and black precipitates appeared during 2 weeks of cultivation . Under these conditions, two main types of deposit were formed extracellularly . These deposits were thought to contain red amorphous selenium and black vitreous selenium . The selenite reduction to elemental selenium in this bacterium was induced by the introduction of light and L-malic acid under anaerobic conditions . These results suggest that selenite reduction is coupled with photosynthesis and L-malic acid can serve as the indirect electron donor for its reduction . Phylogenetic analysis based on the 16S rDNA sequence showed that NKPB0360619 belongs to the alpha subdivision of Proteobacteria and is classified into the Rhodobacter species . The highest similarity of 86.2% was observed with R . sphaeroides.

Lett Appl Microbiol, 1997 Sep, 25(3), 162 - 8
Isolation and characterization of a transposon mutant of Shewanella putrefaciens MR-1 deficient in fumarate reductase; Myers CR et al.; A transposon mutant, designated CMTn-3, of Shewanella putrefaciens MR-1 that was deficient in fumarate reduction was isolated and characterized . In contrast to the wild-type, CMTn-3 could not grow anaerobically with fumarate as the electron acceptor, and it lacked benzyl viologen-linked fumarate reductase activity . Consistent with this, CMTn-3 lacked a 65 kDa c-type cytochrome, which is the same size as the fumarate reductase enzyme . CMTn-3 retained the wild-type ability to use nitrate, iron(III), manganese(IV) and trimethylamine N-oxide (TMAO) as terminal electron acceptors . The results indicate that the loss of the fumarate reductase enzyme does not affect other anaerobic electron transport systems in this bacterium.

Eur J Biochem, 1997 Sep 1, 248(2), 323 - 8
Multiheme cytochromes from the sulfur-reducing bacterium Desulfuromonas acetoxidans; Pereira IA et al.; Two new multiheme cytochromes were isolated from the anaerobic sulfur reducing bacterium Desulfuromonas acetoxidans . They have monomeric molecular masses of 50 and 65 kDa and contain six and eight hemes, respectively . Visible and EPR spectroscopies, in the as-isolated (oxidised) cytochromes, show the presence of only low-spin hemes in the 50-kDa cytochrome, and of high-spin and low-spin hemes in the 65-kDa cytochrome . The EPR spectra of the native 65-kDa cytochrome indicate multiple heme-heme interactions, including integer-spin systems as judged by parallel-mode EPR . The 50-kDa cytochrome has a complex redox pattern, as shown by EPR redox titrations, and contains one heme with unusual characteristics . Both cytochromes cover an extremely wide range of reduction potentials, which go from +100 mV to -375 mV for the 50-kDa cytochrome, and +185 mV to -235 mV for the 65-kDa cytochrome . The two cytochromes were tested for hydroxylamine oxidoreductase activity and polysulfide reductase activity, but neither displayed any activity . In contrast, it was found for the first time that the previously characterised cytochrome c551.5, from the same bacterium is very active in the reduction of polysulfide, which suggests that it acts as a terminal reductase in D . acetoxidans.

Biochem Biophys Res Commun, 1997 Oct 20, 239(2), 626 - 32
Apoptosis in gastric epithelial cells is induced by Helicobacter pylori and accompanied by increased expression of BAK; Chen G et al.; Carriage of the bacterium H . pylori in the human stomach is associated with evidence of increased epithelial cell apoptosis . This may be of significance in the etiology of gastritis, peptic ulcers, and neoplasia . The ability of H . pylori to directly induce epithelial apoptosis was examined in vitro by fluorescence and electron microscopy, flow cytometry, and DNA fragmentation ELISA . The induction of apoptosis by H . pylori was time and concentration-dependent and inhibited by preventing direct bacterial-epithelial cell contact . Apoptosis was accompanied by increased expression of Bak, with little change in expression of other Bcl-2 family proteins . The expression of Bak was also increased in gastric biopsies from patients colonized by H . pylori . Thus, H . pylori induces gastric epithelial cell apoptosis, by a Bak-dependent pathway .

Am J Pathol, 1997 Oct, 151(4), 933 - 41
Increased oxidative DNA damage and hepatocyte overexpression of specific cytochrome P450 isoforms in hepatitis of mice infected with Helicobacter hepaticus; Sipowicz MA et al.; A recently discovered bacterium, Helicobacter hepaticus, infects the intrahepatic bile canaliculi of mice, causing a severe chronic hepatitis culminating in liver cancer . Thus, it affords an animal model for study of bacteria-associated tumorigenesis including H . pylori-related gastric cancer . Reactive oxygen species are often postulated to contribute to this process . We now report that hepatitis of male mice infected with H . hepaticus show significant increases in the oxidatively damaged DNA deoxynucleoside 8-hydroxydeoxyguanosine, with the degree of damage increasing with progression of the disease . Perfusion of infected livers with nitro blue tetrazolium revealed that superoxide was produced in the cytoplasm of hepatocytes, especially in association with plasmacytic infiltrates near portal triads . Contrary to expectations, Kupffer cells, macrophages, and neutrophils were rarely involved . However, levels of cytochrome P450 (CYP) isoforms 1A2 and 2A5 in hepatocytes appeared to be greatly increased, as indicated by the number of cells positive in immunohistochemistry and the intensity of staining in many cells, concomitant with severe hepatitis . The CYP2A5 immunohistochemical staining co-localized with formazan deposits resulting from nitro blue tetrazolium reduction and occurred in nuclei as well as cytoplasm . These findings suggest that CYP2A5 contributes to the superoxide production and 8-hydroxydeoxyguanosine formation, although reactive oxygen species from an unknown source in the hepatocytes leading to CYP2A5 induction or coincidental occurrence of these events are also possibilities . Three glutathione S-transferase isoforms, mGSTP1-1 (pi), mGSTA1-1 (YaYa), and mGSTA4-4, also showed striking increases evidencing major oxidative stress in these livers.

Appl Environ Microbiol, 1997 Oct, 63(10), 4135 - 8
Detection of Mycobacterium ulcerans in environmental samples during an outbreak of ulcerative disease; Ross BC et al.; Mycobacterium ulcerans is an environmental bacterium which causes chronic skin ulcers . Despite significant epidemiological evidence to suggest that water is the source of infection, the organism has never been identified in the environment . Environmental water samples were collected from a small town in which an outbreak of 29 cases had occurred in a 3-year period . These were examined by mycobacterial culture and PCR amplification . Similar to previous studies, M . ulcerans was not cultured from the water samples . However, five samples were positive for M . ulcerans by PCR . These samples were collected from a swamp and a golf course irrigation system within the outbreak area . This is the first time that M . ulcerans has been demonstrated to be present in the environment and supports the postulated epidemiology of disease due to this organism.

Berl Munch Tierarztl Wochenschr, 1997 Jul-Aug, 110(7-8), 267 - 71
{Pilot study on the prevalence of Ornithobacterium rhinotracheale infections in food chickens in northwest Germany}; Ryll M et al.; Samples of sera from broiler chicken and broiler chicken breeder flocks of North Western Germany were examined for presence of antibodies against Ornithobacterium (O.) rhinotracheale with a self-developed ELISA system . A total of 3600 samples of sera from 190 broiler flocks in 70 different farms were tested . In 332 samples of sera (9.4%) from 25 flocks (13.2%) in 22 farms specific antibodies were detected . Additionally, serum samples from the breeder hens, taken between the 9, and 61, weeks of age were tested . In 28 from 29 flocks (96.6%) specific antibodies were detected using this ELISA-system . First detectable antibodies were found between the 14, and 36, week of age in the parent breeder flocks . These results show that infections with O . rhinotracheale are widely distributed in the parent-breeder chicken broiler flocks . The significance of this bacterium as the cause of respiratory diseases in chickens are discussed.

Eur J Epidemiol, 1997 Sep, 13(6), 729 - 31
Copy number of the 16S rRNA gene in Coxiella burnetii; Afseth G et al.; Coxiella burnetii is an obligate intracellular bacterium with a doubling time of 5-7 hours . Chromosomal DNA from C . burnetii was digested with various restriction enzymes previously determined to not cut within the genomic 16S rRNA gene, or with a combination of these noncutting enzymes in conjunction with AflIII, a restriction enzyme that cuts twice within the 16S rRNA gene . Restriction fragments were resolved electrophoretically and probed with a radiolabeled DNA fragment containing the 3' AflIII portion of the C . burnetii 16S rRNA gene . Only a single DNA fragment in these digests hybridized to the probe, indicating that there is a single genomic copy of the 16S rRNA gene in C . burnetii and thus only a single copy of the rRNA operon.

Mol Gen Genet, 1997 Aug, 255(6), 637 - 42
Genes required for assembly and function of the protein synthetic system in Chlamydia trachomatis are expressed early in elementary to reticulate body transformation; Gerard HC et al.; Following binding and internalization into the host cell cytoplasm, elementary bodies (EB) of the obligate intracellular bacterium Chlamydia trachomatis undergo a developmental process resulting in production of reticulate bodies (RB), the vegetative growth form of the organism . EB are metabolically inactive, but EB to RB transformation requires bacterial protein synthesis . Using HeLa cells infected with EB of C . trachomatis serovar C, we examined the time of first appearance of transcripts from several genes whose products are required for assembly and function of the chlamydial protein synthetic system . We monitored appearance of chlamydial RNAs using reverse transcription-polymerase chain reaction assays targeting primary transcripts from the bacterial rRNA operons, and mRNAs encoding the glycyl tRNA synthetase and the ribosomal proteins S5 and L5 . Transcripts from the proximal rRNA promoters, and those from the r-protein and tRNA synthetase genes, are detectable as early as 4 h after EB-host binding; transcripts from distal rRNA promoters do not appear until 6 h post-infection . Thus, expression of bacterial genes whose products are required for protein synthesis begins earlier in chlamydial EB to RB development than previously thought.

EXS, 1997, 83, 135 - 54
Phenotypic and evolutionary adaptation of a model bacterial system to stressful thermal environments; Bennett AF et al.; We studied both phenotypic and evolutionary adaptation to various thermal environments using the bacterium Escherichia coli as an experimental model system . We determined that 42 degrees C was stressful to a bacterial clone adapted to 37 degrees C, based on reductions in both absolute and competitive fitness, as well as induction of a heat stress response . This clone was also used to found replicated populations that were propagated for thousands of generations under several different thermal regimes, including 42 degrees C . Evolutionary adaptation of the populations to 42 degrees C resulted in an increase in both absolute and relative fitness at that temperature, measured respectively as an increase in the number of descendants (and their biovolume) and in competitive ability relative to the ancestral clone . The replicated experimental lineages achieved their evolutionary improvement by several distinct pathways, which produced differential preadaptation to a non-stressful nutrient environment . Adaptation to this stressful temperature entailed neither a change in the ancestral thermal niche nor any pronounced trade-offs in fitness within the thermal niche, contrary to a priori predictions . This study system was several important advantages for evaluating hypotheses concerning the effects of stress on phenotypic and evolutionary adaptation, including the ability to obtain lineages that have evolved in controlled and defined environments, to make direct measurements of fitness and to quantify the degree of stress imposed by different environments.

Biosci Biotechnol Biochem, 1997 Sep, 61(9), 1577 - 9
Orientation of photosynthetic reaction center reconstituted in neutral and charged liposomes; Hara M et al.; The photosynthetic reaction center from the photosynthetic bacterium Rhodobacter sphaeroides was reconstituted into neutral, positively charged, or negatively charged liposomes . About 70% of photosynthetic reaction centers were reconstituted in the proteoliposomes exposing their H-subunit outside with positively charged lipids while only 30-40% of them were in the same topological orientation with neutral or negatively charged lipids.

J Bacteriol, 1997 Oct, 179(19), 6053 - 60
Purification and molecular characterization of the H2 uptake membrane-bound NiFe-hydrogenase from the carboxidotrophic bacterium Oligotropha carboxidovorans; Santiago B et al.; The membrane-bound hydrogenase of Oligotropha carboxidovorans was solubilized with n-dodecyl-beta-D-maltoside and purified 28-fold with a yield of 29% and a specific activity of 173 to 178 micromol of H2 x min(-1) x mg(-1) . It is the first hydrogenase studied in a carboxidotrophic bacterium . The enzyme acts on artificial electron-accepting dyes, such as methylene blue, but is ineffective with pyridine nucleotides or other soluble physiological electron acceptors . Hydrogenase of O . carboxidovorans belongs to class I of hydrogenases and is a heterodimeric 101,692-Da NiFe-protein composed of the polypeptides HoxL and HoxS . Molecular cloning data revealed, that HoxL comprises 604 amino acid residues and has a molecular mass of 67,163 Da . Pre-HoxS comprises 360 amino acid residues and is synthesized as a precursor protein which is cleaved after alanine at position 45, thus producing a mature HoxS of 33,767 Da . The leader sequence corresponds to the signal peptide of small subunits of hydrogenases . The hydropathy plots of HoxL and HoxS were indicative for the absence of transmembranous helices . HoxZ has four transmembranous helices and is considered the potential membrane anchor of hydrogenase in O . carboxidovorans . Hydrogenase genes show the transcriptional order 5' hoxV --> hoxS --> hoxL --> hoxZ 3' . The hox gene cluster as well as the clustered CO dehydrogenase (cox) and Calvin cycle (cbb) genes are arranged within a 30-kb DNA segment of the 128-kb megaplasmid pHCG3 of O . carboxidovorans.

J Physiol Pharmacol, 1997 Sep, 48(3), 345 - 51
Suppression of Helicobacter pylori protease activity towards growth factors by sulglycotide; Piotrowski J et al.; Infection with H . pylori is now recognized as a major factor in the pathogenesis of gastric disease . Here, we examined the susceptibility of epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), transforming growth factor-beta (TGF beta) and platelet derived growth factor (PDGF) to degradation by H . pylori protease, and assessed the effect of a cytoprotective agent, sulglycotide, on this process . The 125I-labeled EGF, bFGF, TGF beta and PDGF were incubatet with H . pylori protease, obtained from the filtrates of saline washes of the bacterium culture, in the presence of 0-100 micrograms sulglycotide . The results showed that, under the assay conditions, H . pylori protease caused only 5% degradation of EGF and 7% degradation of bFGF . However, the protease evoked a 61.7% degradation of PDGF and a 62.3% degradation of TGF beta . Introduction of sulglycotide to the reaction assay system caused a dose-dependent inhibition in PDGF and TGF beta proteolysis by the H . pylori enzyme . The maximal inhibitory effect was obtained with sulglycotide at 100 micrograms/ml, at which dose an 84.4% decrease in PDGF and 88.3% decrease in TGF beta degradation was achieved . The results provide a strong evidence for the effectiveness of sulglycotide in the protection of gastric mucosal growth factors against degradation by H . pylori.

Int J Syst Bacteriol, 1997 Oct, 47(4), 1140 - 4
In vitro culture and phylogenetic analysis of "Candidatus Arsenophonus triatominarum," an intracellular bacterium from the triatomine bug, Triatoma infestans; Hypsa V et al.; An intracellular symbiotic bacterium was isolated from the hemolymph of Triatoma infestans and cultured in an Aedes albopictus cell line . 16S ribosomal DNA sequence analysis revealed that the bacterium was a member of the gamma-3 subgroup of the class Proteobacteria, having 96.2% sequence identity with the most closely related bacterium, Arsenophonus nasoniae, the causative agent of the son-killer trait in the parasitoid wasp Nasonia vitripennis . These bacteria share morphological features and a common tissue distribution and transmission mode . The A . nasoniae-T . infestans symbiont branch represents a lineage of insect symbionts which may be capable of horizontal transmission between phylogenetically distant host insects . We propose that the intracellular symbiont from T . infestans be classified as "Candidatus Arsenophonus triatominarum." The bacterium found in the hemocytes of T . infestans is designated the type strain of this species.

Int J Syst Bacteriol, 1997 Oct, 47(4), 933 - 8
Nocardioides pyridinolyticus sp . nov., a pyridine-degrading bacterium isolated from the oxic zone of an oil shale column; Yoon JH et al.; A bacterial strain which is able to degrade pyridine was previously isolated from the oxic zone of an oil shale column and described as Pimelobacter sp . strain OS4T . However, Pimelobacter species have been transferred to the genera Nocardioides and Terrabacter . Strain OS4T was identified as a member of the genus Nocardioides on the basis of chemotaxonomic analysis and phylogenetic inference based on 16S ribosomal DNA (rDNA) sequence analysis . The G+C content of strain OS4T is 72.5 mol% . The cell wall peptidoglycan contains LL-diaminopimelic acid as the diamino acid . The predominant menaquinone is MK-8(H4) . The cellular fatty acid profile of strain OS4T is similar to that of the genus Nocardioides . The 16S rDNA similarity of strain OS4T with previously described Nocardioides species is 94.5% +/- 0.7%, and a phylogenetic tree based on 16S rDNA sequences revealed a distinct lineage for strain OS4T within the evolutionary radiation enclosed by the genus Nocardioides . Therefore, on the basis of our data, we propose that strain OS4T should be placed in the genus Nocardioides as a member of a new species, Nocardioides pyridinolyticus . The type strain of the new species is strain OS4 (= KCTC 0074BP).

Protein Sci, 1997 Oct, 6(10), 2159 - 65
Dissection of the gene of the bifunctional PGK-TIM fusion protein from the hyperthermophilic bacterium Thermotoga maritima: design and characterization of the separate triosephosphate isomerase; Beaucamp N et al.; Triosephosphate isomerase (TIM), from the hyperthermophilic bacterium Thermotoga maritima, has been shown to be covalently linked to phosphoglycerate kinase (PGK) forming a bifunctional fusion protein with TIM as the C-terminal portion of the subunits of the tetrameric protein (Schurig et al., EMBO J 14:442-451, 1995) . To study the effect of the anomalous state of association on the structure, stability, and function of Thermotoga TIM, the isolated enzyme was cloned and expressed in Escherichia coli, and compared with its wild-type structure in the PGK-TIM fusion protein . After introducing a start codon at the beginning of the tpi open reading frame, the gene was expressed in E.c.BL21(DE3)/ pNBTIM . The nucleotide sequence was confirmed and the protein purified as a functional dimer of 56.5 kDa molecular mass . Spectral analysis, using absorption, fluorescence emission, near- and far-UV circular dichroism spectroscopy were used to compare the separated Thermotoga enzyme with its homologs from mesophiles . The catalytic properties of the enzyme at approximately 80 degrees C are similar to those of its mesophilic counterparts at their respective physiological temperatures, in accordance with the idea that under in vivo conditions enzymes occupy corresponding states . As taken from chaotropic and thermal denaturation transitions, the separated enzyme exhibits high intrinsic stability, with a half-concentration of guanidinium-chloride at 3.8 M, and a denaturation half-time at 80 degrees C of 2 h . Comparing the properties of the TIM portion of the PGK-TIM fusion protein with those of the isolated recombinant TIM, it is found that the fusion of the two enzymes not only enhances the intrinsic stability of TIM but also its catalytic efficiency.

Biophys J, 1997 Oct, 73(4), 2090 - 6
Pump-probe anisotropies of Fenna-Matthews-Olson protein trimers from Chlorobium tepidum: a diagnostic for exciton localization?
Savikhin S, Buck DR, Struve WS.
Exciton calculations on symmetric and asymmetric Fenna-Matthews-Olson (FMO) trimers, combined with absorption difference anisotropy measurements on FMO trimers from the green bacterium Chlorobium tepidum, suggest that real samples exhibit sufficient diagonal energy disorder so that their laser-excited exciton states are noticeably localized . Our observed anisotropies are clearly inconsistent with 21-pigment exciton simulations based on a threefold-symmetric FMO protein . They are more consistent with a 7-pigment model that assumes that the laser-prepared states are localized within a subunit of the trimer . Differential diagonal energy shifts of 50 cm(-1) between symmetry-related pigments in different subunits are large enough to cause sharp localization in the stationary states; these shifts are commensurate with the approximately 95 cm(-1) inhomogeneous linewidth of the lowest exciton levels . Experimental anisotropies (and by implication steady-state linear and circular dichroism) likely arise from statistical averaging over states with widely contrasting values of these observables, in consequence of their sensitivity to diagonal energy disorder.

J Bacteriol, 1997 Oct, 179(20), 6448 - 52
Transcriptional characterization of the Rickettsia prowazekii major macromolecular synthesis operon; Shaw EI et al.; Recent studies have demonstrated that Rickettsia prowazekii can regulate transcription of selected genes at the level of initiation . However, little information concerning the existence of operons and coordinate gene regulation in this obligate intracellular parasitic bacterium is available . To address these issues, we have focused on the rpoD gene linkage group (greA-open reading frame 23 {ORF23}-dnaG-rpoD), which includes the rickettsial analog (ORF23-dnaG-rpoD) of the major macromolecular synthesis operon (MMSO) . The rickettsial MMSO consists of an ORF coding for a protein of unknown function the structural genes for DNA primase (dnaG) and the major sigma factor of RNA polymerase (rpoD) . RNase protection assays (RPA) were used to determine if these genes are organized into an operon controlled by multiple promoters and the quantities of transcripts produced by these genes relative to each other . RPA with a probe spanning the 270-base greA-ORF23 intervening region identified a putative transcriptional promoter within the intervening sequence . Multiple RPA probes spanning the next 4,041 bases of the linkage group demonstrated the presence of a continuous transcript and thus the existence of an operon . A probe spanning the dnaG-rpoD region revealed that two additional mRNA fragments were also protected, which enabled us to identify additional putative promoters for rpoD within dnaG . Primer extension determined that the 5' ends of the three transcripts consist separately of adenine (located 227 bases upstream of ORF23) and uracil and adenine (located 336 and 250 bases upstream of rpoD, respectively) . Quantitation of transcripts produced by the three ORFs determined the relative amounts of transcripts (ORF23 to dnaG to rpoD) to be 1:2.7:5.1.

J Biotechnol, 1997 Oct 2, 58(1), 59 - 66
Overproduction and purification of an agarase of bacterial origin; Parro V et al.; The agarase gene from Streptomyces coelicolor has been cloned in the non-producer bacterium Streptomyces lividans under the control of its own set of promoters and under the control of a heterologous promoter that is functional only during exponential growth . The best level of overproduction was obtained when the strain containing the natural gene was cultivated in fed batch with mannitol as carbon source . The protein, with a relative molecular mass of 32 kDa, has been purified following an affinity purification method . Contaminating activities seem to be absent from the purified enzyme preparation that can be used to purify DNA from agarose gels.

J Exp Med, 1997 Oct 20, 186(8), 1241 - 6
Mycobacterium tuberculosis chaperonin 10 stimulates bone resorption: a potential contributory factor in Pott's disease; Meghji S et al.; Pott's disease (spinal tuberculosis), a condition characterized by massive resorption of the spinal vertebrae, is one of the most striking pathologies resulting from local infection with Mycobacterium tuberculosis (Mt; Boachie-Adjei, O., and R.G . Squillante . 1996 . Orthop . Clin . North Am . 27:95-103) . The pathogenesis of Pott's disease is not established . Here we report for the first time that a protein, identified by a monoclonal antibody to be the Mt heat shock protein (Baird, P.N., L.M . Hall, and A.R.M . Coates . 1989 . J . Gen . Microbiol . 135:931-939) chaperonin (cpn) 10, is responsible for the osteolytic activity of this bacterium . Recombinant Mt cpn10 is a potent stimulator of bone resorption in bone explant cultures and induces osteoclast recruitment, while inhibiting the proliferation of an osteoblast bone-forming cell line . Furthermore, we have found that synthetic peptides corresponding to sequences within the flexible loop and sequence 65-70 of Mt cpn10 may comprise a single conformational unit which encompasses its potent bone-resorbing activity . Our findings suggest that Mt cpn10 may be a valuable pharmacological target for the clinical therapy of vertebral tuberculosis and possibly other bone diseases.

Proc Natl Acad Sci U S A, 1997 Oct 14, 94(21), 11439 - 44
The role of Wolbachia bacteria in reproductive incompatibilities and hybrid zones of Diabrotica beetles and Gryllus crickets; Giordano R et al.; A rickettsial bacterium in the genus Wolbachia is the cause of a unidirectional reproductive incompatibility observed between two major beetle pests of maize, the western corn rootworm, Diabrotica virgifera virgifera, and the Mexican corn rootworm, D . v . zeae . These subspecies are allopatric except for two known regions of sympatry in Texas and Mexico . We demonstrate that populations of D . v . virgifera, with the exception of two populations in southern Arizona, are infected with a strain of Wolbachia . Populations of D . v . zeae are not infected . Treatment of D . v . virgifera with tetracycline eliminated the Wolbachia and removed the reproductive incompatibility . Similar patterns of reproductive incompatibility exist among taxa of the cricket genus Gryllus . Gryllus assimilis, G . integer, G . ovisopis, G . pennsylvanicus, and G . rubens are infected with Wolbachia whereas G . firmus is usually not . Populations of G . rubens and G . ovisopis carry the same Wolbachia strain, which is distinct from that of G . integer . G . pennsylvanicus is infected with two Wolbachia strains, that found in G . rubens and one unique to G . pennsylvanicus . Moreover, a proportion of G . pennsylvanicus individuals harbors both strains . Wolbachia may have influenced speciation in some members of the genus Gryllus by affecting the degree of hybridization between species . Given that Wolbachia infections are relatively common in insects, it is likely that other insect hybrid zones may be influenced by infections with Wolbachia.

Proc Natl Acad Sci U S A, 1997 Oct 14, 94(21), 11216 - 20
CooA, a CO-sensing transcription factor from Rhodospirillum rubrum, is a CO-binding heme protein; Shelver D et al.; Biological sensing of small molecules such as NO, O2, and CO is an important area of research; however, little is know about how CO is sensed biologically . The photosynthetic bacterium Rhodospirillum rubrum responds to CO by activating transcription of two operons that encode a CO-oxidizing system . A protein, CooA, has been identified as necessary for this response . CooA is a member of a family of transcriptional regulators similar to the cAMP receptor protein and fumavate nitrate reduction from Escherichia coli . In this study we report the purification of wild-type CooA from its native organism, R . rubrum, to greater than 95% purity . The purified protein is active in sequence-specific DNA binding in the presence of CO, but not in the absence of CO . Gel filtration experiments reveal the protein to be a dimer in the absence of CO . Purified CooA contains 1.6 mol heme per mol of dimer . Upon interacting with CO, the electronic spectrum of CooA is perturbed, indicating the direct binding of CO to the heme of CooA . A hypothesis for the mechanism of the protein's response to CO is proposed.

J Clin Microbiol, 1997 Oct, 35(10), 2606 - 11
Comparison of major antigenic proteins of six strains of the human granulocytic ehrlichiosis agent by Western immunoblot analysis; Zhi N et al.; The etiologic agent of human granulocytic ehrlichiosis (HGE) is an obligate intracellular bacterium . In 1996, blood specimens from 53 patients suspected of having HGE were examined by indirect fluorescent antibody (IFA) testing with the HGE agent no . 13 isolate as the antigen, by nested PCR, and by culture . All patients resided in Westchester County, N.Y . Twelve patient specimens were positive for IFA (titer > or = 1:40) . Seven of these were also positive by PCR . Of the seven specimens positive by both IFA testing and PCR, the HGE agent was isolated from four (no . 2, 3, 6, and 11) and continuously cultured in HL-60 cells . These were confirmed as the HGE agent by sequencing of 16S rDNA . Both purified whole-cell organisms and the outer membrane fractions of the new isolates were compared with no . 13 isolate and a tick (USG) isolate by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western immunoblot analysis . No . 11 and 13 isolates had identical SDS-PAGE patterns with respect to 49- and 47-kDa proteins . No . 3 and USG isolates lacked the 47-kDa protein, and no . 6 isolate lacked the 49-kDa protein . Both 49- and 47-kDa bands were absent in no . 2 isolate . Western blot results with seven different sera, including five convalescent-phase sera from these patients, one dog anti-USG isolate, and one horse anti-BDS isolate, showed that all major antigens in six isolates were recognized by all sera . However, the molecular sizes and the numbers of major antigens recognized varied among the six isolates . Overall, HGE agent no . 3, 6, 11, and 13, and USG isolates had similar patterns, with 1 or 2 major antigens with molecular masses of around 49 and 47 kDa . No . 2 isolate was quite distinct in having a major antigen of 43 kDa . This indicates that although these antigenic epitopes are all cross-reactive among strains, the HGE agent has a strain pleomorphism in its major antigenic proteins . The major antigen profiles of the outer membrane protein fractions and of whole organisms of six HGE agent isolates were similar, suggesting that 49- and 47-kDa major antigens are the outer membrane proteins of the HGE agent.

Proc Natl Acad Sci U S A, 1997 Sep 30, 94(20), 10606 - 11
Both DNA gyrase and reverse gyrase are present in the hyperthermophilic bacterium Thermotoga maritima; Guipaud O et al.; Like all hyperthermophiles yet tested, the bacterium Thermotoga maritima contains a reverse gyrase . Here we show that it contains also a DNA gyrase . The genes top2A and top2B encoding the two subunits of a DNA gyrase-like enzyme have been cloned and sequenced . The Top2A (type II DNA topoisomerase A protein) is more similar to GyrA (DNA gyrase A protein) than to ParC {topoisomerase IV (Topo IV) C protein} . The difference is especially striking at the C-terminal domain, which differentiates DNA gyrases from Topo IV . DNA gyrase activity was detected in T . maritima and purified to homogeneity using a novobiocin-Sepharose column . This hyperhermophilic DNA gyrase has an optimal activity around 82-86 degrees C . In contrast to plasmids from hyperthermophilic archaea, which are from relaxed to positively supercoiled, we found that the plasmid pRQ7 from Thermotoga sp . RQ7 is negatively supercoiled . pRQ7 became positively supercoiled after addition of novobiocin to cell cultures, indicating that its negative supercoiling is due to the DNA gyrase of the host strain . The findings concerning DNA gyrase and negative supercoiling in Thermotogales put into question the role of reverse gyrase in hyperthermophiles.

Mol Microbiol, 1997 Aug, 25(4), 671 - 81
Localization of the Escherichia coli cell division protein Ftsl (PBP3) to the division site and cell pole; Weiss DS et al.; FtsI, also known as penicillin-binding protein 3, is a transpeptidase required for the synthesis of peptidoglycan in the division septum of the bacterium, Escherichia coli . FtsI has been estimated to be present at about 100 molecules per cell, well below the detection limit of immunoelectron microscopy . Here, we confirm the low abundance of FtsI and use immunofluorescence microscopy, a highly sensitive technique, to show that FtsI is localized to the division site during the later stages of cell growth . FtsI was also sometimes observed at the cell pole; polar localization was not anticipated and its significance is not known . We conclude (i) that immunofluorescence microscopy can be used to localize proteins whose abundance is as low as approximately 100 molecules per cell; and (ii) that spatial and temporal regulation of FtsI activity in septum formation is achieved, at least in part, by timed localization of the protein to the division site.

J Biochem (Tokyo), 1997 Aug, 122(2), 467 - 73
Fatty acid synthesis of an eicosapentaenoic acid-producing bacterium: de novo synthesis, chain elongation, and desaturation systems; Watanabe K et al.; The fatty acid synthesis systems of a Shewanella sp., strain SCRC-2738, that produces a large amount of eicosapentaenoic acid were investigated . Two kinds of fatty acid synthesis system, de novo synthesis and chain elongation ones, were detected in the cytosol . The de novo synthesis system required an acyl carrier protein, and produced palmitoyl- and palmitoleoyl-acyl carrier proteins as final products . The chain elongation system also required an acyl carrier protein, and produced an acyl-acyl carrier protein as a product, using palmitoyl-, palmitoleoyl-, stearoyl-, and oleoyl-CoAs as primers but not eicosanoyl- or eicosenoyl-CoA . There were an anaerobic pathway and an aerobic desaturation one for the production of unsaturated fatty acids . Eicosapentaenoic acid seemed to be produced through the aerobic desaturation pathway and not through the anaerobic one, since the latter pathway produced n-7 type monoenoic fatty acids, which are different from eicosapentaenoic acid in the position of the double bond . The desaturase utilized an acyl-acyl carrier protein as a substrate, and this activity increased in the presence of ferredoxin and ferredoxin NADP+ reductase . Thus, Shewanella sp., strain SCRC-2738, has novel characteristics as to both fatty acid chain elongation and desaturation systems.

Br J Surg, 1997 Sep, 84(9), 1190 - 9
Helicobacter pylori and gastric cancer; McFarlane GA et al.; BACKGROUND: Considerable progress has been made over the past two decades in understanding gastric carcinogenesis . Genetic and environmental factors have been shown to play a part . Infection with Helicobacter pylori leads to chronic gastritis and evidence is accumulating to link this disease with the subsequent development of gastric cancer . METHOD: A literature review was undertaken using Medline (National Library of Medicine, Washington DC, USA) searches of the headings gastrointestinal neoplasms and H . pylori for the years 1993-1997; further relevant references were examined . RESULTS: H . pylori is implicated in gastric carcinogenesis . Proposed mechanisms centre on the inflammatory response of the gastric mucosa to the bacterium . CONCLUSION: Investigation is still required to elucidate the exact role of H . pylori in the development of gastric carcinoma . Eradication of the organism in high-risk groups might lead to a reduction in the incidence of gastric cancer.

Schweiz Rundsch Med Prax, 1997 May 14, 86(20), 856 - 60
{JOMOL--immunostimulation and rejection of tumor cells?}; Allewelt MC et al.; Jomol contains cell wall fragments of the bacterium Nocardia opaca composed of 40% oligopeptides, 40% lipids, 10-15% polysaccharides, and 5-10% peptidoglycans . Technetium labeled--Jomo tech--and Indium-labeled--Jomo in--preparations are used for diagnosing cancer . Jomol is promoted to cure any cancer, except Non-Hodgkin lymphoma and ovarian cancer . It is claimed that Jomol has no side effects, however chills and high fever have been observed . Following initial intensive therapy with daily injections of Jomol, a maintenance therapy is recommended to prevent relapses . One vial of Jomol or Jomo-tech costs 450.--DM . Dr . U . Ehrenfeld, an oral surgeon, developed the aqueous extract of Nocardia opaca and patented it in 1983 under the name of Jomol . Ehrenfeld is promoting Jomo-tech and Jomo-in for qualitative and semi-quantitative cancer diagnosis with whole body scintigraphy . The following action of mechanisms of Jomol are claimed: 1 . Unspecific stimulation of cellular immune mechanisms, 2 . xenogenization of tumor cells towards a bacterial surface by adhesion of Jomol and then attraction and activation of macrophages . Despite the positive claims of the promoters the preclinical and clinical investigations are insufficient or missing, therefore a routine treatment of malignancies with Jomol is not justified except in study protocols . Jomol is not registered at the Intercantonal Office for Drug Control in Switzerland and is not approved by the Public Health Department in Germany and is not reimbursed by most of the state health insurances.

Pediatr Med Chir, 1997 Mar-Apr, 19(2), 143 - 4
{Atypical cat-scratch disease: a case report of splenic granulomatosis}; Dodi I et al.; Generally cat-scratch disease is a benign inflammatory adenopathy . The Authors describe an atypical form of this disease, characterized by persistent fever and splenic granulomatosis requiring a diagnostic and therapeutic prolonged effort . They point out the important role of new immuno-fluorescent techniques to exactly identify the bacterium--Bartonella henselae--causing cat-scratch disease and suggest to include cat-scratch disease among the causes of unknown origin fever.

Bull Math Biol, 1997 Sep, 59(5), 833 - 56
Mathematical modeling of adhesion of bacteria to host cell lines; Galvez J et al.; A mathematical model which describes adhesion of bacteria to host cell lines is presented . The model is flexible enough to account for the following situations: extracellular bacteria are either in exponential or in stationary phase . Adhesion is described as a reversible binding process in which the bacteria attach to or detach from specific receptors uniformly distributed on the cell surface . In turn, attached bacteria can either replicate or, conversely, they are restrained to remain in stationary phase . In the first case, however, we must consider the problem of whether the decrease of unoccupied receptors as adhesion progresses imposes a limit to the replicating capacity of the attached bacteria . The effect exerted by the multiplicity of infection (MOI), i.e . the ratio of the number of bacteria to the number of host cells, on the process of adhesion is also contemplated by the model . This has revealed that experiments performed at the same values of MOI can show completely different levels of adhered bacteria, depending on the number of host cells in the assays . This finding demonstrates that the report of the MOI values is insufficient to characterize comparative studies of bacterial adhesion since it could lead to a misunderstanding of the corresponding data . Simplified models based on the steady-state approximation and in equilibrium analysis by means of a Lagmuir absorption isotherm for the attached bacteria are also discussed . This allows us to define the adhesion coefficient ( beta) in a given bacterium-cell system so that, with the exception of those systems where these coefficients cannot be defined, larger values of beta are related to a greater adhesion capacity . An overview of the procedures to perform quantitative adhesion data analysis is outlined . Finally, theoretical predictions are compared with experimental results from the literature.

