Bacteria

Indian J Ophthalmol, 2002 Mar, 50(1), 35 - 9
Further investigations on the association of Mycobacterium tuberculosis with Eales' disease; Madhavan HN et al.; PURPOSE: To apply polymerase chain reaction (PCR) on vitreous fluid (VF) from Eales' disease to further confirm its association with Mycobacterium tuberculosis . METHODS: Sixty nine VF samples from 69 patients (24 Eales' disease and 45 Non-Eales' as controls) were processed by conventional methods for detection of mycobacteria . Polymerase chain reaction (PCR) specific for IS 6110 and nested PCR (nPCR) using primers coding for MPB 64 gene were applied on all 69 VF . PCR based dot-blot hybridisation was applied on the IS 6110 amplified products of n PCR-positive VFs . RESULTS: Conventional methods (direct smear and culture) did not detect mycobacteria in any of the 69 VF samples . Five (20.8%) of 24 VF from Eales' and 2 (4.2%) of 45 VF from control patients tested positive for M . tuberculosis DNA by nPCR . This difference was statistically significant (P < 0.05) . All 69 VF were negative by PCR for IS 6110 . Two VF of Eales' patients positive by nPCR were also positive by DNA probe dot-blot hybridisation for IS 6110 . CONCLUSION: Detection of M . tuberculosis DNA by PCR in a significant number of VF of Eales' disease patients reemphasizes the association of this bacterium with Eales' disease.

Proc Natl Acad Sci U S A, 2002 Jul 9, 99(14), 9568 - 72 Epub 2002 Jun 27.
Assaying gene content in Arabidopsis; Allen KD; Arabidopsis has been popular as a model plant system for decades . Completion of the Arabidopsis genome and the availability of large expressed sequence-tag collections from other dicot species provides an opportunity to assess gene content in Arabidopsis, specifically by identifying genes from dicot test species that are absent from Arabidopsis . I report here results from these sorts of comparisons, carried out in part to assess the extent to which Arabidopsis is representative of dicot genomes and also the degree to which gene loss and novel gene acquisition has accompanied angiosperm speciation . More than 10% of the contigs from each of three dicot test species have no detectable homologue in Arabidopsis . By means of cross comparison among the test species, 154 specific cases of gene loss in the lineage leading to Arabidopsis were identified, including several well characterized enzymes and a group of proteins with strong homologs in the photosynthetic bacterium Synechocystis . These results show that although Arabidopsis is broadly representative of the other dicot genomes, there seems to be substantial variation even among relatively closely related genera . Further, although we cannot yet draw a causative link, variation in actual gene content seems appears to be a feature of angiosperm speciation.

J Clin Microbiol, 2002 Jul, 40(7), 2626 - 8
Large-scale outbreak of infection with Mycobacterium chelonae subsp . abscessus after penicillin injection; Zhibang Y et al.; An outbreak of infection with Mycobacterium chelonae subsp . abscessus after the injection of penicillin in 86 patients attending a factory hospital is reported . The bacterium was isolated both from lids and from the soil where the drug was stored . Molecular analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of whole-cell proteins and plasmids revealed a pattern identical to that of the strains isolated from the wounds . The source of the infections was soil contamination of the vial lids and was caused by improper use and sterilization of penicillin vials.

J Clin Microbiol, 2002 Jul, 40(7), 2513 - 9
Helicobacter marmotae sp . nov . isolated from livers of woodchucks and intestines of cats; Fox JG et al.; Woodchucks (Marmota monax) have a high incidence of hepatocellular carcinoma (HCC) associated with chronic infection with woodchuck hepatitis virus (WHV) and serve as a model of hepatitis B virus-associated HCC in humans . Helicobacter hepaticus, an enterohepatic helicobacter in mice, is known to cause hepatocellular adenomas and carcinomas in susceptible mouse strains . In long-term chemical bioassays conducted with B6C3F(1) mice, H . hepaticus has been regarded as a confounding factor because of its tumor-promoting activity . In order to determine if woodchucks harbor a Helicobacter sp . that might play a role in potentiating hepatic inflammation or neoplasia, a study was undertaken to determine whether woodchucks' livers were infected with a Helicobacter sp . Frozen liver samples from 20 (17 WHV-infected and 3 noninfected) woodchucks, 10 with WHV-associated hepatic tumors and 10 without tumors, were cultured by microaerobic techniques and analyzed by using genus- and species-specific helicobacter PCR primers . A 1,200-bp Helicobacter sp.-specific sequence was amplified from 14 liver samples . Southern hybridization confirmed the specific identity of the PCR products . Nine of the 10 livers with tumors had positive Helicobacter sp . identified by PCR, whereas 5 of the 10 livers without tumors were positive . By use of 16S rRNA species-specific primers for H . marmotae, two additional liver samples from the nontumor group had positive PCR amplicons confirmed by Southern hybridization . A urease-, catalase-, and oxidase-positive bacterium was isolated from one liver sample from a liver tumor-positive woodchuck . By 16S rRNA analysis and biochemical and phenotypic characteristics, the organism was classified as a novel Helicobacter sp . Subsequently, four additional bacterial strains isolated from feces of cats and characterized by biochemical, phenotypic, and 16S rRNA analysis were determined to be identical to the woodchuck isolate . We propose the name Helicobacter marmotae sp . nov . for these organisms . Further studies are required to ascertain if this novel Helicobacter sp . plays a tumor promotion role in hepadnavirus-associated tumors in woodchucks or causes enterohepatic disease in cats.

J Biol Chem, 2002 Sep 20, 277(38), 35133 - 9 Epub 2002 Jun 27.
A novel family 8 xylanase, functional and physicochemical characterization; Collins T et al.; Xylanases are generally classified into glycosyl hydrolase families 10 and 11 and are found to frequently have an inverse relationship between their pI and molecular mass values . However, we have isolated a psychrophilic xylanase that belongs to family 8 and which has both a high pI and high molecular mass . This novel xylanase, isolated from the Antarctic bacterium Pseudoalteromonas haloplanktis, is not homologous to family 10 or 11 enzymes but has 20-30% identity with family 8 members . NMR analysis shows that this enzyme hydrolyzes with inversion of anomeric configuration, in contrast to other known xylanases which are retaining . No cellulase, chitosanase or lichenase activity was detected . It appears to be functionally similar to family 11 xylanases . It hydrolyzes xylan to principally xylotriose and xylotetraose and is most active on long chain xylo-oligosaccharides . Kinetic studies indicate that it has a large substrate binding cleft, containing at least six xylose-binding subsites . Typical psychrophilic characteristics of a high catalytic activity at low temperatures and low thermal stability are observed . An evolutionary tree of family 8 enzymes revealed the presence of six distinct clusters . Indeed classification in family 8 would suggest an (alpha/alpha)(6) fold, distinct from that of other currently known xylanases.

J Bacteriol, 2002 Jul, 184(14), 3815 - 22
Natural resistance to inhibitors of the ubiquinol cytochrome c oxidoreductase of Rubrivivax gelatinosus: sequence and functional analysis of the cytochrome bc(1) complex; Ouchane S et al.; Biochemical analyses of Rubrivivax gelatinosus membranes have revealed that the cytochrome bc(1) complex is highly resistant to classical inhibitors including myxothiazol, stigmatellin, and antimycin . This is the first report of a strain exhibiting resistance to inhibitors of both catalytic Q(0) and Q(i) sites . Because the resistance to cytochrome bc(1) inhibitors is primarily related to the cytochrome b primary structure, the petABC operon encoding the subunits of the cytochrome bc(1) complex of Rubrivivax gelatinosus was sequenced . In addition to homologies to the corresponding proteins from other organisms, the deduced amino acid sequence of the cytochrome b polypeptide shows (i) an E303V substitution in the highly conserved PEWY loop involved in quinol/stigmatellin binding, (ii) other substitutions that could be involved in resistance to cytochrome bc(1) inhibitors, and (iii) 14 residues instead of 13 between the histidines in helix IV that likely serve as the second axial ligand to the b(H) and b(L) hemes, respectively . These characteristics imply different functional properties of the cytochrome bc(1) complex of this bacterium . The consequences of these structural features for the resistance to inhibitors and for the properties of R . gelatinosus cytochrome bc(1) are discussed with reference to the structure and function of the cytochrome bc(1) complexes from other organisms.

Photochem Photobiol, 2002 Jun, 75(6), 605 - 12
Light- and redox-dependent thermal stability of the reaction center of the photosynthetic Bacterium rhodobacter sphaeroides; Tokaji Z et al.; Irreversible loss of the photochemical activity and damage of the pigments (bacteriochlorophyll {Bchl} monomer, Bchl dimer {P} and bacteriopheophytin) by combined treatment with intense and continuous visible light and elevated temperature have been studied in a deoxygenated solution of reaction center (RC) protein from the nonsulfur purple photosynthetic bacterium Rhodobacter sphaeroides . Both the fraction of RC in the charge-separated redox state (P+Q-, where Q is a quinone electron acceptor) and the degradation of the pigments showed saturation as a function of increasing light intensity up to 400 mW cm(-2) (488/515 nm) or 1100 microE m(-2) s(-1) (white light) . The thermal denaturation curves of the RC in the P+Q- redox state demonstrated broadening and 10-20 degrees C shift to lower temperature (after 30-90 min heat treatment) compared with those in the PQ redox state . Similar but less striking behavior was seen for RC of other redox states (P+Q and PQ-) generated either by light or by electrochemical treatment in the dark . These experiments suggest that it is not the intense light per se but the changes in the redox state of the protein that are responsible for the increased sensitivity to photo- and heat damage . The RC with a charge pair (P+Q-) is more vulnerable to elevated temperature than the RC with (P+Q or PQ-) or without (PQ) a single charge . To reveal both the thermodynamic and kinetic aspects of the denaturation, a simple three-state model of coupled reversible thermal and irreversible kinetic transitions is presented . These effects may have relevance to the heat stability of other redox proteins in bioenergetics.

FEMS Microbiol Lett, 2002 Jun 4, 211(2), 129 - 32
Construction and intergeneric conjugative transfer of a pSG5-based cosmid vector from Escherichia coli to the polyisoprene rubber degrading strain Micromonospora aurantiaca W2b; Rose K et al.; A non-rubber degrading mutant of the polyisoprene rubber degrading bacterium Micromonospora aurantiaca W2b lacking the capability to form halos on latex overlay agar plates was isolated after N-methyl-N-nitro-N-nitrosoguanidine mutagenesis . A 10.3-kb shuttle cosmid vector pGM446 was constructed from the Streptomyces cloning vectors pGM160 and pOJ446 . This vector was transferred by conjugation from Escherichia coli to M . aurantiaca W2b . The frequency of formation of exconjugants with pGM446 was 3.6 x 10(-3) . This vector could be useful for shotgun cloning of genes into the non-rubber degrading mutant L1 from M . aurantiaca W2b.

Int J Food Microbiol, 2002 Jul 25, 77(1-2), 135 - 45
Pasteurization of milk and the heat resistance of Mycobacterium avium subsp . paratuberculosis: a critical review of the data; Lund BM et al.; Mycobacterium avium subsp . paratuberculosis (M . paratuberculosis) causes Johne's disease in ruminants (including cattle, sheep and goats) and other animals, and may contribute to Crohn's disease in humans . This possibility, and the fact that M . paratuberculosis may be present in raw milk, make it important to ensure that the heat treatment specified for pasteurization of milk will give acceptable inactivation of this bacterium, with an adequate margin of safety . Published studies of the heat resistance of this bacterium in milk have given widely differing results . Possible reasons for these differences, and the technical problems involved in the work, are reviewed . It is concluded that there is a need (i) for the adoption of an agreed Performance Criterion for pasteurization of milk in relation to this bacterium, (ii) a need for definitive laboratory experiments to understand and determine the heat resistance of M . paratuberculosis, and (iii) a need for an assessment of whether the minimum heat treatments specified at present for pasteurization of milk (Process Criteria) will meet the Performance Criterion for M . paratuberculosis . Measures are also required to ensure that commercial processes deliver continually the specified heat treatment, and to ensure that post-pasteurization contamination is avoided.

Biochem Biophys Res Commun, 2002 Jun 28, 294(5), 1138 - 43
Keratin degradation: a cooperative action of two enzymes from Stenotrophomonas sp; Yamamura S et al.; A novel keratin-degrading bacterium Stenotrophomonas sp . strain D-1, isolated from deer fur, produced two types of extracellular proteins: proteolytic and disulfide bond-reducing . The results on the biochemical properties suggest that this protease belongs to the serine protease, and the disulfide bond-reducing protein could be the disulfide reductase type . None of these enzymes showed keratinolytic activity independently . However, after mixing of the two enzymes, the keratinolytic activity was increased tremendously (more than 50-fold) over that of the protease only . This keratinolytic activity was more than 2-fold higher than that of the combination with proteinase K (also known for its high keratinolytic activity) . Since the two enzymes discovered in this study acted cooperatively and resulted in higher keratinolytic activity, a new mechanism of keratin degradation has been revealed . To our knowledge, this is the first report on the cooperative action of two enzymes resulting in the effective degradation of keratin.

J Ind Microbiol Biotechnol, 2002 Feb, 28(2), 103 - 11
High-yield actinorhodin production in fed-batch culture by a Streptomyces lividans strain overexpressing the pathway-specific activator gene actll-ORF4; Bruheim P et al.; Streptomyces lividans 1,326 usually does not produce the red/blue colored polyketide actinorhodin in liquid culture even though it carries the entire actinorhodin biosynthesis gene cluster . The bacterium can be forced to produce this secondary metabolite by introducing actII-ORF4, the actinorhodin pathway-specific activator gene from Streptomyces coelicolor, on a multicopy plasmid . The production of actinorhodin by such a strain has been optimized by medium and process manipulations in fed-batch cultures . With high-yield cultivation conditions, 5 g actinorhodin/l are produced during 7 days of cultivation; or approximately 0.1 g actinorhodin/g dry weight (DW)/day in the production phase . The yield in this phase is 0.15 Cmol actinorhodin/Cmol glucose, which is in the range of 25% to 40% of the maximum theoretical yield . This high-level production mineral medium is phosphate limited . In contrast, nitrogen limitation resulted in low-level production of actinorhodin and high production of a-ketoglutaric acid . Ammonium as nitrogen source was superior to nitrate supporting an almost three times higher actinorhodin yield as well as a two times higher specific production rate . The wild-type strain lacking the multicopy plasmid did not produce actinorhodin when cultivated under any of these conditions . This work examines the actinorhodin-producing potential of the strain, as well as the necessity to improve the culture conditions to fully utilize this potential . The overexpression of biosynthetic pathway-specific activator genes seems to be a rational first step in the design of secondary metabolite overproducing strains prior to alteration of primary metabolic pathways for redirection of metabolic fluxes.

Arch Microbiol, 2002 Jul, 178(1), 59 - 64 Epub 2002 Apr 30.
Intracellular localization of the particulate methane monooxygenase and methanol dehydrogenase in Methylomicrobium album BG8; Brantner CA et al.; The methanotrophic bacterium Methylomicrobium album BG8 uses methane as a sole source of carbon and energy . This bacterium forms an extensive intracytoplasmic membrane . The first enzymes of the methane oxidation pathway are the membrane-bound particulate methane monooxygenase and the periplasmic methanol dehydrogenase . Immunoelectron microscopy with specific antibodies was used to localize these enzymes to the intracytoplasmic membrane.

Curr Microbiol, 2002 Aug, 45(2), 123 - 7
Cloning and sequencing of an intracellular D(-)-3-hydroxybutyrate oligomer hydrolase from Acidovorax sp . strain SA1 and purification of the enzyme; Sugiyama A et al.; The gene of an intracellular D(-)-3-hydroxybutyrate oligomer hydrolase (i3HBOH) was cloned and sequenced from a poly(3-hydroxybutyrate) (PHB)-degrading bacterium, Acidovorax sp . strain SA1 . The i3HBOH gene has 876 nucleotides corresponding to the deduced sequence of 292 amino acids . In this amino acid sequence, the general lipase box sequence (G-X(1)-S-X(2)-G) was found, whose serine residue was determined to the active sites serine by site-directed mutagenesis.An i3HBOH was purified to electrophoretical homogeneity from SA1 . The molecular mass of the purified enzyme was estimated to be 32 kDa by SDS-PAGE . The N-terminal amino acid sequence of the purified enzyme corresponded to the deduced N-terminal amino acid sequence in the cloned i3HBOH gene.This is the first cloning and sequencing of an intracellular D(-)-3-hydroxybutyrate oligomer hydrolase gene to date.

Environ Toxicol Chem, 2002 Jun, 21(6), 1191 - 7
Acute toxicity of triorganotin compounds: different specific effects on the energy metabolism and role of pH; Hunziker RW et al.; Triorganotin compounds exhibit several modes of toxic action on the energy metabolism in energy-transducing membranes . The inhibition of the adenosine triphosphate (ATP) synthase and the hydroxide/chloride-antiport have been extensively investigated, but debate still exists on whether further mechanisms are relevant . In this work, two possible further effects have been investigated: inhibition of the bc1 complex and the hydroxide uniport, and in addition, the overall inhibition of the ATP synthesis was investigated in chromatophores of the photosynthetic purple bacterium Rhodobacter sphaeroides at pH = 7.5 and pH = 6.1 . Experimental conditions were chosen in order to exclude the hydroxide/anion antiport as a possible effect . Inhibition of the cytochromes bc1 complex was detected, but at such high concentrations that it is not relevant for acute toxicity . Tributyltin was found to induce a decrease of the membrane potential, which can be attributed to a hydroxide uniport, whereas for triphenyltin no such activity was observed . For both compounds, inhibition of the ATP synthesis was higher at pH = 6.1 than at pH = 7.5 . Also the hydroxide uniport activity of tributyltin was higher at lower pH . The contribution of the hydroxide uniport of tributyltin to the overall inhibition of the ATP synthesis cannot be quantified; however, hydroxide uniport occurred in the same concentration range as inhibition of the ATP synthesis . For triphenyltin, inhibition of the ATP synthesis can be attributed to the inhibition of the ATP synthase . It was concluded that chromatophores of R . sphaeroides are a useful system to discriminate various effects of toxicants on the energy metabolism of a cell.

Biofizika, 2002 May-Jun, 47(3), 490 - 9
{Participation of inorganic phosphates as electron donors in the primary reactions of photosynthesis in Rhodobacter sphaeroides}; Goncharova NV et al.; The formation of ATP during photophosphorylation in chromatophores from purple nonsulfur bacterium Rhodobacter sphaeroides in the presence of phenazine methosulfate and without exogenous electron carriers under constant illumination and by the action of single light flashes was studied . It was shown that the photoinduced transport of electrons to the exogenous electron acceptor depends on phosphate . It was assumed that phosphate ions are electron donors in the reaction center P870; by the action of light, P870 converts the phosphate ion HPO4(2-) into anion radical HPO4-. . In the difference EPR spectra "light minus darkness" at 77 K, an asymmetrical doublet signal with a weak low-field line was observed . The signal had a g-tensor of about 2.014 and a hyperfine coupling constant of about 2.5 mT and belongs probably to the phosphate anion radical.

Microbes Infect, 2002 Jun, 4(7), 693 - 8
The putative haemobartonella that influences Plasmodium falciparum parasitaemia in squirrel monkeys is a haemotrophic mycoplasma; Neimark H et al.; Splenectomised squirrel monkeys (Saimiri sciureus) are increasingly being used as an experimental host for human malaria studies, notably for the assessment of candidate vaccines against Plasmodium falciparum blood-stage infection . Recently, S . sciureus monkeys in our primate-breeding colony were reported to be asymptomatic carriers of a putative Haemobartonella species . Patent haemobartonella infection is frequently activated following splenectomy, and may interfere with studies on the course of P . falciparum parasitaemia in these animals . Here, we show by 16S rRNA gene sequence analysis that this wall-less bacterium is not a rickettsia but, instead, is a haemotrophic mycoplasma . Haemotrophic mycoplasmas are a newly identified group of mycoplasmas that parasitise the surfaces of erythrocytes of a wide variety of vertebrate hosts.