EMBO J, 1997 Oct 1, 16(19), 5827 - 36
Nucleator function of CsgB for the assembly of adhesive surface organelles in Escherichia coli; Bian Z et al.; Curli are surface organelles in Escherichia coli that assemble outside the bacterium through the precipitation of secreted soluble CsgA monomers, requiring the CsgB nucleator protein . Using immunoelectron microscopy and immunoblotting assays, CsgB is shown to be located on the bacterial surface and also as a minor component of wild-type curli . CsgB lacking its 20 N-terminal residues when fused to maltose-binding protein (MBP) can still trigger polymerization of CsgA monomers in vivo . However, the resulting surface organelles are only formed at one of the two bacterial poles and are morphologically distinct from wild-type curli . These Bfco organelles (CsgB-Free Curli-related Organelles) are highly regular structures reacting with anti-CsgA, but not anti-CsgB antibodies . The CsgB of the active MBP-CsgBII fusion is surface exposed but, unlike the native CsgB in wild-type curli, is not detectable in the Bfco organelles . Overexpression of csgB within a csgA mutant results in the formation of short CsgB polymers on the cell surface . It is suggested that in wild-type bacteria, both CsgA and CsgB are secreted proteins . Interaction between CsgA and CsgB triggers wild-type curli formation, resulting in CsgA-CsgB heteropolymers, while surface-anchored CsgB in MBP-CsgBII triggers morphologically distinct, CsgB-free/CsgA Bfco organelles . In the absence of CsgA, CsgB can self-assemble into polymers.

FEMS Microbiol Lett, 1997 Sep 15, 154(2), 201 - 5
Relationship between pathogenicity of Coxiella burnetii isolates and gene sequences of the macrophage infectivity potentiator (Cbmip) and sensor-like protein (qrsA); Masuzawa T et al.; Coxiella burnetii, the Q fever agent, is an obligate intracellular bacterium and survival in phagolysosomes is an important virulence factor . The present study was performed to determine the relationship between its pathogenicity and genes related to its survival in macrophages, i.e . macrophage infectivity potentiator and Q fever agent regulatory sensor-like protein . The sequence similarity was more than 99% among Japanese, European and American strains, and no relationship was found between pathogenicity in guinea pigs and these genes.

Biochimie, 1997 Jun, 79(6), 315 - 22
Artefactual cleavage of E coli H-NS by OmpT; Goldberg MD et al.; In the bacterium Escherichia coli, H-NS-(H1, H1a) is a heat-stable protein with a molecular mass of 15.5 kDa involved in nucleoid organisation and gene regulation linked to certain signal transduction pathways . We have shown that, following addition of preparations of everted inner membrane vesicles, heat-stable cleavage products of approximately 10 kDa of H-NS are formed in vitro from newly synthesised, radio-labelled H-NS and from purified H-NS . The 15.5 kDa protein and its cleavage products were also recovered from a minicell system . These results raised the possibility that cleavage of H-NS is physiologically significant . However, the cleavage of H-NS observed appears to occur during cell breakage and to depend on the method of protein extraction and the presence of the outer membrane protease, OmpT . Nevertheless, the results indicate that H-NS may contain at least two separate domains with cleavage occurring between these domains at a preferred OmpT site . Failure to take account of H-NS cleavage in sample preparation and analysis can lead to serious underestimation of H-NS levels.

Lipids, 1997 Sep, 32(9), 975 - 8
Lipid and fatty acid compositions of a novel docosahexaenoic acid-producing marine bacterium; Watanabe K et al.; An unidentified bacterial strain, SCRC-21406, isolated from the intestine of a marine fish, Glossanodon semifasciatus, produced docosahexaenoic acid at 23% (mol/mol) {= 28% (w/w)} of total fatty acids in a medium containing 0.5% (wt/vol) peptone and 0.1% (wt/vol) yeast extract at 12 degrees C under atmospheric pressure . The cell yield was 0.43 g/L . The major lipids of the strain were phosphatidylethanolamine and phophatidylglycerol . Docosahexaenoic acid was localized at the sn-2 positions of both phospholipids . The amounts of polyunsaturated fatty acids other than docosahexaenoic acid were extremely small {< 3% (mol/mol)} . Monounsaturated fatty acids of the cis-7, cis-9 and cis-11 types were detected.

Curr Microbiol, 1997 Oct, 35(4), 221 - 7
Cellobiose and cellodextrin metabolism by the ruminal bacterium Ruminococcus albus; Lou J et al.; Ruminococcus albus is an important fibrolytic bacterium in the rumen . Cellobiose is metabolized by this organism via hydrolytic and well as phosphorylytic enzymes, but the relative contributions of each pathway were not clear . The cellobiose consumption rate by exponentially growing cells was less than that of crude extracts (75 versus 243 nmol/min/mg protein) . Cellobiose phosphorolytic cleavage was much greater than hydrolytic activity (179 versus 19 nmol/min/mg protein) indicating that phosphorylases were key enzymes in the initial metabolism of the soluble products of cellulose degradation . Cellodextrin phosphorylase appeared to be active against substrates as large as cellohexaose . Phosphorylase activities were cytoplasmic, but hydrolytic activities were associated with both the membrane and cytoplasmic fractions . Free glucose was phosphorylated with a GTP-dependent glucokinase, and this enzyme showed 20-fold higher activity with GTP or ITP (>324 nmol/min/mg protein) than with ATP, UTP, CTP, GDP, or PEP . The activity was decreased at least 57% when mannose, 2-deoxyglucose, or fructose was used as substrate compared with glucose . The Kms for glucose and GTP were 321 and 247 microM, respectively . Since phosphorolytic cleavage conserves more metabolic energy than simple hydrolysis, it is likely that such pathways provide for more efficient growth of R . albus in substrate-limiting conditions like those found in the rumen.

FEBS Lett, 1997 Sep 1, 414(1), 45 - 9
The assimilatory nitrate reductase from the phototrophic bacterium, Rhodobacter capsulatus E1F1, is a flavoprotein; Blasco R et al.; The assimilatory nitrate reductase from the phototrophic bacterium Rhodobacter capsulatus has been purified to electrophoretic homogeneity and its molecular and kinetic parameters determined . The native nitrate reductase is a dimer of 144 kDa composed of two subunits of 46 and 95 kDa . The purified enzyme catalyzes the electron transfer from NADH, reduced bromophenol blue or reduced viologens to nitrate . The nitrate reductase contains 1 mol FAD per mole of enzyme and also reduces cytochrome c or dichlorophenol indophenol with NADH as the electron donor . The diaphorase activity is located in the small subunit.

Aliment Pharmacol Ther, 1997 Aug, 11(4), 699 - 703
A new 1-week therapy for Helicobacter pylori eradication: ranitidine bismuth citrate plus two antibiotics; Savarino V et al.; BACKGROUND: One-week triple regimens are currently the most recommended therapy for the eradication of Helicobacter pylori . No previous study has evaluated the efficacy of a short-term regimen combining ranitidine bismuth citrate with two antibiotics . METHODS: Seventy-two consecutive H . pylori-positive dyspeptic patients were recruited for this randomized, three-centre, open, parallel-group study . They were subdivided into two groups receiving either ranitidine bismuth citrate 400 mg b.d . + clarithromycin 250 mg b.d . and metronidazole 500 mg b.d . (group A) or ranitidine bismuth citrate 400 mg b.d . + clarithromycin 250 mg b.d . and metronidazole 250 mg q.d.s (group B) for 1 week . H . pylori infection was assessed by CLO-test and histology on both antral and corpus biopsies before and at least 4 weeks after the end of therapy . The bacterium was considered eradicated when both tests were negative . Eradication rates and the number of side-effects were evaluated in each group . The Chi-squared test was used for statistical analysis . RESULTS: One patient with only CLO-test positivity was erroneously randomized to group B and four patients dropped out of the study (two in group A and two in group B), mainly because they refused the second endoscopy . In group A, H . pylori was eradicated in 31 of 36 patients (intention-to-treat = 86%; 95% CI = 71-95% and per protocol 31/34 = 91%; 95% CI = 76-98%) . Side-effects occurred in 10 patients (27%) and they were generally mild . In group B, H . pylori was eradicated in 29 of 35 patients (intention-to-treat = 83%; 95% CI = 66-93%; and per protocol 29/33 = 88%; 95% CI = 72-97%) . Seven patients (20%) complained of modest side-effects . There was no significant difference between the two treatment arms (P = N.S.): no severe adverse events occurred and none of the patients was withdrawn from the study because of them . CONCLUSIONS: The co-administration of ranitidine bismuth citrate plus clarithromycin at low dosage and metronidazole in twice daily doses for 1 week is a short, effective and well-tolerated regimen for the eradication of H . pylori . These findings should provide the impetus for large-scale investigations.

Zhonghua Hu Li Za Zhi, 1996 Dec, 31(12), 690 - 1
{Study of surgical instruments contamination by bacteria from air during the operation}; Yin SH et al.; Routinely sterilized surgical instruments were divided into two groups and put on the same instrument table, one group was covered with dressing and the other was exposed to the air . The samples were collected at 30 min, 60 min, and 90 min respectively after operation began and bacterium culture was done . The results showed that the general air contamination rate of the exposed group was 1.18 times higher than that of the covered one . The exposure time had a positive correlation with bacterium contamination rate . This study gave the laboratory evidence for controlling the infection in the operation room.

EMBO J, 1997 Aug 1, 16(15), 4777 - 87
Photooxidative stress stimulates illegitimate recombination and mutability in carotenoid-less mutants of Rubrivivax gelatinosus; Ouchane S et al.; Carotenoids are essential to protection against photooxidative damage in photosynthetic and non-photosynthetic organisms . In a previous study, we reported the disruption of crtD and crtC carotenoid genes in the purple bacterium Rubrivivax gelatinosus, resulting in mutants that synthesized carotenoid intermediates . Here, carotenoid-less mutants have been constructed by disruption of the crtB gene . To study the biological role of carotenoids in photoprotection, the wild-type and the three carotenoid mutants were grown under different conditions . When exposed to photooxidative stress, only the carotenoid-less strains (crtB-) gave rise with a high frequency to four classes of mutants . In the first class, carotenoid biosynthesis was partially restored . The second class corresponded to photosynthetic-deficient mutants . The third class corresponded to mutants in which the LHI antenna level was decreased . In the fourth class, synthesis of the photosynthetic apparatus was inhibited only in aerobiosis . Molecular analyses indicated that the oxidative stress induced mutations and illegitimate recombination . Illegitimate recombination events produced either functional or non-functional chimeric genes . The R . gelatinosus crtB- strain could be very useful for studies of the SOS response and of illegitimate recombination induced by oxidants in bacteria.

Immunology, 1997 Jul, 91(3), 414 - 20
Mycobacterium avium infection in mice is associated with time-related expression of Th1 and Th2 CD4+ T-lymphocyte response; Azouaou N et al.; Disseminated infection caused by organisms of Mycobacterium avium complex is common in acquired immune deficiency syndrome (AIDS) patients . M . avium is an intracellular bacterium that multiplies within macrophages . We examined the effect of M . avium infection on the T-helper cell response in C57/BL/6 black mice . At weekly intervals, CD4+ T-cells were isolated from spleens and lines were created . T-cell lines were exposed to sonicated M . avium in the presence of feeder cells and macrophages and the supernatant were collected to measure the concentrations of interferon-gamma (IFN-gamma and interleukin-10 (IL-10) . Production of IFN-gamma in CD4+ T-cells obtained from uninfected mice did not vary significantly during the 5 weeks . Levels of IFN-gamma produced by T-cell lines of infected mice were similar to the control mice during the first 2 weeks but significantly reduced (approximately 30 ng/ml) thereafter . In contrast, production of IL-10 by T-cell lines of infected mice was in a range of 190 to 342 pg/ml in weeks 1, 2 and 3, but increased to an average of 1300 pg/ml at weeks 4 and 5 . Pre-immunized mice, when infected with M . avium strain 101, showed a different profile of T-cell cytokines, with high IFN-gamma and low IL-10 production . Proteins purified from a number of disease-associated (D-A) and non-D-A strains of M . avium were tested for the ability to induce IL-10 . 65,000 MW and 60,000 MW proteins of M . avium induced significantly more IL-10 than 45,000 MW, 33,000 MW and 27,000 MW proteins . These results showed that M . avium predominantly stimulates either Th1 or Th2 T-helper cells according to the phase of the infection.

Appl Microbiol Biotechnol, 1997 Aug, 48(2), 162 - 7
Production of zeaxanthin in Escherichia coli transformed with different carotenogenic plasmids; Ruther A et al.; Carotenoids are of great commercial interest and attempts are made to produce different carotenoids in transgenic bacteria and yeasts . Development of appropriate systems and optimization of carotenoid yield involves transformation with several new genes on suitable plasmids . Therefore, the non-carotenogenic bacterium Escherichia coli JM101 was transformed in our study with several genes that mediated the biosynthetic production of the carotenoid zeaxanthin in this host . Selection of plasmids for the introduction of five essential genes for zeaxanthin formation showed that a pACYC-derived plasmid was the best . Multiplasmid transformation generally decreased production of zeaxanthin . By cotransformation with different plasmids, limitations in the biosynthetic pathway were found at the level of geranylgeranyl-pyrophosphate synthase and beta-carotene hydroxylase . In our study a maximum zeaxanthin content of 289 micrograms/g dry weight was obtained . This involved the construction of a plasmid that mediate high-level expression of beta-carotene hydroxylase . The level of expression was demonstrated on protein gels and solubilization by the mild detergent Brij 78 revealed that a significant portion of the expressed enzyme is located in the E . coli membranes where it can exert its catalytic function . Based on the results obtained, new strategies for vector construction and strain selection were proposed which could increase the present concentrations drastically . Optimal growth conditions of the transformed E . coli strains for carotenoid formation were found at a temperature of 28 degrees C and a cultivation period of 2 days.

Electrophoresis, 1997 Aug, 18(8), 1335 - 46
Proteome analysis of Spiroplasma melliferum (A56) and protein characterisation across species boundaries; Cordwell SJ et al.; Spiroplasma melliferum (Class: Mollicutes) is a wall-less, helical bacterium with a genome of approximately 1460 kbp encoding 800-1000 gene-products . A two-dimensional electrophoresis gel reference map of S . melliferum was produced by Phoretix 2-D gel software analysis of eight high quality gels . The reference map showed 456 silver-stained and replicated protein spots . 156 proteins (34% of visible protein spots) from S . melliferum were further characterised by one, or a combination, of the following: amino acid analysis, peptide-mass fingerprinting via matrix assisted laser desorption ionisation-time of flight (MALDI-TOF) mass spectrometry, and N-terminal protein microsequencing . Proteins with close relationship to those previously determined from other species were identified across species barriers . Thus, this study represents the first larger-scale analysis of a proteome based upon the attribution of predominantly 'unique numerical parameters' for protein characterisation across species boundaries, as opposed to a sequence-based approach . This approach allowed all database entries to be screened for homology, as is currently the case for studies based on nucleic acid or protein sequence information . Several proteins studied from this organism were identified as hypothetical, or having no close homolog already present in the databases . Gene-products from major families such as glycolysis, translation, transcription, cellular processes, energy metabolism and protein synthesis were identified . Several gene-products characterised in S . melliferum were not previously found in studies of the entire Mycoplasma genitalium and Mycoplasma pneumoniae (both closely related Mollicutes) genomes . The presence of such gene-products in S . melliferum is discussed in terms of genome size as compared with the smallest known free-living organisms . Finally, the levels of expression of S . melliferum gene-products were determined with respect to total optical intensity associated with all visible proteins expressed in exponentially grown cells.

Eur J Oral Sci, 1997 Aug, 105(4), 310 - 7
Relationship of serotype, leukotoxin gene type and lysogeny in Actinobacillus actinomycetemcomitans to periodontal disease status; Barretto Tinoco EM et al.; Actinobacillus actinomycetemcomitans has been associated with different forms of periodontitis, particularly with localized juvenile periodontitis (LJP) . The bacterium possesses several virulence factors which have been shown to interact with the host immune system . Among these factors, leukotoxin, surface antigens (serotype) and bacteriophages have been suggested directly or indirectly to influence the course of infection . However, few studies have been able to show associations between these factors and periodontal disease, alone or in combination . Thus, the purpose of the present study was to investigate possible correlations between periodontal disease status and selected virulence factors (serotype, presence of bacteriophages, and the presence of a 530 bp deletion in the promoter region of the leukotoxin gene) . 36 subjects took part in the study . Serotype c was the most frequently found serotype among periodontally affected subjects, although serotypes a and b were also present . 27 out of 36 strains harbored bacteriophages, and there was strong evidence that some of the bacteriophages were different from the previously characterized phi Aa phage . A . actinomycetemcomitans containing the F-fragment phage were more frequently associated with periodontal disease . Five strains, all serotype b, 3 from LJP patients and 2 from healthy subjects, showed a 530-bp deletion in the promoter region of the leukotoxin gene.

FEMS Microbiol Lett, 1997 Sep 1, 154(1), 73 - 9
Detection of genomic polymorphisms among isolates of the intracellular bacterium Cowdria ruminantium by random amplified polymorphic DNA and Southern blotting; Perez JM et al.; Sixteen primers were successfully used in a RAPD assay to generate reproducible fingerprints for six isolates of Cowdria ruminantium, a tick-transmitted rickettsia of ruminants . Distinction between stocks was possible by using one or at most two primers . Two stocks were very similar although originating from widely distant geographical regions . A genetic distance tree was constructed by analysing 108 fragments in pairwise comparison between stocks . Three amplification fragments probed with C . ruminantium genomic DNA determined a restriction fragment length polymorphism which allowed the distinction between stocks except for the two stocks that had similar RAPD patterns . The potential of RAPD to determine the extent of genetic diversity of C . ruminantium and to develop probes or PCR primers for diagnostic purposes is discussed.

Nature, 1997 Sep 11, 389(6647), 206 - 11
Surface of bacteriorhodopsin revealed by high-resolution electron crystallography; Kimura Y et al.; Bacteriorhodopsin is a transmembrane protein that uses light energy, absorbed by its chromophore retinal, to pump protons from the cytoplasm of bacteria such as Halobacterium salinarium into the extracellular space . It is made up of seven alpha-helices, and in the bacterium forms natural, two-dimensional crystals called purple membranes . We have analysed these crystals by electron cryo-microscopy to obtain images of bacteriorhodopsin at 3.0 A resolution . The structure covers nearly all 248 amino acids, including loops outside the membrane, and reveals the distribution of charged residues on both sides of the membrane surface . In addition, analysis of the electron-potential map produced by this method allows the determination of the charge status of these residues . On the extracellular side, four glutamate residues surround the entrance to the proton channel, whereas on the cytoplasmic side, four aspartic acids occur in a plane at the boundary of the hydrophobic-hydrophilic interface . The negative charges produced by these aspartate residues is encircled by areas of positive charge that may facilitate accumulation and lateral movement of protons on this surface.

Ann Otolaryngol Chir Cervicofac, 1997, 114(3), 80 - 3
{Cervicofacial manifestations of tularemia . Apropos of a familial case}; Benlyazid A et al.; Tularemia is a rare infectious disease, due to Francisella tularensis, a virulent bacterium transmitted by a carrier insect (essentially ticks) or by the meat of an infected animal (generally hares) . We report 3 cases that occurred in the same family, showing the various symptoms of this disease . Revealing head and neck manifestations may mislead diagnosis.

Biochim Biophys Acta, 1997 Aug 16, 1347(2-3), 164 - 76
Polyunsaturated fatty acids in the psychrophilic bacterium Shewanella gelidimarina ACAM 456T: molecular species analysis of major phospholipids and biosynthesis of eicosapentaenoic acid; Nichols DS et al.; The production of eicosapentaenoic acid {20:5omega3; EPA} from Shewanella gelidimarina (ACAM 456T) was investigated with respect to growth temperature and growth on sole carbon sources . The percentage and quantitative yield of EPA remained relatively constant at all growth temperatures within or below the optimal growth temperature region . At higher growth temperatures, these values decreased greatly . Growth on differing sole carbon sources also influenced the percentage and amount of EPA produced, with the fatty acid composition influenced by provision of potential acyl chain primers as sole carbon sources . The highest amounts of EPA occurred from growth on propionic acid and L-leucine respectively, while the highest percentage of EPA occurred from growth on L-proline . Monounsaturated fatty acid components and EPA were concentrated in phosphatidylglycerol (PG), while the proportion of branched-chain fatty acids was elevated in phosphatidylethanolamine (PE); the two major phospholipid classes . Specific associations of EPA with other acyl chains were identified within cellular phospholipid classes . The association of EPA with 17:1 and 18:0 acyl chains in phospholipid species was specific to PG, whereas the association of EPA with i13:0/13:0 and 14:0/i14:0 was specific to PE . Such acyl chain 'tailoring' is indicative of the important role of EPA in bacterial membrane adaptive responses . EPA was also a large component (22%) of a non-esterified fatty acid (NEFA) fraction within the total lipid extract of the bacterium . This may point toward a particular role of NEFA in polyunsaturated fatty acid (PUFA) metabolism . The formation of EPA was investigated by labelling with L-{U-14C}serine and sodium {1-14C}acetate . The accumulation of radiolabel within unsaturated intermediates (di-, tri- and tetraunsaturated fractions) was low, indicating a rapid formation and derivatisation of these components . Similar results were found for the unsaturated fatty acid fractions of both PE and PG using sodium {1-14C}acetate radiolabel . The regulation of triunsaturated fatty acid components may be a potential control site in PUFA biosynthesis.

Biochim Biophys Acta, 1997 Aug 16, 1347(2-3), 151 - 63
Membrane lipids of Rhodopseudomonas viridis; Linscheid M et al.; In search of the precyanobacterial origin of the typical thylakoid lipids found in cyanobacteria and chloroplasts, we analyzed the polar lipids of the anaerobic phototrophic bacterium Rhodopseudomonas viridis . Glycolipids (monogalactosyl-, digalactosyl- and glucuronosyl diacylglycerol), phospholipids (phosphatidyl choline, -ethanolamine, -glycerol and cardiolipin) and an ornithine lipid were isolated and identified by NMR (1H, 13C, 31P) and mass spectrometry . Positional distribution and pairing of fatty acids in molecular species show small, but significant differences between glyco- and phospholipids . In this context, a new enzymatic method is described for assigning the enantiomeric structure of the diacylglycerol moiety in glyco- and phospholipids . 14C-Labelling studies suggest that monogalactosyl diacylglycerol is formed by galactosylation of diacylglycerol as in chloroplasts and not by glucosylation followed by epimerization as in cyanobacteria . The two 1,6-linked galactopyranose residues of digalactosyl diacylglycerol are both in beta-linkage and thus differ from the corresponding chloroplast lipid with its alpha-beta-sequence . R . viridis does not contain the sulfolipid, and even phosphate starvation does not induce the synthesis of this most characteristic thylakoid lipid, which on the other hand is present in other anaerobic phototrophic bacteria.

Diagn Microbiol Infect Dis, 1997 Jul, 28(3), 149 - 52
Modification of Christensen urease test as an inexpensive tool for detection of Helicobacter pylori; Dominguez-Bello MG et al.; About half the world population is infected with Helicobacter pylori . Most live in developing countries where clinical studies face the constraints of high costs of imported rapid diagnostic tests . In this work, we describe and validate a simple local urease test (LUT) to determine the presence of the bacterium in gastric biopsies, and report the incidence of infection among symptomatic patients in Caracas, Venezuela . Statistical comparison of LUT and CLOtest (Delta West, Bentley, Australia) (N = 216 patients) showed that the probability of 95% agreement between the two test was 0.936 . Overall incidence of infection determined by the LUT was 65% (N = 229), and it was higher in patients from public (72%; N = 153) than from private (50%; N = 76) hospitals (p = .001) . Therefore, the incidence of infection differs in two socioeconomic groups that coexist in the same city . LUT may represent an affordable tool in clinical studies needed to identify social factors that increase the risk of infection by H . pylori.

J Bacteriol, 1997 Sep, 179(18), 5892 - 902
Helicobacter pylori ABC transporter: effect of allelic exchange mutagenesis on urease activity; Hendricks JK et al.; Helicobacter pylori urease requires nickel ions in the enzyme active site for catalytic activity . Nickel ions must, therefore, be actively acquired by the bacterium . NixA (high-affinity nickel transport protein)-deficient mutants of H . pylori retain significant urease activity, suggesting the presence of alternate nickel transporters . Analysis of the nucleotide sequence of the H . pylori genome revealed a homolog of NikD, a component of an ATP-dependent nickel transport system in Escherichia coli . Based on this sequence, a 378-bp DNA fragment was PCR amplified from H . pylori genomic DNA and used as a probe to identify an H . pylori lambda ZAPII genomic library clone that carried these sequences . Four open reading frames of 621, 273, 984, and 642 bp (abcABCD) were revealed by sequencing and predicted polypeptides of 22.7, 9.9, 36.6, and 22.8 kDa, respectively . The 36.6-kDa polypeptide (AbcC) has significant homology (56% amino acid sequence identity) to an E . coli ATP-binding protein component of an ABC transport system, while none of the other putative proteins are significantly homologous to polypeptides in the available databases . To determine the possible contribution of these genes to urease activity, abcC and abcD were each insertionally inactivated with a kanamycin resistance (aphA) cassette and allelic exchange mutants of each gene were constructed in H . pylori UMAB41 . Mutation of abcD resulted in an 88% decrease in urease activity to 27 +/- 31 mumol of NH3/min/mg of protein (P < 0.0001), and a double mutant of nixA and abcC resulted in the near abolishment of urease activity (1.1 +/- 1.4 mumol of NH3/min/mg of protein in the double mutant versus 228 +/- 92 mumol of NH3/min/mg of protein in the parent {P < 0.0001}) . Synthesis of urease apoenzyme, however, was unaffected by mutations in any of the abc genes . We conclude that the abc gene cluster, in addition to nixA, is involved in production of a catalytically active urease.

J Bacteriol, 1997 Sep, 179(18), 5712 - 9
Analysis of a chemotaxis operon from Rhodospirillum centenum; Jiang ZY et al.; A chemotaxis gene cluster from the photosynthetic bacterium Rhodospirillum centenum has been cloned, sequenced, and analyzed for the control of transcription during swimmer-to-swarm cell differentiation . The first gene of the operon (cheAY) codes for a large 108-kDa polypeptide with an amino-terminal domain that is homologous to CheA and a carboxyl terminus that is homologous to CheY . cheAY is followed by cheW, an additional homolog of cheY, cheB, and cheR . Sequence analysis indicated that all of the che genes are tightly compacted with the same transcriptional polarity, suggesting that they are organized in an operon . Cotranscription of the che genes was confirmed by demonstrating through Western blot analysis that insertion of a polar spectinomycin resistance gene in cheAY results in loss of cheR expression . The promoter for the che operon was mapped by primer extension analysis as well as by the construction of promoter reporter plasmids that include several deletion intervals . This analysis indicated that the R . centenum che operon utilizes two promoters; one exhibits a sigma 70-like sequence motif, and the other exhibits a sigma 54-like motif . Expression of the che operon is shown to be relatively constant for swimmer cells which contain a single flagellum and for swarm cells that contain multiple lateral flagella.

J Bacteriol, 1997 Sep, 179(18), 5677 - 83
Purification and cloning of a proline 3-hydroxylase, a novel enzyme which hydroxylates free L-proline to cis-3-hydroxy-L-proline; Mori H et al.; Proline 3-hydroxylase was purified from Streptomyces sp . strain TH1, and its structural gene was cloned . The purified enzyme hydroxylated free L-proline to cis-3-hydroxy-L-proline and showed properties of a 2-oxoglutarate-dependent dioxygenase (H . Mori, T . Shibasaki, Y . Uosaki, K . Ochiai, and A . Ozaki, Appl . Environ . Microbiol, 62:1903-1907, 1996) . The molecular mass of the purified enzyme was 35 kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis . The isoelectric point of the enzyme was 4.3 . The optimal pH and temperature were 7.0 and 35 degrees C, respectively . The K(m) values were 0.56 and 0.11 mM for L-proline and 2-oxoglutarate, respectively . The Kcat value of hydroxylation was 3.2 s-1 . Determined N-terminal and internal amino acid sequences of the purified protein were not found in the SwissProt protein database . A DNA fragment of 74 bp was amplified by PCR with degenerate primers based on the determined N-terminal amino acid sequence . With this fragment as a template, a digoxigenin-labeled N-terminal probe was synthesized by PCR . A 6.5-kbp chromosome fragment was cloned by colony hybridization with the labeled probe . The determined DNA sequence of the cloned fragment revealed a 870-bp open reading frame (ORF 3), encoding a protein of 290 amino acids with a calculated molecular weight of 33,158 . No sequence homolog was found in EMBL, GenBank, and DDBJ databases . ORF 3 was expressed in Escherichia coli DH1 . Recombinants showed hydroxylating activity five times higher than that of the original bacterium, Streptomyces sp . strain TH1 . It was concluded that the ORF 3 encodes functional proline 3-hydroxylase.

Biochim Biophys Acta, 1997 Aug 7, 1353(2), 118 - 24
Cloning and sequence of a thermostable multidomain xylanase from the bacterium Rhodothermus marinus; Nordberg Karlsson E et al.; The gene (xyn1) encoding a Rhodothermus marinus xylanase has been cloned and expressed in Escherichia coli . The gene comprises 5 different domains in an unusual combination . The cellulose binding domains (CBDs) encoded by xyn1 are repeated in tandem at the N-terminus and show similarity with the CBD family IV . The xyn1-gene is the first example encoding a CBD family IV in combination with a xylan hydrolyzing catalytic domain of the glycosyl hydrolase family 10.

Appl Environ Microbiol, 1997 Sep, 63(9), 3707 - 10
Purification and characterization of a haloalkane dehalogenase of a new substrate class from a gamma-hexachlorocyclohexane-degrading bacterium, Sphingomonas paucimobilis UT26; Nagata Y et al.; The linB gene product (LinB), 1,3,4,6-tetrachloro-1,4-cyclohexadiene halidohydrolase, which is involved in the degradation of gamma-hexachlorocyclohexane in Sphingomonas paucimobilis UT26 (Y . Nagata, T . Nariya, R . Ohtomo, M . Fukuda, K . Yano, and M . Takagi, J . Bacteriol . 175:6403-6410, 1993), was overproduced in E . coli and purified to homogeneity . The molecular mass of LinB was deduced to be 30 kDa by gel filtration chromatography and 32 kDa by electrophoresis on sodium dodecyl sulfate-polyacrylamide gel, indicating that LiuB is a monomeric enzyme . The optimal pH for activity was 8.2 . Not only monochloroalkanes (C3 to C10) but also dichloroalkanes, bromoalkanes, and chlorinated allphatic alcohols were good substrates for LinB, suggesting that LinB shares properties with another haloalkane dehalogenase, DhlA (S . Keuning, D.B . Janssen, and B . Witholt, J . Bacteriol . 163:635-639, 1985), which shows significant similarity to LinB in primary structure (D . B . Janssen, F . Pries, J . van der Ploeg, B . Kazemier, P . Terpstra, and B . Witholt, J . Bacteriol . 171:6791-6799, 1989) but not in substrate specificity . Principal component analysis of substrate activities of various haloalkane dehalogenases suggested that LinB probably constitutes a new substrate specificity class within this group of enzymes.

Appl Environ Microbiol, 1997 Sep, 63(9), 3600 - 6
Diversity of the ribulose bisphosphate carboxylase/oxygenase form I gene (rbcL) in natural phytoplankton communities; Pichard SL et al.; The phytoplankton of the world's oceans play an integral part in global carbon cycling and food webs by conversion of carbon dioxide into organic carbon . They accomplish this task through the action of the Calvin cycle enzyme ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) . Here we have investigated the phylogenetic diversity in the form I rbcL locus in natural phytoplankton communities of the open ocean and representative clones of marine autotrophic picoplankton by mRNA or DNA amplification and sequencing of a 480 to 483 bp internal fragment of this gene . Five gene sequences were recovered from nucleic acids of natural phytoplankton communities of the Gulf of Mexico . The rbcL genes of two Prochlorococcus isolates and one Synechococcus strain (WH8007) were also sequenced . Sequences were aligned with the database of rbcL genes and subjected to both neighbor-joining and parsimony analyses . The five sequences from the natural phytoplankton community spanned nearly the entire diversity of characterized form I rbcL genes, with some sequences closely related to isolates such as Synechococcus and Prochlorococcus (forms IA and I) and prymnesiophyte algae (form ID), while other sequences were deeply rooted . Unexpectedly, the deep euphotic zone contained an organism that possesses a transcriptionally active rbcL gene closely related to that of a recently characterized manganese-oxidizing bacterium, suggesting that such chemoautotrophs may contribute to the diversity of carbon-fixing organisms in the marine euphotic zone.

J Biol Chem, 1997 Sep 5, 272(36), 22502 - 8
Studies on the redox centers of the terminal oxidase from Desulfovibrio gigas and evidence for its interaction with rubredoxin; Gomes CM et al.; Rubredoxin-oxygen oxidoreductase (ROO) is the final component of a soluble electron transfer chain that couples NADH oxidation to oxygen consumption in the anaerobic sulfate reducer Desulfovibrio gigas . It is an 86-kDa homodimeric flavohemeprotein containing two FAD molecules, one mesoheme IX, and one Fe-uroporphyrin I per monomer, capable of fully reducing oxygen to water . EPR studies on the native enzyme reveal two components with g values at approximately 2.46, 2.29, and 1.89, which are assigned to low spin hemes and are similar to the EPR features of P-450 hemes, suggesting that ROO hemes have a cysteinyl axial ligation . At pH 7.6, the flavin redox transitions occur at 0 +/- 15 mV for the quinone/semiquinone couple and at -130 +/- 15 mV for the semiquinone/hydroquinone couple; the hemes reduction potential is -350 +/- 15 mV . Spectroscopic studies provided unequivocal evidence that the flavins are the electron acceptor centers from rubredoxin, and that their reduction proceed through an anionic semiquinone radical . The reaction with oxygen occurs in the flavin moiety . These data are strongly corroborated by the finding that rubredoxin and ROO are located in the same polycistronic unit of D . gigas genome . For the first time, a clear role for a rubredoxin in a sulfate-reducing bacterium is presented.

Healthc Demand Dis Manag, 1997 Oct, 3(10), 154 - 5
Use new diagnostic know-how to improve prevention, treatment of peptic ulcer disease; Biochemical and spectroscopic characterization of two new cytochromes isolated from Desulfuromonas acetoxidans; Laboratoire de Bioenergetique et Ingenierie des Proteines, IFR 1, C.N.R.S., Marseille, France . bruschi@ibsm.cnrs-mrs.fr

The multimeric cytochromes described to date in sulfate- and sulfur-reducing bacteria are associated with diverse respiratory modes involving the use of elemental sulfur or oxidized sulfur compounds as terminal acceptors . They exhibit no structural similarity with the other cytochrome c classes and are characterized by a bis-histidinyl axial iron coordination and low redox potentials . We have purified two new cytochromes c with markedly different molecular masses (10 000 and 50 000) from the bacterium Desulfuromonas acetoxidans, which uses anaerobic sulfur respiration as its sole energy source . The characterization by electrochemistry and optical and EPR spectroscopies revealed the cytochrome c (Mr = 10 000) to be the first monohemic cytochrome c exhibiting a bis-histidinyl axial coordination and a low redox potential (-220 mV) . The cytochrome c (Mr = 50 000) contains four hemes of low potential (-200, -210, -370, and -380 mV) with the same axial coordination . The N-terminal amino acid sequences were compared with that of the trihemic cytochrome c7, previously described in D . acetoxidans and which is related to tetrahemic cytochrome c3 from sulfate reducing bacteria . Some homology was found between cytochrome c (Mr = 10 000) and cytochrome c7 . Both D . acetoxidans cytochromes c are located in the periplasmic space and their biochemical and spectroscopic properties indicate that they belong to the class III cytochromes.

J Bacteriol, 1997 Sep, 179(17), 5422 - 8
Differential levels of specific cytochrome c biogenesis proteins in response to oxygen: analysis of the ccl operon in Rhodobacter capsulatus; Gabbert KK et al.; The photosynthetic bacterium Rhodobacter capsulatus synthesizes c-type cytochromes under a variety of growth conditions . For example, under aerobic growth, c-type cytochromes are synthesized as part of an electron transport pathway, using oxygen as the terminal electron acceptor . Anaerobically in the light, R . capsulatus requires cytochrome bc1 and other c-type cytochromes for the photosynthetic electron transport pathway . It is shown here that the ccl1 and ccl2 genes of R . capsulatus are required for the synthesis of all c-type cytochromes, including the cytochrome c' protein of unknown function but of structural similarity to cytochrome b562 . Polar and nonpolar mutations constructed in each gene demonstrated that the ccl12 genes form an operon . Expression of the ccl12 genes was examined by using lacZ and phoA fusions as translational reporters . Primer extension analysis was used to determine transcriptional control and the start site of the ccl12 promoter . Finally, antiserum to the Ccl2 protein was used to quantitate levels of Ccl2 under six different growth conditions . The Ccl2 protein is present at 20-fold-higher levels under conditions where oxygen is present . In contrast, other cytochromes c biogenesis proteins, HelA and HelX, previously shown to be part of an helABCDX operon, are at relatively similar levels under these six growth conditions . This discovery is discussed in terms of the physiology and evolution of cytochromes c biogenesis, with particular attention to oxidative environments.

Curr Biol, 1997 Sep 1, 7(9), R538 - 40
Bacterial pathogenesis: a variation on variation in Lyme disease; Koomey M; The discovery of antigenic variation in Borrelia burgdorferi, the bacterium that causes Lyme disease, provides a potential explanation for the chronic nature of infection as well as new insights into the genetic structure of highly recombinogenic loci responsible for combinatorial genetic diversification.

Infect Immun, 1997 Sep, 65(9), 3622 - 30
Role of adherence in interleukin-8 induction in Helicobacter pylori-associated gastritis; Rieder G et al.; Active Helicobacter pylori-associated gastritis is characterized by a dense mucosal infiltration with granulocytes . Since H . pylori is noninvasive, secondary signals must induce the accumulation of granulocytes . Interleukin-8 (IL-8) has been shown to play a key role in this event . Using competitive reverse transcriptase-PCR on mRNA from gastric biopsies, we could show a clear correlation between the amount of IL-8 transcripts and the activity of H . pylori gastritis . Due to the inability of the bacterium to invade host cells, the epithelial layer is a potential candidate as an IL-8 source . To study the mechanism of IL-8 induction, established gastric carcinoma epithelial cell lines (AGS and Kato III) and well-defined H . pylori strains were used in a modified in vitro system . The experimental design enabled us to prevent direct contact of bacteria with epithelial cells by use of a filter membrane which did not block secreted bacterial products crossing the membrane . The data clearly showed that the direct contact of the bacterial cell with the epithelial cell is necessary for optimal IL-8 production because not only live bacteria, but also metabolically inactive bacteria, increased IL-8 secretion . Neither purified lipopolysaccharide nor water-soluble protein fractions of H . pylori NCTC 11637 and Tx30a nor the cytotoxin of H . pylori was able to increase IL-8 production significantly by the epithelial cells used . Furthermore, preparations of total membrane and outer membrane proteins of H . pylori were not able to stimulate IL-8 release in vitro . Accumulatively, these results imply that active metabolism is not necessary for stimulation as long as there is an intact membrane aiding the presentation of a stimulating membrane complex or aggregate on the surface of the bacteria . From these results, we conclude that whole bacteria and their direct contact with epithelial cells may be critical for IL-8 induction in vivo.

Bull Math Biol, 1997 Sep, 59(5), 857 - 79
Penetration of host cell lines by bacteria . Characteristics of the process of intracellular bacterial infection; Galvez J et al.; A model which describes the characteristics of the penetration of the cells by bacteria is presented . Since the process of invasion is preceded necessarily by the step in which the bacteria adhere to the cells, the proposed model is based on the expressions previously derived for the process of adhesion, which allow us to determine the number of attached bacteria under different conditions . Thus, the model considers that invasion occurs irreversibly from attached bacteria to specific receptors located on the cell surface with a rate coefficient = ki so that the invasive capacity in a given bacterium-host cell system is mainly determined by the value of this coefficient . Once internalized, the bacteria can follow three different time courses, namely: 1) intracellular growth is hindered so that the bacteria remain in stationary phase, 2) there is a lag phase during which the bacteria stay in stationary phase before they are able to grow exponentially with a rate coefficient = kc, and 3) the bacteria exhibit a growth exponential phase as they enter the cells . In turn, the time course followed by extracellular bacteria also has a decisive influence on the process of invasion and, in this regard, unbound bacteria are considered either in stationary or in exponential phase . Expressions for these different situations have been derived, and from them, procedures to determine the levels of bacterial infection and for quantitative invasive data analysis are presented.

J Biol Chem, 1997 Aug 22, 272(34), 21558 - 64
Purification, characterization, and reconstitution of DNA-dependent RNA polymerases from Caulobacter crescentus; Wu J et al.; Cell differentiation in the Caulobacter crescentus cell cycle requires differential gene expression that is regulated primarily at the transcriptional level . Until now, however, a defined in vitro transcription system for the biochemical study of developmentally regulated transcription factors had not been available in this bacterium . We report here the purification of C . crescentus RNA polymerase holoenzymes and resolution of the core RNA polymerase from holoenzymes by chromatography on single-stranded DNA cellulose . The three RNA polymerase holoenzymes Esigma54, Esigma32, and Esigma73 were reconstituted exclusively from purified C . crescentus core and sigma factors . Reconstituted Esigma54 initiated transcription from the sigma54-dependent fljK promoter of C . crescentus in the presence of the transcription activator FlbD, and active Esigma32 specifically initiated transcription from the sigma32-dependent promoter of the C . crescentus heat-shock gene dnaK . For reconstitution of the Esigma73 holoenzyme, we overexpressed the C . crescentus rpoD gene in Escherichia coli and purified the full-length sigma73 protein . The reconstituted Esigma73 recognized the sigma70-dependent promoters of the E . coli lacUV5 and neo genes, as well as the sigma73-dependent housekeeping promoters of the C . crescentus pleC and rsaA genes . The ability of the C . crescentus Esigma73 RNA polymerase to recognize E . coli sigma70-dependent promoters is consistent with relaxed promoter specificity of this holoenzyme previously observed in vivo.

Proc Natl Acad Sci U S A, 1997 Aug 19, 94(17), 9091 - 5
Parallel-up structure evidences the molecular directionality during biosynthesis of bacterial cellulose; Koyama M et al.; The "parallel-up" packing in cellulose Ialpha and Ibeta unit cells was experimentally demonstrated by a combination of direct-staining the reducing ends of cellulose chains and microdiffraction-tilting electron crystallographic analysis . Microdiffraction investigation of nascent bacterial cellulose microfibrils showed that the reducing end of the growing cellulose chains points away from the bacterium, and this provides direct evidence that polymerization by the cellulose synthase takes place at the nonreducing end of the growing cellulose chains . This mechanism is likely to be valid also for a number of processive glycosyltransferases such as chitin synthases, hyaluronan synthases, and proteins involved in the synthesis of nodulation factor backbones.

J Exp Med, 1997 Aug 18, 186(4), 537 - 47
Identification of a Gal/GalNAc lectin in the protozoan Hartmannella vermiformis as a potential receptor for attachment and invasion by the Legionnaires' disease bacterium; Venkataraman C et al.; The Legionnaire's disease bacterium, Legionella pneumophila, is a facultative intracellular pathogen which invades and replicates within two evolutionarily distant hosts, free-living protozoa and mammalian cells . Invasion and intracellular replication within protozoa are thought to be major factors in the transmission of Legionnaire's disease . Although attachment and invasion of human macrophages by L . pneumophila is mediated in part by the complement receptors CR1 and CR3, the protozoan receptor involved in bacterial attachment and invasion has not been identified . To define the molecular events involved in invasion of protozoa by L . pneumophila, we examined the role of protein tyrosine phosphorylation of the protozoan host Hartmannella vermiformis upon attachment and invasion by L . pneumophila . Bacterial attachment and invasion were associated with a time-dependent tyrosine dephosphorylation of multiple host cell proteins . This host cell response was highly specific for live L . pneumophila, required contact with viable bacteria, and was completely reversible following washing off the bacteria from the host cell surface . Tyrosine dephosphorylation of host proteins was blocked by a tyrosine phosphatase inhibitor but not by tyrosine kinase inhibitors . One of the tyrosine dephosphorylated proteins was identified as the 170-kD galactose/N-acetylgalactosamine-inhibitable lectin (Gal/GalNAc) using immunoprecipitation and immunoblotting by antibodies generated against the Gal/GalNAc lectin of the protozoan Entamoeba histolytica . This Gal/GalNAc-inhibitable lectin has been shown previously to mediate adherence of E . histolytica to mammalian epithelial cells . Uptake of L . pneumophila by H . vermiformis was specifically inhibited by two monovalent sugars, Gal and GalNAc, and by mABs generated against the 170-kD lectin of E . histolytica . Interestingly, inhibition of invasion by Gal and GalNAc was associated with inhibition of bacterial-induced tyrosine dephosphorylation of H . vermiformis proteins . High stringency DNA hybridization confirmed the presence of the 170-kD lectin gene in H . vermiformis . We conclude that attachment of L . pneumophila to the H . vermiformis 170-kD lectin is required for invasion and is associated with tyrosine dephosphorylation of the Gal lectin and other host proteins . This is the first demonstration of a potential receptor used by L . pneumophila to invade protozoa.

Anal Chem, 1997 Aug 15, 69(16), 3380 - 4
Sensing antimonite and arsenite at the subattomole level with genetically engineered bioluminescent bacteria; Ramanathan S et al.; A highly sensitive and selective optical sensing system for antimonite has been developed using genetically engineered bacteria . The basis of this system is the ability of certain bacteria to survive in environments that are contaminated with antimonite, arsenite, and arsenate . The survival is conferred to the bacteria by the ars operon, which consists of five genes that code for three structural proteins, ArsA, ArsB, and ArsC, and two regulatory proteins, ArsD and ArsR . ArsA, ArsB, and ArsC form a protein pump system that extrudes antimonite, arsenite, and arsenate once these anions reach the cytoplasm of the bacterium . A method was developed for monitoring antimonite and arsenite by using a single plasmid that incorporates the regulatory gene of the extrusion system, arsR, and the genes of bacterial luciferase, luxA and luxB . In the designed plasmid, ArsR regulates the expression of bacterial luciferase in a manner that is dependent on the concentration of antimonite and arsenite in the sample . Thus, the bioluminescence emitted by luciferase can be related to the concentration of antimonite and arsenite in the sample . Concentrations for antimonite and arsenite in the order of 10(-5) M, which corresponds to subattomole levels, can be detected . This bacterial-based sensing system is highly selective for antimonite and arsenite.

Proc Natl Acad Sci U S A, 1997 Aug 5, 94(16), 8456 - 61
Purification and characterization of acetone carboxylase from Xanthobacter strain Py2; Sluis MK et al.; Acetone metabolism in the aerobic bacterium Xanthobacter strain Py2 proceeds by a carboxylation reaction forming acetoacetate as the first detectable product . In this study, acetone carboxylase, the enzyme catalyzing this reaction, has been purified to homogeneity and characterized . Acetone carboxylase was comprised of three polypeptides with molecular weights of 85,300, 78,300, and 19,600 arranged in an alpha2beta2gamma2 quaternary structure . The carboxylation of acetone was coupled to the hydrolysis of ATP and formation of 1 mol AMP and 2 mol inorganic phosphate per mol acetoacetate formed . ADP was also formed during the course of acetone consumption, but only accumulated at low, substoichiometric levels ( approximately 10% yield) relative to acetoacetate . Inorganic pyrophosphate could not be detected as an intermediate or product of acetone carboxylation . In the absence of CO2, acetone carboxylase catalyzed the acetone-dependent hydrolysis of ATP to form both ADP and AMP, with ADP accumulating to higher levels than AMP during the course of the assays . Acetone carboxylase did not have inorganic pyrophosphatase activity . Acetone carboxylase exhibited a Vmax for acetone carboxylation of 0.225 micromol acetoacetate formed min-1.mg-1 at 30 degrees C and pH 7.6 and apparent Km values of 7.80 microM (acetone), 122 microM (ATP), and 4 . 17 mM (CO2 plus bicarbonate) . These studies reveal molecular properties of the first bacterial acetone-metabolizing enzyme to be isolated and suggest a novel mechanism of acetone carboxylation coupled to ATP hydrolysis and AMP and inorganic phosphate formation.

Eur J Biochem, 1997 Aug 1, 247(3), 833 - 42
Molecular characterization of Fdx1, a putidaredoxin-type {2Fe-2S} ferredoxin able to transfer electrons to the dioxin dioxygenase of Sphingomonas sp . RW1; Armengaud J et al.; Bacterium Sphingomonas sp . strain RW1 is, under aerobic conditions, able to degrade dibenzofuran and dibenzo-p-dioxin . The first step of the pathway is performed by a ring-dihydroxylating enzyme . Bunz and Cook have reported the purification and characterization of this dioxin dioxygenase and a ferredoxin able to transfer electrons to the dioxygenase {Bunz, P . V . & Cook, A . M . (1993) J . Bacteriol . 175, 6467-6475} . The gene encoding this {2Fe-2S} ferredoxin was identified by screening a genomic library constructed in pLAFR3 with a probe generated by a nested-PCR amplification . Primers for the amplification were designed based on the N-terminus sequence of the purified ferredoxin and on sequence comparisons with related proteins . Several cosmids were obtained and the ferredoxin gene, fdx1, was subcloned from one of them . The nucleotide sequence of a 4.6-kb DNA fragment encompassing the ferredoxin gene was determined . While in the case of all known multi-component dioxygenases, genes encoding the alpha and beta subunits are found to be contiguous with the gene of the specific electron carrier, the fdx1 gene in Sphingomonas sp . RW1 does not appear to be directly linked with the dioxin dioxygenase genes . Rather, it is clustered with genes apparently encoding two atypical decarboxylases/isomerases and a glutathione S-transferase . The ferredoxin gene was hyperexpressed and the recombinant ferredoxin was purified . Spectroscopic characterization of Fdx1 demonstrated the presence of a putidaredoxin-type {2Fe-2S} cluster in this protein . Its redox potential was determined to be -245 (+/- 5) mV versus the normal hydrogen electrode at 25 degrees C, pH 8.0 . Therefore, the protein is closely related to {2Fe-2S} ferredoxins known to be electron donors to monooxygenases involved in hydroxylation of aromatic compounds . Thus, this report provides clear evidence that a putidaredoxin-type {2Fe-2S} ferredoxin, namely Fdx1, is able to transfer electrons to the dioxin dioxygenase of Sphingomonas sp . RW1.

J Appl Microbiol, 1997 Aug, 83(2), 227 - 35
Diversity and partial characterization of putative virulence determinants in Pasteuria penetrans, the hyperparasitic bacterium of root-knot nematodes (Meloidogyne spp.); Davies KG et al.; Antigens recognized by monoclonal antibodies (Mabs) raised to the surface of the obligate nematode hyperparasite Pasteuria penetrans were characterized . Using the attachment of spores of the bacterium to host nematodes to determine the biological variability present on the spore surface greatly underestimated the amount of surface heterogeneity present compared with estimates from immunological techniques . This heterogeneity differed not only between different individual spores from the same population but also between different spore populations . None of the Mabs completely inhibited any spore population from attaching to the nematode cuticle, suggesting that the mechanism of attachment may be more complex than previously supposed . Chemical degradation of one particular epitope recognized by monoclonal antibody PP1/117, and designated ep117, occurred after treatment with NaOH, periodate or Proteinase K, suggesting that an O-linked glycoprotein may be involved . Fibronectin, which had been found to bind to Pasteuria spores through hydrophobic interactions, also prohibited the Mab from recognizing ep117 . However, SDS-PAGE of spore extracts followed by immunoblotting showed that none of the Mabs could detect this epitope and so ep117 may be conformational in nature . Thus, the conformation of any particular epitope recognized by a Mab may be important in determining to which nematode a particular spore will attach . The distribution of a particular epitope within a population of spores will in turn therefore determine its virulence on a particular nematode.

J Dairy Sci, 1997 Aug, 80(8), 1651 - 5
Malate content of forage varieties commonly fed to cattle; Callaway TR et al.; The objective of this study was to determine the concentration of malate in forage varieties at different stages of maturity . Five alfalfa varieties (Alfagraze, Apollo Supreme Cimarron, Crockett, and Magnum III) and three bermudagrass varieties (Coastal, Tifton-78, and Tifton-85) were collected at different stages of maturity . Samples were collected from replicate plots (n = 3) of each alfalfa variety at 9, 18, 28, 35, and 42 d of maturity; bermudagrass hay samples were composited from six bales of each variety from two cuttings staged to be harvested at 27 and 41 d of maturity . Malate was extracted from the samples and quantitated by high performance liquid chromatography using an organic acid column . As maturity increased, the concentration of malate declined in both plant species . Concentrations of malate were numerically higher in two alfalfa varieties (Crockett and Magnum III) at 35 and 42 d of maturity than in all other alfalfa varieties . Concentrations of malate in bermudagrass at 41 d of maturity were lower than concentrations of malate in all alfalfa varieties at 42 d of maturity . Malate declined as maturity increased in the Coastal and Tifton-78 varieties . Because malate stimulates the utilization of lactate by the predominant ruminal bacterium Selenomonas ruminantium, some of the benefits associated with alfalfa in the diets of dairy cattle may be due to the malate in this forage.

Microbiology, 1997 Aug, 143 ( Pt 8), 2725 - 31
Expression of the eicosapentaenoic acid synthesis gene cluster from Shewanella sp . in a transgenic marine cyanobacterium, Synechococcus sp; Takeyama H et al.; The eicosapentaenoic acid (EPA) synthesis gene cluster isolated from a marine bacterium, Shewanella putrefaciens strain SCRC-2738, was cloned and expressed in the marine cyanobacterium Synechococcus sp . A broad-host-range cosmid vector, pJRD215 (10.2 kb, Smr Kmr), was used to clone a 38 kb insert, pEPA, containing the EPA synthesis gene cluster, creating plasmid pJRDEPA (approx . 48 kb) . This plasmid was transferred to the cyanobacterial host at a frequency of 2.2 x 10(-7) . Cyanobacterial transconjugants grown at 29 degrees C produced 0.12 mg EPA (g dry weight)-1, whereas those grown at 23 degrees C produced 0.56 mg EPA (g dry weight)-1 . The yield was further improved to 0.64 mg (g dry weight)-1 by incubation for 1 d at 17 degrees C . This is believed to be the first successful cloning and expression of such a large heterologous gene cluster in a marine cyanobacterium.

J Bacteriol, 1997 Aug, 179(16), 5094 - 103
Domain-swapping analysis of FtsI, FtsL, and FtsQ, bitopic membrane proteins essential for cell division in Escherichia coli; Guzman LM et al.; FtsI, FtsL, and FtsQ are three membrane proteins required for assembly of the division septum in the bacterium Escherichia coli . Cells lacking any of these three proteins form long, aseptate filaments that eventually lyse . FtsI, FtsL, and FtsQ are not homologous but have similar overall structures: a small cytoplasmic domain, a single membrane-spanning segment (MSS), and a large periplasmic domain that probably encodes the primary functional activities of these proteins . The periplasmic domain of FtsI catalyzes transpeptidation and is involved in the synthesis of septal peptidoglycan . The precise functions of FtsL and FtsQ are not known . To ask whether the cytoplasmic domain and MSS of each protein serve only as a membrane anchor or have instead a more sophisticated function, we have used molecular genetic techniques to swap these domains among the three Fts proteins and one membrane protein not involved in cell division, MalF . In the cases of FtsI and FtsL, replacement of the cytoplasmic domain and/or MSS resulted in the loss of the ability to support cell division . For FtsQ, MSS swaps supported cell division but cytoplasmic domain swaps did not . We discuss several potential interpretations of these results, including that the essential domains of FtsI, FtsL, and FtsQ have a role in regulating the localization and/or activity of these proteins to ensure that septum formation occurs at the right place in the cell and at the right time during the division cycle.

Protein Sci, 1997 Aug, 6(8), 1718 - 26
Xylanase XynA from the hyperthermophilic bacterium Thermotoga maritima: structure and stability of the recombinant enzyme and its isolated cellulose-binding domain; Wassenberg D et al.; The hyperthermophilic bacterium Thermotoga maritima is capable of gaining metabolic energy utilizing xylan . XynA, one of the corresponding hydrolases required for its degradation, is a 120-kDa endo-1,4-D-xylanase exhibiting high intrinsic stability and a temperature optimum approximately 90 degrees C . Sequence alignments with other xylanases suggest the enzyme to consist of five domains . The C-terminal part of XynA was previously shown to be responsible for cellulose binding (Winterhalter C, Heinrich P, Candussio A, Wich G, Liebl W . 1995 . Identification of a novel cellulose-binding domain within the multi-domain 120 kDa Xylanase XynA of the hyperthermophilic bacterium Thermotoga maritima . Mol Microbiol 15:431-444) . In order to characterize the domain organization and the stability of XynA and its C-terminal cellulose-binding domain (CBD), the two separate proteins were expressed in Escherichia coli . CBD, because of its instability in its ligand-free form, was expressed as a glutathione S-transferase fusion protein with a specific thrombin cleavage site as linker . XynA and CBD were compared regarding their hydrodynamic and spectral properties . As taken from analytical ultracentrifugation and gel permeation chromatography, both are monomers with 116 and 22 kDa molecular masses, respectively . In the presence of glucose as a ligand, CBD shows high intrinsic stability . Denaturation/renaturation experiments with isolated CBD yield > 80% renaturation, indicating that the domain folds independently . Making use of fluorescence emission and far-UV circular dichroism in order to characterize protein stability, guanidine-induced unfolding of XynA leads to biphasic transitions, with half-concentrations c1/2 (GdmCl) approximately 4 M and > 5 M, in accordance with the extreme thermal stability . At acid pH, XynA exhibits increased stability, indicated by a shift of the second guanidine-transition from 5 to 7 M GdmCl . This can be tentatively attributed to the cellulose-binding domain . Differences in the transition profiles monitored by fluorescence emission and dichroic absorption indicate multi-state behavior of XynA . In the case of CBD, a temperature-induced increase in negative ellipticity at 217 nm is caused by alterations in the environment of aromatic residues that contribute to the far-UV CD in the native state.

Biophys J, 1997 Aug, 73(2), 972 - 82
Coupling of cytochrome and quinone turnovers in the photocycle of reaction centers from the photosynthetic bacterium Rhodobacter sphaeroides; Osvath S et al.; A minimal kinetic model of the photocycle, including both quinone (Q-6) reduction at the secondary quinone-binding site and (mammalian) cytochrome c oxidation at the cytochrome docking site of isolated reaction centers from photosynthetic purple bacteria Rhodobacter sphaeroides, was elaborated and tested by cytochrome photooxidation under strong continuous illumination . The typical rate of photochemical excitation by a laser diode at 810 nm was 2.200 s-1, and the rates of stationary turnover of the reaction center (one-half of that of cytochrome photooxidation) were 600 +/- 70 s-1 at pH 6 and 400 +/- 50 s-1 at pH 8 . The rate of turnover showed strong pH dependence, indicating the contribution of different rate-limiting processes . The kinetic limitation of the photocycle was attributed to the turnover of the cytochrome c binding site (pH < 6), light intensity and quinone/quinol exchange (6 < pH < 8), and proton-coupled second electron transfer in the quinone acceptor complex (pH > 8) . The analysis of the double-reciprocal plot of the rate of turnover versus light intensity has proved useful in determining the light-independent (maximum) turnover rate of the reaction center (445 +/- 50 s-1 at pH 7.8).

Appl Environ Microbiol, 1997 Aug, 63(8), 3010 - 3
Positive selection systems for discovery of novel polyester biosynthesis genes based on fatty acid detoxification; Kranz RG et al.; The photosynthetic bacterium Rhodobacter capsulatus can grow with short- to long-chain fatty acids as the sole carbon source (R . G . Kranz, K . K . Gabbert, T . A . Locke, and M . T . Madigan, Appl . Environ . Microbiol . 63:3003-3009, 1997) . Concomitant with growth on fatty acids is the production to high levels of the polyester storage compounds called polyhydroxyalkanoates (PHAs) . Here, we describe colony screening and selection systems to analyze the production of PHAs in R . capsulatus . A screen with Nile red dissolved in acetone distinguishes between PHA producers and nonproducers . Unlike the wild type, an R . capsulatus PhaC- strain with the gene encoding PHA synthase deleted is unable to grow on solid media containing high concentrations of certain fatty acids . It is proposed that this deficiency is due to the inability of the PhaC- strain to detoxify the surrounding medium by consumption of fatty acids and their incorporation into PHAs . This fatty acid toxicity phenotype is used in selection for the cloning and characterization of heterologous phaC genes.

J Bacteriol, 1997 Aug, 179(15), 4942 - 5
Structural and functional analysis of the phosphoenolpyruvate carboxylase gene from the purple nonsulfur bacterium Rhodopseudomonas palustris No . 7; Inui M et al.; The ppc gene, encoding phosphoenolpyruvate carboxylase (PEPC), from Rhodopseudomonas palustris No . 7 was cloned and sequenced . Primer extension analysis identified a transcriptional start site 42 bp upstream of the ppc initiation codon . An R . palustris No . 7 PEPC-deficient strain showed a slower doubling time compared with the wild-type strain either anaerobically in the light or aerobically in the dark, when pyruvate was used as a carbon source.

Arch Microbiol, 1997 Aug, 168(2), 136 - 42
Isolation of O-demethylase, an ether-cleaving enzyme system of the homoacetogenic strain MC; Kaufmann F et al.; The O-demethylase of the methylotrophic homoacetogenic bacterium strain MC was purified to apparent homogeneity . The enzyme system consisted of four different components that were designated A, B, C, and D according to their elution sequence from the anionic-exchange chromatography column . All four components were essentially required for catalysis of the transfer of the methyl group from phenyl methyl ethers to tetrahydrofolate . According to gel filtration and SDS-PAGE, components A and B were monomers with apparent molecular masses of approximately 26 kDa (subunit 25 kDa) and 36 (subunit 41 kDa), respectively; component C appeared to be a trimeric protein (195 kDa, subunit 67 kDa); and component D was probably a dimer (64 kDa, subunit 30 kDa) . Component A contained one corrinoid per monomer . In crude extracts, component D appeared to be the rate-limiting protein for the complete methyl transfer reaction . Additional requirements for the reaction were ATP and low-potential reducing equivalents supplied by either titanium(III) citrate or H2 plus hydrogenase purified from strain MC.

Infect Immun, 1997 Aug, 65(8), 3132 - 7
Identification and characterization of a metalloprotease activity from Helicobacter pylori; Windle HJ et al.; Helicobacter pylori produces a metalloprotease with a native molecular size of approximately 200 kDa, as determined by size-exclusion chromatography . Subcellular distribution studies demonstrated that the activity was associated with the outer membrane fraction of the bacterium . In addition, the protease was secreted by the bacterium when grown in liquid culture . The enzyme activity was measured by hydrolysis of azocasein and biotinylated casein and exhibited optimal caseinolytic activity at pH 8.0 (37 degrees C) . The activity was inhibited by EDTA, 1,10-phenanthroline, phosphoramidon, pyridine-2,6-dicarboxylic acid, and 8-hydroxyquinoline-5-sulfonic acid (HQSA) . Inhibition by HQSA was reversed by zinc, whereas inhibition due to EDTA was reversed by excess calcium, thus indicating that the enzyme was a zinc-dependent, calcium-stabilized endoproteinase . Furthermore, titration with Zn2+ of a desalted, active-site zinc-chelated preparation of the protease demonstrated that Zn2+ was essential for activity . Leupeptin, phenylmethylsulfonyl fluoride, E-64, pepstatin A, dithiothreitol, and 2-mercaptoethanol had no effect on enzymatic activity . Addition of Ca2+ or Mg2+ to the incubation medium resulted in approximately a twofold stimulation of the azocaseinolytic activity of the enzyme . The protease was stably expressed since it was active even after repeated subculture of the bacterium . Bovine serum albumin, hide powder azure, and elastin-Congo red remained intact even after prolonged exposure to the enzyme . The surface expression of this metalloprotease activity raises the possibility that this enzyme may be involved in the proteolysis of a variety of host proteins in vivo and thereby contributes to gastric pathology.

J Clin Microbiol, 1997 Aug, 35(8), 1988 - 95
Phylogenetic placement and characterization of a new alpha-2 proteobacterium isolated from a patient with sepsis; Blomqvist G et al.; An alpha-2 proteobacterium, previously unknown as determined by its phylogenetic characteristics and the DNA sequence of its 16S rRNA gene, was isolated from a patient who presented an unusual clinical picture, including high remitting fever and multiorgan involvement . The bacterium was detected in multiple plasma samples, obtained during the acute phase of the disease, after cocultivation in cell culture media . Electron microscopy of the organism showed a three-layer laminar cell wall and electron-dense granules within the cytoplasm, as well as a polar flagellum . By means of PCR followed by sequencing of amplified 16S ribosomal DNA fragments, the bacterium was found to differ from all species for which ribosomal sequence information is available . It is here provisionally named the Rasbo bacterium . At a subsequent relapse, the bacterium was identified in pericardial fluid both by PCR/sequencing and by direct electron microscopy . At a second relapse, it was again cultured from plasma . After in vitro adaptation to solid media, the MICs of various antibiotics could be determined . A transient immunoglobulin M (IgM) but no IgG response to the bacterium was found by an indirect immunofluorescence test, as well as by an immobilization test during the acute phase of the disease.

Biochemistry, 1997 Jul 29, 36(30), 9267 - 72
Viscosity dependence of the electron transfer rate from bound cytochrome c to P840 in the photosynthetic reaction center of the green sulfur bacterium Chlorobium tepidum; Oh-oka H et al.; Anomalous high viscosity dependence was found in the rate of reaction between the bound cytochrome c and the primary donor bacteriochlorophyll dimer (P840) of the reaction center complex purified from the green sulfur bacterium Chlorobium tepidum . The cytochrome has a primary structure with the N-terminal three membrane-spanning helices connected to the extended C-terminal heme-containing hydrophilic moiety . The rate constant of the reaction decreased from 5.0 x 10(3) s-1 to 1.0 x 10 s-1 as the glycerol concentration increased from 0 to 60% (v/v) at 295 K, showing a linear dependence on the -2.4th power of the specific viscosity . The glycerol effect was fully reversible . The extraordinary high viscosity dependence cannot be explained by the simple diffusive Brownian fluctuation model and suggests that the electron transfer mechanism is dependent on the unique conformational fluctuations of the heme-containing moiety of cytochrome c.

Biochim Biophys Acta, 1997 Jul 19, 1336(1), 28 - 32
N-acyl amino acid biosynthesis in marine bacterium, Deleya marina; Yagi H et al.; We reported previously that the marine bacterium, Deleya marina (ATCC 25374), produced N-acyl leucine and isoleucine, in which nonhydroxy fatty acid was linked to alpha-amino group of amino acid . Further analysis of bacterium lipids revealed the additional production of N-acyl ornithine . The N-acyl ornithine had a 3-hydroxy fatty acid linked by an amide bond to a-amino group of ornithine and a nonhydroxy fatty acid esterified to the hydroxy group of the 3-hydroxy fatty acid . N-acyl ornithine was located in the cell membrane and N-acyl leucine and isoleucine in cytoplasm . N-acyl ornithine is thought to be a functional analogue of phosphatidylethanolamine (PE) because of their similar structure . PE replacement into N-acyl ornithine in the cell membrane under phosphate-limited conditions was observed with other bacteria, so we anticipated the nonbiosynthesis of N-acyl ornithine under phosphate-sufficient conditions . We did not anticipate that N-acyl leucine and isoleucine in cytoplasm, whose structure is dissimilar to that of PE, would be replaced into PE in the cell membrane . Neither N-acyl leucine, N-acyl isoleucine, nor N-acyl ornithine was biosynthesized under phosphate-sufficient condition . Thus, we report here for the first time that N-acyl amino acids in cytoplasm were not biosynthesized under phosphate-sufficient conditions.

Ugeskr Laeger, 1997 Jul 7, 159(28), 4400 - 1
{Erysipelothrix rhusiopathiae bacteremia after dog bite}; Abedini S et al.; A case of erysipeloid with bacteraemia caused by Erysipelothrix rhusiopathiae (ER) in a previously healthy 41-year old man is presented . The bacterium was probably introduced by the bite of a dog . He was treated successfully with penicillin V . The ER bacteraemia occurred without complications of endocarditis.

Can J Gastroenterol, 1997 Jul-Aug, 11(5), 437 - 40
Evaluation of salivary antibodies to detect infection with Helicobacter pylori; Loeb MB et al.; Helicobacter pylori infection is an important cause of peptic ulcer disease and chronic gastritis . Infection with this bacterium stimulates the production of immunoglobulin (Ig) G antibody . Salivary IgG antibody tests to detect H pylori infection offer a convenient and noninvasive method of diagnosis . To evaluate an IgG salivary antibody kit, saliva was collected from 157 out-patients with dyspepsia referred for endoscopy to a tertiary centre . A salivary IgG ELISA antibody assay was performed using the Helisal Helicobacter pylori (IgG) assay kit, and at least four gastric biopsies were obtained . H pylori infection was confirmed by demonstration of the organism on Warthin-Starry silver stain (sensitivity 85%, specificity 55%) . The prevalence of infection with H pylori was 30% . When the analysis was redone, excluding those treated with eradication therapy, the results were similar (sensitivity 86%, specificity 58%) . The positive predictive value of the assay was 45% and the negative predictive value was 90% . Despite the ease of sampling, the assay used has limited diagnostic utility, lacking the predictive value to indicate which patients referred with dyspeptic symptoms to a tertiary care setting are infected with H pylori.

Biol Chem, 1997 Jul, 378(7), 679 - 85
The PGK-TIM fusion protein from Thermotoga maritima and its constituent parts are intrinsically stable and fold independently; Beaucamp N et al.; In the hyperthermophilic bacterium Thermotoga maritima, the two glycolytic enzymes phosphoglycerate kinase (PGK) and triosephosphate isomerase (TIM) are covalently connected forming a tetrameric single-chain PGK-TIM fusion protein . A frameshift allows the translation of PGK alone, whereas TIM activity exclusively resides in the fusion protein (Schurig et al., 1995) . Cloning the pgk-tim gene from Thermotoga maritima in Escherichia coli, yields monomeric PGK and tetrameric PGK-TIM fusion protein as authentic recombinant proteins (Beaucamp et al., 1995) . Both exhibit high intrinsic stability . The thermal transitions at approximately 80 degrees C are irreversible, rendering determination of thermodynamic data impossible . The half-concentrations, (cGdmCl)1/2, of the guanidinium-chloride induced unfolding transitions are 3.0 and 3.9 M GdmCl for PGK and the PGK-TIM fusion protein, respectively . Monitoring denaturation by activity, fluorescence emission and circular dichroism, deactivation and unfolding of the two-domain PGK is found to precede the transitions of the TIM domain . With increasing temperature, (cGdmCl)1/2 is shifted to lower denaturant concentrations; at the same time, the transitions change from bimodal to unimodal . As indicated by the incomplete reversibility of the deactivation/unfolding/dissociation transitions, misfolding, as well as wrong domain interactions seem to interfere with the correct folding and association of the bienzyme complex.

Plant J, 1997 Jul, 12(1), 133 - 42
Transformation of Arabidopsis thaliana with the codA gene for choline oxidase; accumulation of glycinebetaine and enhanced tolerance to salt and cold stress; Hayashi H et al.; Glycinebetaine is one of the compatible solutes that accumulate in the chloroplasts of contain halotolerant plants when these plants are exposed to salt or cold stress . The codA gene for choline oxidase, the enzyme that converts choline into glycinebetaine, has previously been cloned from a soil bacterium, Arthrobacter globiformis . Transformation of Arabidopsis thaliana with the cloned codA gene under the control of the 35S promoter of cauliflower mosaic virus enabled the plant to accumulate glycinebetaine and enhanced its tolerance to salt and cold stress . At 300 mM NaCl, considerable proportions of seeds of transformed plants germinated well, whereas seeds of wild-type plants failed to germinate . At 100 mM NaCl, transformed plants grew well whereas wild-type plants did not do so . The transformed plants tolerated 200 mM NaCl, which was lethal to wild-type plants . After plants had been incubated with 400 mM NaCl for two days, the photosystem II activity of wild-type plants had almost completely disappeared, whereas that of transformed plants remained at more than 50% of the original level . When exposed to a low temperature in the light, leaves of wild-type plants exhibited symptoms of chlorosis, whereas those of transformed plants did not . These observations demonstrate that the genetic modification of Arabidopsis thaliana that allowed it to accumulate glycinebetaine enhanced its ability to tolerate salt and cold stress.