Infect Immun, 2002 Jul, 70(7), 3816 - 23
Inhibition of fusion of Chlamydia trachomatis inclusions at 32 degrees C correlates with restricted export of IncA; Fields KA et al.; Chlamydia trachomatis is an obligate intracellular bacterium that develops within a parasitophorous vacuole termed an inclusion . The inclusion is nonfusogenic with lysosomes but intercepts lipids from a host cell exocytic pathway . Initiation of chlamydial development is concurrent with modification of the inclusion membrane by a set of C . trachomatis-encoded proteins collectively designated Incs . One of these Incs, IncA, is functionally associated with the homotypic fusion of inclusions . Inclusions also do not fuse when cultures are multiply infected with C . trachomatis and cultivated at 32 degrees C . We obtained evidence linking these experimental observations by characterizing IncA localization in 32 degrees C cultures . Analysis of inclusions by light and transmission electron microscopy confirmed that HeLa cells infected with multiple C . trachomatis elementary bodies and cultivated at 32 degrees C for 24 h contained multiple, independent inclusions . Reverse transcriptase PCR and immunoblot analyses of C . trachomatis-infected HeLa cells demonstrated the presence of IncA at 24 h in 32 degrees C cultures . When parallel cultures were probed with IncA-specific antibodies in indirect immunofluorescence assays, IncA was detectable in intracellular chlamydiae but not within the inclusion membrane . In addition, analysis of purified reticulate bodies from 37 and 32 degrees C cultures showed that bacterium-associated pools of IncA are enriched in cultures grown at 32 degrees C . Microscopic observation of infected cells revealed that some vacuoles had fused by 48 h postinfection, and this finding was correlated with the detection of IncA in inclusion membranes by immunofluorescence microscopy . The data are consistent with a requirement for IncA in fusions of C . trachomatis inclusions and suggest that the effect of incubation at 32 degrees C is manifested by restricted export of IncA to the inclusion membrane.

Infect Immun, 2002 Jul, 70(7), 3649 - 55
Regulation of proinflammatory cytokines in human lung epithelial cells infected with Mycoplasma pneumoniae; Yang J et al.; Mycoplasma pneumoniae is a small bacterium without a cell wall that causes tracheobronchitis and atypical pneumonia in humans . It has also been associated with chronic conditions, such as arthritis, and extrapulmonary complications, such as encephalitis . Although the interaction of mycoplasmas with respiratory epithelial cells is a critical early phase of pathogenesis, little is known about the cascade of events initiated by infection of respiratory epithelial cells by mycoplasmas . Previous studies have shown that M . pneumoniae can induce proinflammatory cytokines in several different study systems including cultured murine and human monocytes . In this study, we demonstrate that M . pneumoniae infection also induces proinflammatory cytokine expression in A549 human lung carcinoma cells . Infection of A549 cells resulted in increased levels of interleukin-8 (IL-8) and tumor necrosis factor alpha mRNA, and both proteins were secreted into culture medium . IL-1 beta mRNA also increased after infection and IL-1 beta protein was synthesized, but it remained intracellular . In contrast, levels of IL-6 and gamma interferon mRNA and protein remained unchanged or undetectable . Using protease digestion and antibody blocking methods, we found that M . pneumoniae cytoadherence is important for the induction of cytokines . On the other hand, while M . pneumoniae protein synthesis and DNA synthesis do not appear to be prerequisites for the induction of cytokine gene expression, A549 cellular de novo protein synthesis is responsible for the increased cytokine protein levels . These results suggest a novel role for lung epithelial cells in the pathogenesis of M . pneumoniae infection and provide a better understanding of M . pneumoniae pathology at the cellular level.

Infect Immun, 2002 Jul, 70(7), 3637 - 48
Macrophage-induced genes of Legionella pneumophila: protection from reactive intermediates and solute imbalance during intracellular growth; Rankin S et al.; A promoter-probe strategy was devised to identify genes specifically expressed by Legionella pneumophila during growth within the macrophage . Random fragments from the L . pneumophila chromosome were inserted upstream of a promoterless phage T4 td gene, and fragments that led to complementation of thymine auxotrophy during intracellular growth of the bacterium were identified . Two different selection strategies were employed to eliminate promoters that were also active during extracellular growth of the bacterium . Some of these genes were identified independently by using both of the selection strategies . The factors identified include orthologs of efflux-mediated resistance determinants and transporters, a transporter involved in protection from osmotic stress, a stress response GTP-binding protein, a response regulator, a sensor kinase, and two systems that increase the reducing potential of the bacterium, one of which encodes the L . pneumophila ortholog of ahpC . Five of the clones analyzed here were fusions to promoters that were closely linked to genes encoding three-component chemiosmotic efflux pumps that export heavy metals or toxic organic compounds . Analysis of ahpC gene expression indicates that levels increased at least sevenfold during intracellular growth of the bacterium . Inactivation of several of the genes at their chromosomal loci had no effect on the intracellular growth rate of L . pneumophila in cultured macrophages . This suggests that a number of genes with increased expression during intracellular growth may be part of redundant systems that allow survival and growth under the conditions encountered within host cells.

J Med Entomol, 2002 May, 39(3), 534 - 40
Rickettsiella-like bacteria in Ixodes woodi (Acari: Ixodidae); Kurtti TJ et al.; We examined a parthenogenetic strain of the hard tick Ixodes woodi Bishopp for the presence of endosymbiotic bacteria . Electron microscopic examination revealed the ovarian tissues and Malpighian tubules were infected with pleomorphic bacteria . Two basic types were observed: a larger granular cell and a smaller condensed cell . Cloning and sequence analysis of polymerase chain reaction (PCR) amplified 16S rRNA gene yielded a single sequence from bacteria present in I . woodi tissues . Phylogenetic analysis of the nearly complete 16S rDNA indicated that the ticks were infected with an endosymbiont belonging to the gamma subdivision of the Proteobacteria . It clustered with the insect pathogenic species Rickettsiellagrylli (Vago and Martoja 1963) and the animal pathogen Coxiella burnetii (Derrick 1939) Philip 1948 . Our results suggest that the I . woodi females harbored a single endosymbiotic bacterium related to selected Rickettsiella species and to C burnetii.

Naturwissenschaften, 2002 Apr, 89(4), 167 - 70
Feminization of genetic males by a symbiotic bacterium in a butterfly, Eurema hecabe (Lepidoptera: Pieridae); Hiroki M et al.; Wolbachia are symbiotic bacteria found in many arthropods and filarian nematodes . They often manipulate the reproduction of host arthropods . In the present study, female-biased sex-ratio distortion in the butterfly Eurema hecabe was investigated . Breeding experiments showed that this distorted sex ratio is maternally inherited . When treated with tetracycline, adult females of the thelygenic line produced male progeny only . After PCR using Wolbachia-specific primers for the ftsZ gene a positive result was seen in the thelygenic females, but not in male progeny from tetracycline-treated females, or individuals from a Tokyo population with normal sex ratio and reproduction . Cytological observations showed that thelygenic females lack the sex chromatin body (W chromosome) . The results strongly suggest that the sex-ratio distortion in E . hecabe is due to feminization of genetic males by Wolbachia.

Biochem Biophys Res Commun, 2002 Jun 14, 294(3), 567 - 73
Evidence for a wide occurrence of proton-translocating pyrophosphatase genes in parasitic and free-living protozoa; Perez-Castineira JR et al.; Proton-translocating inorganic pyrophosphatases (H(+)-PPase, EC 3.6.1.1) are integral membrane proteins that have been extensively studied in higher plants, the photosynthetic bacterium Rhodospirillum rubrum and, more recently, in some human pathogenic protozoa . By using a PCR-based approach, fragments of genes coding for H(+)-PPases in a number of protists, both free-living and parasites of animals and plants, that belong to diverse taxonomic groups (trypanosomatids, ciliates, apicomplexans, euglenoids, amoeboid mycetozoa, heterokonts) have been isolated . The experimental procedure involved the use of degenerate oligonucleotides designed from protein domains conserved in H(+)-PPases from plants and bacteria . The PCR-amplified DNA fragments exhibited the characteristic genomic structure and codon usage of the corresponding protozoan group . Paralogous genes were found in some species suggesting the occurrence of protein isoforms . These results indicate that H(+)-PPases are more widely distributed among protozoa than previously thought.

J Mol Biol, 2002 May 10, 318(4), 1085 - 95
Assembly of light-harvesting bacteriochlorophyll in a model transmembrane helix in its natural environment; Braun P et al.; The transmembrane, bacteriochlorophyll-binding region of a bacterial light-harvesting complex, (LH2-alpha from the photosynthetic bacterium Rhodobacter sphaeroides) was redesigned and overexpressed in a mutant of Rb . sphaeroides lacking LH2 . Bacteriochlorophyll served as internal probe for the fitness of this new region for the assembly and energy transfer function of the LH2 complex . The ability to absorb and transfer light energy is practically undisturbed by the exchange of the transmembrane segment, valine -7 to threonine +6, of LH2-alpha with a 14 residue Ala-Leu sequence . This stretch makes up the residues of the transmembrane helix that are in close contact (< or =4.5 A) with the bacteriochlorophyll molecules that are coordinated through His of both the alpha and beta-subunits . In this Ala-Leu stretch, neither alpha-His0, which binds the bacteriochlorophyll, nor the adjacent alpha-Ile-1, were replaced . Novel LH2 complexes composed of LH2-alpha with a model transmembrane sequence and a normal LH2-beta are assembled in vivo into a complex, the biochemical and spectroscopic properties of which closely resemble the native one . In contrast, the additional insertion of four residues just outside the C-terminal end of the model transmembrane helix leads to complete loss of functional antenna complex . The results suggest that light energy can be harvested and transferred efficiently by bacteriochlorophyll molecules attached to only few key residues distributed over the polypeptide, while residues at the bacteriochlorophyll-helix interface seem to be largely dispensable for the functional assembly of this membrane protein complex . This novel antenna with a simplified transmembrane domain and a built-in probe for assembly and function provides a powerful model system for investigation of the factors that contribute to the assembly of chromophores in membrane-embedded proteins . (c) 2002 Elsevier Science Ltd.