Acta Cytol, 1997 Jul-Aug, 41(4), 1178 - 82
A triple stain for the detection of Helicobacter pylori in gastric brushing cytology; Ghoussoub RA et al.; OBJECTIVE: Helicobacter pylori is a surface bacterium associated with gastritis . The triple stain (TS), combining silver, hematoxylin and eosin, and alcian blue at pH 2.5, is valuable in the detection of H pylori in tissue . We evaluated the usefulness of TS as compared with Papanicolaou stain in detecting H pylori in gastric brushings and biopsy specimens . STUDY DESIGN: Gastric brushings and biopsy specimens were obtained from 21 patients . The Papanicolaou-stained slides were restained with TS, and the brushings and biopsy specimens were independently evaluated by the authors . RESULTS: H pylori was found in 9 of 21 randomly selected cases using TS . Of these, two cases were positive on brushings alone and three on biopsy alone . Only three of the nine cases were positive by Papanicolaou stain . None of the 21 samples were positive with Papanicolaou stain yet negative with TS . Detection of H pylori was restricted to patients with gastric ulcerations but was not limited by paucity of glands . CONCLUSION: TS is useful for detecting H pylori in gastric brushings and provides excellent cytologic detail . Furthermore, TS is superior to Papanicolaou stain for H pylori detection and is a valuable adjunct to biopsy.

Microbiology, 1997 Jul, 143 ( Pt 7), 2373 - 9
An outer-membrane porin inducible by short-chain amides and urea in the methylotrophic bacterium Methylophilus methylotrophus; Mills J et al.; The fmdA and fmdB genes encoding formamidase and a putative regulatory protein, respectively, from the methylotrophic bacterium Methylophilus methylotrophus were recloned with additional flanking DNA (pSW1) . fmdC, encoding a weakly hydrophilic protein containing an N-terminal signal sequence, was identified upstream of fmdAB . The derived amino acid sequence of mature FmdC (M(r) 39204) showed that it was rich in beta-sheet and aromatic amino acids, and exhibited significant similarities to several outer-membrane porins from other bacteria . Cell fractionation studies showed that the protein was located in the outer membrane . Mature FmdC was purified and shown to consist of a single type of subunit (M(r) 40,000) with the predicted N-terminal amino acid sequence (GATISF-) . SDS-PAGE and Western blotting of cells grown in continuous culture under various conditions showed that mature FmdC was induced by formamide, acetamide and urea, repressed by excess ammonia, and over-expressed during prolonged growth under formamide limitation . It is concluded that mature FmdC is a porin involved in the transport of short-chain amides and urea through the outer membrane of M . methylotrophus under conditions where these nitrogen sources are present at very low concentration.

Int J Syst Bacteriol, 1997 Jul, 47(3), 899 - 903
Characterization of some Actinomyces-like isolates from human clinical specimens: reclassification of Actinomyces suis (Soltys and Spratling) as Actinobaculum suis comb . nov . and description of Actinobaculum schaalii sp . nov; Lawson PA et al.; Five strains of a hitherto unknown Actinomyces-like bacterium were isolated from human clinical sources, including blood cultures . Biochemical and chemotaxonomic characterization indicated that the strains were distinct from previously described Actinomyces and Arcanobacterium species . A comparative 16S rRNA gene sequence analysis demonstrated that the undescribed strains constitute a new subline within the Actinomyces-Arcanobacterium species complex . The closest known relative of the isolates was found to be Actinomyces suis, although a 16S rRNA sequence divergence value of approximately 6% clearly demonstrated that the unknown bacterium represents a distinct species . Based on the results of the present and earlier phylogenetic investigations, it is proposed that Actinomyces suis should be reclassified in a new genus, the genus Actinobaculum, as Actinobaculum suis comb . nov . In addition, a new species, Actinobaculum schaalii, is proposed for the Actinomyces-like bacterium from human sources . The type strain of Actinobaculum schaalii is CCUG 27420.

Int J Syst Bacteriol, 1997 Jul, 47(3), 635 - 9
Comparison of the 16S ribosomal DNA sequences from the intracellular agents of proliferative enteritis in a hamster, deer, and ostrich with the sequence of a porcine isolate of Lawsonia intracellularis; Cooper DM et al.; Proliferative enteritis is an enteric disease that affects a variety of animals . The causative agent in swine has been determined to be an obligate intracellular bacterium, Lawsonia intracellularis, related to the sulfate-reducing bacterium Desulfovibrio desulfuricans . The intracellular agents found in the lesions of different animal species are antigenically similar . In addition, strains from the pig, ferret, and hamster have been shown to be genetically similar . In this study we performed a partial 16S ribosomal DNA sequence analysis on the intracellular agent of proliferative enteritis from a hamster, a deer, and an ostrich and compared these sequences to that of the porcine L . intracellularis isolate . Results of this study indicate that the intracellular agents from these species with proliferative enteritis have high sequence similarity, indicating that they are all in the genus Lawsonia and that they may also be the same species, L . intracellularis.

Int J Syst Bacteriol, 1997 Jul, 47(3), 627 - 34
Helicobacter rodentium sp . nov., a urease-negative Helicobacter species isolated from laboratory mice; Shen Z et al.; A spiral-shaped bacterium with bipolar, single, nonsheathed flagella was isolated from the intestines of laboratory mice . The organism grew at 37 and 42 degrees C under microaerobic and anaerobic conditions, did not hydrolyze urea, was weakly positive for catalase and oxidase, reduced nitrate to nitrite, did not hydrolyze indoxyl acetate or hippurate, and was resistant to cephalothin and nalidixic acid . This is the first urease-negative, murine Helicobacter spp . isolated from intestines . Also, Helicobacter pullorum and this bacterium are unique among the genus Helicobacter in having nonsheathed flagella . The new bacterium appears to be part of the normal intestinal flora; although its pathogenic potential is unknown, this organism was also isolated from scid mice with diarrhea that were co-infected with Helicobacter bilis . On the basis of 16S rRNA gene sequence analysis data and biochemical and phenotypic criteria, the new organism is classified as a novel helicobacter, for which we propose the name Helicobacter rodentium . The type strain is MIT 95-1707 (= ATCC 700285).

Appl Environ Microbiol, 1997 Jul, 63(7), 2625 - 30
Nested PCR for ultrasensitive detection of the potato ring rot bacterium, Clavibacter michiganensis subsp . sepedonicus; Lee IM et al.; Oligonucleotide primers derived from sequences of the 16S rRNA gene (CMR16F1, CMR16R1, CMR16F2, and CMR16R2) and insertion element IS1121 of Clavibacter michiganensis subsp . sepedonicus (CMSIF1, CMSIR1, CMSIF2, and CMISR2) were used in nested PCR to detect the potato ring rot bacterium C . michiganensis subsp . sepedonicus . Nested PCR with primer pair CMSIF1-CMSIR1 followed by primer pair CMSIF2-CMSIR2 specifically detected C . michiganensis subsp . sepedonicus, while nested PCR with CMR16F1-CMR16R1 followed by CMR16F2-CMR16R2 detected C . michiganensis subsp . sepedonicus and the other C . michiganensis subspecies . In the latter case, C . michiganensis subsp . sepedonicus can be differentiated from the other subspecies by restriction fragment length polymorphism (RFLP) analyses of the nested PCR products (16S rDNA sequences) . The nested PCR assays developed in this work allow ultrasensitive detection of very low titers of C . michiganensis subsp . sepedonicus which may be present in symptomiess potato plants or tubers and which cannot be readily detected by direct PCR (single PCR amplification) . RFLP analysis of PCR products provides for an unambiguous confirmation of the identify of C . michiganensis subsp . sepedonicus.

Arch Microbiol, 1997 Jul, 168(1), 16 - 23
Thiorhodococcus minus, gen . nov., sp . nov., A new purple sulfur bacterium isolated from coastal lagoon sediments; Guyoneaud R et al.; A new marine phototrophic purple sulfur bacterium (strain CE2203) was isolated in pure culture from a man-made coastal lagoon located on the Atlantic coast (Arcachon Bay, France) . Single cells were coccus-shaped, did not contain gas vesicles, and were highly motile . Intracellular photosynthetic membranes were of the vesicular type . Bacteriochlorophyll a and carotenoids of the normal spirilloxanthin series were present as photosynthetic pigments . Hydrogen sulfide, thiosulfate, elemental sulfur, and molecular hydrogen were used as electron donors during photolithotrophic growth under anoxic conditions, while carbon dioxide was utilized as carbon source . Acetate, propionate, lactate, glycolate, pyruvate, fumarate, succinate, fructose, sucrose, ethanol, and propanol were photoassimilated in the presence of hydrogen sulfide . During growth on sulfide, elemental sulfur globules were stored inside the cells . Chemotrophic growth under microoxic conditions in the dark was possible . The DNA base composition was 66.9 mol% G+C . Comparative sequence analysis of the 16S rRNA gene confirmed the membership of strain CE2203 in the family Chromatiaceae . Morphological characteristics of strain CE2203 indicated a close affiliation to the genera Thiocystis and Thiocapsa . However, the phylogenetic treeing revealed no closer relationship to Thiocystis spp . than to Thiocapsa roseopersicina or other known members of the Chromatiaceae . Consequently, strain CE2203 is proposed as the type strain of a new genus and species, Thiorhodococcus minus gen . nov., sp . nov.

J Bacteriol, 1997 Jul, 179(13), 4190 - 4
Uridylylation of the P(II) protein in the photosynthetic bacterium Rhodospirillum rubrum; Johansson M et al.; The regulatory protein P(II) has been studied in great detail in enteric bacteria; however, its function in photosynthetic bacteria has not been clearly established . As a number of these bacteria have been shown to regulate nitrogenase activity by a metabolic control system, it is of special interest to establish the role of P(II) in these diazotrophs . In this study, we show that P(II) in Rhodospirillum rubrum is modified in response to the N status in the cell and that addition of ammonium or glutamine leads to demodification . We also provide evidence that P(II) is uridylylated . In addition, we show that not only these compounds but also NAD+ promotes demodification of P(II), which is of particular interest as this pyridine nucleotide has been shown to act as a switch-off effector of nitrogenase . Demodification of P(II) by ammonium or NAD+ did not occur in cultures treated with an inhibitor of glutamine synthetase (methionine sulfoximine), whereas treatment with the glutamate synthase inhibitor 6-diazo-5-oxo-norleucine led to total demodification of P(II) without any other addition . The results indicate that P(II) probably is not directly involved in darkness switch-off of nitrogenase but that a role in ammonium switch-off cannot be excluded.

Infect Immun, 1997 Jul, 65(7), 2914 - 24
Differences in the association of Chlamydia trachomatis serovar E and serovar L2 with epithelial cells in vitro may reflect biological differences in vivo; Davis CH et al.; Chlamydia trachomatis serovar E is one of the most common bacterial sexually transmitted pathogens . Since it is an obligate intracellular bacterium, efficient colonization of genital mucosal epithelial cells is crucial to the infectious process . Serovar E elementary bodies (EB) metabolically radiolabeled with 35S-Cys-Met and harvested from microcarrier bead cultures, which significantly improves the infectious EB-to-particle ratio, provided a more accurate picture of the parameters of attachment of EB to human endometrial epithelial cells (HEC-1B) than did less infectious 14C-EB harvested from flask cultures . Binding of serovar E EB was (i) equivalent at 35 and 4 degrees C, (ii) decreased by preexposure of EB to heat or the topical microbicide C31G, (iii) comparable among common eukaryotic cell lines (HeLa, McCoy), and (iv) significantly increased to the apical surfaces of polarized cells versus nonpolarized cells . In parallel experiments with C . trachomatis serovar L2, serovar E attachment was not affected by heparin or heparan sulfate whereas these glucosaminoglycans dramatically reduced serovar L2 attachment . These data were confirmed by competitive inhibition of serovar E binding and infectivity by excess unlabeled live and UV-inactivated serovar E EB but not by excess serovar L2 EB . The noninvasive serovar E strains in the lumen of the genital tract enter and exit the apical domains of target columnar epithelial cells to spread canalicularly in an ascending fashion from the lower to the upper genital tract . In contrast, the invasive serovar L2 strains are primarily submucosal pathogens and likely use the glucosaminoglycans concentrated in the extracellular matrix to colonize the basolateral domains of mucosal epithelia to perpetuate the infectious process.

Infect Immun, 1997 Jul, 65(7), 2890 - 7
Anti-Ehrlichia chaffeensis antibody complexed with E . chaffeensis induces potent proinflammatory cytokine mRNA expression in human monocytes through sustained reduction of IkappaB-alpha and activation of NF-kappaB; Lee EH et al.; Ehrlichia chaffeensis is an obligatory intracellular bacterium that infects monocytes and macrophages and is the etiologic agent of human ehrlichiosis in the United States . Our previous studies showed that the exposure of human monocytes to E . chaffeensis induces the expression of interleukin-1beta (IL-1beta), IL-8, and IL-10 genes in vitro but not the expression of tumor necrosis factor alpha (TNF-alpha) and IL-6 mRNAs . In this study, the effect of anti-E . chaffeensis antibody complexed with E . chaffeensis on the expression of major proinflammatory cytokines in human monocytes was examined . Human monocytic cell line THP-1 was treated with E . chaffeensis which had been preincubated with human anti-E . chaffeensis serum for 2 h, and the levels of cytokine mRNAs were evaluated by competitive reverse transcription-PCR . Anti-E . chaffeensis antibody complexed with E . chaffeensis significantly enhanced mRNA expression of IL-1beta in THP-1 cells . The expression of TNF-alpha and IL-6 mRNAs was also induced . The levels of secreted IL-1beta, TNF-alpha, and IL-6 during 24 h of stimulation were comparable to those induced by Escherichia coli lipopolysaccharide at 1 microg/ml . Fab fragment of anti-E . chaffeensis immunoglobulin G complexed with E . chaffeensis did not induce any of these three cytokines, indicating that ehrlichial binding is required for IL-1beta mRNA expression and that binding of the immune complex to the Fc gamma receptor is required for TNF-alpha and IL-6 mRNA expression and enhanced IL-1beta mRNA expression . Furthermore, prolonged degradation of IkappaB-alpha and activation of NF-kappaB were demonstrated in THP-1 cells exposed to anti-E . chaffeensis serum and E . chaffeensis . This result implies that development of anti-E . chaffeensis antibody in patients can result in the production of major proinflammatory cytokines, which may play an important role in the pathophysiology of ehrlichiosis and immune responses to it.

Infect Immun, 1997 Jul, 65(7), 2668 - 75
Antigenic characterization and analysis of the human immune response to outer membrane protein E of Branhamella catarrhalis; Bhushan R et al.; Outer membrane protein E (OMP E) is a 50-kDa major OMP of Branhamella catarrhalis . Polyclonal antisera and four monoclonal antibodies (MAbs) to OMP E were generated to study its antigenic structure . All antibodies recognized epitopes in all 19 B . catarrhalis strains tested by immunoblot assays . By flow cytometry, it was determined that MAbs 1B3 and 9G10d recognized epitopes which are expressed on the surface of the intact bacterium, while MAbs IC11 and 7C10 recognized epitopes which were buried within the outer membrane . A competitive enzyme-linked immunosorbent assay showed that MAbs 1B3 and 9G10d recognize the same or closely related epitopes . Proteinase K treatment of whole bacterial cells revealed that MAbs 1B3 and 9G10d recognize a surface-exposed epitope located in the 17-kDa region towards the amino terminus of OMP E . The human serum and mucosal antibody responses to OMP E in adults with chronic bronchitis were studied . A majority of these patients had immunoglobulin A to OMP E in sputum supernatants . None of ten adults who experienced lower respiratory tract infections due to B . catarrhalis demonstrated a clear-cut rise in antibody titer to OMP E in serum or sputum supernatant . This study has demonstrated that OMP E has at least one surface-exposed epitope which is highly conserved among strains of B . catarrhalis and which is located in the amino-terminal 184 amino acids of the molecule.

J Clin Microbiol, 1997 Jul, 35(7), 1696 - 700
Development of a PCR assay for rapid diagnosis of Mycobacterium ulcerans infection; Ross BC et al.; The diagnosis of Mycobacterium ulcerans infection is hampered by the slow growth of the bacterium in culture, resulting in a delay of several months before a specific diagnosis can be obtained . In addition, M . ulcerans cannot be isolated from water even when there is convincing epidemiological evidence implicating this as the source of infection . The aim of the present study was to develop a PCR assay to circumvent the problems of delayed diagnosis and insensitivity of standard bacterial culture for M . ulcerans . For the PCR, we isolated an M . ulcerans-specific DNA fragment, 1,109 bp long, which is repeated at least 50 times throughout the genome . Use of this sequence as a target for PCR allowed us to detect as few as 2 molecules of genomic DNA in vitro . The PCR was used to detect M . ulcerans DNA in fresh tissue and paraffin-embedded sections from all seven patients with culture-confirmed cases of infection.

Curr Microbiol, 1997 Jul, 35(1), 22 - 7
Construction of a Fibrobacter succinogenes genomic map and demonstration of diversity at the genomic level; Ogata K et al.; The genomic cleavage map of the type strain Fibrobacter succinogenes S85 was constructed . The restriction enzymes AscI, AvrII, FseI, NotI, and SfiI generated DNA fragments of suitable size distribution that could be resolved by pulsed-field gel electrophoresis (PFGE) . An average genome size of 3.6 Mb was obtained by summing the total fragment sizes . The linkages between the 15 AscI fragments of the genome were determined by combining two approaches: isolation of linking clones and cross-hybridization of restriction fragments . The genome of F . succinogenes was found to be represented by the single circular DNA molecule . Southern hybridization with specific probes allowed the eight genetic markers to be located on the restriction map . The genome of this bacterium contains at least three rRNA operons . PFGE of the other three strains of F . succinogenes gave estimated genome sizes close to that of the type strain . However, RFLP patterns of these strains generated by AscI digestion are completely different . Pairwise comparison of the genomic fragment distribution between the type strain and the three isolates showed a similarity level in the region of 14.3% to 31.3% . No fragment common to all of these F . succinogenes strains could be detected by PFGE . A marked degree of genomic heterogeneity among members of this species makes genomic RFLP a highly discriminatory and useful molecular typing tool for population studies.

Cell, 1997 Jun 27, 89(7), 1111 - 9
Plasminogen is required for efficient dissemination of B . burgdorferi in ticks and for enhancement of spirochetemia in mice; Coleman JL et al.; The role of the host plasminogen activation system in transmission of and invasion by Borrelia burgdorferi, the tick-borne spirochetal agent of Lyme disease, was investigated using plasminogen (Plg)-knockout mice . PLG was not detected in spirochetes from unfed ticks, but binding occurred as ticks fed on the host's blood . Plasminogen activators were derived from the host blood meal . PLG was required for efficient dissemination of B . burgdorferi within the tick and for enhancement of spirochetemia in mice but was not critical for transmission and infection . These results provide evidence for a bacterium using a vertebrate protease to disseminate in an invertebrate vector and underscores the interplay among vector, pathogen, and host in promoting the life cycle and disease.

J Biol Chem, 1997 Jun 27, 272(26), 16256 - 61
Functional analysis of nodulin 26, an aquaporin in soybean root nodule symbiosomes; Rivers RL et al.; Upon infection of soybean roots, nitrogen-fixing bacteria become enclosed in a specific organelle known as the symbiosome . The symbiosome membrane (SM) is a selectively permeable barrier that controls metabolite flux between the plant cytosol and the symbiotic bacterium inside . Nodulin 26 (NOD 26), a member of the aquaporin (AQP) water channel family, is a major protein component of the SM . Expression of NOD 26 in Xenopus oocytes gave a mercury-sensitive increase in osmotic water permeability (Pf) . To define the biophysical properties of NOD 26 water channels in their native membranes, symbiosomes were isolated from soybean root nodules and the SM separated as vesicles from the bacteria . Permeabilities were measured using stopped-flow fluorimetry in SM vesicles with entrapped carboxyfluorescein . Osmotic water permeability (Pf) of SM was high, with a value of 0.05 +/- 0.003 cm/s observed at 20 degrees C (mean +/- S.E.; n = 15) . Water flow exhibited a low activation energy, was inhibited by HgCl2 (0.1 mM), and exhibited a unit conductance of 3.2 +/- 1.3 x 10(-15) cm3/s, a value 30-fold lower than that of AQP 1, the red blood cell water channel . Diffusive water permeability (Pd) was 0.0024 +/- 0.0002 cm/s, and the resulting Pf to Pd ratio was 18.3, indicating that water crosses the SM in single file fashion via the NOD 26 water channel . In addition to high water permeability, SM vesicles also show high mercury-sensitive permeability to glycerol and formamide, but not urea, suggesting that NOD 26 also fluxes these solutes . Overall, we conclude that NOD 26 acts as a water channel with a single channel conductance that is 30-fold lower than AQP 1 . Because the solutes that permeate NOD 26 are far larger than water, and water appears to cross the channel via a single file pathway, solute flux across NOD 26 appears to occur by a pathway that is distinct from that for water.

J Biol Chem, 1997 Jun 20, 272(25), 16062 - 7
Activation of blood coagulation factor X by arginine-specific cysteine proteinases (gingipain-Rs) from Porphyromonas gingivalis; Imamura T et al.; The effect of two arginine-specific cysteine proteinases (gingipain Rs) from Porphyromonas gingivalis, a causative bacterium of adult periodontitis, on human blood coagulation was investigated . Activated partial thromboplastin time and prothrombin time were shortened by these proteinases, with a 95-kDa gingipain R containing adhesin domains being 5-fold more efficient in comparison to a 50-kDa gingipain R containing the catalytic domain alone . The 50-kDa enzyme reduced each coagulation time in several plasmas deficient in various coagulation factors, while it was ineffective in factor X-deficient plasma unless reconstituted with this protein . Each proteinase activated factor X in a dose- and time-dependent manner, with Michaelis constants (Km) being found to be lower than the normal plasma factor X concentration, strongly suggesting that factor X activation by gingipain Rs, especially the 95-kDa form which is strongly activated by phospholipids, could occur in plasma . This is the first report of factor X activation by bacterial proteinases and indicates that the gingipain Rs could be responsible for the production of thrombin and, indirectly, with the generation of prostaglandins, interleukin-1, etc., which have been found to be associated with the development of periodontitis induced by P . gingivalis infections . Furthermore, the data support the hypothesis that induction of blood coagulation by bacterial proteinases may be a causative agent in the pathogenesis of disseminated intravascular coagulation in sepsis.

Ann N Y Acad Sci, 1997 Jun 17, 816, 404 - 10
Sexually transmitted diseases and oral contraceptive use during adolescence; Creatsas G; Sexually transmitted diseases (STD) cause lower genital tract infections (cervicitis, vaginitis) or ascending infections of the fallopian tubes, and, possibly, pelvic inflammatory disease (PID) . The syphilis bacterium, human immunodeficiency virus (HIV), and the hepatitis virus cause systematic disease . Although oral contraceptives (OCs) are the most reliable contraceptive method, they have limited anti-STD properties and their relationship with STDs remain unclear . Various mechanisms explain a protective role of OCs against STDs; however, in no way can OCs be considered a safe anti-STD contraceptive method, when compared to specific barrier methods, which provide both contraception and anti-STD protection . The above has been confirmed by a recent study performed in our institution where 10.3% and 6.9% of OC users presented a prevalence of Chlamydia trachomatis and Mycoplasma, respectively, when compared to 0% and 4.5% infection rates found among condom users . It is concluded that although OCs possess some anti-STD properties, mainly in the prevention of PID, they should be used in combination with a barrier method.

FEBS Lett, 1997 Jun 16, 409(3), 343 - 6
A single mutation in the M-subunit of Rhodospirillum rubrum confers herbicide resistance; Sopp G et al.; Cells of the photosynthetic bacterium Rhodospirillum rubrum were rendered resistant against the inhibitor 2-(1-phenyl)ethylamino-3-propionylamino-4-cyano-thiazole (PPCTH) . Electron transport in reaction centers prepared from one of the mutants (M6) was neither inhibited by PPCTH and other NH-thiazoles nor terbutryn . These inhibitors are known to bind at the Q(B) site of the L-subunit . Compared to the wild type, chromatophores from M6 exhibited strongly altered Q(B)- Fe2+ and Q(A)- Fe2+ EPR signals . Inhibitor resistance is due to a mutation in the bacterial reaction center M-subunit, where Glu234 is exchanged against Lys . This is the first example of an inhibitor resistance in the Q(B) site caused by a mutation in the M-subunit.

Biochim Biophys Acta, 1997 Jun 12, 1326(2), 307 - 18
Outer membrane cytochromes of Shewanella putrefaciens MR-1: spectral analysis, and purification of the 83-kDa c-type cytochrome; Myers CR et al.; The metal-reducing bacterium Shewanella putrefaciens MR-1 is known to localize a majority of its membrane-bound cytochromes to its outer membrane when grown under anaerobic conditions . In this study, pyridine hemochrome spectra confirmed that these outer membrane cytochromes are c-type, and electrophoretic data demonstrated the presence of four distinct outer membrane cytochromes, with apparent molecular masses of 150, 83, 65, and 53 kDa . Fourth-order derivative analysis of 77 K spectra of the outer membrane revealed four spectrally distinct c-type hemes, with peaks at 545.4, 548.0, 550.6, and 552.6 nm . Outer membrane cytochromes in the reduced state were rapidly re-oxidized by oxidized iron and manganese, which have previously been shown to serve as electron acceptors for anaerobic respiration in this bacterium . The 83-kDa outer membrane cytochrome was purified and a specific polyclonal antibody was generated against this protein . Western blot analysis demonstrated that the vast majority of this protein was localized to the outer membrane and an intermediate density membrane fraction of similar composition . Its levels, but not its subcellular distribution, were somewhat influenced by the electron acceptor used to support anaerobic growth, with levels higher in fumarate-grown cells relative to iron(III)- or trimethylamine N-oxide-grown cells . Its specific content in cells grown under aerobic conditions was only 14% of that of fumarate-grown cells, suggesting that a switch to anaerobic conditions significantly increases the de novo synthesis of this outer membrane cytochrome.

Eur J Epidemiol, 1997 Jun, 13(4), 471 - 5
Study of the 16S-23S ribosomal DNA internal spacer of Coxiella burnetii; Stein A et al.; The complete 16S-23S ribosomal DNA (rDNA) internal transcribed spacer (ITS) of 22 isolates of the obligate intracellular bacterium Coxiella burnetii, the agent of Q fever, were amplified by the polymerase chain reaction (PCR) and sequenced using an automated laser fluorescent DNA sequencer . The ITS measured 497 base pairs (bp) and encoded isoleucine-tRNA and alanine-tRNA . The comparison of the sequence alignments of the 22 C . burnetii strains revealed very high levels of sequence similitary (> 99%) although they had different geographic origins and phenotypic characteristics . Sequencing of the 16S-23S rDNA ITS of C . burnetii could be utilized for identification of the bacterium but is not applicable to studies of epidemiology, virulence and taxonomy.

Trends Microbiol, 1997 Jun, 5(6), 224 - 8
The role of Actinobacillus actinomycetemcomitans in the pathogenesis of periodontal disease; Meyer DH et al.; Periodontal disease consists of a constellation of complex bacterium-host cell interactions . One example of these oral pathogens, Actinobacillus actinomycetemcomitans, has an arsenal of putative virulence determinants that account for its potent periodontopathogenicity . Of these determinants, invasion of host cells and leukocytotoxicity have been studied extensively.

Microbiology, 1997 Jun, 143 ( Pt 6), 1815 - 26
Targeted gene-replacement mutagenesis of dcrA, encoding an oxygen sensor of the sulfate-reducing bacterium Desulfovibrio vulgaris Hildenborough; Fu R et al.; A gene-replacement mutagenesis method has been developed for the anaerobic, sulfate-reducing bacterium Desulfovibrio vulgaris Hildenborough and used to delete dcrA, encoding a potential oxygen or redox sensor with homology to the methyl-accepting chemotaxis proteins . A suicide plasmid, containing a cat-marked dcrA allele and a counter-selectable sacB marker was transferred from Escherichia coli S17-1 to D . vulgaris by conjugation . Following plasmid integration the desired dcrA deletion mutant (D . vulgaris F100) was obtained in media containing sucrose and chloramphenicol . Southern blot screening was required to distinguish D . vulgaris F100 from strain in which the sacB marker was inactivated by transposition of an endogenous IS element . No anaerotactic deficiency has so far been detected in D . vulgaris F100, which was found to be more resistant to inactivation by oxygen that the wild-type . Increased transcription of the rbo-rub operon, located immediately downstream from dcrA, was demonstrated by Northern blotting and may be the cause of this unusual phenotype, in view of the recent discovery that Rbo can complement the deleterious effects of superoxide dismutase deficiency in E . coli.

Scand J Gastroenterol, 1997 Jun, 32(6), 617 - 22
Reversal of long-standing iron deficiency anaemia after eradication of Helicobacter pylori infection; Marignani M et al.; Helicobacter pylori has been proposed as a major determinant in multiple gastric disorders . We describe the case of a young adult with a long-standing medical history of sideropenic anaemia and of oral iron consumption dependence with a chronic superficial H . pylori-positive gastritis . All other causes of sideropenic anaemia were carefully excluded . Histology showed a peculiar pattern of non-active H . pylori-positive gastritis . The bacterium was a non-VacA-producing strain . The first attempt at eradication caused a reduction in bacterial load and led to a partial normalization of haematologic variables without improving the ferritin level . A successful second course of eradication therapy completely reversed the anaemia and restored the iron deposit, which persisted at the 29-month follow-up . H . pylori infection can be involved in unexplained cases of iron deficiency anaemia in adults, and its cure can normalize the haematologic picture.

FEMS Microbiol Lett, 1997 Jun 1, 151(1), 77 - 81
Interaction between Actinobacillus actinomycetemcomitans lipopolysaccharides and human hemoglobin; Grenier D et al.; Actinobacillus actinomycetemcomitans, a bacterium associated with juvenile periodontitis, was found to use human hemoglobin as a source of iron for cell growth . Cultivation in the presence of hemoglobin had only a slight effect on the cellular protein pattern, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis . Lipopolysaccharides obtained from A . actinomycetemcomitans were found to have a strong capacity to bind hemoglobin . This interaction appears not to involve the lipid A portion of the molecule as no inhibition was obtained when lipopolysaccharides were pre-treated with polymyxin B . This interaction between hemoglobin and lipopolysaccharides of A . actinomycetemcomitans may facilitate iron acquisition by this bacterium.

Biochem Mol Biol Int, 1997 Jun, 42(1), 155 - 61
Suppression of Helicobacter pylori urease activity by sucralfate and sulglycotide; Slomiany BL et al.; The effect of gastroprotective agents, sucralfate and sulglycotide, on the in vitro activity of H . pylori urease was investigated . The bacterium was subjected to sonication, centrifuged, and the supernatant used as an enzyme source . The assays revealed that the rate of urea degradation was proportional to enzyme protein up to 100 micrograms and remained constant with time for 10 min . Introduction of sucralfate or sulglycotide to the assay system led to the reduction in the rate of ammonia production . With both drugs the optimal inhibition was attained at 10 micrograms/ml, at which dose a 63.1% decrease in urease activity occurred with sucralfate and a 70.2% inhibition was obtained with sulglycotide . The findings demonstrate that the inhibitory action of sucralfate and sulglycotide on H . pylori urease activity may be of value in the treatment of gastric disease associated with H . pylori infection.

Biochem Mol Biol Int, 1997 Jun, 42(1), 21 - 7
Functioning of oligomeric-type light-harvesting antenna; Timpmann KE et al.; In our previous work, we developed, for the first time, a theory of excitation energy transfer within an oligomeric-type light-harvesting antenna and, in particular, within the chlorosome of green bacteria (Biophys.J., 1996, vol.71, pp.995-1010) . The theory was recently developed for a new original exciton model of aggregation of chlorosomal pigments, bacteriochlorophylls (BCh1) c/d/e (Biochem, Mol.Biol.Int., 1996, vol.40, No.2, pp . 243-252) . In this paper, it was demonstrated with picosecond fluorescence spectroscopy that this theory explains the antenna-size-dependent kinetics of fluorescence decay in chlorosomal antenna, measured for intact cells of different cultures of the green bacterium Chlorobium limicola with different chlorosomal antenna size determined by electron microscopic examination of the ultrathin sections of the cells . According to our model, the energy transfer dynamics within the chlorosome imply the formation of a cylindrical exciton, delocalized over a tubular aggregate of BCh1 c chains, and inductive-resonance-type transfer of such a cylindrical exciton between the nearest tubular BCh1 c aggregates and to BCh1 a of the chlorosome.

J Bacteriol, 1997 Jun, 179(12), 4003 - 12
Comparative characterization of SecA from the alpha-subclass purple bacterium Rhodobacter capsulatus and Escherichia coli reveals differences in membrane and precursor specificity; Helde R et al.; We have cloned the secA gene of the alpha-subclass purple bacterium Rhodobacter capsulatus, a close relative to the mitochondrial ancestor, and purified the protein after expression in Escherichia coli . R . capsulatus SecA contains 904 amino acids with 53% identity to E . coli and 54% identity to Caulobacter crescentus SecA . In contrast to the nearly equal partitioning of E . coli SecA between the cytosol and plasma membrane, R . capsulatus SecA is recovered predominantly from the membrane fraction . A SecA-deficient, cell-free synthesis-translocation system prepared from R . capsulatus is used to demonstrate translocation activity of the purified R . capsulatus SecA . This translocation activity is then compared to that of the E . coli counterpart by using various precursor proteins and inside-out membrane vesicles prepared from both bacteria . We find a preference of the R . capsulatus SecA for the homologous membrane vesicles whereas E . coli SecA is active with either type of membrane . Furthermore, the two SecA proteins clearly select between distinct precursor proteins . In addition, we show here for the first time that a bacterial c-type cytochrome utilizes the canonical, Sec-dependent export pathway.

Cancer Epidemiol Biomarkers Prev, 1997 Jun, 6(6), 387 - 400
Cancer and infection: estimates of the attributable fraction in 1990; Pisani P et al.; Various estimates of the proportion of all cancers attributable to infections have been proposed but none have been numerically justified . We have reviewed the evidence for "causality" with respect to infectious agents linked with cancer and estimated the fraction of all cancer cases concerned that are attributable to it . The etiological fraction was applied to the estimated annual incidence of cancer at each specific site in 1990, and the number of attributable cases was obtained . We estimate that 15.6% (1,450,000 cases) of the worldwide incidence of cancer in 1990 can be attributed to infection with either the hepatitis B and C viruses, the human papillomaviruses, EBV, human T-cell lymphotrophic virus I, HIV, the bacterium Helicobacter pylori, schistosomes, or liver flukes . There would be 21% fewer cases of cancer in developing countries (1,000,000 fewer cases per year) and 9% fewer cases in developed countries (375,000 fewer cases per year) if these infectious diseases were prevented . The attributable fraction at the specific sites varies from 89% of cervix cancers attributable to the papillomaviruses to 1% of all leukemias attributable to human T-cell lymphotrophic virus I.

Appl Environ Microbiol, 1997 Jun, 63(6), 2449 - 53
Catalytic properties of the cellulose-binding endoglucanase F from Fibrobacter succinogenes S85; Malburg SR et al.; The celF gene from the predominant cellulolytic ruminal bacterium Fibrobacter succinogenes encodes a 118.3-kDa cellulose-binding endoglucanase, endoglucanase F (EGF) . This enzyme possesses an N-terminal cellulose-binding domain and a C-terminal catalytic domain . The purified catalytic domain displayed an activity profile typical of an endoglucanase, with high catalytic activity on carboxymethyl cellulose and barley beta-glucan . Immunoblotting of EGF and the formerly characterized endoglucanase 2 (EG2) from F . succinogenes with antibodies prepared against each of the enzymes demonstrated that EGF and EG2 contain cross-reactive epitopes . This data in conjunction with evidence that the proteins are the same size, share a 19-residue internal amino acid sequence, possess similar catalytic properties, and both bind to cellulose allows the conclusion that celF codes for EG2.

Appl Environ Microbiol, 1997 Jun, 63(6), 2273 - 80
Cloning, sequencing, and overexpression of the Anaerobiospirillum succiniciproducens phosphoenolpyruvate carboxykinase (pckA) gene; Laivenieks M et al.; The phosphoenolpyruvate (PEP) carboxykinase-encoding gene from the anaerobic, CO2-fixing, succinate-producing bacterium Anaerobiospirillum succiniciproducens was cloned, sequenced, and expressed in Escherichia coli . The gene encoded a 532-residue polypeptide with a calculated molecular mass of 58.7 kDa . The sequence of the A . succiniciproducens PEP carboxykinase was similar to those of all known ATP/ADP-dependent PEP carboxykinases . In particular, the A . succiniciproducens enzyme was 67.3% identical and 79.2% similar to the E . coli enzyme . The A . succiniciproducens pckA transcription start site was determined, and putative promoter regions were identified . The recombinant enzyme was overexpressed in E . coli . The purified enzyme was indiscernible from the native enzyme by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and had the same activity as the native enzyme.

J Clin Microbiol, 1997 Jun, 35(6), 1348 - 52
Detection of Treponema pallidum by a sensitive reverse transcriptase PCR; Centurion-Lara A et al.; Syphilis is diagnosed by serologic testing or by identification of the causative agent, Treponema pallidum . The bacterium has historically been detected in clinical specimens by dark-field microscopy, immunostaining with polyclonal or monoclonal antibodies, or the rabbit inoculation test (RIT) . RIT is considered to be very sensitive and specific, although it is available only in research settings and is not clinically useful due to the length of time required to obtain a result . In recent years, several PCR methods have been developed for the detection of T . pallidum, but none of these has shown a clear advantage in sensitivity over RIT . We have developed a specific and highly sensitive reverse transcriptase PCR (RT-PCR) that targets a 366 bp region of the 16S rRNA of T . pallidum . This RT-PCR can detect a single organism by Southern analysis when whole organisms are diluted and 10(-2) to 10(-3) T . pallidum organisms when RNA equivalents are used to make cDNA . The test was demonstrated to detect 10(-2) T . pallidum RNA equivalents in cerebrospinal fluid . Twenty different strains of T . pallidum, isolated from cerebrospinal fluids, aqueous humor, blood, and chancres, were shown to be detectable by this test . This efficient and sensitive technique could be more useful than existing methods for detecting very low numbers of organisms in clinical samples.

Presse Med, 1997 May 17, 26(16), 761 - 3
{Biological diagnosis of whooping cough: contribution of gene amplification}; Prevel A et al.; An upsurge in pertussis infections, despite mandatory vaccination in France since 1966, has occurred again in developed countries due to progressive loss of vaccinal immunity and wider circulation of the causal bacterium, Bordetella pertussis . Unfortunately, the classical culture method is insufficiently sensitive and serology can only confirm diagnosis retrospectively . New techniques are needed for rapid diagnosis, and subsequent treatment and preventive measures . One new method, gene amplification using polymerase chain reaction (PCR), has been particularly useful in detecting Bordetella pertussis . PCR is highly specific and more sensitive than culture . It is thus quite useful in case of atypical clinical presentations and in previously treated or vaccinated patients . Less restrictions on sample transportation and preservation make PCR a technique which general practitioners can use for rapid easy diagnosis of pertussis.

EMBO J, 1997 May 15, 16(10), 2955 - 67
The human RNA 3'-terminal phosphate cyclase is a member of a new family of proteins conserved in Eucarya, Bacteria and Archaea; Genschik P et al.; RNA 3'-terminal phosphate cyclase catalyses the ATP-dependent conversion of the 3'-phosphate to a 2',3'-cyclic phosphodiester at the end of RNA . The physiological function of the cyclase is not known, but the enzyme could be involved in the maintenance of cyclic ends in tRNA splicing intermediates or in the cyclization of the 3' end of U6 snRNA . In this work, we describe cloning of the human cyclase cDNA . The purified bacterially overexpressed protein underwent adenylylation in the presence of {alpha-32P}ATP and catalysed cyclization of the 3'-terminal phosphate in different RNA substrates, consistent with previous findings . Comparison of oligoribonucleotides and oligodeoxyribonucleotides of identical sequence demonstrated that the latter are approximately 500-fold poorer substrates for the enzyme . In Northern analysis, the cyclase was expressed in all analysed mammalian tissues and cell lines . Indirect immunofluorescence, performed with different transfected mammalian cell lines, showed that this protein is nuclear, with a diffuse nucleoplasmic localization . The sequence of the human cyclase has no apparent motifs in common with any proteins of known function . However, inspection of the databases identified proteins showing strong similarity to the enzyme, originating from as evolutionarily distant organisms as yeast, plants, the bacterium Escherichia coli and the archaeon Methanococcus jannaschii . The overexpressed E . coli protein has cyclase activity similar to that of the human enzyme . The conservation of the RNA 3'-terminal phosphate cyclase among Eucarya, Bacteria and Archaea argues that the enzyme performs an important function in RNA metabolism.

Biochemistry, 1997 May 13, 36(19), 5912 - 20
Spectral equilibration and primary photochemistry in Heliobacillus mobilis at cryogenic temperature; Liebl U et al.; We performed multicolor femtosecond transient absorption measurements on membranes of the photosynthetic bacterium Heliobacillus mobilis at 20 K, by selective excitation at either the red or the blue extreme of the bacteriochlorophyll g Q(Y) band, which is split in three spectral forms (Bchl g 778, 793, and 808) at low temperature . In contrast to room temperature, there is no observable uphill energy transfer upon excitation at the red extreme . This provides a direct experimental confirmation of the expected strong temperature dependence of uphill energy transfer in multichromophore systems . Upon excitation at the blue edge, downhill energy transfer is observed on time ranges varying over 2 orders of magnitude and is discussed in terms of four distinct energy transfer processes: Bchl g 778* --> Bchl g 793* (approximately 50 fs); Bchl g 778* --> Bchl g 808* (approximately 400 fs); Bchl g 793* --> Bchl g 808* (approximately 1.4 ps); and within Bchl g 808* (approximately 7 ps) . Surprisingly, the amount of oxidized primary donor P798+ formed on the time scale of picoseconds and tens of picoseconds was found to depend on the excitation conditions: trapping occurs mainly in approximately 80 ps and slower from directly excited Bchl g 808* and can additionally occur in a few picoseconds from Bchl g 778* and Bchl g 793* upon blue excitation . This finding implies that spectral equilibration is not complete prior to charge separation and furthermore is inconsistent with a funnel model, in which P798 is surrounded by long-wavelength pigments . More generally, we discuss to what extent our data bring constraints on the spatial distribution of the different spectral forms of the pigments.

Biochim Biophys Acta, 1997 May 2, 1352(1), 18 - 22
Cloning and sequencing of the gene encoding the high potential iron-sulfur protein (HiPIP) from the purple sulfur bacterium Chromatium vinosum; Bruser T et al.; The gene encoding the high potential iron-sulfur protein (HiPIP) of Chromatium vinosum strain D (DSM 180T) was cloned from an EcoRI-HindIII digest of genomic DNA . A nucleotide sequence of 648 bp length was determined which contained the coding region and putative promoter and termination sites . The gene codes for a 122 residue 12761 Da protein . The C-terminal 85 residues are those of the previously biochemically determined sequence, whereas the N-terminal 37 residues constitute a leader peptide which shows characteristics of the double arginine signal sequences of complex cofactor containing periplasmic proteins.

Recenti Prog Med, 1997 May, 88(5), 232 - 6
{The modes of transmission of Helicobacter pylori infection}; Tursi A et al.; Helicobacter pylori plays an essential role in the development of several both acid-related and neoplastic gastroduodenal pathologies . There are still uncertainties about the transmission routes and the sources of H . pylori infection . Man is the only well established "reservoir" of H . pylori, while the role of other mammalians (cat, pig, primates), as sources of infection, is still controversy . Literature data suggest four different modalities of transmission of the infection: faeco-oral, oro-oral, gastro-oral, gastro-gastric . By faeco-oral route, the bacterium, excreted with faeces, might colonize water sources, becoming so available to be transmitted to man and other mammalians . By oro-oral route, H . pylori, which colonizes dental plaque and saliva, may be transmitted by saliva to other individuals . The gastro-oral route is the typical modality of transmission in the childhood, when H . pylori uses the mucous achlorhydric vomitus of the children to infect a new host . Finally, by gastro-gastric route the bacterium might be transmitted by endoscopic procedures . In conclusion, we believe the different modalities of transmission may be contemporaneously involved, since none per se is able to explain the widespread occurrence of H . pylori infection.

Eur J Gastroenterol Hepatol, 1997 May, 9(5), 447 - 50
Role of gastric mucosal cytokines in the immunopathogenesis of Helicobacter pylori infection: new hypotheses but still few certitudes; Glupczynski Y et al.; Since the recognition of Helicobacter pylori as a pathogen involved in chronic gastritis, peptic ulcers and gastric cancer, many studies have shown that clinical manifestations of H . pylori infection occur only in a minority of infected patients . Studies of the genomic diversity of this bacterium show relations of some bacterial characteristics with pathology . Imbalanced host response to infection may also play a major role in the clinical expression of H . pylori infection . Gastric epithelial cells are involved in the process, as well as lymphocytes and other immune cells of the underlying gastric tissue . A better understanding of the immunopathogenesis of H . pylori infection is required to understand the exact role of both the strain and the host.

Int Immunol, 1997 May, 9(5), 713 - 20
Human CD30+ cells are induced by Mycobacterium tuberculosis and present in tuberculosis lesions; Munk ME et al.; CD30 is a member of the tumor necrosis factor/nerve growth factor receptor family and evidence has been presented that activated CD4+ CD45Ro+ T cells of Th2 type selectively express CD30 . Mycobacterium tuberculosis, a facultative intracellular bacterium capable of replicating in resting macrophages, is a potent inducer of IFN-gamma secretion by Th1 cells . We find increased CD30 expression by M . tuberculosis-stimulated alpha beta and gamma delta T cells, and elevated numbers of CD30+ alpha beta T cells in tuberculosis pleuritis and affected lung tissue . Furthermore, surface CD30 was associated with intracytoplasmic IFN-gamma expression and IFN-gamma production by M . tuberculosis-stimulated alpha beta and gamma delta T cells . Thus, our results indicate that M . tuberculosis is a potent inducer of CD30 expression in Th1 cells and argue against exclusive correlation of CD30 expression with Th2 cell responses.

J Invertebr Pathol, 1997 May, 69(3), 212 - 7
Pathogenicity of axenic Steinernema feltiae, Xenorhabdus bovienii, and the bacto-helminthic complex to larvae of Tipula oleracea (Diptera) and Galleria mellonella (Lepidoptera); Ehlers RU et al.; The pathogenicity of the nematode-bacterium complex Steinernema feltiae-Xenorhabdus bovienii to larvae of Tipula oleracea and Galleria mellonella was investigated by injection of dauer juvenile nematodes carrying their bacterial symbiont cells (monoxenic nematodes) . Axenic nematodes (free of bacteria) and the symbiotic bacteria themselves were tested . The LC50 of X . bovienii in T . oleracea was 15,700 colony forming units (CFU)/larva compared to < or = 8 CFU in G . mellonella . Xenorhabdus bovienii is apparently removed from the tipulids hemolymph, possibly by cellular defense mechanisms . Axenic nematodes were less pathogenic than monoxenic nematodes for both insects . The difference was less pronounced in G . mellonella larvae: one axenic nematode was sufficient to kill 80% in 1 day . The remaining insects found dead after 50 days were developmentally arrested . In T . oleracea 20 axenic nematodes caused 39% whereas 20 monoxenic dauer juveniles caused 90% mortality within 8 days . The data indicate that the virulence of the S . feltiae/X . bovienii complex is greater than the additive effect of the nematodes and their bacteria, further evidence for the synergistic activity of the symbiotic bacto-helminthic complex during pathogenesis.

Can J Microbiol, 1997 May, 43(5), 477 - 85
An endoglucanase from the anaerobic fungus Orpinomyces joyonii: characterization of the gene and its product; Liu JH et al.; An endoglucanase gene (celA) was isolated from a genomic library of the ruminal fungus Orpinomyces joyonii . DNA sequence analysis of celA revealed an intronless gene encoding a typical signal sequence, an N-terminal catalytic domain, two repeated regions linked by a short Ser/Thr-rich linker and a domain of unknown function . The deduced amino acid sequence of the catalytic domain showed homology with the family 5 cellulases . While the catalytic domain of CelA was not homologous to the catalytic domain of the endoglucanase gene (EG3) from the ruminal bacterium Fibrobacter succinogenes, the repeated regions of CelA were very similar to the noncatalytic domain of EG3 . This suggests that evolutionary shuffling of endoglucanase domains might occur among bacteria and fungi within the anaerobic ecosystem of the rumen . The celA gene was expressed in Escherichia coli, and the periplasmic endoglucanase was used for the characterization studies of the enzyme . CelA exhibited both endoglucanase and xylanase activities . Its pH optimum was 4 and the temperature optimum was 40 degrees C . Deletion analysis showed that the repeated sequences and C-terminal domain of CelA were not required for enzyme activity.

FEMS Microbiol Lett, 1997 May 1, 150(1), 27 - 32
Genetic relationship among isolates of Helicobacter pylori: evidence for the existence of a Helicobacter pylori species-complex; Hazell SL et al.; We investigated the population genetics of 23 isolates of H . pylori by allozyme electrophoresis using 16 enzyme loci . Isolates were obtained from adult patients of whom 48% were of Greek extraction . Eight patients (35%) had an active duodenal ulcer . Allelic variation per loci ranged from 2 to 11 alleles . Four major genetic clusters were apparent, having > 75% fixed genetic differences . There was no distinct clustering (clonal structure) on the basis of the geographical origin of the persons from whom isolates were obtained, indicating that this bacterium has not recently jumped a species barrier into humans . Isolates associated with ulcer disease were not monophyletic, with isolates from ulcer patients being found in phylogenetically diverse branches of the dendogram derived from the data . Based on the genetic diversity of H . pylori isolates, we propose that isolates should be classified as belonging not to a single species but to a 'Helicobacter pylori species-complex'.

J Bacteriol, 1997 May, 179(10), 3378 - 81
DNA sequencing and expression of the formyl coenzyme A transferase gene, frc, from Oxalobacter formigenes; Sidhu H et al.; Oxalic acid, a highly toxic by-product of metabolism, is catabolized by a limited number of bacterial species utilizing an activation-decarboxylation reaction which yields formate and CO2 . frc, the gene encoding formyl coenzyme A transferase, an enzyme which transfers a coenzyme A moiety to activate oxalic acid, was cloned from the bacterium Oxalobacter formigenes . DNA sequencing revealed a single open reading frame of 1,284 bp capable of encoding a 428-amino-acid protein . A presumed promoter region and a rho-independent termination sequence suggest that this gene is part of a monocistronic operon . A PCR fragment containing the open reading frame, when overexpressed in Escherichia coli, produced a product exhibiting enzymatic activity similar to the purified native enzyme . With this, the two genes necessary for bacterial catabolism of oxalate, frc and oxc, have now been cloned, sequenced, and expressed.

J Bacteriol, 1997 May, 179(10), 3350 - 3
Na+-induced structural change of a soil bacterium, S34, and Ca2+ requirement for preserving its original structure; Mitsui H et al.; A drastic change in the outer membrane structure of a salt-sensitive soil bacterium, S34, related to the genus Deinococcus was induced by 0.2 to 0.4% (wt/vol) NaCl . The change was relieved by 6 mM CaCl2 and induced by 1 mM EGTA . The results indicate the strong dependence of the organism on calcium.

J Bacteriol, 1997 May, 179(10), 3304 - 9
A {2Fe-2S} ferredoxin (FdVI) is essential for growth of the photosynthetic bacterium Rhodobacter capsulatus; Armengaud J et al.; The physiological function of Rhodobacter capsulatus FdVI, a {2Fe-2S} ferredoxin, was investigated by the cloning, sequence analysis, and mutagenesis of its structural gene, called fdxE . The DNA region surrounding fdxE was mapped, and the nucleotide sequence of a 4.2-kb fragment was determined . fdxE is preceded by a sequence that is very similar to a sigma54 recognition site and is followed by a putative transcription stop signal, suggesting that fdxE forms a separate cistron . Two open reading frames were identified upstream and downstream of fdxE and were named ORFE0 and ORFE1, respectively . The former may encode a polypeptide having 34% similarity with HtrA, a serine protease found in enteric bacteria . ORFE1 is homologous to purU, a gene involved in purine biosynthesis . Interposon mutagenesis of fdxE was unsuccessful when attempted on the wild-type strain B10 . Disruption of fdxE could be achieved only in strains harboring an additional copy of fdxE on a plasmid . Mutants obtained in this way and carrying a plasmid-borne copy of fdxE under the control of the nifH promoter grew only in N-free medium, thus demonstrating that fdxE expression is required for growth . Nevertheless, such mutants were found to spontaneously revert at a frequency of 5 x 10(-6) to an apparent wild-type phenotype, although they contained no detectable amount of FdVI . Taken together, the results indicate that FdVI is required for an essential metabolic function in R . capsulatus and that this FdVI dependence could be relieved by a single-mutation event . In accordance, FdVI biosynthesis was found to be constitutive in R . capsulatus.

J Bacteriol, 1997 May, 179(10), 3270 - 6
The actinomycete Thermobispora bispora contains two distinct types of transcriptionally active 16S rRNA genes; Wang Y et al.; Here we present the first description of the presence of two distinct types of 16S rRNA genes in the genome of a (eu)bacterium, Thermobispora bispora . We cloned and determined the nucleotide sequences of all four rRNA operons of T . bispora . Sequence comparisons revealed that the genome of T . bispora contains two distinct types of 16S rRNA genes, each type consisting of two identical or nearly identical copies, and three identical copies of the 23S RNA gene . The nucleotide sequences of the two types of 16S rRNA genes differ at 98 nucleotide positions (6.4% of total nucleotides) together with six regions of deletion-insertions . None of the base substitutions or insertion-deletions corresponds to any of the approximately 600 evolutionarily invariable or rarely variable nucleotides, indicating that both genes are functional . Both types of 16S rRNA genes are transcribed and processed as determined by Northern (RNA) hybridization and reverse transcriptase-mediated PCR.

Parasitology, 1997 May, 114 ( Pt 5), 483 - 8
Mechanisms of specificity of association between the nematode Steinernema scapterisci and its symbiotic bacterium; Grewal PS et al.; We suggest a new mechanism for the maintenance of specificity of the association between the entomopathogenic nematode Steinernema scapterisci and its symbiotic bacteria . We evaluated the development and reproduction of infective and non-infective juvenile S . scapterisci in monoxenic combinations with its symbiotic bacteria, Xenorhabdus sp . 'S' and with the bacterial symbiont of Steinernema carpocapsae and Steinernema riobravis . Although development of non-infective stages occurred on all Xenorhabdus spp., the development of infective juveniles to the 4th stage ('dauer' recovery) was significantly delayed and reduced with X . nematophilus and Xenorhabdus sp . 'R', the bacterial symbionts of S . carpocapsae and S . riobravis, respectively . 'Dauer' recovery improved significantly when the cultures of X . nematophilus and Xenorhabdus sp . 'R' were supplemented with cell-free filtrates from Xenorhabdus sp . 'S' . The infective juvenile S . scapterisci produced in all 3 cultures were virulent to Galleria mellonella larvae, confirming successful retention of Xenorhabdus from other steinernematids in their intestine . In fact, S . scapterisci infective juveniles containing X . nematophilus or Xenorhabdus sp . 'R' were more virulent to G . mellonella than those containing their natural symbiont, Xenorhabdus sp . 'S' . We believe that this is the first demonstration of the symbiont-specific exit of infective juveniles from the 'dauer' phase which represents the finest level of specificity of bacteria-nematode association . This is also the first report of successful isolation of the natural symbiont of S . scapterisci.

Appl Environ Microbiol, 1997 May, 63(5), 2089 - 91
Transformation of Escherichia coli with a large plasmid of Acidiphilium multivorum AIU 301 encoding arsenic resistance; Suzuki K et al.; Acidiphilium multivorum AIU 301 isolated from acid mineral water had strong arsenic resistance . This bacterium harbored a number of plasmids with different molecular sizes . A plasmid of 56 kbp, named pKW301, was isolated from A . multivorum AIU 301 . When pKW301 was transferred into Escherichia coli JM109 by electroporation, an E . coli transformant carrying pKW301 exhibited resistance to sodium arsenite, sodium arsenate, and mercuric (II) chloride.

Appl Environ Microbiol, 1997 May, 63(5), 2047 - 53
Resuscitation of viable but nonculturable Legionella pneumophila Philadelphia JR32 by Acanthamoeba castellanii; Steinert M et al.; Legionella pneumophila is an aquatic bacterium and is responsible for Legionnaires' disease in humans . Free-living amoebae are parasitized by legionellae and provide the intracellular environment required for the replication of this bacterium . In low-nutrient environments, however, L . pneumophila is able to enter a non-replicative viable but nonculturable (VBNC) state . In this study, L . pneumophila Philadelphia I JR 32 was suspended in sterilized tap water at 10(4) cells/ml . The decreasing number of bacteria was monitored by CFU measurements, acridine orange direct count (AODC), and hybridization with 16S rRNA-targeted oligonucleotide probes . After 125 days of incubation in water, the cells were no longer culturable on routine plating media; however, they were still detectable by AODC and by in situ hybridization . The addition of Acanthamoeba castellanii to the dormant bacteria resulted in the resuscitation of L . pneumophila JR 32 to a culturable state . A comparison of plate-grown legionellae and reactivated cells showed that the capacity for intracellular survival in human monocytes and intraperitoneally infected guinea pigs, which is considered a parameter for virulence, was not reduced in the reactivated cells . However, reactivation of dormant legionellae was not observed in the animal model.

Head Neck, 1997 May, 19(3), 216 - 8
Lack of serologic evidence for Helicobacter pylori infection in head and neck cancer; Grandis JR et al.; BACKGROUND: Several epidemiologic investigations have established a link between Helicobacter pylori infection and gastric malignancies . Because the stomach is in continuity with the oral cavity and the bacterium has been isolated from dental plaque and saliva, we hypothesized that H . pylori infection of the upper aerodigestive tract might result in mucosal disruption, allowing for subsequent transformation by known carcinogens such as tobacco and alcohol . METHODS: To test this hypothesis, we assayed for the presence of IgG antibodies to H . pylori in the serum of 21 patients with squamous cell carcinoma of the head and neck (SCCHN) and 21 matched controls without a history of head and neck cancer . RESULTS: The incidence of seropositivity in the SCCHN patients was 57% and in the controls, 62% (p > 0.05) . CONCLUSIONS: These data do not support an etiologic role for H . pylori infection in head and neck cancer.

Gastroenterology, 1997 May, 112(5), 1482 - 6
MALT-type lymphoma of the stomach is associated with Helicobacter pylori strains expressing the CagA protein; Eck M et al.; BACKGROUND & AIMS: Helicobacter pylori is considered to be involved in the pathogenesis of gastric lymphoma of mucosa-associated lymphoid tissue (MALT) type . Strains expressing the CagA protein (CagA+ strains) have been strongly associated with severe gastritis, duodenal ulceration, and gastric adenocarcinoma . The aim of this study was to determine the presence of H . pylori as well as incidence of CagA+ strains in gastric MALT-type lymphoma . METHODS: Sera of 68 patients with gastric MALT-type lymphoma (22 with low grade, 36 with high grade, and 10 with secondary high grade) were obtained, and the serological response to CagA was studied by immunoblotting using a purified recombinant CagA protein, a CagA+ strain, and the corresponding isogenic CagA- mutant . RESULTS: Of the patients with MALT-type lymphoma, 98.5% (67 of 68 patients) were H . pylori seropositive . In the only seronegative patient, the bacterium was detected histologically by Warthin-Starry staining . Of the seropositive patients, 95.5% had serum immunoglobulin G antibodies to CagA compared with 67% of an H . pylori-positive control group (33 of 49 patients; P = 0.000037) with chronic active gastritis . CONCLUSIONS: These results indicate infection of almost all patients with MALT-type lymphoma by CagA+ H . pylori strains . Strains expressing the CagA protein seem to play a crucial role in the pathogenesis of gastric MALT-type lymphoma.

Biophys J, 1997 May, 72(5), 2304 - 19
The interaction of quinone and detergent with reaction centers of purple bacteria . I . Slow quinone exchange between reaction center micelles and pure detergent micelles; Shinkarev VP et al.; The kinetics of light-induced electron transfer in reaction centers (RCs) from the purple photosynthetic bacterium Rhodobacter sphaeroides were studied in the presence of the detergent lauryldimethylamine-N-oxide (LDAO) . After the light-induced electron transfer from the primary donor (P) to the acceptor quinone complex, the dark re-reduction of P+ reflects recombination from the reduced acceptor quinones, QA- or QB- . The secondary quinone, QB, which is loosely bound to the RC, determines the rate of this process . Electron transfer to QB slows down the return of the electron to P+, giving rise to a slow phase of the recovery kinetics with time tau P approximately 1 s, whereas charge recombination in RCs lacking QB generates a fast phase with time tau AP approximately 0.1 s . The amount of quinone bound to RC micelles can be reduced by increasing the detergent concentration . The characteristic time of the slow component of P+ dark relaxation, observed at low quinone content per RC micelle (at high detergent concentration), is about 1.2-1.5 s, in sharp contrast to expectations from previous models, according to which the time of the slow component should approach the time of the fast component (about 0.1 s) when the quinone concentration approaches zero . To account for this large discrepancy, a new quantitative approach has been developed to analyze the kinetics of electron transfer in isolated RCs with the following key features: 1) The exchange of quinone between different micelles (RC and detergent micelles) occurs more slowly than electron transfer from QB- to P+; 2) The exchange of quinone between the detergent "phase" and the QB binding site within the same RC micelle is much faster than electron transfer between QA- and P+; 3) The time of the slow component of P+ dark relaxation is determined by (n) > or = 1, the average number of quinones in RC micelles, calculated only for those RC micelles that have at least one quinone per RC (in excess of QA) . An analytical function is derived that relates the time of the slow component of P+ relaxation, tau P, and the relative amplitude of the slow phase . This provides a useful means of determining the true equilibrium constant of electron transfer between QA and QB (LAB), and the association equilibrium constant of quinone binding at the QB site (KQ+) . We found that LAB = 22 +/- 3 and KQ = 0.6 +/- 0.2 at pH 7.5 . The analysis shows that saturation of the QB binding site in detergent-solubilized RCs is difficult to achieve with hydrophobic quinones . This has important implications for the interpretation of apparent dependencies of QB function on environmental parameters (e.g . pH) and on mutational alterations . The model accounts for the effects of detergent and quinone concentration on electron transfer in the acceptor quinone complex, and the conclusions are of general significance for the study of quinone-binding membrane proteins in detergent solutions.

Infect Immun, 1997 May, 65(5), 1644 - 52
Down regulation of intimin expression during attaching and effacing enteropathogenic Escherichia coli adhesion; Knutton S et al.; Enteropathogenic Escherichia coli (EPEC) produces attaching and effacing (A/E) lesions in the intestinal mucosa . The intimate bacterial adhesion associated with A/E lesion formation is promoted by intimin, a 94-kDa EPEC surface protein . Anti-intimin antisera raised in rabbits by using the purified 280-amino-acid cell binding domain of intimin as the immunogen were employed in immunofluorescence and immunoelectron microscopical studies to investigate the expression of intimin by classical EPEC strain E2348/69 (O127:H6) and defined E2348/69 derivatives during culture growth and A/E bacterium adhesion to cultured HEp-2 cells . In stationary-phase broth cultures, only a small fraction of E2348/69 bacteria expressed intimin, and of those that did, immunolabelling revealed a uniform distribution of intimin over the bacterial surface; increased numbers of bacteria expressing intimin were detected when E2348/69 was grown in tissue culture medium, an effect not seen with strain JPN15, a virulence plasmid-cured derivative of E2348/69 . Strain CVD206, an eaeA mutant of E2348/69, did not stain with the anti-intimin antisera, but strain CVD206(pCVD438), containing a functional eaeA gene, stained uniformly . After a 3-h incubation of HEp-2 cells with strain E2348/69, double immunofluorescence labelling of intimin and cellular actin revealed strong intimin expression by all A/E bacteria, but after 6 h of incubation, intimin expression by most E2348/69 bacteria was greatly reduced or not detected . This effect on intimin expression was not observed with strain JPN15 but was restored for strain JPN15(pCVD450) harboring the virulence plasmid-encoded per genes . These results indicate that surface expression of intimin is regulated by environmental factors during bacterial growth and following A/E lesion formation and that virulence plasmid-encoded genes participate in these regulation processes.

Curr Microbiol, 1997 May, 34(5), 309 - 13
Endo-beta-Glucanase from Acetobacter xylinum: Purification andCharacterization
Oikawa T, Kamatani T, Kaimura T, Ameyama M, Soda K.
A cellulose-producing acetic acid bacterium,Acetobacter xylinum KU-1, abundantly produces an extracellularendo-beta-glucanase (EC 3.2.1.4) in the culture broth . The enzyme was purifiedto homogeneity by DEAE- and CM- Toyopearl 650M ion-exchange chromatography,Butyl-Toyopearl 650M hydrophobic chromatography, and Toyopearl HW-50 gelfiltration . The purified enzyme showed the maximum activity at pH 5 and50°C: it was stable up to 50°C at pH 5, activated by Co2+, andcompetitively inhibited by Hg2+; the apparentKi was 7 &mgr;M . The molecular weight of the enzyme wasdetermined to be about 39,000 by sodium dodesyl sulfate/polyacrylamide gelelectrophoresis, and about 41,000 by Toyopearl HW-50 gel filtration; theenzyme is monomeric . The enzyme hydrolyzed carboxymethylcellulose with anapparent Km of 30 mg/ml and Vmax of 1.2&mgr;M/min . It hydrolyzed cellohexaose to cellobiose, cellotriose andcellotetraose, and also cellopentaose to cellobiose and cellotriose, but didnot act on cellobiose, cellotriose, or cellotetraose.

Mol Gen Genet, 1997 Apr 28, 254(4), 456 - 63
Genetic organization and transposition properties of IS511; Mullin DA et al.; IS511 is an endogenous insertion sequence (IS) of the bacterium Caulobacter crescentus strain CB15 and it is the first Caulobacter IS to be characterized at the molecular level . We determined the 1266-bp nucleotide sequence of IS511 and investigated its genetic organization, relationship to other ISs, and transposition properties . IS511 belongs to a distinct branch of the IS3 family that includes ISR1, IS476, and IS1222, based on nucleotide sequence similarity . The nucleotide sequence of IS511 encodes open reading frames (orfs) designated here as orfA and orfB, and their relative organization and amino acid sequences of the predicted protein products are very similar to those of orfAs and orfBs of other IS3 family members . Nuclease S1 protection assays identified an IS511 RNA, and its 5' end maps approximately 16 nucleotides upstream of orfA and about six nucleotides downstream of a sequence that is similar to the consensus sequence of C . crescentus housekeeping promoters . Evidence is presented that IS511 is capable of precise excision from the chromosome, and transposition from the chromosome to a plasmid . Transpositional insertions of IS511 occurred within sequences with a relatively high G + C content, and they were usually, but not always, flanked by a 4-bp direct repeat that matches a sequence at the site of insertion . We also determined the nucleotide sequence flanking the four endogenous IS511 elements that reside in the chromosome of C . crescentus . Our findings demonstrate that IS511 is a transposable IS that belongs to a branch of the IS3 family.

J Biotechnol, 1997 Apr 25, 54(2), 139 - 48
Improvement of expression and secretion of a fungal xylanase in the rumen bacterium Butyrivibrio fibrisolvens OB156 by manipulation of promoter and signal sequences; Xue GP et al.; Promoters and signal sequences for expression and secretion of a fungal xylanase encoded by a modified Neocallimastix patriciarum xynA cDNA in the rumen bacterium, Butyrivibrio fibrisolvens OB156, were investigated . Successful expression of the fungal xylanase in OB156 was obtained using the putative xylanase promoter from B . fibrisolvens strain 49 . Replacing the putative -35 region sequence (TTGCAC) of the xylanase promoter with the sequence TTGACA by mutagenesis reduced the fungal xylanase expression level 4-fold in OB156, indicating that this B . fibrisolvens strain did not efficiently recognise the E . coli consensus -35 sequence . Reduction of the spacer length between the -35 and -10 regions of the xylanase promoter from 18 to 17 base-pairs (bp) considerably increased the expression levels of the fungal enzyme in both E . coli and OB156 . Insertion of a pUB110 mob promoter upstream of the xylanase promoter also significantly improved the fungal xylanase expression . Secretion of the fungal xylanase mediated by the alpha-amylase signal peptide from B . fibrisolvens strain H17c was efficient in E . coli, but very poor in OB156 . An increase in the hydrophobicity of the signal sequence resulted in a 4-fold increase in the extracellular portion of the fungal xylanase in OB156, indicating marked improvement in xylanase secretion efficiency . The recombinant plasmids and xylanase expression/secretion cassettes were found to be stable in OB156 after prolonged cultivation (100 generations) in the absence of antibiotic selection . These results suggest that the rumen bacterium B . fibrisolvens can be manipulated to produce and secrete a eukaryotic extracellular protein with stable maintenance of the expression cassette in plasmid form.

Rev Prat, 1997 Apr 15, 47(8), 848 - 54
{Gastric lymphoma}; Ruskone-Fourmestraux A; The stomach is the most common site involved in primary gastrointestinal lymphoma . Gastric lymphoma originates from the mucosa-associated lymphoid tissue so called MALT . It comprises a group of distinctive clinicopathological entities which are important to take in account for clinical behavior . In recent years, new diagnostic tools and modern modes of treatment have improved their overall prognosis . One of the most exciting recent discoveries is the hypothesis that an infection by a bacterium . Helicobacter pylori has a decisive role in gastric lymphoma.

Arch Biochem Biophys, 1997 Apr 15, 340(2), 219 - 30
Accumulation of sulfoquinovosyl-1-O-dihydroxyacetone in a sulfolipid-deficient mutant of Rhodobacter sphaeroides inactivated in sqdC; Rossak M et al.; The biosynthesis of the sulfolipid sulfoquinovosyl diacylglycerol in the purple bacterium Rhodobacter sphaeroides requires at least four genes:sqdA, sqdB, sqdC, and sqdD . As part of our strategy aimed at the elucidation of the function of the different sqd gene products, we insertionally inactivated sqdC of R . sphaeroides . The resulting sqdC null mutant showed only a 90% reduction in sulfolipid content . Apparently, the sqdC gene product is required for optimal sulfolipid biosynthesis, but either catalyzes no essential reaction in the pathway or can be functionally replaced to a certain extent by a different protein . The mutant accumulated a 35S-labeled compound that was purified to homogeneity from cell extracts . Matrix-assisted laser desorption mass spectrometry and nuclear magnetic resonance spectroscopy provided conclusive structural evidence to identify the compound as alpha-D-sulfoquinovosyl-1-O-dihydroxyacetone that exists in two interconvertible, keto and hemiacetal forms . Incubation of wild-type protein extracts with the labeled compound did not result in the incorporation into sulfolipid as would be expected for an intermediate of the pathway . Based on our results we propose that the sqdC gene product mediates the substrate specificity of the UDP-sulfoquinovose:diacylglycerol sulfoquinovosyltransferase that is encoded by sqdD and that catalyzes the final reaction of sulfolipid biosynthesis.

Arch Microbiol, 1997 Apr 15, 167(5), 295 - 301
A novel membrane-bound flavocytochrome c sulfide dehydrogenase from the colourless sulfur bacterium Thiobacillus sp . W5
Visser JM, de Jong GAH, Robertson LA, Kuenen JG.
A novel membrane-bound sulfide-oxidizing enzyme was purified 102-fold from the neutrophilic, obligately chemolithoautotrophic Thiobacillus sp . W5 by means of a six-step procedure . Spectral analysis revealed that the enzyme contains haem c and flavin . SDS-PAGE showed the presence of two types of subunit with molecular masses of 40 and 11 kDa . The smaller subunit contains covalently bound haem c, as was shown by haem staining . A combination of spectral analysis and the pyridine haemochrome test indicated that the sulfide-oxidizing heterodimer contains one molecule of haem c and one molecule of flavin . It appeared that the sulfide-oxidizing enzyme is a member of a small class of redox proteins, the flavocytochromes c, and is structurally most related to the flavocytochrome c sulfide dehydrogenase of the green sulfur bacterium Chlorobium limicola . The pH optimum of the enzyme is 8.6 . At pH 9, the Vmax was 2.1 ± 0.1 μmol cytochrome c (mg protein)-1 min-1, and the Km values for sulfide and cytochrome c were 1.7 ± 0.4 μM and 3.8 ± 0.8 μM, respectively . Cyanide inhibited the enzyme by the formation of an N-5 adduct with the flavin moiety of the protein . On the basis of electron transfer stoichiometry, it seems likely that sulfur is the oxidation product.