Int J Syst Evol Microbiol, 2002 May, 52(Pt 3), 983 - 6
Anaeroglobus geminatus gen . nov., sp . nov., a novel member of the family Veillonellaceae; Carlier JP et al.; A hitherto unknown anaerobic coccus isolated from a post-operative fluid collection was characterized by phenotypic and phylogenetic methods . 16S rDNA sequence analysis revealed an affiliation of this isolate to the family Veillonellaceae . Also, a high level of sequence similarity was observed to some oral clone sequences of Megasphaera spp . contained in the GenBank database under designations BB166, CS025 and BS073 . These clones and the unknown bacterium form a well-separated phylogenetic branch that may represent a novel lineage within the family Veillonellaceae . Based on phenotypic and phylogenetic evidence, a new genus, Anaeroglobus gen . nov., is proposed for the unknown bacterium, with one species, Anaeroglobus geminatus gen . nov., sp . nov . The type strain of Anaeroglobus geminatus is strain AIP 313.00T (= CIP 106856T = CCUG 44773T) . It is also suggested that the oral clones BB166, CS025 and BS073 belong to the genus Anaeroglobus.

Int J Syst Evol Microbiol, 2002 May, 52(Pt 3), 831 - 4
Weissella soli sp . nov., a lactic acid bacterium isolated from soil; Magnusson J et al.; Phylogenetic analysis of the 16S rRNA gene of bacterial isolates from garden soil showed relatedness to Weissella kandleri and Weissella confusa . However, the sequences had notable differences, and DNA-DNA hybridizations confirmed that the isolates are separate from these two species . The isolates could be further distinguished from all previously described Weissella species by electrophoretic analysis of whole-cell proteins, as well as by the results from different biochemical tests . The name Weissella soli is proposed for the new species, the type strain being Mi268T (= LMG 20113T = DSM 14420T).

Int J Syst Evol Microbiol, 2002 May, 52(Pt 3), 813 - 8
Kozakia baliensis gen . nov., sp . nov., a novel acetic acid bacterium in the alpha-proteobacteria; Lisdiyanti P et al.; Four bacterial strains were isolated from palm brown sugar and ragi collected in Bali and Yogyakarta, Indonesia, by an enrichment culture approach for acetic acid bacteria . Phylogenetic analysis based on 16S rRNA gene sequences showed that the four isolates constituted a cluster separate from the genera Acetobacter, Gluconobacter, Acidomonas, Gluconacetobacter and Asaia with a high bootstrap value in a phylogenetic tree . The isolates had high values of DNA-DNA similarity (78-100%) between one another and low values of the similarity (7-25%) to the type strains of Acetobacter aceti, Gluconobacter oxydans, Gluconacetobacter liquefaciens and Asaia bogorensis . The DNA base composition of the isolates ranged from 56.8 to 57.2 mol% G+C with a range of 0-4 mol% . The major quinone was Q-10 . The isolates oxidized acetate and lactate to carbon dioxide and water, but the activity was weak, as with strains of Asaia bogorensis . The isolates differed from Asaia bogorensis strains in phenotypic characteristics . The name Kozakia baliensis gen . nov., sp . nov., is proposed for the four isolates . Strain Yo-3T (= NRIC 0488T = JCM 11301T = IFO 16664T = DSM 14400T) was isolated from palm brown sugar collected in Bali, Indonesia, and was designated as the type strain.

Int J Syst Evol Microbiol, 2002 May, 52(Pt 3), 1017 - 21
Arthrobacter roseus sp . nov., a psychrophilic bacterium isolated from an antarctic cyanobacterial mat sample; Reddy GS et al.; Strain CMS 90rT, a red-pigmented bacterium, was isolated from a cyanobacterial mat sample from a pond located in McMurdo, Antarctica . Based on its chemotaxonomic and phylogenetic properties, strain CMS 90r(T) was identified as a member of group I of Arthrobacter . It shared 16S rDNA similarity of 98% with Arthrobacter oxydans ATCC 14358T and Arthrobacter polychromogenes ATCC 15216T, while DNA-DNA similarities determined for these three organisms were less than 70% . It also differed from all 17 reported Arthrobacter species with A3alpha-variant peptidoglycan in that it possessed a unique peptidoglycan (Lys-Gly-Ala3) and contained galactose, glucose, ribose and rhamnose as cell-wall sugars . These data and the presence of diagnostic phenotypic traits support the description of CMS 90r(T) as a novel species of Arthrobacter, for which the name Arthrobacter roseus sp . nov . is proposed . The type strain is strain CMS 90r(T) (= MTCC 3712T = DSM 14508T).

Biotechnol Prog, 2002 May-Jun, 18(3), 652 - 6
Expression of double Vitreoscilla hemoglobin enhances growth and alters ribosome and tRNA levels in Escherichia coli; Roos V et al.; In several organisms, expression of a gene encoding dimeric hemoglobin (VHb) from the obligate aerobic bacterium Vitreoscilla stercoraria has been shown to increase microaerobic cell growth and enhance oxygen-dependent cell metabolism . In an attempt to further improve these effects of VHb, a gene encoding two vhb genes connected by a short linker of six base pairs was constructed and expressed in Escherichia coli(double VHb) . Escherichia coli cells expressing double VHb reached a cell density 19% higher than that of cells expressing native VHb . The protein production per cell remained constant since the increase in cell growth was accompanied by an increase in protein content by 16% . Investigation of ribosome and tRNA content revealed that cells expressing double VHb reached their maximal capacity of protein synthesis later during cultivation than cells expressing native VHb, and furthermore they reached considerably higher levels of ribosome and tRNA compared to that of the VHb-expressing cells.

J Mol Biol, 2002 May 31, 319(2), 501 - 15
X-ray structure determination of the cytochrome c2: reaction center electron transfer complex from Rhodobacter sphaeroides; Axelrod HL et al.; In the photosynthetic bacterium Rhodobacter sphaeroides, a water soluble cytochrome c2 (cyt c2) is the electron donor to the reaction center (RC), the membrane-bound pigment-protein complex that is the site of the primary light-induced electron transfer . To determine the interactions important for docking and electron transfer within the transiently bound complex of the two proteins, RC and cyt c2 were co-crystallized in two monoclinic crystal forms . Cyt c2 reduces the photo-oxidized RC donor (D+), a bacteriochlorophyll dimer, in the co-crystals in approximately 0.9 micros, which is the same time as measured in solution . This provides strong evidence that the structure of the complex in the region of electron transfer is the same in the crystal and in solution . X-ray diffraction data were collected from co-crystals to a maximum resolution of 2.40 A and refined to an R-factor of 22% (R(free)=26%) . The structure shows the cyt c2 to be positioned at the center of the periplasmic surface of the RC, with the heme edge located above the bacteriochlorophyll dimer . The distance between the closest atoms of the two cofactors is 8.4 A . The side-chain of Tyr L162 makes van der Waals contacts with both cofactors along the shortest intermolecular electron transfer pathway . The binding interface can be divided into two domains: (i) A short-range interaction domain that includes Tyr L162, and groups exhibiting non-polar interactions, hydrogen bonding, and a cation-pi interaction . This domain contributes to the strength and specificity of cyt c2 binding . (ii) A long-range, electrostatic interaction domain that contains solvated complementary charges on the RC and cyt c2 . This domain, in addition to contributing to the binding, may help steer the unbound proteins toward the right conformation .

Expert Rev Mol Diagn, 2002 May, 2(3), 257 - 66
Moving to nucleic acid-based detection of genital Chlamydia trachomatis; Tong CY et al.; Laboratory diagnosis of the bacterium Chlamydia trachomatis has gone through a complete phase of evolution since it was first identified as a significant cause of sexually transmitted infection . As a fragile, obligatory intracellular organism, it was initially only grown in eggs . Subsequently, diagnosis relied on culture in continuous cell lines . To address the limitations of culture, immunological methods were developed and direct antigen detection using enzyme immunoassay and immunofluorescence flourished . With the advent of molecular technologies, nucleic acid-based amplification techniques became the methods of choice, offering improved standard of care for diagnosis and opening up the possibility of screening using noninvasive, patient-acceptable specimens . In this article, the various currently available molecular methods are examined, some of the existing problems discussed and a view on what we think might happen in the next 5 years to the technology and requirement in diagnosis and screening is given.

Proc Natl Acad Sci U S A, 2002 Jun 11, 99(12), 7917 - 21 Epub 2002 Jun 04.
The RecA proteins of Deinococcus radiodurans and Escherichia coli promote DNA strand exchange via inverse pathways; Kim JI et al.; The RecA protein of Escherichia coli, and all filament-forming homologues identified to date, promote DNA strand exchange by a common, ordered pathway . A filament is first formed on single-stranded DNA, followed by uptake of the duplex substrate . These proteins are thereby targeted to single-strand gaps and tails where recombinational DNA repair is required . The observed course of DNA strand exchange promoted by the RecA protein from the extremely radioresistant bacterium Deinococcus radiodurans is the exact inverse of this established pathway . This reaction lies at the heart of a remarkably efficient system for the repair of DNA damage.

Microbes Infect, 2002 May, 4(6), 591 - 8
Mouse resident peritoneal macrophages partially control in vitro infection with Coxiella burnetii phase II; Zamboni DS et al.; Coxiella burnetii, the agent of Q fever in man and of coxiellosis in other species, is a small, dimorphic, obligate intracellular bacterium, sheltered within large, acidified, and hydrolase-rich phagosomes . Although several primary and established cell lines, macrophage-like cells, and primary macrophages from other species have been infected with C . burnetii, the infection of mouse primary macrophages has not been sufficiently characterized . In this report quantification of DAPI (4', 6-diamino-2-phenylindole) fluorescence images acquired by confocal microscopy, and transmission electron microscopy were used to compare the infection of three mouse-derived cells, L929 fibroblasts, J774 macrophage-like cells, and resident peritoneal macrophages, with a phase II clone of C . burnetii known to be non-virulent for mammals . Infected peritoneal phagocytes differed from L929 or J774 cells in that: (a) large vacuoles took longer to appear (3-5 d instead of 2), and were only found in a subset (20-30%) of macrophages, as opposed to in more than 70% of the other cells; (b) total and vacuole-associated relative bacterial loads in L929 and J774 cells were several-fold higher than in peritoneal macrophages; (c) estimated doubling times of the bacteria were about 68 h in the primary macrophages, 18 h in J774 and 22 h in L929 cells . Thus, mouse resident peritoneal macrophages control both the formation of the large vacuoles and the intracellular proliferation of C . burnetii phase II.