J Biol Chem, 1997 Apr 11, 272(15), 9890 - 4
Sulfide-quinone reductase from Rhodobacter capsulatus . Purification, cloning, and expression; Schutz M et al.; A sulfide-quinone oxidoreductase (SQR, EC 1.8.5.'.) has been purified to homogeneity from chromatophores of the non-sulfur purple bacterium Rhodobacter capsulatus DSM 155 . It is composed of a single polypeptide with an apparent molecular mass of about 55 kDa, exhibiting absorption and fluorescence spectra typical for a flavoprotein and similar to the SQR from the cyanobacterium Oscillatoria limnetica . From N-terminal and tryptic peptide sequences of the pure protein a genomic DNA clone was obtained by polymerase chain reaction amplification . Its sequence contains an open reading frame of 1275 base pairs (EMBL nucleotide sequence data base, accession no . X97478X97478) encoding the SQR of R . capsulatus . The deduced polypeptide consists of 425 amino acid residues with a molecular mass of 47 kDa and a net charge of +9 . The high similarity (72%)/identity (48%) between the N termini of the cyanobacterial and the bacterial enzyme was confirmed and extended . Both enzymes exhibit the FAD/NAD(P) binding betaalphabeta-fold (Wierenga, R . K., Terpstra, P., and Hol, W . G . S . (1986) J . Mol . Biol . 187, 101-107) . The complete sequence of the SQR from R . capsulatus shows further similarity to flavoproteins, in particular glutathione reductase and lipoamide dehydrogenase . The cloned sqr was expressed in Escherichia coli in a functional form.

Biochemistry (Mosc), 1997 Apr, 62(4), 343 - 9
Site-specific endonuclease AbaI from Azospirillum brasilense UQ 1796 is an isoschizomer of endonuclease BclI; Zabaznaya EV et al.; The site-specific endonuclease AbaI was isolated and purified to functional purity from the soil nitrogen-fixing bacterium Azospirillum brasilense UQ 1796 . Purification included successive chromatography on columns with phosphocellulose, heparin-Sepharose, and hydroxyapatite . The purified enzyme recognizes the palindromic DNA sequence 5'-T decreases ATCA-3' and cleaves it as shown by the arrow . The isolated enzyme belongs to class II restriction endonuclease and is an isoschizomer of endonuclease BclI . The enzyme of AbaI is active at 26-56 degrees C . The optimal temperature is 48 degrees C and the optimal buffer is LRB.

Aliment Pharmacol Ther, 1997 Apr, 11 Suppl 1, 109 - 15
The future of Helicobacter pylori eradication: a personal perspective; Marshall BJ; Future changes in the use of Helicobacter pylori eradication therapy first will involve a decision-making process to determine which individuals require testing . Clearly, only persons who require therapy need to be diagnosed and, at present, the indications for therapy are constantly expanding . The author takes the view that everyone with H . pylori would be better off without the bacterium, but accepts that in many countries resources are inadequate to achieve this goal . Where antibiotic therapy for H . pylori fails due to a resistant organism, second treatment must include a different class of antibiotic . When a third therapy is contemplated, antibiotic sensitivity studies are usually necessary . In developing counties where reinfection with H . pylori is common, lesser goals than permanent cure might be appropriate . Thus, selected patients could have H . pylori suppressive therapy to prevent full expression of H . pylori-associated disease, or to prevent reinfection after an initial eradicative therapy . After considering all these alternatives, one must conclude that a vaccination strategy, if safe and cost effective, is the ideal future therapy.

Biosci Biotechnol Biochem, 1997 Apr, 61(4), 710 - 5
A novel metalloproteinase, almelysin, from a marine bacterium, Alteromonas sp . No . 3696: purification and characterization; Shibata M et al.; We have discovered a novel metalloproteinase, which has high activity at low temperatures, from the culture supernatant of a marine bacterium . The strain was identified as Alteromonas sp . No . 3696 . The metalloproteinase, named almelysin, was purified to homogeneity from the cultured supernatant at 10 degrees C by two column chromatographies . About 20 mg of purified almelysin was obtained from 18.4 liters of the culture supernatant . The molecular mass of almelysin was estimated to be 28 kDa by SDS-PAGE and the isoelectric point was 4.3 . The optimum pH for activity of almelysin was pH 8.5-9.0 and 6.5 using casein and (7-methoxycoumarin-4-yl)acetyl(MOCAc)-Pro-Leu-Gly-Leu-(N3- {2,4-dinitrophenyl}-L-2,3-diaminopropionyl) {A2pr(Dnp)}-Ala-Arg-NH2 as substrates, respectively . Almelysin was stable between pH 7.5-8.0 and below 40 degrees C . The optimum temperature for the activity was observed to be 40 degrees C using both casein and MOCAc-Pro-Leu-Gly-Leu-A2pr(Dnp)-Ala-Arg-NH2 as substrates . The activity of almelysin was inhibited by such metallo chelators as EDTA and o-phenanthroline, while talopeptin, phosphoramidon, and SMPI, typical metalloproteinase inhibitors, had no effect . Almelysin primarily cleaved the Ala14-Leu15 bond and Phe24-Phe25 bond, and secondarily the Tyr16-Leu17 bond in oxidized insulin B-chain . However, almelysin could not cleave the His5-Leu6, His10-Leu11, and Gly23-Phe24 bonds, which were cleaved by other metalloproteinases . These results indicate that the substrate specificity of almelysin is different from other metalloproteinases . Interestingly, Alteromonas sp . No . 3696 strain produced another proteinase as well as almelysin at 25 degrees C.

Hybridoma, 1997 Apr, 16(2), 183 - 7
Characterization and application of a strain-specific monoclonal antibody against the rhizosphere bacterium Azospirillum brasilense Wa5; Schloter M et al.; A hybridoma cell line producing a rat monoclonal antibody (MAb Bo-33) directed against lipopolysaccharide of Azospirillum brasilense Wa5 has been established and characterized . Whole bacteria were used as immunogens . The number of antigens per cell was about 1500 . The number of antigens per cell of reisolates from the rhizosphere of what was similar to the number of antigens of bacteria cultivated in rich medium . The sensitivity of detection using MAb Bo-33 was about 100 bacteria/ml . Therefore, the MAb was suitable for in situ immunofluorescence detection and a sensitive direct quantification of Azospirillum brasilense Wa5 in rhizosphere extracts.

Microbiology, 1997 Apr, 143 ( Pt 4), 1203 - 10
Expression of a Butyrivibrio fibrisolvens E14 gene (cinB) encoding an enzyme with cinnamoyl ester hydrolase activity is negatively regulated by the product of an adjacent gene (cinR); Dalrymple BP et al.; A second cinnamoyl ester hydrolase (CEH) encoding gene (cinB) has been characterized from the ruminal bacterium Butyrivibrio fibrisolvens E14 . CinB is more similar to CinA (previously named CinI) (28% amino acid identify), the first CEH described from B . fibrisolvens E14, than either of the enzymes are to any other member of the family of hydrolases to which they belong . Upstream of cinB, and in the opposite orientation, is a gene (cinR) encoding a protein with substantial similarity to members of the MarR family of negative regulators of bacterial gene expression . By alignment of these sequences, a possible helix-turn-helix DNA-binding domain has been identified . CinR was expressed at a high level in Escherichia coli using the lac promoter . In E . coli CinR repressed the expression of CinB, but had no effect on the expression of CinA . In gel mobility-shift assays, CinR bound specifically to the cinR-cinB intergenic region . Two identical 16 nucleotide inverted repeats adjacent to the putative PcinR and PcinB promoters are likely binding sites for CinR . The addition of FAXX (O-{5-O-(trans-feruloyl)-alpha-L-arabinofuranosyl}-(1,3)- O-beta-D-xylopyranosyl-(1,4)-D-xylopyranose) and Fara {5-O-(trans-feruloyl)-arabinofuranose}, but not xylobiose, ferulic acid and a number of other soluble components of hemicellulose, inhibited the binding of CinR to DNA.

J Periodontal Res, 1997 Apr, 32(3), 279 - 86
Secretion of IL-1 beta, TNF-alpha, IL-8 and IL-1ra by human polymorphonuclear leukocytes in response to lipopolysaccharides from periodontopathic bacteria; Yoshimura A et al.; Polymorphonuclear leukocytes (PMN) are the first cells that migrate into periodontal tissues and gingival crevices in response to invading pathogens . It was recently demonstrated that PMN have the ability to synthesize and release cytokines following appropriate stimulation, while it is not clear whether these capacities are directly related to periodontal destructive processes . We therefore investigated the amounts of the cytokines interleukin-1 beta (IL-1 beta), tumor necrosis factor alpha (TNF-alpha), IL-8 and IL-1 receptor antagonist (IL-1ra) secreted by PMN from healthy donors following stimulation with lipopolysaccharide (LPS) from 4 periodontopathic bacteria, Porphyromonas gingivalis, Actinobacillus actinomycetemcomitans, Capnocytophaga ochracea and Fusobacterium nucleatum, and the non-oral bacterium Escherichia coli . A actinomycetemcomitans, F . nucleatum and E . coli LPS stimulated the release of significantly greater amounts of IL-1 beta, TNF-alpha and IL-8 than the control unstimulated PMN (p < 0.01) . The levels of IL-1 beta, TNF-alpha and IL-8 released from cells stimulated with P . gingivalis or C . ochracea LPS were significantly lower than those of cells stimulated with A . actinomycetemcomitans or E . coli LPS (p < 0.05) . On the other hand, substantially greater amounts of IL-1ra were released from PMN stimulated with each LPS and from control unstimulated PMN during the first 6 h, and then significantly greater amounts of IL-1ra were secreted by PMN stimulated with A . actinomycetemcomitans and E.coli LPS during the following 12 h (p < 0.01) . The inhibitory effects of IL-1ra on the biological activity of IL-1 in the supernatants of PMN were examined by the thymocyte comitogen proliferation assay . The supernatants of PMN stimulated with each LPS showed less biological IL-1 activity as compared with the same doses of recombinant human IL-1 beta detected by enzyme-linked immunosorbent assay . Furthermore, no activity was detected in the supernatants of PMN stimulated with P . gingivalis or C . ochracea LPS . These findings demonstrated that LPS from periodontopathic bacteria were capable of stimulating PMN to release not only pro-inflammatory cytokines but also their inhibitors such as IL-1ra . Different secretion levels of these cytokines and their biological activities induced by the various LPS might be important in the onset and progression of periodontal diseases.

J Appl Microbiol, 1997 Apr, 82(4), 422 - 30
Alteromonas infernus sp . nov., a new polysaccharide-producing bacterium isolated from a deep-sea hydrothermal vent; Raguenes GH et al.; A deep-sea, aerobic, mesophilic and heterotrophic new bacterium was isolated from a sample of fluid collected among a dense population of Riftia pachyptila, in the vicinity of an active hydrothermal vent of the Southern depression of the Guaymas basin (Gulf of California) . On the basis of phenotypic and phylogenetic analyses and DNA/DNA relatedness, the strain GY785 was recognized as a new species of the genus Alteromonas and the name of Alteromonas infernus is proposed . During the stationary phase in batch cultures in the presence of glucose, this bacterium secreted two unusual polysaccharides . The water-soluble exopolysaccharide-1 produced contained glucose, galactose, galacturonic and glucuronic acids as monosaccharides . The gel-forming exopolysaccharide-2 was separated from the bacterial cells by dialysis against distilled water and partially characterized.

Infect Immun, 1997 Apr, 65(4), 1455 - 61
Ehrlichia chaffeensis inclusions are early endosomes which selectively accumulate transferrin receptor; Barnewall RE et al.; Ehrlichia chaffeensis is an obligatory intracellular bacterium which infects macrophages and monocytes . Double immunofluorescence labeling was used to characterize the nature of E . chaffeensis inclusion in the human promyelocytic leukemia cell line THP-1 . E . chaffeensis was labeled with dog anti-E . chaffeensis serum and fluorescein isothiocyanate-conjugated anti-dog immunoglobulin G (IgG) . Lissamine rhodamine-conjugated anti-mouse IgG was used to label various mouse monoclonal antibodies . Ehrlichial inclusions did not fuse with lysosomes, since they were not labeled with anti-CD63 or anti-LAMP-1 . The ehrlichial inclusions were slightly acidic, since they weakly accumulated 3-(2,4-dinitroanilino)-3'-amino-N-methyldipropylamine and stained weakly positive for vacuolar type H+ ATPase . Some ehrlichial inclusions were labeled positive with antibodies against HLA-DR, HLA-ABC, and beta2 microglobulin, while other inclusions in the same cell were labeled negative . The inclusions were labeled strongly positive for transferrin receptors (TfRs) and negative for the clathrin heavy chain . Time course labeling for TfRs showed that up to 3 h postinfection, most of the ehrlichial inclusions were negative for TfRs . After 6 h postinfection, 100% of the ehrlichial inclusions became TfR positive and the intensity of labeling was increased during the subsequent 3 days . Reverse transcription-PCR showed a gradual increase in the level of TfR mRNA postinfection, which reached a peak at 24 h postinfection . These results suggest that ehrlichial inclusions are early endosomes which selectively accumulate TfRs and that the ehrlichiae up-regulate TfR mRNA expression.

Infect Immun, 1997 Apr, 65(4), 1147 - 51
Role of CD14 molecules in internalization of Actinobacillus actinomycetemcomitans by macrophages and subsequent induction of apoptosis; Muro M et al.; We report the evidence for apoptosis in J774.1 cells by the periodontopathic bacterium Actinobacillus actinomycetemcomitans, suggesting that the ability of A . actinomycetemcomitans to promote apoptosis might be important in the initiation and development of periodontitis . In this study, we examined the role of macrophage CD14, anchored by a glycerophosphatidylinositol tail, in the induction of apoptosis by A . actinomycetemcomitans infection by using the parent J774.1 cells and CD14-defective mutant (LR-9) cells . A small number of A . actinomycetemcomitans Y4 cells inside the LR-9 cells compared with the number in J774.1 cells was detected by confocal scanning microscopy . We found that LR-9 cells showed a weak cytotoxic effect after being infected with A . actinomycetemcomitans Y4 . Apoptotic cell death of LR-9 cells infected with A . actinomycetemcomitans Y4, compared with that of the parent J774.1 cells was almost undetectable, as shown by the proportion of fragmented DNA in agarose gel electrophoresis and by the terminal deoxynucleotidyl transferase-mediated dUTP end-labeling method . Flow cytometric cell cycle analysis of J774.1 cells infected with A . actinomycetemcomitans Y4 revealed the increased percentage of apoptotic cells with hypodiploid DNA . However, LR-9 cells infected with A . actinomycetemcomitans Y4 showed no increase in population of apoptotic nuclei compared with the noninfected cells . These findings suggest that the CD14 molecules may contribute to the phagocytosis of A . actinomycetemcomitans by J774.1 cells and regulate, at least in part, apoptotic cell death of macrophages infected with A . actinomycetemcomitans.

Am Fam Physician, 1997 Apr, 55(5), 1783 - 9, 1793-4
Cat-scratch disease and related clinical syndromes; Smith DL; Bartonella (Rochalimaea) henselae is a common cause of cat-scratch disease . This newly identified bacterium is also the cause of several other clinical syndromes, including bacillary angiomatosis, bacillary peliosis hepatitis and splenitis, and acute and relapsing bacteremia . A high percentage of young cats carry B . henselae . Fortunately, serious complications of B . henselae infections are rare in immunocompetent patients . Cat-scratch disease is usually a self-limited illness that does not necessarily require antibiotic therapy . Severe or persistent cases respond well to several antibiotics, including erythromycin and doxycycline . Cat-scratch disease should be included in the differential diagnosis of serious neurologic disease, particularly when regional lymphadenopathy develops suddenly in a previously healthy patient who owns a cat . Treatment of uncomplicated central nervous system disease is generally supportive . Antibiotic therapy is reserved for patients with atypical or severe involvement, including encephalopathy and retinitis . Other internal and cutaneous manifestations of B . henselae infection have recently been described . These potentially life-threatening infections respond well to antibiotic therapy, even in immunocompromised patients.

Int J Syst Bacteriol, 1997 Apr, 47(2), 566 - 8
Transfer of Blastobacter natatorius (Sly 1985) to the genus Blastomonas gen . nov . as Blastomonas natatoria comb . nov; Sly LI et al.; The budding bacterium Blastobacter natatorius belongs to the alpha-4 group of the Proteobacteria and clusters phylogenetically on a deep branch with Sphingomonas capsulata, with which it shares 93.9% 16S rRNA sequence similarity . On phylogenetic, phenotypic, and chemotaxonomic grounds a proposal is made to transfer B . natatorius to the genus Blastomonas gen . nov . as Blastomonas natatoria comb . nov.

Int J Syst Bacteriol, 1997 Apr, 47(2), 381 - 4
Anaerobiospirillum thomasii sp . nov., an anaerobic spiral bacterium isolated from the feces of cats and dogs and from diarrheal feces of humans, and emendation of the genus Anaerobiospirillum; Malnick H; Thirty-seven similar strains isolated from feces of cats and dogs and from human diarrheal feces had characteristics of the genus Anaerobiospirillum . These organisms were distinguished from the only previously described Anaerobiospirillum species, Anaerobiospirillum succiniciproducens, by producing acid from adonitol but not from fructose, raffinose, or sucrose and by the lack of alpha-glucosidase . The G + C contents of the DNAs of the new strains were 39 to 42 mol% . The results of morphological, physiological, DNA G + C content, and DNA homology studies support the proposal that the description of the genus Anaerobiospirillum should be emended so that a new species can be included in the genus . The new species Anaerobiospirillum thomasii is proposed, with strain A273/88 (= NCTC 12467) as the type strain.

Int J Syst Bacteriol, 1997 Apr, 47(2), 252 - 61
Citrate synthase gene comparison, a new tool for phylogenetic analysis, and its application for the rickettsiae; Roux V et al.; Using PCR and an automated laser fluorescent DNA sequencer, we amplified and sequenced a 1,234-bp fragment of the citrate synthase-encoding gene (gltA) of 28 bacteria belonging to the genus Rickettsia . Comparative sequence analysis showed that most of the spotted fever group (SFG) rickettsiae belonged to one of two subgroups . The first subgroup included Rickettsia massiliae, strain Bar 29, Rickettsia rhipicephali, "Rickettsia aeschlimanni," and Rickettsia montana, which have been isolated only from ticks . The second subgroup was larger and included the majority of the human pathogens and also rickettsiae isolated only from ticks; the members of this subgroup were strain S, Rickettsia africae, "Rickettsia monglotimonae," Rickettsia sibirica, Rickettsia parkeri, Rickettsia conorii, Rickettsia rickettsii, the Thai tick typhus rickettsia, the Israeli tick typhus rickettsia, the Astrakhan fever rickettsia, "Rickettsia slovaca," and Rickettsia japonica . The sequence analysis also showed that the tick-borne organisms Rickettsia helvetica and Rickettsia australis and the mite-borne organism Rickettsia akari were associated with the SFG cluster, that Rickettsia prowazekii and Rickettsia typhi, two representatives of the typhus group, clustered together, and that Rickettsia canada; Rickettsia bellii, and the AB bacterium probably represent three new groups . We compared the phylogenetic trees inferred from citrate synthase gene sequences and from 16S ribosomal DNA (rDNA) sequences . For rickettsial phylogeny, the citrate synthase approach was more suitable, as demonstrated by significant bootstrap values for all of the nodes except those in the larger subgroup defined above . We also compared phylogenetic analysis results obtained in a comparison of the sequences of both genes for all of the representatives of the domain Bacteria for which the gltA sequence was determined . We believe that comparison of gltA sequences could be a complementary approach to 16S rDNA sequencing for inferring bacterial evolution, especially when unstable phylogenetic models are obtained from ribosomal sequences because of high levels of sequence similarity between the bacteria studied.

Gastroenterology, 1997 Apr, 112(4), 1386 - 97
A standardized mouse model of Helicobacter pylori infection: introducing the Sydney strain; Lee A et al.; BACKGROUND & AIMS: Currently available Helicobacter pylori models show variable and, in some instances, poor colonization . There is a need for a strain with high colonizing ability to act as a standard for animal studies . METHODS: After screening a range of fresh clinical isolates and long-term adaptation in mice, a strain of H . pylon has been isolated with a very good colonizing ability . RESULTS: This strain, named the Sydney strain of H . pylori (strain SS1), is cagA and vacA positive . High levels of colonization (10(6)-10(7) colony-forming units/g tissue) were achieved consistently in C57BL/6 mice . Colonization levels varied depending on the mouse strain used with BALB/c, DBA/2, and C3H/He, all being colonized but in lower numbers . In all strains of mice, bacteria were clearly visible at the junctional zone between the antrum and the body . The phenotype was stable with colonizing ability remaining after 20 subcultures in vitro . The bacterium attached firmly to gastric epithelium . During 8 months, a chronic active gastritis slowly developed, progressing to severe atrophy in both C57BL/6 and BALB/c mice . CONCLUSIONS: The Sydney strain of H . pylori is available to all and will provide a standardized mouse model for vaccine development, compound screening, and studies in pathogenesis.

J Bacteriol, 1997 Apr, 179(7), 2319 - 30
Transcription of genes encoding DNA replication proteins is coincident with cell cycle control of DNA replication in Caulobacter crescentus; Roberts RC et al.; DNA replication in the dimorphic bacterium Caulobacter crescentus is tightly linked to its developmental cell cycle . The initiation of chromosomal replication occurs concomitantly with the transition of the motile swarmer cell to the sessile stalked cell . To identify the signals responsible for the cell cycle control of DNA replication initiation, we have characterized a region of the C . crescentus chromosome containing genes that are all involved in DNA replication or recombination, including dnaN, recF, and gyrB . The essential dnaN gene encodes a homolog of the Escherichia coli beta subunit of DNA polymerase III . It is transcribed from three promoters; one is heat inducible, and the other two are induced at the transition from swarmer to stalked cell, coincident with the initiation of DNA replication . The single gyrB promoter is induced at the same time point in the cell cycle . These promoters, as well as those for several other genes encoding DNA replication proteins that are induced at the same time in the cell cycle, share two sequence motifs, suggesting that they represent a family whose transcription is coordinately regulated.

Biochem Biophys Res Commun, 1997 Mar 27, 232(3), 777 - 82
Cross-linking studies and membrane localization and assembly of radiolabelled large mechanosensitive ion channel (MscL) of Escherichia coli; Hase CC et al.; The gene encoding the large conductance mechanosensitive ion channel (MscL) of Escherichia coli and several deletion mutants of mscL were cloned under the control of the T7 RNA polymerase promoter . Transformation of these constructs into an E . coli strain carrying an inducible T7 RNA polymerase gene allowed the specific production and labelling of MscL with {35S}methionine . Preparation of membrane fractions of E . coli cells by sucrose gradient centrifugation indicated that the radiolabelled MscL was present in the inner cytoplasmic membrane in agreement with results of several studies . However, treatment of the labelled cells and cell membrane vesicles with various cross-linkers resulted in the majority of labelled protein migrating as a monomer with a small proportion of molecules (approximately 25%) migrating as dimers and higher order multimers . This result is in contrast with a finding of a study suggesting that the channel exclusively forms hexamers in the cell membrane of E . coli (1) and therefore may have profound implication for the activation and/or "multimerization" of the channel by mechanical stress exerted to the membrane . In addition, from the specific activity of the radiolabelled protein and the amount of protein in the cytoplasmic membrane fraction we estimated the number of MscL ion channels expressed under these conditions to be approximately 50 channels per single bacterium.

Biochemistry, 1997 Mar 25, 36(12), 3671 - 9
Evaluation of structure-function relationships in the core light-harvesting complex of photosynthetic bacteria by reconstitution with mutant polypeptides; Davis CM et al.; Seven mutant LH1 polypeptides of Rhodobactor sphaeroides have been isolated, and their behaviors in in vitro reconstitution of LH1 and its subunit complex have been characterized . Two mutants were selected to address the increased stability of the subunit complex of Rb . sphaeroides compared with that of Rhodobacter capsulatus . We found that this difference can be largely ascribed to the existence of Tyr at position +4 in the beta-polypeptide (the numbering system used assigns position 0 to the His which provides the coordinating ligand to bacteriochlorophyll) of the former bacterium compared to Met in that position in the latter . The amount of energy involved in the increased interaction was 1.6 kcal/mol, which would be consistent with a hydrogen bond involving Tyr . Mutation of the His at position 0 to Asn allows an estimate of the binding energy for subunit formation contributed by coordination of the imidazole group of His to the Mg atom of bacteriochlorophyll of >4.5 kcal/mol per BChl . Finally, an evaluation of the role of amino acids in the C-terminal region of the alpha-polypeptide was begun . Reconstitution of a mutant alpha-polypeptide in which Trp at position +11 was changed to Phe resulted in optimal formation of an LH1-type complex whose lambda(max) was blue-shifted to 853 nm, the same as observed in the intact bacterium harboring this mutation . These results provide further confirmation that the environment of BChl in reconstituted LH1 complexes is the same as in vivo and support the assignment of this residue to a role in hydrogen bonding with the C3(1) carbonyl group of BChl . Two other mutants of the alpha-polypeptide in which 5 and 14 amino acids in the C-terminus were deleted were also examined . These were of interest because the latter mutant, unlike the former, resulted in a low level of expression of LH1 in intact cells . However, with both of these mutant polypeptides, reconstitution appeared identical to that of the native system . In the case of the mutant shortened by 14 amino acids, a small blue-shift in lambda(max) to 861 nm was observed, again reproducing the blue-shift exhibited by the intact cells . Thus, these results suggest that the lowered levels of in vivo expression observed in these two mutants are due to reduced incorporation of the alpha-polypeptide into the membrane or its increased degradation, rather than to decreased stabilization of the LH1 complex.

Gene, 1997 Mar 18, 187(2), 225 - 9
Recombination between a resident plasmid and the chromosome following irradiation of the radioresistant bacterium Deinococcus radiodurans; Daly MJ et al.; Interplasmidic and intrachromosomal recombination in Deinococcus radiodurans has been studied recently and has been found to occur at high frequency following exposure to ionizing radiation . In the current work, we document plasmid-chromosome recombination following exposure of D . radiodurans to 1.75 Mrad (17.5 kGy) 60Co, when the plasmid is present in the cell at the time of irradiation . Recombination is assayed using both physical and allelic polymorphisms of homologous genes in the plasmid and chromosome . Recombination was found to be largely, but not entirely, recA-dependent . Crossovers occur frequently, and a significant fraction of these are non-reciprocal.

Toxicol Lett, 1997 Mar 14, 91(1), 63 - 71
Evidence for arylamine N-acetyltransferase activity in the bacterium Helicobacter pylori; Chung JG et al.; N-Acetyltransferase activities with p-aminobenzoic acid and 2-aminofluorene were determined in Helicobacter pylori from gastroduodenal disease patients . The N-acetyltransferase activity was determined using an acetyl CoA recycling assay and high pressure liquid chromatography . The N-acetyltransferase activities from a number of Helicobacter pylori samples were found to be 0.91 +/- 0.12 nmole/min/mg protein for the acetylation of 2-aminofluorene and 0.75 +/- 0.22 nmole/min/mg protein for the acetylation of p-aminobenzoic acid . The apparent K(m) and V(max) values obtained were 1.10 +/- 0.08 mM and 2.34 +/- 0.14 nmol/min/mg protein for 2-aminofluorene, and 0.92 +/- 0.09 mM and 2.08 +/- 0.16 nmol/min/mg protein for p-aminobenzoic acid . The optimal pH value for the enzyme activity was 6.0 for both substrates tested . The optimal temperature for enzyme activity was 37 degrees C for both substrates . The N-acetyltransferase activity was inhibited by iodacetamide: at 0.25 mM iodacetamide, activity was reduced 50% and 1.0 mM iodacetamide inhibited activity more than 90% . Among a series of divalent cations and salts, Cu2+ and Zn2+ were demonstrated to be the most potent inhibitors . Among the protease inhibitors, only ethylenediaminetetraacetic acid significantly protected N-acetyltransferase . Iodoacetic acid, in contrast to the other agents, markedly inhibited N-acetyltransferase . This is the first demonstration of acetyl CoA:arylamine N-acetyltransferase activity in Helicobacter pylori.

Carbohydr Res, 1997 Mar 13, 298(4), 311 - 8
Structure of the repeating oligosaccharide from the lipopolysaccharide of the nitrogen-fixing bacterium Acetobacter diazotrophicus strain PAL 5; Previato JO et al.; Acetobacter diazotrophicus is an acid-tolerant nitrogen-fixing bacterium found in roots, rhizosphere, stems, and leaves of sugar cane (Saccharum officinarum) cultivated in Brazil . The O-polysaccharide from the lipopolysaccharide of the root isolate strain PAL 5 has been determined by a combination of methylation analysis and two-dimensional high field NMR spectroscopy . The pentasaccharide repeat has the structure: {formula: see text} Minor resonances in the NMR spectra are consistent with the presence of a proportion of repeating units which lack the beta-D-Glc side-chain.

Biochemistry, 1997 Mar 11, 36(10), 3027 - 36
Influence of Asn/His L166 on the hydrogen-bonding pattern and redox potential of the primary donor of purple bacterial reaction centers; Ivancich A et al.; The primary electron donor (P) of the photosynthetic reaction center (RC) from the purple bacterium Rhodobacter (Rb.) sphaeroides is constituted of two bacteriochlorophyll molecules in excitonic interaction . The C2 acetyl carbonyl group of one of the two bacteriochlorophyll molecules (PL), the one more closely associated with the L polypeptide subunit, is engaged in a hydrogen bond with histidine L168, while the other pi-conjugated carbonyl groups of P are free from such hydrogen-bonding interactions . The three-dimensional X-ray crystal structures of the RC from several strains of Rb . sphaeroides reveal that asparagine L166 probably interacts indirectly with P through His L168 . Such an interaction is expected to modulate the hydrogen bond between P and His L168, a residue which is highly conserved in purple bacteria . RC mutants of Rb . sphaeroides where asparagine L166 was genetically replaced by leucine {NL(L166)}, histidine {NH(L166)}, and aspartate {ND(L166)} were studied using Fourier transform (FT) Raman spectroscopy . All of these mutations resulted in an increase in the strength of the hydrogen bond between His L168 and the acetyl carbonyl group of P(L), as observed in the FT Raman spectrum, by the 2-4 cm(-1) decrease in vibrational frequency of the 1620 cm(-1) band which has been assigned to this specific acetyl carbonyl group {Mattioli, T . A., Lin, X., Allen, J . P., & Williams, J . C . (1995) Biochemistry 34, 6142-6152} . At pH 8, the NH(L166) mutation showed the greatest change in the P0/P.+ redox midpoint potential (515 mV), increasing it by ca . 30 mV compared to that of wild type (485 mV) . A similar increase in P0/P.+ redox midpoint potential for NH(L166) compared to that of wild type is also observed at pH 5, 6, and 9.5 . The p0/P.+ midpoint potential of the NL(L166) mutant was comparable to that of wild type at all pH values . In contrast, for the ND(L166) mutant, the midpoint potential shows a markedly different pH dependency, being 25 mV higher than wild type at pH 5 but 20 mV lower than wild type at pH 9.5 . The hydrogen bond interactions of the primary electron donor from Rhodospirillum (Rsp.) centenum were determined from the FT Raman vibrational spectrum which exhibits a 1616 cm(-1) band similar to what is seen in the NH(L166) and ND(L166) Rb . sphaeroides mutants . Comparison of the sequence of the L subunit determined for the Rsp . centenum RC with that of other species indicates that positions L166 and L168 are occupied by His residues . The stronger hydrogen bond between the conserved His L168 and the acetyl carbonyl group of P(L), observed in the primary donor of Rsp . centenum and of several bacterial species which are known to possess a histidine residue at the analogous L166 position, is proposed to be due to interactions between these two histidine residues.

FEBS Lett, 1997 Mar 10, 404(2-3), 143 - 7
Terminal oxidases of Azoarcus sp . BH72, a strictly respiratory diazotroph; Reinhold-Hurek B et al.; The nitrogen-fixing, grass-associated bacterium Azoarcus sp . BH72 was characterized with respect to its terminal oxidases . Inhibitory respiratory analysis revealed the presence of at least one cytochrome c oxidase and one quinol oxidase . The cytochrome c oxidase was preferably used by the cells under aerobic, whereas the quinol oxidase seemed to be dominant under microaerobic, nitrogen-fixing conditions . Differential spectroscopy and heme analysis of the membrane preparations indicated that the cytochrome c oxidase is probably of the cb type.

Gene, 1997 Mar 10, 187(1), 99 - 105
Genomic organization of the metal-mobilizing bacterium Thiobacillus cuprinus; Marin I et al.; The genomic organization of Thiobacillus cuprinus, a facultative chemolithotrophic bacterium that preferentially solubilizes copper from complex ores, has been studied by Pulsed Field Gel Electrophoresis (PFGE) . It has been determined that T . cuprinus has a circular chromosome of about 3.8 Mb in size as concluded by analysis of gamma-irradiated total DNA and restriction analysis . Macrorestriction patterns for several restriction enzymes have been generated . Restriction enzymes AseI, DraI, SpeI, SwaI and XbaI give a number of fragments that can be adequately resolved by PFGE and then be used for electrophoretic karyotyping and for the construction of physical maps of the chromosome . Such a map has been constructed for the endonucleases SpeI and SwaI . The localization of several heterologous and homologous genes on the physical map, including those for rRNA, has paved the way for the construction of a genetic map.

Cell, 1997 Mar 7, 88(5), 675 - 84
Cell cycle-dependent polar localization of chromosome partitioning proteins in Caulobacter crescentus; Mohl DA et al.; In the bacterium C . crescentus, the cellular homologs of plasmid partitioning proteins, ParA and ParB, localize to both poles of the predivisional cell following the completion of DNA replication . ParB binds to DNA sequences adjacent to the origin of replication suggesting that this region of the genome is tethered to the poles of the cell at a specific time in the cell cycle . Increasing the cellular levels of ParA and ParB disrupts polar localization and results in defects in both cell division and chromosome partitioning . These results suggest that ParA and ParB are involved in partitioning newly replicated chromosomes to the poles of the predivisional cell and may function as components of a bacterial mitotic apparatus.

Arch Microbiol, 1997 Mar 7, 167(2/3), 99 - 105
The effect of respiration on the phototactic behavior of the purple nonsulfur bacterium Rhodospirillum centenum
Romagnoli S, Hochkoeppler A, Damgaard L, Zannoni D.
The effect of respiration on the positive phototactic movement of swarming agar colonies of the facultative phototroph Rhodospirillum centenum was studied . When the electron flow was blocked at the bc1 complex level by myxothiazol, the oriented movement of the colonies was totally blocked . Conversely, inhibition of respiration via the cytochrome c oxidase stimulated the phototactic response . No phototaxis was observed in a photosynthesis deficient mutant (YB707) lacking bacteriochlorophylls . Analyses of the respiratory activities as monitored by a oxygen microelectrode in single agar colonies during light/dark transitions showed a close functional correlation between the photosynthetic and respiratory apparatuses . The respiratory chain of Rsp . centenum was formed by two oxidative pathways: one branch leading to a cytochrome c oxidase inhibited by low cyanide concentrations and a second pathway formed by an oxidase less-sensitive to cyanide that also catalyzes the light-driven respiration . These results were interpreted to indicate that (1) there is a cyclic electron transport, and (2) photoinduced cyclic electron flow is required for the phototactic response of Rsp . centenum . Furthermore, under oxic conditions in the light, reducing equivalents may switch from photosynthetic to respiratory components so as to reduce both the membrane potential and the rate of locomotion.

Genetika, 1997 Mar, 33(3), 321 - 7
{Isolation and properties of mutants devoid of pseudo-HPr activity of the fructose transfer system in Escherichia coli K12}; Sergeev KV et al.; Mutants without pseudo-HPr activity intrinsic to the H domain of the FruB protein were obtained in Escherichia coli K12 through insertional mutagenesis by means of the MudIlac phage and TnPhoA transposon . For isolating these mutants, double mutants of enteric bacterium (ptsH fruR of ptsH fruS) were used as original strains . These double mutants were inactive with respect to the total HPr protein of the phosphoenolpyruvate-carbohydrate phosphotransferase system and could not provide constitutive synthesis of fructose-specific proteins of this system . The mutants obtained were designated fruH, mapped, and genetically characterized . Insertions were shown to be located in exactly the portion of the fruB gene that specifies synthesis of the H domain of the FruB protein . The fruH mutations decreased motility of bacterial both in a medium free of carbohydrates and in a medium with glucose, fructose, or maltose . In addition, fruH mutants with the MudIlac genome insertion had sharply decreased activity of beta-galactosidase.

Curr Opin Pulm Med, 1997 Mar, 3(2), 105 - 10
Bacterial infections of the bronchial tree; Wilson R; In health, bacteria colonize the upper respiratory tract, but the lower respiratory tract defenses keep the lung sterile from the first bronchial division, despite being regularly challenged by bacteria from the nasopharynx and the environment . Bronchial infections reflect a failure that can be ascribed either to the virulence of the bacterium or to a deficiency of one or more of these defenses . An abnormality of the airway defenses, which may be inherited (e.g., cystic fibrosis) or acquired (e.g., following a viral infection or cigarette smoking), is present in most patients with bronchial infection, and this fact should be considered when planning investigation.

Biol Chem, 1997 Mar-Apr, 378(3-4), 303 - 8
Organization of the DMSO respiratory operon of Rhodobacter capsulatus and its consequences for homologous expression of DMSOR/TMAOR; Knablein J et al.; In this report we describe the organization of the respiratory operon from the non-sulfur photosynthetic bacterium Rhodobacter capsulatus and its consequences for homologous expression of recombinant dimethylsulfoxide/trimethylamine N-oxide reductase (DMSOR/TMAOR) . This enzyme is of special interest since molybdopterin dinucleotide is its only cofactor . Overexpression of the dmsA gene and production of active enzyme is not possible in E . coli because this bacterium is unable to supply the required complex molybdopterin cofactor . To investigate the catalytic mechanism and binding of the cofactor by structure-based site-directed mutagenesis, and to ensure sufficient production and incorporation of the complex cofactor, we designed a system for homologous expression in R . capsulatus.

Biol Chem, 1997 Mar-Apr, 378(3-4), 293 - 302
Isolation, cloning, sequence analysis and X-ray structure of dimethyl sulfoxide/trimethylamine N-oxide reductase from Rhodobacter capsulatus; Knablein J et al.; The periplasmic enzyme dimethyl sulfoxide/trimethylamine N-oxide reductase (DMSOR/TMAOR) from the photosynthetic purple bacterium Rhodobacter capsulatus functions as the terminal electron acceptor in its respiratory chain . The enzyme catalyzes the reduction of highly oxidized substrates like dimethyl sulfoxide (DMSO) or trimethylamine N-oxide (TMAO) . At a molybdenum redox centre, two single electrons are transferred from cytochrome c556 to the substrate, e.g . DMSO, generating dimethyl sulfide (DMS) and water . The operon encoding this enzyme was isolated, cloned and sequenced, and its chromosomal location determined . It was shown by analytical and crystallographic data that DMSOR and TMAOR are identical enzymes . Degenerate primers were derived from short peptide sequences and a 700 bp fragment was amplified by nested PCR, subsequently cloned and radioactively labeled to screen a prepared lambda DASH library . Positive lambda clones were subcloned into pBluescript and subsequently transformed into Escherichia coli to sequence the DMSOR/TMAOR operon . By an optimized protein purification high yields (5 mg protein/l culture) with a specific activity of 30 U/mg were obtained . The molecular mass was experimentally determined by electrospray mass spectroscopy (MS) to be 85034 Da and from the deduced amino acid sequence of the apoenzyme to be 85033 Da . The enzyme was crystallized in space group P4(1)2(1)2 with unit cell dimensions of a = b = 80.7 A and c = 229.2 A diffracting beyond 1.8 A . The three-dimensional structure was solved by a combination of multiple isomorphous replacement (MIR) and molecular replacement techniques . The atomic model was refined to an R-factor of 0.169 for 57394 independent reflections . The spherical protein consists of four domains with a funnel-like cavity that leads to the freely accessible metal-ion redox center . The sole bis(molybdopterin guanine dinucleotide)molybdenum cofactor (1541 Da) of the single chain protein has the molybdenum ion bound to the cis-dithiolene group of only one molybdopterin guanine dinucleotide (MGD) molecule . In addition, two oxo ligands and the oxygen of a serine side chain are bound to the molybdenum ion.

Biol Chem, 1997 Mar-Apr, 378(3-4), 223 - 30
Titration and mapping of the active site of cysteine proteinases from Porphyromonas gingivalis (gingipains) using peptidyl chloromethanes; Potempa J et al.; Porphyromonas gingivalis is one of the major pathogens associated with periodontal disease and releases powerful cysteine proteinases known as the gingipains, which are key virulence factors for this organism . The three forms of gingipains, gingipain R1, gingipain R2 (gingipain Rs) and gingipain K, which cleave specifically after arginine (R) or lysine (K) residues, were characterized in terms of the kinetics of their interaction with a wide range of synthetic peptidyl chloromethane inhibitors and a peptidyl (acyloxy)methane . Chloromethane inhibitors were found to inhibit all the enzymes to varying degree dependent on the peptidyl components of the inhibitor . Thus, inhibitors containing a basic residue at P1 rapidly inactivated the gingipains and some specificity could be seen at the P2 site . The (acyloxy)methane inhibitor, Cbz-Phe-Lys-CH2OCO-2,4,6-Me3-Ph, was very specific in its rapid inhibition of gingipain K over the gingipains R . This inhibitor, together with the peptidyl chloromethanes, D-Phe-Pro-Arg-CH2Cl and D-Phe-Phe-Arg-CH2Cl, which reacted most rapidly with the Arg-specific proteinases, could be used to active site titrate purified forms of the enzymes and enzymes found in crude fractions such as intact P . gingivalis cells, vesicles or membrane fractions . From these titrations it was evident that gingipains R were always in an excess of about 3-fold over gingipain K and that the gingipains as a whole made up 85% of the proteolytic activity associated with the bacterium . The elucidation of the kinetics of inhibition by the range of compounds and the development of the titration method for gingipains will considerably aid in future studies on the proteases elaborated by P . gingivalis.

Protein Eng, 1997 Mar, 10(3), 197 - 206
Three-dimensional model of sensory rhodopsin I reveals important restraints between the protein and the chromophore; Lin SL et al.; A structural model is constructed for the integral membrane protein, sensory rhodopsin I (SRI), the phototaxis receptor of the archaeon Halobacterium salinarium . The model is built on the template of the homologous bacteriorhodopsin (BR) . The modeling procedure includes sequence alignment, a side chain rotamer search and simulated annealing by restricted molecular dynamics . The structure is in general agreement with previous results from mutagenesis experiments, chromophore substitution and room and cryogenic temperature spectroscopy . In particular, a residue near the beta-ionone ring of the retinylidene chromophore is found to be critical in maintaining the proper isomeric conformation of the chromophore; a layer of residues lying on the cytoplasmic side of the chromophore pocket is found to modulate the restraints around the C13 region of the chromophore, affecting the isomerizations around its 13 = 14 bond that are important to the protein's activity . The restraints in these regions are more stringent in SRI than in BR . The tightened restraints are chiefly due to van der Waals interactions, where the attractive and repulsive components play separable roles . Aromatic residues account for a majority of the restrictive interactions . It is hypothesized that the enhanced barriers due to these restrictions regulate the progress of SRI's photocycle, so that it can couple with the phototaxis reaction chain in the bacterium . A possibility is also suggested that conformational changes of the protein provide the signal recognized by the transducer.

Vet Microbiol, 1997 Mar, 54(3-4), 385 - 92
In-vitro interactions of Lawsonia intracellularis with cultured enterocytes; McOrist S et al.; Strains of the obligately intracellular bacterium Lawsonia intracellularis, the etiologic agent of porcine proliferative enteropathy, were co-cultured in rat enterocyte cell cultures (IEC-18) and examined ultrastructurally . No regular surface arrays typical of surface or S-layers were visible on any bacterial strain, with or without Triton-X-100 detergent treatment . In separate experiments, there was no difference in the ability of L . intracellularis to attach and enter enterocytes with or without the presence of added bovine plasma fibronectin, or the peptide Arg-Gly-Ser . Interestingly, there was an increase in the invasiveness of L . intracellularis in the presence of the peptide Arg-Gly-Asp (RGD), in a dose-related manner . A reduction was observed in the ability of L . intracellularis to invade enterocytes in the presence of monovalent fragments of IgG monoclonal antibodies to an outer surface component of L . intracellularis . This neutralization showed an antibody concentration-dependent titration effect and was not apparent with co-cultures incorporating control antibodies . The exact nature of ligand and cell receptor interactions for L . intracellularis remain to be determined.

Semin Respir Infect, 1997 Mar, 12(1), 2 - 6
Q fever pneumonia; Antony SJ et al.; Pneumonia is one of the principal manifestations of Q fever, a disease caused by Coxiella burnetii . This bacterium can replicate only within cells, yet it is capable of surviving in the environment because it can withstand drying and substantial temperature variations . Livestock, especially sheep, goats and cattle, are a major reservoir of C burnetii . The organism is transmitted to humans by direct contact with animal products, especially during parturition . Aerosols have transmitted infection over considerable distances . The illness is characterized by an influenza-like syndrome with patchy pulmonary infiltrates . The pneumonia may be accompanied by biochemical evidence of mild hepatitis . The diagnosis is established serologically . Tetracycline or doxycycline provide effective therapy.

Eur J Gastroenterol Hepatol, 1997 Mar, 9(3), 231 - 3
Extraintestinal pathology associated with Helicobacter infection; Gasbarrini A et al.; Helicobacter pylori appears to be the most important cause of gastritis and gastric and duodenal ulcer . Its eradication, in fact, promotes healing of these diseases and significantly reduces ulcer recurrence . In the last few years, moreover, besides a local tissue damage, an association between H . pylori infection and various extraintestinal pathologies has also been described . The presence of more toxic strains of H . pylori and the multiplicity of the substances produced, directly or indirectly, in response to the bacterium, besides a genetic predisposition of the individual, are all factors which may enhance the ability of Helicobacter to generate both local and systemic damage.

Scand J Gastroenterol, 1997 Mar, 32(3), 285 - 7
The growth of primary low-grade B-cell gastric lymphoma is sustained by Helicobacter pylori; Cammarota G et al.; BACKGROUND: The growth of primary low-grade B-cell gastric mucosa-associated lymphoid tissue (MALT) lymphoma is an antigen-dependent process CASE: We were able to document the influence of Helicobacter pylori on the natural history of primary low-grade B-cell gastric MALT lymphoma in a case investigated by means of polymerase chain reaction for IgH rearrangement . In this case the presence or absence of the bacterium appeared to affect both clinical manifestations and histologic features of the neoplasia, with an acceleration or a slowing down of the neoplastic expansion, respectively . CONCLUSIONS: We identified H . pylori as an antigenic stimulus that supports the growth of MALT-type lymphoma . This finding enables us to make some considerations about the management of the disease.

Lett Appl Microbiol, 1997 Mar, 24(3), 221 - 5
Replication of plasmids with the p15A origin in Shewanella putrefaciens MR-1; Myers CR et al.; The plasmid pACYC184 was introduced into Shewanella putrefaciens MR-1 by electroporation . In 100% of the transformants examined, the plasmid was maintained as a free replicon outside the chromosome . This was the case whether or not the plasmid contained a 224-bp DNA insert derived from an open-reading frame of MR-1 genomic DNA . Therefore, in contrast to a report in the literature, plasmids containing the p15A origin of replication can replicate freely in S . putrefaciens MR-1, and do not make convenient vectors for gene replacement in this bacterium . However, we found that plasmids with the pMB1 origin of replication (e.g . pBR322) cannot replicate in MR-1 and could therefore have potential as vectors for gene replacement.

Mol Microbiol, 1997 Mar, 23(5), 1043 - 52
A gene involved in both protein secretion during growth and starvation-induced development encodes a subunit of the NADH:ubiquinone oxidoreductase in Myxococcus xanthus; Laval-Favre K et al.; The secretion of numerous proteins during vegetative growth of Myxococcus xanthus, and the multicellular development cycle induced upon starvation of these bacteria, are partially interrelated in so far as mutants impaired in extracellular protein production are unable to undergo development . We have cloned and sequenced a gene in which a Tn5 insertion leads to a decrease in the production of most, if not all, extracellular proteins, and prevents development and sporulation . The deduced protein is homologous to the putative ubiquinone-binding subunit of bacterial and mitochondrial NADH:ubiquinone oxidoreductases (complex I) . This is the first example of the presence of this complex in a bacterium from subclass delta of the proteobacteria . This gene is expressed during growth and during early development . As its disruption by Tn5 does not impair growth of the mutant strain, we assume the presence of a second alternative NADH oxidoreductase, and suggest that the phenotypic alterations caused by the mutation are due to a decrease in the proton-motive force.

Microb Pathog, 1997 Mar, 22(3), 133 - 42
Ultrastructural and molecular analyses of the persistence of Chlamydia trachomatis (serovar K) in human monocytes; Koehler L et al.; Previous studies have suggested that monocytes may play a role in the dissemination of Chlamydia trachomatis, and in establishment of persistent infection with this bacterium . Infection of cultured human peripheral blood monocytes with C . trachomatis serovar K produced persistent, nonproductive infection . Transmission electron microscopy of such infected cultures revealed single or multiple Chlamydia in monocyte inclusions over a culture period of 10 days . Those inclusions were aberrant, and normal reticulate bodies within the inclusions were not observed . Immunoelectron microscopy showed the chlamydial major outer membrane protein and lipopolysaccharide to be associated with the bacterial plasma membrane . Lipopolysaccharide was also identified in the monocyte cytoplasm . Molecular analyses of primary chlamydial rRNA transcripts demonstrated that the organism is viable and metabolically active within monocyte inclusions . However, attempts to overcome chlamydial growth arrest by incubation of Chlamydia-infected monocytes with tryptophan, and antibodies against alpha interferon, gamma interferon, or tumor necrosis factor, were all ineffective, suggesting that known mechanisms of growth inhibition do not hold in human monocytes . These observations indicate that infection of human peripheral blood monocytes with C . trachomatis may be involved in the genesis/maintenance of extra-urogenital inflammation, since non-culturable, metabolically active bacteria persist in those cells.

J Bacteriol, 1997 Mar, 179(6), 1946 - 50
Genomic organization of the acidophilic chemolithoautotrophic bacterium Thiobacillus ferrooxidans ATCC 21834; Irazabal N et al.; The genomic organization of the acidophilic chemolithoautotrophic bacterium Thiobacillus ferrooxidans ATCC 21834 has been studied by pulsed-field gel electrophoresis (PFGE) . Analysis of its intact DNA, as well as the restriction patterns obtained with several endonucleases, allowed the characterization of one circular chromosome of 2.9 Mb and one plasmid of 8.6 kb . The first complete and highly resolved physical map (86 restriction sites) of the chromosome of an acidophilic obligate chemolithoautotrophic bacterium has been constructed by using endonucleases PmeI, SwaI, XbaI, and SpeI . The rRNA and str operons have been located on the chromosomal physical map.

Am J Gastroenterol, 1997 Mar, 92(3), 387 - 93
Do ulcers burn out or burn on? Managing duodenal ulcer diathesis in the Helicobacter pylori era . Ad Hoc Committee on FDA-Related Matters; Neil GA; Now that the Food and Drug Administration is examining various treatment regimens for eradication of Helicobacter pylori (H . pylori) infection, the question of whom to treat has come to the forefront . Widespread attempts to eradicate the bacterium are not without risk, including the possibility of accelerating the emergence of resistant strains of the organism, in addition to possible adverse events of agents used to cure the infection . These risks are of concern to the Agency in the process of granting marketing approvals for various therapies . The association of H . pylori and duodenal ulcer (DU) is no longer disputed; however, data to date have been generated in studies of patients with active (acute) DU . In these patients, the benefits of eradication therapy are clear . Patients with a documented history of DU who do not have an ulcer crater at the time of presentation have not been well studied in controlled trials . If they are at similar risk for recurrence and complications, it follows that they, too, should be candidates for H . pylori testing and treatment . However, if, as some believe, duodenal ulcer disease becomes "inactive" or "burns out" with time, an argument could be made for expectant treatment for this subgroup of patients . The present review examines the available literature on the natural history of DU disease and explores the validity of the hypothesis of duodenal ulcer "burn out." Analysis of the data shows that there is little support for the phenomenon of duodenal ulcer disease "burn out," and that, in fact, DU disease resulting from H . pylori infection is a chronic, relapsing condition, lasting for decades, if not a lifetime . The literature is compatible with the view that, in the majority of patients, DU occurs on a background of an "ulcer diathesis" that is fueled by H . pylori infection, and "burns on, not out", until and unless the infection is extinguished . The true incidence of ulcer relapse is difficult to predict due to the common occurrence of asymptomatic ulcers and the poor correlation of symptoms with the presence of ulcer . Therefore, treatment of patients with a history of duodenal ulcer disease should be advocated, even if their ulcer disease is not "active," because the risk of recurrence and complications does not diminish unless the H . pylori bacterium is eradicated.

Am J Pathol, 1997 Mar, 150(3), 971 - 9
Role of neutrophils in a rat model of gastric ulcer recurrence caused by interleukin-1 beta; Watanabe T et al.; The production of several cytokines such as interleukin (IL)-1 in gastric mucosa is increased in subjects infected with Helicobacter pylori, a bacterium associated with ulcer recurrence . This study was performed to determine whether the administration of IL-1 beta can cause recurrence of gastric ulcers in rats . Rats with healed ulcers received an injection of IL-1 beta (0.01 to 1 microgram/kg) or vehicle alone . Some rats received an injection of antiserum to rat neutrophils at the same time as 1 microgram/kg IL-1 beta or an injection of monoclonal antibodies against adhesion molecules (anti-intercellular adhesion molecule-1, anti-CD11a, and anti-CD11b) at 0, 12, and 24 hours after the initial injection . At this dose of IL-1 beta, the numbers of neutrophils and monocytes/macrophages infiltrating the scarred mucosa were higher at 12 and 24 hours than without injection of IL-1 beta . By 48 hours, seven of the eight bealed ulcers in the group treated with 1 microgram/kg IL-1 beta had recurred, as had one of the seven healed ulcers in the group given 0.1 microgram/kg IL-1 beta . No recurrence was found in the rats treated with 0.01 microgram/kg IL-1 beta or vehicle alone . Treatment with antiserum to neutrophils or antibodies to adhesion molecules inhibited both neutrophil infiltration into the scarred mucosa and the ulcer recurrence caused by IL-1 beta . These findings suggest possible mechanisms of recurrence of human peptic ulcers.

Appl Environ Microbiol, 1997 Mar, 63(3), 962 - 8
Role of the histidine kinase, EnvZ, in the production of outer membrane proteins in the symbiotic-pathogenic bacterium Xenorhabdus nematophilus; Forst SA et al.; We show that inactivation of envZ, the gene encoding the histidine kinase sensor protein, EnvZ, of Xenorhabdus nematophilus, affected the production of several outer membrane proteins (Opns) . X . nematophilus produced five major Opns during exponential growth . Insertional inactivation of envZ led to a decrease in the production of OpnP, the OmpF-like pore-forming protein which constitutes approximately 50% of the total outer membrane protein in X . nematophilus . OpnA production was also reduced, while the remaining Opns were produced normally . During the transition to stationary phase, three new outer membrane proteins, OpnB, OpnS, and OpnX, were induced in the wild-type strain . The envZ-minus strain, ANT1, did not produce OpnB and OpnX, while OpnS was induced at markedly reduced levels . These results suggest that EnvZ was required for the high-level production of OpnP during exponential growth and may be involved in the production of OpnB, OpnS, and OpnX during stationary-phase growth . We also show that ANT1 was more pathogenic than the wild-type strain when as few as five cells were injected into the hemolymph of the larval stage of the tobacco hornworm (Manduca sexta) . The larvae died before significant numbers of bacteria were detectable in the hemolymph . These results are discussed in relation to the role of EnvZ in the life cycle of X . nematophilus.

J Bacteriol, 1997 Mar, 179(5), 1460 - 8
Regulation of synthesis of pyruvate carboxylase in the photosynthetic bacterium Rhodobacter capsulatus; Yakunin AF et al.; The synthesis of pyruvate carboxylase (PC) was studied by using quantitative immunoblot analysis with an antibody raised against PC purified from Rhodobacter capsulatus and was found to vary 20-fold depending on the growth conditions . The PC content was high in cells grown on pyruvate or on carbon substrates metabolized via pyruvate (lactate, D-malate, glucose, or fructose) and low in cells grown on tricarboxylic acid (TCA) cycle intermediates or substrates metabolized without intermediate formation of pyruvate (acetate or glutamate) . Under dark aerobic growth conditions with lactate as a carbon source, the PC content was approximately twofold higher than that found under light anaerobic growth conditions . The results of incubation experiments demonstrate that PC synthesis is induced by pyruvate and repressed by TCA cycle intermediates, with negative control dominating over positive control . The content of PC in R . capsulatus cells was also directly related to the growth rate in continuous cultures . The analysis of intracellular levels of pyruvate and TCA cycle intermediates in cells grown under different conditions demonstrated that the content of PC is directly proportional to the ratio between pyruvate and C4 dicarboxylates . These results suggest that the regulation of PC synthesis by oxygen and its direct correlation with growth rate may reflect effects on the balance of intracellular pyruvate and C4 dicarboxylates . Thus, this important enzyme is potentially regulated both allosterically and at the level of synthesis.

J Clin Microbiol, 1997 Mar, 35(3), 780 - 2
Recurrent CDC group IVc-2 bacteremia in a human with AIDS; Anderson RR et al.; A 30-year-old man with AIDS developed separate episodes of bacteremia with multiple organisms including CDC group IVc-2, related to an indwelling central venous catheter . Recovery at each interval followed antibiotic therapy and replacement of the central venous access . This is the first reported case of infection with this organism in a patient with AIDS . A review of the biochemical features of this bacterium as well as pertinent literature is presented.

J Clin Microbiol, 1997 Mar, 35(3), 724 - 9
Simple diagnosis of Encephalitozoon sp . microsporidial infections by using a panspecific antiexospore monoclonal antibody; Enriquez FJ et al.; Microsporidia (phylum Microsproa) have recently become recognized as common opportunistic protozoans in the United States and worldwide, particularly affecting immunodeficient patients . Microsporidian organisms within the genus Encephalitozoon are the cause of nephrologic, ophthalmic, pneumologic, gastroenteric, and systemic infections . However, diagnosis of the small spores by light microscopy is difficult, even with newly developed and improved staining techniques . We have developed an anti-Encephalitozoon species monoclonal antibody-based immunoassay for easy diagnosis . A hybridoma was produced and selected following one main criterion: recognition by immunofluorescence of all known Encephalitozoon spores affecting humans . The selected monoclonal antibody-secreting hybridomas were characterized by enzyme-linked immunosorbent assay, immunofluorescence, Western blot, and immunoelectron microscopy using Encephalitozoon species from fresh and fixed samples from patients and from in vitro cultures . In the immunofluorescence assay, one monoclonal antibody, termed 3B6, strongly recognized Encephalitozoon cuniculi, E . hellem, and E . intestinalis . Monoclonal antibody 3B6 bound to other microsporidia (Nosema and Vairimorpha spp.) without cross-reacting with any other parasite, including Enterocytozoon bieneusi, fungus, or bacterium tested . In immunoelectron microscopy assays, monoclonal antibody 3B6 bound to the exospore of Encephalitozoon species, while in Western blot assays, it recognized three to seven antigens with molecular masses ranging from 34 to 117 kDa . We have developed a sensitive and specific monoclonal antibody-based immunoassay to diagnose common microsporidian infections, particularly with Encephalitozoon species . This is a new tool for identifying spores in bodily fluids and biopsy samples and is an efficient diagnostic test . Additionally, monoclonal antibody 3B6 can serve to assess the prevalence of microsporidial infections in immunodeficient and immunocompetent patients.

Biochim Biophys Acta, 1997 Feb 28, 1350(3), 235 - 9
Cloning of dnaK and dnaJ homologous genes from a purple non-sulfur bacterium Rhodopseudomonas species; Momma K et al.; The dnaK and dnaJ genes were isolated as a cluster from a purple non-sulfur phototrophic bacterium, Rhodopseudomonas species No . 7 by a polymerase chain reaction (PCR) based method . The deduced products of dnaK (631 amino acids) and dnaJ (379 amino acids) were 67% and 56% identical to the respective Escherichia coli gene products . The functions of DnaK and DnaJ could be confirmed by complementation of the respective E . coli mutants.

Biochemistry, 1997 Feb 25, 36(8), 2178 - 87
Site-directed mutations near the L-subunit D-helix of the purple bacterial reaction center: a partial model for the primary donor of photosystem II; Coleman WJ et al.; We have engineered a photosynthetically competent mutant of the purple non-sulfur bacterium Rhodobacter capsulatus which seeks to mimic the behavior of the primary electron donor (P) of the plant photosystem II (PS II) reaction center (RC) . To construct this mutant (denoted D1-ILMH), four residues in the bacterial L subunit were mutagenized, such that an 11-residue segment was made identical to the analogous segment from the D1 subunit of PS II . The electronic properties of the bacteriochlorophyll (Bchl) dimer which constitutes the primary donor are substantially altered by these modifications, to the degree that the dimer becomes functionally much more "monomeric" . The changes include (1) an increase in the values of the zero-field splitting (ZFS) parameters, as measured by electron paramagnetic resonance (EPR), for the spin-polarized triplet state, 3P, from /D/ = 185 x 10(-4) cm(-1) and /E/ = 31 x 10(-4) cm(-1) in wild-type (WT) chromatophore membranes to /D/ = 200 x 10(-4) cm(-1) and /E/ = 44 x 10(-4) cm(-1) in the mutant and (2) an increase in the EPR line width of the oxidized state, P+, from 0.97 mT in WT to 1.09 mT in D1-ILMH RCs . However, unlike the PS II primary donor (P680), the orientation of 3P in the D1-ILMH mutant is the same as in WT bacteria and does not display the unusual orientation found for PS II . And whereas the redox couple P/P+ has a very high midpoint potential in PS II, P/P+ in the D1-ILMH mutant has a lower midpoint (90 mV more negative) than in WT Rb . capsulatus . In addition, Raman measurements indicate that the hydrogen bond between HisL168 and the C2 acetyl carbonyl oxygen of the Bchl on the active electron transfer pathway (P(A)) is absent in the mutant, due to the fact that HisL168 in the WT sequence has been replaced by a leucine in D1-ILMH . However, the Raman data also reveal the presence of a new hydrogen bond in the D1-ILMH RCs, between the C9 keto carbonyl oxygen of P(A) and an unknown hydrogen-bond donor . Thus, although the protein environment around one of the Bchls of the special pair is significantly changed in D1-ILMH, the chimeric RC does not, as a result of these changes, have a primary donor that is oriented like the one in PS II.

Biochemistry, 1997 Feb 18, 36(7), 1927 - 32
Membrane-associated c-type cytochromes from the green sulfur bacterium Chlorobium limicola forma thiosulfatophilum: purification and characterization of cytochrome c553; Albouy D et al.; Tetraheme cytochromes involved in photosynthetic electron transport have previously been described associated with the reaction centers of purple photosynthetic bacteria; however, similar heme proteins have not until now been characterized in the phylogenetically distinct green sulfur bacteria . In this paper we describe the first isolation and characterization of a multitheme, membrane-associated cytochrome from a green sulfur bacterium, Chlorobium limicola forma thiosulfatophilum . We show that this cytochrome contains a single polypeptide of 32 kDa apparent molecular mass on SDS-PAGE and has a characteristic broad alpha-band absorption at 553 nm . By both low-temperature absorption and electron paramagnetic resonance spectroscopy, we demonstrate that there are at least four distinct heme groups.

Structure, 1997 Feb 15, 5(2), 217 - 25
The structure of an energy-coupling protein from bacteria, IIBcellobiose, reveals similarity to eukaryotic protein tyrosine phosphatases; van Montfort RL et al.; BACKGROUND: . The bacterial phosphoenolpyruvate-dependent phosphotransferase system (PTS) mediates the energy-driven uptake of carbohydrates and their concomitant phosphorylation . In addition, the PTS is intimately involved in the regulation of a variety of metabolic and transcriptional processes in the bacterium . The multiprotein PTS consists of a membrane channel and at least four cytoplasmic proteins or protein domains that sequentially transfer a phosphoryl group from phosphoenolpyruvate to the transported carbohydrate . Determination of the three-dimensional structure of the IIB enzymes within the multiprotein complex would provide insights into the mechanisms by which they promote efficient transport by the membrane channel IIC protein and phosphorylate the transported carbohydrate on the inside of the cell . RESULTS: . The crystal structure of the IIB enzyme specific for cellobiose, IIBcellobiose (molecular weight 11.4 kDa), has been determined to a resolution of 1.8 and refined to an R factor of 18.7% (Rfree of 24 . 1%) . The enzyme consists of a single four-stranded parallel beta sheet flanked by helices on both sides . The phosphorylation site (Cys 10) is located at the C-terminal end of the first beta strand . No positively charged residues, which could assist in phosphoryl-transfer, can be found in or near the active site . The fold of IIBcellobiose is remarkably similar to that of the mammalian low molecular weight protein tyrosine phosphatases . CONCLUSIONS: . A comparison between IIBcellobiose and the structurally similar low molecular weight protein tyrosine phosphatases provides insight into the mechanism of the phosphoryltransfer reactions in which IIBcellobiose is involved . The differences in tertiary structure and active-site composition between IIBcellobiose and the glucose-specific IIBglucose give a structural explanation why the carbo-hydrate-specific components of different families cannot complement each other.

J Biol Chem, 1997 Feb 14, 272(7), 4467 - 73
Characterization of the Bradyrhizobium japonicum CycY protein, a membrane-anchored periplasmic thioredoxin that may play a role as a reductant in the biogenesis of c-type cytochromes; Fabianek RA et al.; A new member of membrane-anchored periplasmic thioredoxin-like proteins was identified in Bradyrhizobium japonicum . It is the product of cycY, the last gene in a cluster of cytochrome c biogenesis genes . Mutational analysis revealed that cycY is essential for the biosynthesis of all c-type cytochromes in this bacterium . The CycY protein was shown to be exported to the periplasm by its N-terminal signal sequence-like domain . Results from Western blot analyses of membrane and soluble fractions indicated that the CycY protein remains bound to the membrane . A soluble version of the protein devoid of its N-terminal membrane anchor (CycY*) was expressed in Escherichia coli and purified to homogeneity from the periplasmic fraction . The protein showed redox reactivity and properties similar to other thioredoxins such as fluorescence quenching in the oxidized form . Its equilibrium constant with glutathione was determined to be 168 mM, from which a standard redox potential of -0.217 V was calculated, suggesting that CycY might act as a reductant in the otherwise oxidative environment of the periplasm . This is in agreement with our hypothesis that CycY is required, directly or indirectly, for the reduction of the heme-binding site cysteines in the CXXCH motif of c-type apocytochromes before heme attachment occurs.

Biochemistry, 1997 Feb 11, 36(6), 1428 - 40
Structure and function of cytochrome c2 in electron transfer complexes with the photosynthetic reaction center of Rhodobacter sphaeroides: optical linear dichroism and EPR; Drepper F et al.; The photosynthetic reaction center (RC) and its secondary electron donor the water-soluble cytochrome (cyt) c2 from the purple bacterium Rhodobacter sphaeroides have been used in cross-linked and non-cross-linked complexes, oriented in compressed gels or partially dried multilayers, to study the respective orientation of the primary donor P (BChl dimer) and of cyt c2 . Three methods were used: (i) Polarized optical absorption spectra at 295 and 10 K were measured and the linear dichroism of the two individual transitions (Qx, Qy), which are nearly degenerate within the alpha-band of reduced cyt c2, was determined . Attribution of the polarization directions to the molecular axes within the heme plane yielded the average cyt orientation in the complexes . (ii) Time-resolved flash absorption measurements using polarized light allowed determination of the orientation of cyt c2 in complexes which differ in their kinetics of electron transfer . (iii) EPR spectroscopy of ferricyt c2 in cross-linked RC-cyt c2 complexes was used to determine the angle between the heme and the membrane plane . The results suggest the following structural properties for the docking of cyt c2 to the RC: (i) In cross-linked complexes, the two cytochromes displaying half-lives of 0.7 and 60 micros for electron transfer to P+ are similarly oriented (difference < 10 degrees) . (ii) For cross-linked cyt c2 the heme plane is parallel to the symmetry axis of the RC (0 degrees +/- 10 degrees) . Moreover, the Qy transition, which is assumed to be polarized within the ring III-ring I direction of the heme plane, makes an angle of 56 degrees +/- 1 degree with the symmetry axis . (iii) The dichroism spectrum for the fast phase (0.7 micros) for the non-cross-linked cyt c2-RC complex suggests an orientation similar to that of cross-linked cyt c2, but the heme plane is tilted about 20 degrees closer to the membrane . An alternative model is that two or more bound states of cyt c2 with heme plane tilt angles between 0 degrees and 30 degrees allow the fast electron transfer . Zero-length cross-linking of cyt c2 may take place in one of these bound states . These orientations of cyt c2 are compared to different structural models of RC-cyt c2 complexes proposed previously . The relation of the two kinetic phases observed in cross-linked cyt c2 complexes to biphasic kinetics of the mobile reaction partners is discussed with respect to the dynamic electrostatic interactions during the formation of a docking complex and its dissociation . A mechanism is proposed in which a pre-orientation of cyt c2 relative to the membrane plane occurs by interaction of its strong electrostatic dipole with the negative surface charges of the RC . The optimal matching of the oppositely charged surfaces of the two proteins necessitates further rotation of the cyt around its dipole axis.

FEBS Lett, 1997 Feb 10, 403(1), 10 - 4
Anaerobic carotenoid biosynthesis in Rhodobacter sphaeroides 2.4.1: H2O is a source of oxygen for the 1-methoxy group of spheroidene but not for the 2-oxo group of spheroidenone; Yeliseev AA et al.; Anaerobic biosynthesis of carotenoids in the purple facultative photosynthetic bacterium Rhodobacter sphaeroides was studied using mass spectrometry . We have demonstrated that (18)O from H2(18)O was incorporated into the 1-methoxy group of spheroidene and spheroidenone, the two major carotenoids produced by this bacterium during photosynthetic growth . Neither water nor CO2 was shown to provide an oxygen atom for the 2-oxo group of spheroidenone in R . sphaeroides 2.4.1 grown photosynthetically in the absence of molecular oxygen . Possible mechanisms for the anaerobic biosynthesis of spheroidenone in R . sphaeroides are discussed.

Proc Natl Acad Sci U S A, 1997 Feb 4, 94(3), 940 - 5
5-Methylcytosine is not a mutation hot spot in nondividing Escherichia coli; Lieb M et al.; Spontaneous deamination of 5-methylcytosine (5meC) causes hot spots of CxG --> TxA mutations in Escherichia coli and in human cells . In E . coli, the resulting TxG mispairs can be corrected to CxG by very short patch (VSP) repair, which requires the product of gene vsr . Mutation hot spots in genes of replicating vsr+ bacteria are attributable to low Vsr activity . To determine the rate of deamination of 5meC and the efficiency of VSP repair in nondividing bacteria, we used kanamycin-sensitive (KanS) lysogens containing a lambda kan- prophage . Deamination of a 5meC in the kan- gene resulted in mutation to kanamycin resistance (KanR) . Lysogens containing a single lambda kan- prophage per bacterial genome were grown in synthetic medium with limiting amino acids and stored at 15 degrees C or 37 degrees C . In the absence of VSP repair, KanR mutants accumulated at the rate of approximately 1.3 x 10(-7) per bacterium per day at 37 degrees C . This is similar to the 5meC --> T mutation rate reported for DNA in solution . In vsr+ bacteria, the KanR accumulation rate was 3 x 10(-9) per bacterium per day, which is not significantly higher than the rate observed when the target cytosine was unmethylated . The increase in KanR mutants was barely detectable in vsr+ cultures stored at 15 degrees C for 4 months . It is likely that mutation hot spots at 5meC in rapidly dividing cells are attributable to insufficient time for TxG correction in the interval between deamination of 5meC and subsequent DNA replication . DNA synthesis occurred in bacteria starved for amino acids and this synthesis was not highly mutagenic.

Mutat Res, 1997 Feb 3, 373(2), 277 - 84
On the origin of spontaneous somatic mutations and sectored plaques detected in transgenic mice; Paashuis-Lew Y et al.; The use of transgenic mice with bacterial genes that can be readily recovered and analysed for mutation has made it possible to measure mutant frequencies in many tissues . The mutations are detected by packaging the murine DNA into lambda phage and then growing these phage on a bacterial lawn under conditions such that the mutants are distinguishable from nonmutants . In the lacI mouse assay, the mutant plaques are blue whereas the nonmutant plaques are clear . The mutations detected in the lacI Big Blue Mouse are, in principle, a mixture of mutations that arose in the mouse (in vivo mutations), mutations that arose in the bacterium from lesions pre-existing in the murine DNA (ex vivo mutations), and mutations that arose during growth of the phage on the bacteria (in vitro mutations) . It has been suggested that plaque morphology can be used to visually distinguish in vivo mutations (which would be seen as wholly blue plaques) from ex vivo mutations (which would produce blue and white sectored plaques) and in vitro mutations (which would produce sectored or pin-point plaques) . We show here that this is not the case: ex vivo mutations produce plaques that are a homogeneous blue . By superinfection of bacteria with mutant and nonmutant phage and by in vitro mutagenesis, we found that blue plaques may contain large proportions of nonmutant phage . None of the mutant plaques seen after in vitro mutagenesis were sectored but most contained nonmutant phage . In addition, we show that most spontaneous mutations from the small intestine, which has a higher than normal mutant frequency, arose in vivo, since 17 of 17 mutants were homogeneous mutants . Sectored plaques must arise in some way other than by ex vivo mutation, like perhaps by the confluence of a mutant and nonmutant plaque.