Sheng Wu Hua Xue Yu Sheng Wu Wu Li Xue Bao (Shanghai), 2001, 33(5), 519 - 524
Expression of Choline Oxidase Gene (codA) Enhances Salt Tolerance of the Tobacco; He PM et al.; The codA gene for choline oxidase, which converts choline into betaine . This enzyme, cloned from a soil bacterium Arthrobactor globiformis, has been transferred and expressed in tobacco by the Agrobatcterium-method transformation through the binary plasmid pGAH/codA . The pGAH/codA carried with Km(R) and Hyg(R),and coding sequence for a transit peptide from Rubisco small subunit gene (rbcS) was inserted between 35 S promoter and codA, so that the COD could be introduced into the chloroplast by this transit peptide . The transformed plants were screened on the medium containing the Km and Hyg . PCR, Western and gold immunolocalization tests showed that the codA gene has integrated into the tobacco DNA genome and its protein was expressed and the mature peptide has gone into the chloroplasts by the transit peptide . The results of salt-tolerance measuring for transgenic plants showed that the transgenic plants were more tolerance to the salt than the control plants . The young transgenic plants (1.0--1.5 cm) could survive at 400 mmol/L NaCl MS medium for more than 30 days . From them the higher tolerance plant T4-400 was obtained, which could grow at the 300 mmol/L NaCl MS medium well . The transgenic plants (6--8cm) could grow normally at the 400 mmol/L NaCl MS medium while wild plants failed to do so . So the transferred plant with codA enhanced its tolerance to the salt stress.

Appl Environ Microbiol, 2002 Jun, 68(6), 2781 - 93
Genetic complementation of an outer membrane cytochrome omcB mutant of Shewanella putrefaciens MR-1 requires omcB plus downstream DNA; Myers JM et al.; Anaerobically grown cells of the metal-reducing bacterium Shewanella putrefaciens MR-1 contain multiple outer membrane (OM) cytochromes . A gene replacement mutant (strain OMCB1) lacking the OM cytochrome OmcB is markedly deficient in the reduction of MnO2 and exhibits reduced rates of Fe(III) reduction . The levels of other OM cytochromes are also decreased in OMCB1 . Complementation of OMCB1 with wild-type omcB did not restore any of these defects . However, a 21-kb genomic fragment from MR-1, which included omcB and 19 kb of downstream DNA, fully restored MnO2 and Fe(III) reduction and the full complement of OM cytochromes to OMCB1 . A 14.7-kb DNA fragment, including omcB and 12 kb of downstream DNA, provided only a modest increase in MnO2 reduction and OM cytochrome content, but it fully restored Fe(III) citrate reduction and partially restored FeOOH reduction . While omcB mRNA was readily detected in this complement, the OmcB protein was not detected in any cellular compartment . The restoration of Fe(III) reduction despite the absence of OmcB suggests that OmcB itself is not required for Fe(III) reduction . Another OM cytochrome, OmcA, was mislocalized to the cytoplasmic membrane of OMCB1 . Only the 21-kb genomic fragment was able to restore proper localization of OmcA to the OM . This 21-kb fragment does not contain omcA, but it does contain several open reading frames (ORFs) downstream from omcB . The most downstream of these ORFs (altA) encodes a putative AraC-like transcriptional regulator . However, a gene replacement mutant of altA resembled the wild type with respect to MnO2 reduction, OM cytochrome content, and the localization of OmcA and OmcB to the OM . Since OMCB1 continues to express genes immediately downstream from omcB, the lack of expression of this downstream DNA does not explain its phenotype or the need for the large complementing fragment . The results suggest that the DNA downstream of omcB must be present in cis in order to restore Fe(III) reduction, MnO2 reduction, OM cytochrome content, and the localization of OmcA and OmcB to the OM.

Nature, 2002 May 30, 417(6888), 533 - 5
Quantum control of energy flow in light harvesting; Herek JL et al.; Coherent light sources have been widely used in control schemes that exploit quantum interference effects to direct the outcome of photochemical processes . The adaptive shaping of laser pulses is a particularly powerful tool in this context: experimental output as feedback in an iterative learning loop refines the applied laser field to render it best suited to constraints set by the experimenter . This approach has been experimentally implemented to control a variety of processes, but the extent to which coherent excitation can also be used to direct the dynamics of complex molecular systems in a condensed-phase environment remains unclear . Here we report feedback-optimized coherent control over the energy-flow pathways in the light-harvesting antenna complex LH2 from Rhodopseudomonas acidophila, a photosynthetic purple bacterium . We show that phases imprinted by the light field mediate the branching ratio of energy transfer between intra- and intermolecular channels in the complex's donor acceptor system . This result illustrates that molecular complexity need not prevent coherent control, which can thus be extended to probe and affect biological functions.

Biosci Biotechnol Biochem, 2002 Apr, 66(4), 866 - 8
Response of the ice-nucleating bacterium Pantoea ananas KUIN-3 during cold acclimation; Koda N et al.; The ice-nucleating bacterium Pantoea ananas KUIN-3 accumulated glucose in cells following a shift in temperature (10 degrees C) from the optimum growth temperature (30 degrees C) . This accumulation might be caused by the activation of glucose-6-phosphatase . Although this strain after culturing at 30 degrees C was harmed by freezing, the cryotolerance of this strain was reached about 80% after cold acclimation at 10 degrees C.

Biosci Biotechnol Biochem, 2002 Apr, 66(4), 737 - 42
Heterogeneity of dehydrogenases of Stenotrophomonas maltophilia showing dye-linked activity with polypropylene glycols; Tachibana S et al.; Distinct enzyme activities were found in extracts from Stenotrophomonas maltophilia showing dye-linked oxidation of polypropylene glycols . The activities were induced when polypropylene glycols served as sole carbon and energy sources for the bacterium . In the logarithmic phase, most of the enzyme activities (88%) were found in the cytoplasm . In the stationary phase, more than half of the activities (54%) were found on the membrane, but significant activities were also distributed in the periplasm (34%) and the cytoplasm (12%) . The enzyme activities differed from each other in their localization, time of induction in the growth cycle, specificity toward electron acceptor, and electrophoresis mobility.

Sheng Wu Hua Xue Yu Sheng Wu Wu Li Xue Bao (Shanghai), 2001, 33(6), 607 - 614
Isolation and Analysis of hupR Gene Required for the Expression of Hydrogenase in Rhodobacter sphaeroides; Xu DQ et al.; Cosmid 1 containing the hup genes isolated from the photosynthetic bacterium Rhodobacter sphaeroides was studied . The hupR gene from cosmid 1 was cloned and sequenced (EMBL accession number AJ243734) . It encoded a 54.031 kD protein homologous to transcriptional regulators belonging to the superfamily of two-component regulatory systems . The HupR protein was overexpressed in Escherichia coli in the form of His6-tagged HupR . The cloned hupR gene could restore hydrogenase activity in R.sphaeroides hupR mutants and activate hupSL gene transcription.

J Mol Model (Online), 2002 Feb, 8(2), 58 - 64
Homology modeling reveals the structural background of the striking difference in thermal stability between two related {NiFe}hydrogenases; Szilagyi A et al.; Hydrogenases are redox metalloenzymes in bacteria that catalyze the uptake or production of molecular hydrogen . Two homologous nickel-iron hydrogenases, HupSL and HydSL from the photosynthetic purple sulfur bacterium Thiocapsa roseopersicina, differ substantially in their thermal stabilities despite the high sequence similarity between them . The optimum temperature of HydSL activity is estimated to be at least 50 degrees C higher than that of HupSL . In this work, homology models of both proteins were constructed and analyzed for a number of structural properties . The comparison of the models reveals that the higher stability of HydSL can be attributed to increased inter-subunit electrostatic interactions: the homology models reliably predict that HydSL contains at least five more inter-subunit ion pairs than HupSL . The subunit interface of HydSL is more polar than that of HupSL, and it contains a few extra inter-subunit hydrogen bonds . A more optimized cavity system and amino acid replacements resulting in increased conformational rigidity may also contribute to the higher stability of HydSL . The results are in accord with the general observation that with increasing temperature, the role of electrostatic interactions in protein stability increases . Electronic supplementary material to this paper can be obtained by using the Springer Link server located at http://dx.doi.org/10.1007/s00894-001-0071-8.

J Bacteriol, 2002 Jun, 184(12), 3396 - 400
Gas channels for NH(3): proteins from hyperthermophiles complement an Escherichia coli mutant; Soupene E et al.; Ammonium transport (Amt) proteins appear to be bidirectional channels for NH(3) . The amt genes of the hyperthermophiles Aquifex aeolicus and Methanococcus jannaschii complement enteric amtB mutants for growth at 25 nM NH(3) at 37 degrees C . To our knowledge, Amt proteins are the first hyperthermophilic membrane transport proteins shown to be active in a mesophilic bacterium . Despite low expression levels, His-tagged Aquifex Amt could be purified by heating and nickel chelate affinity chromatography . It could be studied genetically in Escherichia coli.

J Bacteriol, 2002 Jun, 184(12), 3368 - 76
Chlorobium tepidum mutant lacking bacteriochlorophyll c made by inactivation of the bchK gene, encoding bacteriochlorophyll c synthase; Frigaard NU et al.; The gene encoding bacteriochlorophyll (BChl) c synthase was identified by insertional inactivation in the photosynthetic green sulfur bacterium Chlorobium tepidum and was named bchK . The bchK mutant of C . tepidum was rusty-orange in color and completely lacked BChl c . Because of the absence of the BChl c antenna, the mutant grew about seven times slower than the wild type at light intensities that were limiting to the wild type (< 90 micromol m(-2) s(-1)) . Various pheophorbides, which probably represent precursors of BChl c which had lost magnesium, accumulated in the mutant cells . A small fraction of these pheophorbides were apparently esterified by the remaining chlorophyll (Chl) a and BChl a synthases in cells . The amounts of BChl a, Chl a, isoprenoid quinones, carotenoids, Fenna-Matthews-Olson protein, and chlorosome envelope protein CsmA were not significantly altered on a cellular basis in the mutant compared to in the wild type . This suggests that the BChl a antennae, photosynthetic reaction centers, and remaining chlorosome components were essentially unaffected in the mutant . Electron microscopy of thin sections revealed that the mutant lacked normal chlorosomes . However, a fraction containing vestigial chlorosomes, denoted "carotenosomes," was partly purified by density centrifugation; these structures contained carotenoids, isoprenoid quinones, and a 798-nm-absorbing BChl a species that is probably protein associated . Because of the absence of the strong BChl c absorption found in the wild type, the bchK mutant should prove valuable for future analyses of the photosynthetic reaction center and of the roles of BChl a in photosynthesis in green bacteria . An evolutionary implication of our findings is that the photosynthetic ancestor of green sulfur bacteria could have evolved without chlorosomes and BChl c and instead used only BChl a-containing proteins as the major light-harvesting antennae.

Proc R Soc Lond B Biol Sci, 2002 May 7, 269(1494), 931 - 6
Antagonistic coevolution between a bacterium and a bacteriophage; Buckling A et al.; Antagonistic coevolution between hosts and parasites is believed to play a pivotal role in host and parasite population dynamics, the evolutionary maintenance of sex and the evolution of parasite virulence . Furthermore, antagonistic coevolution is believed to be responsible for rapid differentiation of both hosts and parasites between geographically structured populations . Yet empirical evidence for host-parasite antagonistic coevolution, and its impact on between-population genetic divergence, is limited . Here we demonstrate a long-term arms race between the infectivity of a viral parasite (bacteriophage; phage) and the resistance of its bacterial host . Coevolution was largely driven by directional selection, with hosts becoming resistant to a wider range of parasite genotypes and parasites infective to a wider range of host genotypes . Coevolution followed divergent trajectories between replicate communities despite establishment with isogenic bacteria and phage, and resulted in bacteria adapted to their own, compared with other, phage populations.

Cell Microbiol, 2002 May, 4(5), 273 - 83
Determination of the physical environment within the Chlamydia trachomatis inclusion using ion-selective ratiometric probes; Grieshaber S et al.; Chlamydia trachomatis is an obligate intracellular bacterium with a biphasic life cycle that takes place entirely within a membrane-bound vacuole termed an inclusion . The chlamydial inclusion is non-fusogenic with endosomal or lysosomal compartments but intersects a pathway involved in transport of sphingomyelin from the Golgi apparatus to the plasma membrane . The physical conditions within the mature chlamydial inclusion are unknown . We used ratiometric imaging with membrane-permeant, ion-selective fluorescent dyes for microanalyis of the physical environment within the inclusion . Determination of H+, Na+, K+ and Ca(2+) concentrations using CFDA (carboxy fluorescein diacetate) or BCECF-AM (2',7'-bis (2-carboxyethyl)-5,6-carboxyfluorescein acetoxymethyl ester, SBFI-AM, PBFI-AM and fura-PE3-acetomethoxyester (Fura-PE3-AM), respectively, indicated that all ions assayed within the lumenal space of the inclusion approximated the concentrations within the cytoplasm . Stimulation of purinergic receptors by addition of extracellular ATP triggered a dynamic Ca(2+) response that occurred simultaneously within the cytoplasm and interior of the inclusion . The chlamydial inclusion thus appears to be freely permeable to cytoplasmic ions . These results have implications for nutrient acquisition by chlamydiae and may contribute to the non-fusogenicity of the inclusion with endocytic compartments.

Environ Sci Technol, 2002 May 1, 36(9), 1971 - 9
Baseline toxicity (narcosis) of organic chemicals determined by in vitro membrane potential measurements in energy-transducing membranes; Escher BI et al.; Baseline toxicity of a selection of industrial chemicals and pharmaceuticals is determined experimentally with a new in vitro test system (Kinspec) using membrane vesicles isolated from a photosynthetic bacterium, Rhodobacter sphaeroides . This test system is selective and more sensitive than other mechanistic test systems for baseline toxicity . The only concomitantly determined mechanism is uncoupling, which can be distinguished from baseline toxicity by pH-dependent measurements . Because the tests system contains only the target site for baseline toxicants, the biological membrane, effective target site concentrations can be directly related to observed effects by combining the in vitro test with membrane-water partition experiments . No differences were found between the effective membrane concentrations of nonpolar and polar compounds, confirming the earlier hypothesis that differences in lethal body burdens are primarily caused by unequal distribution of the compounds between target and nontarget lipids and not by different mechanisms . A selection of pharmaceuticals with various specific modes of toxic action exhibited the same constant effective membrane concentrations as found for pure baseline toxicants . In mixtures of four to six components, the pharmaceuticals were concentration-additive with each other and with the pure baseline toxicants . A potential application of the proposed test system lies, therefore, in assessing the cumulative baseline toxicity in complex environmental mixtures.

Environ Sci Technol, 2002 May 1, 36(9), 1939 - 46
Enhancement of biological reduction of hematite by electron shuttling and Fe(II) complexation; Royer RA et al.; Natural organic matter (NOM) enhancement of the biological reduction of hematite (alpha-Fe2O3) by the dissimilatory iron-reducing bacterium Shewanella putrefaciens strain CN32 was investigated under nongrowth conditions designed to minimize precipitation of biogenic Fe(II) . Hydrogen served as the electron donor . Anthraquinone-2,6-disulfonate (AQDS), methyl viologen, and methylene blue {quinones with an Ew0 (pH 7) of 0.011 V or less}, ferrozine {a strong Fe(II) complexing agent}, and characterized aquatic NOM (Georgetown NOM or Suwannee River fulvic acid) enhanced bioreduction in 5-day experiments whereas 1,4-benzoquinone (Ew0 value = 0.280 V) did not . A linear relationship existed between total Fe(II) produced and concentrations of ferrozine or NOM but not quinones, except in the case of methylene blue . Such a linear relationship between Fe(II) and methylene blue concentrations could be due to the systems being far undersaturated with respect to methylene blue or the loss of the thermodynamic driving force . A constant concentration of AQDS and variable concentrations of ferrozine produced a linear relationship between total Fe(II) produced and the concentration of ferrozine . Enhancement effects of both AQDS and ferrozine were additive . NOM may serve as both an electron shuttle and an Fe(II) complexant; however, the concentration dependence of hematite reduction with NOM was more similar to ferrozine than quinones . NOM likely enhances hematite reduction initially by electron shuttling and then further by Fe(II) complexation, which prevents Fe(II) sorption to hematite and cell surfaces.

Microb Ecol, 2001 Dec, 42(4), 562 - 571
Loss of Estuarine Bacteria by Viral Infection and Predation in Microcosm Conditions; Almeida MA et al.; The bacterioplankton density in Ria de Aveiro, a shallow estuarine ecosystem, varied in the broad range of 1.9-10.6 x 109 cells L-1 . The range of values was about 2 times higher in brackish water than in marine water . At high tide bacterial abundance was 2-3 times lower than at low tide . The overall variation in virioplankton was in the range of 2.4-25.0 x 1010 particles L-1 . Brackish water was about 2 times richer in viral particles than the marine water . Near low tide the virioplankton was 2-3 times higher that at high tide . Viral density followed the pattern of bacterial abundance (it explained 40% of virioplankton variation) . The viruses to bacterium ratio varied, throughout tidal cycles, by a factor of about 10 establishing the range 4.7-55.6 (average 17.6) . This ratio was rather similar in the two estuarine zones . We compared the effects of infection and predation on the control of bacterioplankton size in the two zones of the estuary . The approach to this question was conducted in experimental microcosms, set up in six combinations of plankton variables affecting the presence/absence of predators, virus-to-bacterium ratio (10-fold increase), virus-to-bacterium distance (2.2-fold increase), and bacterial growth rate . The results showed that predation was similar, in a percent basis, in marine (69%) and brackish water (73%) . Viral infection was, however, higher in brackish water (59%) than in the marine water (36%) . We conclude that the bacterioplankton along the salinity gradient evolves under biological pressures that are in different balance in the marine and brackish water zones . The effect of viral lysis on bacterial communities with enhanced growth (after yeast extract addition) was masked even when the initial ratio was 10-fold greater than in the natural samples . The high density of the virioplankton did not preclude the large and rapid increase in bacterial density . We suggest that the dynamics of the equilibrium between bacteria and viruses in the environment is driven to higher numerical levels during periods of intensive bacterial growth . On the contrary, at low bacterial growth rates the temporarily increased virus-to-bacterium ratio may drive the equilibrium to its lowest levels.

J Comput Biol, 2002, 9(2), 261 - 76
Predicting the beta-helix fold from protein sequence data; Cowen L et al.; A method is presented that uses beta-strand interactions to predict the parallel right-handed beta-helix super-secondary structural motif in protein sequences . A program called BetaWrap implements this method and is shown to score known beta-helices above non-beta-helices in the Protein Data Bank in cross-validation . It is demonstrated that BetaWrap learns each of the seven known SCOP beta-helix families, when trained primarily on beta-structures that are not beta-helices, together with structural features of known beta-helices from outside the family . BetaWrap also predicts many bacterial proteins of unknown structure to be beta-helices; in particular, these proteins serve as virulence factors, adhesins, and toxins in bacterial pathogenesis and include cell surface proteins from Chlamydia and the intestinal bacterium Helicobacter pylori . The computational method used here may generalize to other beta-structures for which strand topology and profiles of residue accessibility are well conserved.

Protein Sci, 2002 Jun, 11(6), 1309 - 19
Electrostatic properties of the anion selective porin Omp32 from Delftia acidovorans and of the arginine cluster of bacterial porins; Zachariae U et al.; The functional properties of the anion-selective porin Omp32 from the bacterium Delftia acidovorans, formerly Comamonas acidovorans, are determined by the particularly narrow channel constriction and the electrostatic field inside and outside the pore . A cluster of arginines (Arg 38, Arg 75, and Arg 133) determines the electrostatic field close to the constriction zone . Stacked amino acids carrying charges are prone to drastic pK(a) shifts . However, optimized calculations of the titration behavior of charged groups, based on the finite-difference Poisson-Boltzmann technique, suggest that all the arginines are charged at physiological pH . Protonation of the clustered arginines is stabilized by one buried glutamate residue (Glu 58), which is strongly interacting with Arg 75 and Arg 38 . This functional arrangement of three charged amino acid residues is of general significance because it is found in the constriction zones of all known 16-stranded porins from the alpha-, beta-, and gamma-proteobacteria.