Zoolog Sci, 1997 Feb, 14(1), 69 - 75
Cloning and sequencing of gene coding for a periplasmic 5.4 kDa peptide of the macronucleus-specific symbiont Holospora obtusa of the ciliate Paramecium caudatum; Dohra H et al.; We purified a 5.4 kDa peptide which is present in the intermediate and infectious form, but not in the reproductive form of a symbiotic bacterium Holospora obtusa of the ciliate Paramecium caudatum . Sequencing of its gene revealed that it encodes a peptide composed of 49 amino acids, and that the peptide is preceded by a putative signal peptide of 19 amino acids . We determined the transcription start point for the gene by primer extension analysis, indicating that the transcription starts with a G nucleotide located 33 nucleotides upstream from the translational initiation codon . Northern blot hybridization showed that the gene is highly expressed in the intermediate form, a transitional stage from the reproductive to infectious form of the bacterium . Immunoelectron microscopy with anti-5.4 kDa peptide antiserum revealed that the 5.4 kDa peptide is localized in the periplasm of the infectious form.

Arch Microbiol, 1997 Feb-Mar, 167(2-3), 99 - 105
The effect of respiration on the phototactic behavior of the purple nonsulfur bacterium Rhodospirillum centenum; Romagnoli S et al.; The effect of respiration on the positive phototactic movement of swarming agar colonies of the facultative phototroph Rhodospirillum centenum was studied . When the electron flow was blocked at the bc1 complex level by myxothiazol, the oriented movement of the colonies was totally blocked . Conversely, inhibition of respiration via the cytochrome c oxidase stimulated the phototactic response . No phototaxis was observed in a photosynthesis deficient mutant (YB707) lacking bacteriochlorophylls . Analyses of the respiratory activities as monitored by a oxygen microelectrode in single agar colonies during light/dark transitions showed a close functional correlation between the photosynthetic and respiratory apparatuses . The respiratory chain of Rsp . centenum was formed by two oxidative pathways: one branch leading to a cytochrome c oxidase inhibited by low cyanide concentrations and a second pathway formed by an oxidase less-sensitive to cyanide that also catalyzes the light-driven respiration . These results were interpreted to indicate that (1) there is a cyclic electron transport, and (2) photoinduced cyclic electron flow is required for the phototactic response of Rsp . centenum . Furthermore, under oxic conditions in the light, reducing equivalents may switch from photosynthetic to respiratory components so as to reduce both the membrane potential and the rate of locomotion.

Glycoconj J, 1997 Feb, 14(2), 231 - 9
Screening for the presence of polyglycosylceramides in various tissues: partial characterization of blood group-active complex glycosphingolipids of rabbit and dog small intestines; Miller-Podraza H et al.; Twenty different human and animal tissues were investigated for the presence of polyglycosylceramides . The glycolipids were isolated by peracetylation of dry tissue residues left after conventional lipid extraction, followed by extraction with chloroform and subsequent Sephadex LH-20, Sephadex LH-60 and silica gel chromatography . In most of the cases only trace amounts of complex glycolipids were found . Distinct bands of glycosphingolipids migrating on TLC plates in a region of brain gangliosides and below were observed in bovine erythrocytes, human leukocytes and human colon mucosa . Definite fractions of polyglycosylceramides were isolated from rabbit small intestine, dog small intestine, human placenta and human leukocytes . The polyglycosylceramides of dog and rabbit intestine were characterized by colorimetric analysis, methylation analysis, mass spectrometry and immunological assays . The dog material contained branched carbohydrate chains with repeated fucosylated N-acetyllactosamine units . Rabbit intestine polyglycosylceramides resembled rabbit erythrocyte polyglycosylceramides with Hex-Hex- terminal determinants but were more complex in respect of sugar composition and structure . The material isolated from dog intestine showed A, H, Le(x) and Le(y) blood group activities . Polyglycosylceramides of human erythrocytes, placenta and leukocytes showed strong binding affinity for Helicobacter pylori, while polyglycosylceramide fractions from rabbit and dog intestine were receptor-inactive for this bacterium or displayed only weak and poorly reproducible binding.

Eur J Oral Sci, 1997 Feb, 105(1), 2 - 8
Presence of bacteriophage Aa phi 23 correlates with the population genetic structure of Actinobacillus actinomycetemcomitans; Haubek D et al.; Several bacteriophages associated with the oral bacterium Actinobacillus actinomycetemcomitans have been identified . Lysogeny might affect the virulence of this bacterium, which has been implicated in the etiology of juvenile and adult periodontitis . We have determined the presence of bacteriophage Aa phi 23-related DNA sequences among 185 A . actinomycetemcomitans strains belonging to 2 well-characterized collections and have related the findings to the population genetic structure of the collections . 2 cloned Aa phi 23-specific DNA probes were used in Southern blot hybridization experiments to detect homologous sequences in whole-cell DNA of the strains . DNA from 65 (35%) of the 185 strains hybridized to either of the DNA probes . The majority (74%) of the hybridizing strains showed an identical hybridization pattern, indicating presence of phage Aa phi 23 . Whole-cell DNA from the remaining hybridizing strains hybridized to the probes with different patterns, indicating that DNA sequences related to but different from phage Aa phi 23 occur in these strains . The majority (81%) of the strains which harbored phage Aa phi 23 were of serotype a, whereas serotype d strains appeared to be resistant to infection with this phage . There was a clear correlation between hybridization patterns and genetic subdivisions based on our previous population genetic analyses of A . actinomycetemcomitans . However, there was no significant correlation between occurrence of Aa phi 23 among A . actinomycetemcomitans strains and the periodontal status of the patients from whom the isolates were obtained, suggesting that this bacteriophage does not significantly influence the virulence of A . actinomycetemcomitans.

Gut, 1997 Feb, 40(2), 196 - 203
Epithelium related deposition of activated complement in Helicobacter pylori associated gastritis; Berstad AE et al.; BACKGROUND AND AIMS: It is unknown whether Helicobacter pylori infection activates complement in vivo . Mucosal deposition of various activation products of the complement system may contribute to the pathogenesis of chronic gastritis and was therefore studied by immunohistochemistry . PATIENTS AND METHODS: Ethanol fixed antrum or body gastric tissue sections from 24 patients infected with H pylori (determined by bacterial immunohistochemistry) and 22 uninfected patients were examined by immunofluorescence with monoclonal antibodies to activation neoepitopes in C3b and in the terminal complex (TCC) . As a control group, biopsy samples from the gastric stump of 23 Billroth II operated patients were studied . RESULTS: Patchy, bright staining for TCC occurred below the surface epithelium and around the glands in H pylori positive and negative gastritis as well as in stump gastritis but seldom in normal mucosa . Activated C3 was present at the apical face of the surface epithelium, significantly more often in the antrum and body from patients with than without H pylori infection (p = 0.05 and p = 0.03 respectively), and particularly in samples with granulocyte infiltration (p = 0.04) . Many bacteria were coated with activated C3 towards the pit openings but seldom within the foveolae . CONCLUSIONS: Local complement activation was shown to take place in simple chronic gastritis, associated as well as unassociated with H pylori infection, and also in stump gastritis . The fact that activated C3 was seldom seen on H pylori within the foveolae, suggested that the bacterium evades complement attack in this location.

Biosci Biotechnol Biochem, 1997 Feb, 61(2), 365 - 6
Effect of acetic acid bacterium on ethanol oxidation in vivo; Nomura Y et al.; We investigated the possible effects of acetic acid bacterium on ethanol oxidation in vivo by monitoring the blood ethanol level after injecting 5% ethanol with (treated group) or without (control group) a freeze-dried bacterial cell suspension directly into the stomach of anesthesized rats . Paired comparison t-tests of the results indicate that the blood ethanol concentration of the rats in the treated group was significantly (p < 0.07) lower than that in the control group . When measured 10 min after administering ethanol into the gullet, the concentration in the stomach of the rats that received acetic acid bacterium simultaneously with ethanol was significantly (p < 0.10) lower than that of the rats that received ethanol alone . We consider that freeze-dried cells of acetic acid bacterium oxidized ethanol in the stomach and could be effective for reducing the blood ethanol level after drinking.

Immunol Cell Biol, 1997 Feb, 75(1), 35 - 40
Regulation of the expression of Mycobacterium avium complex proteins differs according to the environment within host cells; Bermudez LE et al.; Mycobacterium avium is an intracellular organism that can infect a number of cell types such as macrophages and epithelial cells . Each one of these cells represents a different environment that requires specific adaptation from the bacterium . The effect of uptake of M . avium and M . smegmatis by both human monocyte-derived macrophages in culture for 6 days, and HT-29 intestinal mucosal cell line on the bacterial synthesis of proteins were comparatively examined . Incorporation of {35S}-methionine by the bacterium was measured at 30 min, 2, 4, and 24 h after infection . Effect of the uptake by cells was compared with bacteria not exposed to cells and bacteria submitted to different stresses such as heat, hyperosmolarity and acid pH . Uptake of M . avium by macrophages triggered the synthesis of 93, 65, 55 and 33 kDa, among other proteins in the bacteria . Between 2 and 4 h of exposure to the intracellular millieu, a number of additional proteins have their synthesis up-regulated such as 39, 31, 43, 42, 61 and 70 kDa . In contrast, uptake by epithelial cells is associated with the up-regulation of 27, 65, 71 and 72 kDa proteins, among others . In this case, exposure to the intracellular environment was associated with expression of a number of proteins that do not vary with time . The results of this study suggest that regulation of the expression of proteins in M, avium varies according to the mammalian cell bacteria they are exposed to, and is influenced by the stage of intracellular infection.

Microbiology, 1997 Feb, 143 ( Pt 2), 519 - 22
Oligonucleotide-mediated genetic transformation of Borrelia burgdorferi; Samuels DS et al.; We have used short oligonucleotides to genetically transform the Lyme disease spirochaete Borrelia burgdorferi . The oligonucleotides are derived from the sequence of an Arg-133 to Ile mutant gyrB (chromosomal) gene that confers resistance to the antibiotic coumermycin A1 . Oligonucleotides were about 10,000-fold less efficient at transformation, on a molar basis, than longer PCR-generated substrates . All of the transformants tested contained the predicted site-directed silent mutation in their gyrB genes . Antisense oligonucleotides were more efficient at transformation than either sense or double-stranded oligonucleotides . This is the first demonstration of oligonucleotides used to introduce site-directed mutations directly into the genome of a bacterium.

FEMS Microbiol Lett, 1997 Feb 1, 147(1), 51 - 6
Methanoplanus petrolearius sp . nov., a novel methanogenic bacterium from an oil-producing well; Ollivier B et al.; A disc-shaped methanogenic bacterium designated strain SEBR 4847T (T = type strain) was isolated from a sample collected from an African offshore oil field . Strain SEBR 4847T was non-motile, had a G + C content of 50 mol% and produced methane from H2 + CO2, formate, and CO2 + propanol . Strain SEBR 4847T grew optimally at 37 degrees C; no growth was observed at 25 degrees C or 45 degrees C . It grew in the presence of up to 50 g/l NaCl; 10-30 g/l was required for optimal growth . The optimum pH for growth was 7.0 . Doubling time was about 10 h under optimal conditions . Based on 16S rRNA sequence analysis, the isolate was identified as a new species of the genus Methanoplanus and designated Methanoplanus petrolearius sp . nov . The type strain is SEBR 4847T (= OCM 486).

Appl Environ Microbiol, 1997 Feb, 63(2), 448 - 53
Use of rRNA gene restriction patterns to evaluate lactic acid bacterium contamination of vacuum-packaged sliced cooked whole-meat product in a meat processing plant; Bjorkroth KJ et al.; Molecular typing was applied to an in-plant lactic acid bacterium (LAB) contamination analysis of a vacuum-packaged sliced cooked whole-meat product . A total of 982 LAB isolates from the raw mass, product, and the environment at different production stages were screened by restriction endonuclease (EcoRI and HindIII) analysis . rRNA gene restriction patterns were further determined for different strains obtained from each source . These patterns were used for recognizing the spoilage-causing LAB strains from the product on the sell-by day and tracing the sources and sites of spoilage LAB contamination during the manufacture . LAB typing resulted in 71 different ribotypes, of which 27 were associated with contamination routes . Raw material was distinguished as the source of the major spoilage strains . Contamination of the product surfaces after cooking was shown to be airborne . The removal of the product from the cooking forms was localized as a major site of airborne LAB contamination . Food handlers and some surfaces in contact with the product during the manufacture were also contaminated with the spoilage strains . Some LAB strains were also able to resist cooking in the core of the product bar . These strains may have an effect on the product shelf life by contaminating the slicing machine . The air in the slicing department and adjacent cold room contained very few LAB . Surface-mediated contamination was detected during the slicing and packaging stages . Food handlers also carried strains later found in the packaged product . Molecular typing provided useful information revealing the LAB contamination sources and sites of this product . The production line will be reorganized in accordance with these results to reduce spoilage LAB contamination.

Infect Immun, 1997 Feb, 65(2), 571 - 8
Topology of Legionella pneumophila DotA: an inner membrane protein required for replication in macrophages; Roy CR et al.; The Legionella pneumophila dotA gene is required for intracellular growth of the bacterium in macrophages . In this study, a structure-function analysis of the DotA protein was conducted to elucidate the role of this protein in L . pneumophila pathogenesis . Translational fusions of dotA to the Escherichia coli phoA and lacZ genes indicated that DotA is an integral cytoplasmic membrane protein with eight membrane-spanning domains . DotA contains two large periplasmic domains of approximately 503 and 73 amino acids and a carboxyl-terminal cytoplasmic domain of 122 amino acids . Protein fractionation studies were consistent with DotA residing in the inner membrane . An alkaline phosphatase fusion located 9 amino acids upstream from the C terminus of DotA still retained function and was able to restore intracellular growth when harbored by two L . pneumophila dotA mutants . A hybrid protein from which the carboxyl-terminal 48 amino acids of DotA were deleted was unable to complement the intracellular growth defect in the dotA mutants, indicating that this cytoplasmic region is required for function.

J Bacteriol, 1997 Feb, 179(3), 941 - 8
Properties and gene structure of the Thermotoga maritima alpha-amylase AmyA, a putative lipoprotein of a hyperthermophilic bacterium; Liebl W et al.; Thermotoga maritima MSB8 has a chromosomal alpha-amylase gene, designated amyA, that is predicted to code for a 553-amino-acid preprotein with significant amino acid sequence similarity to the 4-alpha-glucanotransferase of the same strain and to alpha-amylase primary structures of other organisms . Upstream of the amylase gene, a divergently oriented open reading frame which can be translated into a polypeptide with similarity to the maltose-binding protein MalE of Escherichia coli was found . The T . maritima alpha-amylase appears to be the first known example of a lipoprotein alpha-amylase . This is in agreement with observations pointing to the membrane localization of this enzyme in T . maritima . Following the signal peptide, a 25-residue putative linker sequence rich in serine and threonine was found . The amylase gene was expressed in E . coli, and the recombinant enzyme was purified and characterized . The molecular mass of the recombinant enzyme was estimated at 61 kDa by denaturing gel electrophoresis (63 kDa by gel permeation chromatography) . In a 10-min assay at the optimum pH of 7.0, the optimum temperature of amylase activity was 85 to 90 degrees C . Like the alpha-amylases of many other organisms, the activity of the T . maritima alpha-amylase was dependent on Ca2+ . The final products of hydrolysis of soluble starch and amylose were mainly glucose and maltose . The extraordinarily high specific activity of the T . maritima alpha-amylase (about 5.6 x 10(3) U/mg of protein at 80 degrees C, pH 7, with amylose as the substrate) together with its extreme thermal stability makes this enzyme an interesting candidate for biotechnological applications in the starch processing industry.

Nature, 1997 Jan 30, 385(6615), 461 - 4
Crystal structure of colicin Ia; Wiener M et al.; The ion-channel forming colicins A, B, E1, Ia, Ib and N all kill bacterial cells selectively by co-opting bacterial active-transport pathways and forming voltage-gated ion conducting channels across the plasma membrane of the target bacterium . The crystal structure of colicin Ia reveals a molecule 210 A long with three distinct functional domains arranged along a backbone of two extraordinarily long alpha-helices . A central domain at the bend of the hairpin-like structure mediates specific recognition and binding to an outer-membrane receptor . A second domain mediates translocation across the outer membrane via the TonB transport pathway; the TonB-box recognition element of colicin Ia is on one side of three 80 A-long helices arranged as a helical sheet . A third domain is made up of 10 alpha-helices which form a voltage-activated and voltage-gated ion conducting channel across the plasma membrane of the target cell . The two 160 A-long alpha-helices that link the receptor-binding domain to the other domains enable the colicin Ia molecule to span the periplasmic space and contact both the outer and plasma membranes simultaneously during function.

Arch Microbiol, 1997 Jan 29, 167(1), 1 - 5
Differential cytochrome content and reductase activity in Geospirillum barnesii strain SeS3
Stolz JF, Gugliuzza T, Blum JS, Oremland R, Murillo FM.
The protein composition, cytochrome content, and reductase activity in the dissimilatory selenate-reducing bacterium Geospirillum barnesii strain SeS3, grown with thiosulfate, nitrate, selenate, or fumarate as the terminal electron acceptor, was investigated . Comparison of seven high-molecular-mass membrane proteins (105.3, 90.3, 82.6, 70.2, 67.4, 61.1, and 57.3 kDa) by SDS-PAGE showed that their detection was dependent on the terminal electron acceptor used . Membrane fractions from cells grown on thiosulfate contained a 70.2-kDa c-type cytochrome with absorbance maxima at 552, 522, and 421 nm . A 61.1-kDa c-type cytochrome with absorption maxima at 552, 523, and 423 nm was seen in membrane fractions from cells grown on nitrate . No c-type cytochromes were detected in membrane fractions of either selenate- or fumarate-grown cells . Difference spectra, however, revealed the presence of a cytochrome b554 (absorption maxima at 554, 523, and 422 nm) in membrane fractions from selenate-grown cells and a cytochrome b556 (absorption maxima at 556, 520, and 416 nm) in membrane fractions from fumarate-grown cells . Analysis of reductase activity in the different membrane fractions showed variability in substrate specificity . However, enzyme activity was greatest for the substrate on which the cells had been grown (e.g., membranes from nitrate-grown cells exhibited the greatest activity with nitrate) . These results show that protein composition, cytochrome content, and reductase activity are dependent on the terminal electron acceptor used for growth.

Proc Natl Acad Sci U S A, 1997 Jan 21, 94(2), 559 - 64
Inactivation of FtsI inhibits constriction of the FtsZ cytokinetic ring and delays the assembly of FtsZ rings at potential division sites; Pogliano J et al.; A universally conserved event in cell division is the formation of a cytokinetic ring at the future site of division . In the bacterium Escherichia coli, this ring is formed by the essential cell division protein FtsZ . We have used immunofluorescence microscopy to show that FtsZ assembles early in the division cycle, suggesting that constriction of the FtsZ ring is regulated and supporting the view that FtsZ serves as a bacterial cytoskeleton . Assembly of FtsZ rings was heterogeneously affected in an ftsI temperature-sensitive mutant grown at the nonpermissive temperature, some filaments displaying a striking defect in FtsZ assembly and others displaying little or no defect . By using low concentrations of the beta-lactams cephalexin and piperacillin to specifically inhibit FtsI (PBP3), an enzyme that synthesizes peptidoglycan at the division septum, we show that FtsZ ring constriction requires the transpeptidase activity of FtsI . Unconstricted FtsZ rings are stably trapped at the midpoint of the cell for several generations after inactivation of FtsI, whereas partially constricted FtsZ rings are less effectively trapped . In addition, FtsZ rings are able to assemble in newborn cells in the presence of cephalexin, suggesting that newborn cells contain a site at which FtsZ can assemble (the nascent division site) and that the transpeptidase activity of FtsI is not required for assembly of FtsZ at these sites . However, aside from this first round of FtsZ ring assembly, very few additional FtsZ rings assemble in the presence of cephalexin, even after several generations of growth . One interpretation of these results is that the transpeptidase activity of FtsI is required, directly or indirectly, for the assembly of nascent division sites and thereby for future assembly of FtsZ rings.

FEBS Lett, 1997 Jan 20, 401(2-3), 153 - 7
Very fast electron transfer from cytochrome to the bacterial photosynthetic reaction center at low temperature; Ortega JM et al.; Electron transfer from the proximal heme c-559 to the primary donor P has been studied in reaction centers of the photosynthetic bacterium Rhodopseudomonas viridis in which the tyrosine residue L162 was replaced by threonine . In the wild type, when the two high-potential hemes of the tetraheme cytochrome are reduced before flash excitation, a rapid electron transfer (t1/2 = 190 ns) observed at ambient temperature disappears below 190 K . In the mutant, the reaction is partly maintained down to 8 K, leading to irreversible charge separation . The reaction rate is nearly temperature-independent between 294 K and 8 K (t1/2 approximately 450 ns) . The different behavior of wild type and mutant reaction centers is attributed to differences in a network of water molecules, the freezing of which may block structural reorganizations associated with cytochrome oxidation, in the wild type but not in the mutant.

Chem Phys Lipids, 1997 Jan 17, 85(1), 75 - 89
Cryo-TEM and NMR studies of a micelle-forming phosphoglucolipid from membranes of Acholeplasma laidlawii A and B; Danino D et al.; The chemical structure of a phosphoglucolipid from the membrane of the bacterium Acholeplasma laidlawii strain B-PG9 has been determined by high resolution NMR to be 1,2-diacyl-3-O-{glycerophosphoryl-6-O-(alpha-D-glucopyranosyl-(1 -->2)-O-alpha-D-glucopyranosyl)}-sn-glycerol (GPDGlcDAG) . It was concluded that this lipid has exactly the same structure as one of the phosphoglucolipids from A . laidlawii strain A-EF22 . By cryo transmission electron microscopy (cryo-TEM) and NMR diffusion techniques it was shown that, in highly diluted aqueous solutions, this membrane lipid forms long thread-like micelles in equilibrium with lipid vesicles . The cause of the occurrence of these different aggregates is discussed in terms of the varying molecular shapes of the lipid because of a heterogeneous composition of the acyl chains . A second membrane phosphoglucolipid from the bacterium, namely 1,2-diacyl-3-O-{glycerophosphoryl-6-O-(alpha-D- glucopyranosyl-(1 -->2)-monoacylglycerophosphoryl-6-O-alpha-D-glucopyranosyl)}-sn-gl ycerol (MABGPDGlcDAG), was found to form only a lamellar liquid crystalline phase coexisting with water.

J Mol Biol, 1997 Jan 17, 265(2), 107 - 11
Two-dimensional crystallization of the light-harvesting I-reaction centre photounit from Rhodospirillum rubrum; Walz T et al.; A stoichiometric unit of the light-harvesting complex I and the reaction centre (LHI-RC complex) has been isolated from a carotenoid-less mutant of the purple non-sulphur bacterium Rhodospirillum rubrum by mild solubilization of photosynthetic membranes with the phospholipid detergent diheptylphosphatidylcholine . Dialysis of the isolated LHI-RC complexes in the presence of added dioleoyl-sn-phosphatidylcholine produced ordered two-dimensional crystals . Digital image processing revealed that the LHI-RC are packed together in a square lattice (a = b = 16.3 nm) . The dimensions of the LHI ring are essentially identical with those determined from two-dimensional (2D) crystals of purified carotenoid-containing light-harvesting I complexes after analysis by cryo-electron microscopic techniques or from negatively stained 2D crystals of purified LHI complexes from a carotenoid-less mutant . Each LHI ring of the LHI-RC complex contains a central diffuse stain-excluding region, which is assigned to the reaction centre . The analysis of the LHI-RC 2D crystals strongly suggests that the geometry and subunit stoichiometry of the LHI ring is unaffected by the presence of a reaction centre that can probably assume various orientations within the ring.

Gene, 1997 Jan 15, 184(2), 149 - 54
Cloning and sequencing of the gene for a 120-kDa immunodominant protein of Ehrlichia chaffeensis; Yu XJ et al.; Ehrlichia chaffeensis is the tick-borne, obligately intracellular bacterium that causes human monocytic ehrlichiosis . A 120-kDa protein is one of the immunodominant proteins of E . chaffeensis that stimulates production of specific antibodies in infected humans . A genomic library of E . chaffeensis was constructed in a lambda ZAP II phage vector, and a clone expressing the 120-kDa protein of E . chaffeensis was identified using canine anti-E . chaffeensis serum . DNA sequence analysis of the cloned 120-kDa protein gene of E . chaffeensis identified a 1884-bp open reading frame with an ehrlichial promoter . Five identical 240-bp tandem repeat units were identified in the 120-kDa protein gene of E . chaffeensis, comprising 60% of the entire gene . Aside from the first repeat unit, all the other repeat units are identical . In the first repeat unit there are four nucleotides that are different from the other repeats . Hydropathy analysis of the deduced amino-acid sequence demonstrated that the repeat domain contains highly hydrophilic segments . The 120-kDa protein should be evaluated for a role in stimulating protective immunity.

Eur J Biochem, 1997 Jan 15, 243(1-2), 374 - 83
NMR solution structure of an oxidised thioredoxin h from the eukaryotic green alga Chlamydomonas reinhardtii; Mittard V et al.; NMR solution structures of a cytosolic plant thioredoxin h (112 amino acids, 11.7 kDa) from the green alga Chlamydonmonas reinhardtii have been calculated on the basis of 1904 NMR distance restraints, which include 90 distances used to restrain 45 hydrogen bonds, and 44 phi dihedral restraints . The structure of C . reinhardtii thioredoxin h was solved in its oxidised form, and the ensemble of 23 converged structures superpose to the geometric average structure with an atomic rmsd of 0.080 nm +/- 0.016 for the (N, C(alpha), C) backbone atoms of residues 4-110 . Comparisons with other thioredoxins, such as thioredoxin from the bacterium Escherichia coli, thioredoxin 2 from a cyanobacterium of the Anabaena genus, and human thioredoxin, showed that thioredoxin h models share more structural features with human thioredoxin than with other bacterial thioredoxins . Examination of the accessible surface around the redoxactive peptide sequence indicates that a potent thioredoxin-h-substrate interaction could be similar to the vertebrate thioredoxin-substrate interactions.

Eur J Biochem, 1997 Jan 15, 243(1-2), 322 - 7
Imidase, a dihydropyrimidinase-like enzyme involved in the metabolism of cyclic imides; Ogawa J et al.; Imidase, which preferably hydrolyzed cyclic imides to monoamidated dicarboxylates, was purified to homogeneity from a cell-free extract of Blastobacter sp . A17p-4 . Cyclic imides are known to be hydrolyzed by mammalian dihydropyrimidinases . However, imidase was quite different from known dihydropyrimidinases in structure and substrate specificity . The enzyme has a relative molecular mass of 105 000 and consists of three identical subunits . The purified enzyme showed higher activity and affinity toward cyclic imides, such as succinimide (Km = 0.94 mM; Vmax = 910 micromol x min(-1) x mg(-1)), glutarimide (Km = 4.5 mM; Vmax = 1000 micromol min (-1) x mg (-1) and maleimide (Km = 0.34 mM; Vmax = 5800 micromol x min(-1)x mg(-1)), than toward cyclic ureides, which are the substrates of dihydropyrimidinases, such as dihydrouracil and hydantoin . Sulfur-containing cyclic imides, such as 2,4-thiazolidinedione and rhodanine, were also hydrolyzed . The enzyme catalyzed the reverse reaction, cyclization, but with much lower activity and affinity . The enzyme was non-competitively inhibited by succinate, which was found to be a key compound in cyclic-imide transformation in relation with the tricarboxylic acid cycle in this bacterium, suggesting that the role of imidase is to catalyze the initial step of cyclic-imide degradation.

Biochemistry, 1997 Jan 7, 36(1), 93 - 102
Targeted mutations in the psaC gene of Chlamydomonas reinhardtii: preferential reduction of FB at low temperature is not accompanied by altered electron flow from photosystem I to ferredoxin; Fischer N et al.; The terminal part of the electron pathway within the photosystem I (PSI) complex includes two {4Fe-4S} centers, FA and FB, which are coordinated by the PsaC subunit . To gain new insights into the electron transfer mechanisms through PsaC, we have generated three mutant strains of the alga Chlamydomonas reinhardtii in which two positively charged residues, K52 and R53, near the FA center have been altered in different ways . The mutations K52S/R53D and K52P/R53D lead to a strong destabilization of PSI . The third mutant K52S/R53A accumulates PSI to 30% of wild-type levels and shares the same residues between two of the cysteine ligands of FA as the PsaC homologue in the green sulfur bacterium Chlorobium limicola, in which FB has a higher redox potential than FA {Nitschke, W., Feiler, U., & Rutherford, A . W . (1990) Biochemistry 29, 3834-3842} . Low-temperature electron paramagnetic resonance (EPR) studies reveal that, in contrast to wild type, FB is preferentially photoreduced in this mutant, as was also observed for C . limicola . The preferential photoreduction of FB could be due to changes in the redox potential of FA and/or to slight structural modifications of the PsaC subunit . However, room temperature optical measurements show that stable charge separation still occurs and, surprisingly, that electron transfer from PSI to ferredoxin proceeds at normal rates in the mutant . As C . limicola, the K52S/R53A and K52S/R53D C . reinhardtii mutants are photosensitive when grown aerobically, but can grow photoautotrophically under anaerobic conditions.

Vestn Ross Akad Med Nauk, 1997, (6), 30 - 2
{Tularemia vaccine strain is a potential vector}; Pavlov VM et al.; The immunological and genetic properties of Francisella tularensis vaccine strain are discussion with regard to its use to produce recombinant vaccines . This bacterium-based vector is supposed to be an excellent object for investigating the role of protective antigens in the development of immunity against intracellular bacteria.

Plasmid, 1997, 37(3), 169 - 79
Cloning and sequence analysis of a novel insertion element from plasmids harbored by the carbofuran-degrading bacterium, Sphingomonas sp . CFO6; Feng X et al.; Sphingomonas sp . CFO6 (a member of the alpha group of Proteobacteria) was isolated from a Washington soil by enrichment on the insecticide carbofuran as a sole source of carbon and energy . This strain has been shown to harbor five plasmids, at least some of which are required for catabolism of carbofuran . Rearrangements, deletions, and loss of individual plasmids resulting in the loss of the carbofuran-degrading phenotype were observed following treatment with heat or introduction of Tn5 . Several putative insertion sequence elements of different sizes were cloned from these plasmids by trapping in pUCD800, a positive selection vector for isolation of transposable elements . Three of the most common putative IS elements (designated IS1412, IS1487, and IS1488) in the clone library were of different sizes and cross-hybridize with each other . An element hybridizing with IS1412, IS1487, and IS1488 was mobilized during growth of CFO6 at 42 degrees C and inserted into one of CFO6's plasmids (pCFO4), corresponding to a deletion in the plasmid and a loss of catabolic function . IS1412 was completely sequenced and its sequence analyzed . IS1412 is 1656 bp in length and possesses terminal partially matched inverted repeats of unequal length (17 and 18 bp) . In addition, IS1412 contains an open reading frame which encodes a putative transposase with significant homology to the putative transposases of IS1380 from Acetobacter pasteurianus, HRS1 from Bradyrhizobium japonicum, and IS1247 from Xanthobacter autotrophicus . These related IS elements form part of a family of common IS elements distributed among members of the alpha group of the Proteobacteria.

Ciba Found Symp, 1997, 207, 131 - 40; discussion 141-51
The cost of antibiotic resistance--from the perspective of a bacterium; Lenski RE; The possession of an antibiotic resistance gene clearly benefits a bacterium when the corresponding antibiotic is present . But does the resistant bacterium suffer a cost of resistance (i.e . a reduction in fitness) when the antibiotic is absent? If so, then one strategy to control the spread of resistance would be to suspend the use of a particular antibiotic until resistant genotypes declined to low frequency . Numerous studies have indeed shown that resistant genotypes are less fit than their sensitive counterparts in the absence of antibiotic, indicating a cost of resistance . But there is an important caveat: these studies have put antibiotic resistance genes into naive bacteria, which have no evolutionary history of association with the resistance genes . An important question, therefore, is whether bacteria can overcome the cost of resistance by evolving adaptations that counteract the harmful side-effects of resistance genes . In fact, several experiments have shown that the cost of antibiotic resistance may be substantially diminished, even eliminated, by evolutionary changes in bacteria over rather short periods of time . As a consequence of this adaptation of bacteria to their resistance genes, it becomes increasingly difficult to eliminate resistant genotypes simply by suspending the use of antibiotics.

Kansenshogaku Zasshi, 1997 Jan, 71(1), 46 - 55
{Isolation of enterohemorrhagic Escherichia coli (O157:H7) by an immunomagnetic separation method}; Asai Y et al.; Three sporadic cases of enterohemorrhagic Escherichia coli (EHEC) O157 infection which occurred in Kanagawa in 1996 were investigated . In an attempt to determine sources of the infection, a novel method of immunomagnetic separation (IMS) was employed to isolate the bacterium from feces, foods, and other associated items . In the first case, strains of EHEC O157:H7 producing Vero toxin (VT) 2 were isolated from both feces of the patient and suspected food (cattle liver) kept at a restaurant, and the strains were found to be genotypically identical through an analysis of pulsed-field gel electrophoresis (PFGE) . Subsequent investigation in the meat processing store, from which the above cattle liver had been retailed to the restaurants revealed that the store was contaminated with EHEC O157:H7 producing both VT1 and VT2 . In the second case, a strain isolated from the patient was EHEC O157:H7 producing both VT1 and VT2 while strains isolated from the patient's family (without apparent symptom) and the suspected facility were O137:NM producing VT2 . PFGE analysis indicated that the latter two strains were genotypically identical, suggesting that the facility thus contaminated with EHEC O157 caused the infection in question . In the third case, EHEC O157:NM producing VT2 was isolated from 4 out of 7 family members including the patient, and these strains were found to be genotypically identical by subsequent PFGE analysis . Source of the infection was, however, not determined due to lack of suspected food items . In this context, four slaughterhouses in Kanagawa Prefecture were investigated for presence of EHEC O157 . As a result, strains of EHEC O157:H7 producing VT1 and VT2 were isolated from the contents of cattle's distal colon and surface of the skinned carcasses . Additional attempt was also made to determine a possibility of river water being contaminated with EHEC O157 . The bacterium was, however, not isolated from water samples collected from 4 major rivers in the prefecture (at 10 collecting sites) . Experiments were undertaken in order to evaluate the use of IMS in isolation of EHEC O157 from food items, with different pre-enrichment media and conditions . The results indicated that pre-enrichment by trypticase soy broth at 36 degrees C for 6 h followed by inoculating onto sorbitol MacConkey agar plate containing cefixime and tellurite was most appropriate to isolate EHEC O157 strains.

What Is Water Purification?, What Is Genetics?, What Is Botulism?, What Is Nitrification?, What Is Biotechnology?, c, Microbiology, n, Microbe, n, Bacterium, e, Microorganisms, r, Microbes, c, Pasteurella, e, Antibiotics, e, Bacillus subtilis, o, Pseudomonas aeruginosa, a, Thermophiles, e, Escherichia coli, s, Microorganisms, c, Staphylococcus, c, Multidrug resistant, o, Bioreactor, n, Growth media, r, E coli O157, r, Staphylococcus, e, Haemophilus, o, Growth media, o, Growth media, i, Minimal inhibiting concentration, s, Escherichia coli, a, Antibiotics, s, Microorganism, n, Salmonella typhimurium

Scientific Publications - Work Done by Microbiology Reader Bioscreen C
Agricultural Microbiology Anaerobic Microbiology Antimicrobial Susceptibility Artificial Atmosphere Bioassay of Antibiotics Biofilm Microbiology Bioreactor Technology Biotechnology Cell Biology Clinical Microbiology Environmental Microbiology Experiments with Yeast	Fermentation Food Microbiology Functional Genomics Gene Technology Growth Media Development Growth Rate and Lag Time Industrial Microbiology Medical/Pharmaceutical Field Microbiological Assay Microbiological Research Microbiology of Cosmetics go to a specific theme...	Military Microbiology Molecular Microbiology Mutagenicity and Genotoxicity Oral Microbiology Patents Postantibiotic Studies Soil Microbiology Spore Microbiology Veterinary Microbiology Waste/Wastewater Treatment Water Microbiology Wine Microbiology

� 2005 Transgalactic Ltd (manufacturer of Bioscreen C software) | Privacy Statement | P.O. Box 1393, 00101 Helsinki, Finland, phone: +358 9 85172920, fax: +358 9 8749481, e-mail: microbiology@bionewsonline.com

Last modified: May 25, 2005