J Vet Med B Infect Dis Vet Public Health, 2002 Apr, 49(3), 135 - 41
A simple random amplified polymorphic DNA genotyping method for field isolates of Dermatophilus congolensis; Larrasa J et al.; Dermatophilus congolensis is the pathogenic actinomycete that causes dermatophilosis in cattle, lumpy wool in sheep and rain scald in horses . Phenotypic variation between isolates has previously been described, but its genetic basis, extent and importance have not been investigated . Standard DNA extraction methods are not always successful for D . congolensis due to its complex life cycle, one stage of which is encapsulated . Here we describe the development of rapid and reliable DNA extraction and random amplified polymorphic DNA (RAPD) methods that can be used for genotyping D . congolensis field isolates . Our results suggest that genotypic variation between isolates correlates with host species . Several DNA extraction methods and RAPD protocols were compared . An extraction method based on incubation of the bacterium in lysozyme, sodium dodecyl sulphate (SDS) and proteinase K treatments and phenolic extraction yielded high-quality DNA, which was used to optimize RAPD-polymerase chain reaction (PCR) protocols for two random primers . An alternative rapid, non-phenolic extraction method based on proteinase K treatment and thermal shock was selected for routine RAPD typing of isolates . DNA extracted from reference strains from cattle, sheep and horse using either method gave reproducible banding patterns with different DNA batches and different thermal cyclers . The rapid DNA extraction method and RAPD-PCR were applied to 38 D . congolensis field isolates . The band patterns of the field and type isolates correlated with host species but not with geographical location.

Indian J Exp Biol, 2001 Jul, 39(7), 673 - 7
Mechanism of removal of aflatoxin B1 by a resistant bacterium; Vaithiyanathan S et al.; A bacterium resistant to aflatoxin B1 isolated from the soil was shown to remove a maximum 26% of toxin at 3 microg ml(-1) . A comparison of spectrophotometric and radioisotope methods showed that a maximum of 48 hr was sufficient to remove the toxin . Radioisotope analysis showed that the radioactivity decreased in the chloroform phase while it increased in the aqueous phase during the time course of the experiment . An analysis of the supernatant of culture medium showed that the bacterium had converted aflatoxin B1 to water soluble compounds with lambda(max) of 338 and 374.

New Microbiol, 2002 Apr, 25(2), 253 - 7
Isolation of Bartonella henselae from domestic cats in an Italian urban area; Cabassi CS et al.; Bartonella henselae is the causative agent of Cat Scratch Disease (CSD) in humans . Cat is considered the reservoir of the bacterium . Identification of bacteriemic cats is the basic tool in the prophylaxis of CSD . Blood samples were collected between January 1999-December 2000 from 248 domestic cats living in an urban area (Reggio Emilia) in Northern Italy and tested for Bartonella henselae bacteriemia . Cultural and PCR methods were used . PCR was used directly on cat blood as well as to identify the Bartonella strain growth in culture . 24 (9.7 %) cats were found bacteriemic, most of which aged <1 year . A higher sensitivity was demonstrated by cultural method compared with PCR.

Am J Vet Res, 2002 May, 63(5), 757 - 62
Chemotaxis, phagocytosis, and oxidative metabolism in bovine macrophages exposed to a novel interdigital phlegmon (foot rot) lesion isolate, Porphyromonas levii; Walter MR et al.; OBJECTIVE: To examine the host response toward Porphyromonas levii, by evaluating chemotaxis, phagocytosis, and oxidative burst of bovine macrophages in vitro . SAMPLE POPULATION: Cultured bovine macrophages obtained from monocytes harvested from blood samples of 15 Holstein steers . Porphyromonas levii was isolated from the foot rot lesion of an acutely affected feedlot steer . PROCEDURE: Monocytes were cultured for macrophage differentiation over 7 days . Porphyromonas levii was cultured in strict anaerobic conditions for experimentation . Chemotaxis was evaluated by quantifying macrophage migration toward P . levii in Boyden chambers . Phagocytosis was assessed by quantification of macrophages engulfing P . levii following incubation with or without anti-P . levii serum or purified IgG . Oxidative burst was measured by use of the nitroblue tetrazolium reduction assay . RESULTS: Chemotaxis toward P . levii was not significantly different from control values at any of the tested bacterial concentrations . Phagocytosis of P . levii was approximately 10% at a 10:1 bacterium to macrophage ratio and did not change significantly over time . When higher proportions of P . levii were tested for phagocytosis, the 1,000:1 bacterium to macrophage ratio had a significant increase, compared with the 10:1 test group . Opsonization of P . levii with high-titeranti-P . levii serum or anti-P . levii IgG produced a significant increase in macrophage phagocytosis . Oxidative production significantly increased compared with control in the 1,000:1 test group only . CONCLUSIONS AND CLINICAL RELEVANCE: Porphyromonas levii may evade host detection by decreased chemotaxis, phagocytosis, and oxidative burst by macrophages . Acquired immunity may be beneficial for clearance of P . levii in foot rot lesions in cattle.

Extremophiles, 2002 Apr, 6(2), 89 - 95
Transcriptional regulation under pressure conditions by RNA polymerase sigma54 factor with a two-component regulatory system in Shewanella violacea; Nakasone K et al.; Deep-sea bacteria have unique systems for gene and protein expression controlled by hydrostatic pressure . One of the sigma factors, sigma54, was found to play an important role in pressure-regulated transcription in a deep-sea piezophilic bacterium, Shewanella violacea . A glutamine synthetase gene (glnA) has been targeted as a model for the pressure-regulated promoter to investigate transcriptional regulation by the sigma54 factor . Recognition sites for sigma54 and sigma70 factors were observed at an upstream region of the glnA, and NtrC-binding sites were also identified at the same region . Primer extension analyses revealed that the transcription initiation sites of both promoters were determined and that transcription from the sigma54 site was regulated by elevated pressure . The sigma54 promoter is known to be activated by a two-component signal transduction system, the NtrB-NtrC phosphorylation relay . Our results suggested that this system might be regulated by deep-sea conditions and that the gene expression controlled by the sigma54 promoter was actually regulated by pressure . We propose a possible model of the molecular mechanisms for pressure-regulated transcription.

Infect Immun, 2002 Jun, 70(6), 3304 - 7
Proteolysis of CD14 on human gingival fibroblasts by arginine-specific cysteine proteinases from Porphyromonas gingivalis leading to down-regulation of lipopolysaccharide-induced interleukin-8 production; Tada H et al.; Arginine-specific cysteine proteinases (gingipains-R) from periodontopathic Porphyromonas gingivalis cleaved CD14, a bacterial pattern recognition receptor, on human gingival fibroblasts (HGF) . Consequently, gingipains-R reduced lipopolysaccharide-induced interleukin-8 production by HGF, indicating that gingipains-R inhibited CD14-dependent HGF activation and are involved in immune evasion by the bacterium in periodontal tissues.

Infect Immun, 2002 Jun, 70(6), 2899 - 907
LsaA, an antigen involved in cell attachment and invasion, is expressed by Lawsonia intracellularis during infection in vitro and in vivo; McCluskey J et al.; Lawsonia intracellularis has been identified recently as the etiological agent of proliferative enteropathies, which are characterized by intestinal epithelial hyperplasia and associated moderate immune responses . This disease complex has been reported in a broad range of animals, prevalently in pigs, and L . intracellularis has been linked with ulcerative colitis in humans . L . intracellularis is an obligate intracellular bacterium, and the pathogenic mechanisms used to cause disease are unknown . Using in vitro-grown organisms as a source of genomic DNA, we identified a Lawsonia gene which encodes a surface antigen, LsaA (for Lawsonia surface antigen), associated with attachment to and entry into cells . The deduced amino acid sequence of this protein showed some similarity to members of a novel protein family identified in a number of other bacterial pathogens but for which roles are not fully defined . Transcription of this gene was detected by reverse transcription-PCR in L . intracellularis grown in vitro in IEC18 cells and in bacteria present in ileal tissue from infected animals . Immunohistochemistry with specific monoclonal antibody and immunoblotting with sera from infected animals demonstrated that LsaA protein is synthesized by L . intracellularis during infection . Expression of this gene during infection in vitro and in vivo suggests that this surface antigen is involved during infection, and phenotypic analysis indicated a role during L . intracellularis attachment to and entry into intestinal epithelial cells

Biochem J, 2002 Sep 1, 366(Pt 2), 573 - 83
Isoprenoid biosynthesis in higher plants and in Escherichia coli: on the branching in the methylerythritol phosphate pathway and the independent biosynthesis of isopentenyl diphosphate and dimethylallyl diphosphate; Hoeffler JF et al.; In the bacterium Escherichia coli, the mevalonic-acid (MVA)-independent 2-C-methyl-d-erythritol 4-phosphate (MEP) pathway is characterized by two branches leading separately to isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP) . The signature of this branching is the retention of deuterium in DMAPP and the deuterium loss in IPP after incorporation of 1-{4-(2)H}deoxy-d-xylulose ({4-(2)H}DX) . Feeding tobacco BY-2 cell-suspension cultures with {4-(2)H}DX resulted in deuterium retention in the isoprene units derived from DMAPP, as well as from IPP in the plastidial isoprenoids, phytoene and plastoquinone, synthesized via the MEP pathway . This labelling pattern represents direct evidence for the presence of the DMAPP branch of the MEP pathway in a higher plant, and shows that IPP can be synthesized from DMAPP in plant plastids, most probably via a plastidial IPP isomerase.

Biochim Biophys Acta, 2002 May 20, 1597(1), 123 - 32
Purification and characterization of ferredoxin-NAD(P)(+) reductase from the green sulfur bacterium Chlorobium tepidum; Seo D et al.; Ferredoxin-NAD(P)(+) reductase {EC 1.18.1.3, 1.18.1.2} was isolated from the green sulfur bacterium Chlorobium tepidum and purified to homogeneity . The molecular mass of the subunit is 42 kDa, as deduced by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) . The molecular mass of the native enzyme is approximately 90 kDa, estimated by gel-permeation chromatography, and is thus a homodimer . The enzyme contains one FAD per subunit and has absorption maxima at about 272, 385, and 466 nm . In the presence of ferredoxin (Fd) and reaction center (RC) complex from C . tepidum, it efficiently catalyzes photoreduction of both NADP(+) and NAD(+) . When concentrations of NADP(+) exceeded 10 microM, NADP(+) photoreduction rates decreased with increased concentration . The inhibition by high concentrations of substrate was not observed with NAD(+) . It also reduces 2,6-dichlorophenol-indophenol (DPIP) and molecular oxygen with either NADPH or NADH as efficient electron donors . It showed NADPH diaphorase activity about two times higher than NADH diaphorase activity in DPIP reduction assays at NAD(P)H concentrations less than 0.1 mM . At 0.5 mM NAD(P)H, the two activities were about the same, and at 1 mM, the former activity was slightly lower than the latter.

Environ Pollut, 2002, 118(3), 427 - 33
Role of loosely bound humic substances and humin in the bioavailability of phenanthrene aged in soil; Nam K et al.; A study was conducted to determine a possible role of loosely bound humic substances (i.e., humic and fulvic acids) in bioavailability of aged phenanthrene with time . In this study, long-term residence of phenanthrene in soil is defined as aging or sequestration, and the effect was determined by the declined bioavailability to bacteria of the polycyclic aromatic hydrocarbon with increased residence time . After 1, 7, and 100 days of aging of phenanthrene in Lima loam, about 90-93% of initial phenanthrene was recovered from the humin-mineral fraction of Lima loam whereas less than 12% was found in humic and fulvic acids of the same soil . Mineralization rates of phenanthrene aged in the humin-mineral fraction significantly decreased with time by the test bacterium P5-2 . In terms of extents of mineralization, the difference with time was not appreciable, but still significant at P<0.05 . Additional decreases in the rates and extents of mineralization were observed with the whole soil (i.e . Lima loam) to which phenanthrene had been aged . Data suggest that major sequestration sites for phenanthrene may reside in the humin-mineral fraction, and probably humic and fulvic acids may act as a physico-chemical barrier to bacterial degradation so that the compound's bioavailability may be limited.

Biosci Biotechnol Biochem, 2002 Mar, 66(3), 489 - 500
Cloning, sequencing, and expression of a gene encoding the monomeric isocitrate dehydrogenase of the nitrogen-fixing bacterium, Azotobacter vinelandii; Sahara T et al.; Isocitrate dehydrogenase (IDH: EC 1.1.1.42) of Azotobacter vinelandii was purified to an electrophoretically homogeneous state, and a gene (icd) encoding this enzyme was cloned and sequenced . The N-terminal amino acid sequence of the purified enzyme was consistent with that deduced from the nucleotide sequence of the icd gene . The deduced amino acid sequence of this gene showed high identity (62-66%) to those of the other bacterial monomeric IDHs . Expression of the icd gene in Escherichia coli was examined by measuring the enzyme activity and mRNA level . Primer extension analyses revealed that two species of mRNAs with different lengths of 5'-untranslated regions (TS-1 and TS-2) were present, of which the 5'-terminals (TS-1 and TS-2 sites) were cytosines located at 244 bp and 101 bp upstream of translational initiation codon, respectively . Conserved promoter elements were present at -35 and -10 regions from the TS-1 site, whereas no such a common motif was found in the upstream region of the TS-2 site . Deletion of the promoter elements upstream of the TS-1 site resulted in complete loss of IDH activity in the E . coli transformant . When the promoter elements upstream of the TS-1 site were intact, the levels of TS-1 and TS-2 were varied greatly by altering exogenous nutrients for growth . The cells grown in a nutrient-rich medium produced large amounts of TS-1 and had a low level of IDH activity . In a nutrient-poor medium, the cells contained large amounts of TS-2 and high levels of IDH activity.

Science, 2002 May 10, 296(5570), 1124 - 6
Role of delayed nuclear envelope breakdown and mitosis in Wolbachia-induced cytoplasmic incompatibility; Tram U et al.; The bacterium Wolbachia manipulates reproduction in millions of insects worldwide; the most common effect is cytoplasmic incompatibility (CI) . We found that CI resulted from delayed nuclear envelope breakdown of the male pronucleus in Nasonia vitripennis . This caused asynchrony between the male and female pronuclei and, ultimately, loss of paternal chromosomes at the first mitosis . When Wolbachia were present in the egg, synchrony was restored, which explains suppression of CI in these crosses . These results suggest that Wolbachia target cell cycle regulatory proteins . A striking consequence of CI is that it alters the normal pattern of reciprocal centrosome inheritance in Nasonia.

Curr Drug Metab, 2002 Apr, 3(2), 159 - 73
Tetrahydrobiopterin biosynthesis, utilization and pharmacological effects; Werner-Felmayer G et al.; Tetrahydrobiopterin (H4-biopterin) is an essential cofactor of a set of enzymes that are of central metabolic importance, i.e . the hydroxylases of the three aromatic amino acids phenylalanine, tyrosine, and tryptophan, of ether lipid oxidase, and of the three nitric oxide synthase (NOS) isoenzymes . As a consequence, H4-biopterin plays a key role in a vast number of biological processes and pathological states associated with neurotransmitter formation, vasorelaxation, and immune response . In mammals, its biosynthesis is controlled by hormones, cytokines and certain immune stimuli . This review aims to summarize recent developments concerning regulation of H4-biopterin biosynthetic and regulatory enzymes and pharmacological effects of H4-biopterin in various conditions, e.g . endothelial dysfunction or apoptosis of neuronal cells . Also, approaches towards gene therapy of diseases like the different forms of phenylketonuria or of Parkinson's disease are reviewed . Additional emphasis is given to H4-biopterin biosynthesis and function in non-mammalian species such as fruit fly, zebra fish, fungi, slime molds, the bacterium Nocardia as well as to the parasitic protozoan genus of Leishmania that is not capable of pteridine biosynthesis but has evolved a sophisticated salvage network for scavenging various pteridine compounds, notably folate and biopterin.

Photochem Photobiol, 2002 Apr, 75(4), 433 - 6
Spectral heterogeneity in single light-harvesting chlorosomes from green sulfur photosynthetic bacterium chlorobium tepidum; Saga Y et al.; The fluorescence emission properties of single chlorosomes from the green sulfur photosynthetic bacterium Chlorobium (Chl.) tepidum are studied for the first time, using a total internal reflection fluorescence microscope . The fluorescence peak positions of bacteriochlorophyll (BChl)-c self-aggregates in a single chlorosome of Chl . tepidum were widely distributed in the wavelength region between 750 and 768 nm, and the standard deviation (s.d . = 4.1 nm, n = 51) was larger than that of single chlorosomes of Chloroflexus (Cfl.) (s.d . = 1.9 nm, n = 50) . The spectral heterogeneity among single chlorosomes from Chl . tepidum was in sharp contrast to those from Cfl . aurantiacus . The difference of chlorosomal spectral properties between Chl . tepidum and Cfl . aurantiacus at the single-unit level would be ascribed to the homolog composition of BChl-c--chlorosomes of Chl . tepidum have BChl substituted with various alkyl groups at both the 8- and 12-positions, whereas light-harvesting BChl-c molecules in Cfl . chlorosomes have the same substituents at the 8- (ethyl group) and 12- (methyl group) positions.

J Voice, 2002 Mar, 16(1), 87 - 91
The prevalence of Helicobacter pylori infection in benign laryngeal disorders; Rubin JS et al.; Helicobacter pylori (HP) is an accepted cause of chronic active gastritis and has a major causative role in peptic ulcers . It is a gastric carcinogen . Its role in nonulcer dyspepsia (NUD) is less clear, yet 50% of patients with NUD are infected with HP, and some recent literature demonstrates long-term improvement of symptoms following eradication . HP has been investigated in several other organ systems, but has not been investigated to any major degree in laryngeal disorders, a region that could be directly exposed to the bacterium from pharyngolaryngeal reflux . This study represents one arm of a larger study designed to investigate such a relationship . Of 101 patients with nonmalignant voice disorders presenting to our voice clinics, 54.5% tested positive for the H . pylori organism . Of the controls, 47.1% tested positive . When striated into age groups of < 45 years, 46-61 years, and > 62 years, and then age-matched with the controls, the likelihood of infection with the H . pylori organism was greater in both the experimental middle group, and in the middle group when combined with the elder group, than in the matched controls, and this difference demonstrated a trend approaching statistical significance . This finding is discussed in the light of other studies on HP and on gastroesophageal reflex (GER).

Proc Nutr Soc, 2002 Feb, 61(1), 137 - 43
Salivary antioxidants and periodontal disease status; Sculley DV et al.; Periodontal disease is a common chronic adult condition . The bacterium Porphyromonas gingivalis has been implicated in the aetiology of this disease, which causes destruction of the connective tissue and bone around the root area of the tooth . It has been observed that invading P . gingivalis bacteria trigger the release of cytokines such as interleukin 8 and tumour necrosis factor a, leading to elevated numbers and activity of polymorphonucleocytes (PMN) . As a result of stimulation by bacterial antigens, PMN produce the reactive oxygen species (ROS) superoxide via the respiratory burst as part of the host response to infection . Patients with periodontal disease display increased PMN number and activity . It has been suggested that this proliferation results in a high degree of ROS release, culminating in heightened oxidative damage to gingival tissue, periodontal ligament and alveolar bone . Antioxidant constituents in plasma have been well-documented, being chiefly ascorbate, albumin and urate, and these are known to display sensitivity to dietary antioxidant intakes . The concentration of antioxidants in saliva does not appear to mirror those of plasma . The extent of dietary influence upon salivary antioxidant status is unclear . Urate is the predominant salivary antioxidant, with albumin and ascorbate providing minor contributions . Previous research has found reduced salivary antioxidant activity in patients suffering from periodontal disease . An improved understanding of the role antioxidants play in periodontitis, and the influence of nutrition on antioxidant status, may lead to a possible nutritional strategy for the treatment of periodontal disease.

Life Sci Space Res, 1973, 11, 247 - 59
The radiobiological effects of heavy ions on mammalian cells and bacteria; Grigoryev YG et al.; The radiobiological effects of heavy ions have been studied in experiments with mouse corneal epithelium, liver cells of rats irradiated in vivo, and the bacterium Escherichia coli B . From exposure of E . coli B to radiations with different LET, the effectiveness of the modifying influence of anoxia and some other radioprotectors has been determined.

Curr Microbiol, 2002 Jun, 44(6), 406 - 10
Methanogenesis from furfural by defined mixed cultures; Boopathy R; Methanogenesis from furfural by defined mixed cultures was studied . Under sulfate-reducing conditions, a Desulfovibrio strain was used as the furfural-degrading species producing acetic acid . This sulfate-reducing bacterium (SRB) Desulfovibrio strain B is an incomplete oxidizer, unable to carry out the terminal oxidation of organic substrates, leaving acetic acid as the end product . Introduction of acetate-utilizing methanogenic archaeon Methanosarcina barkeri 227 converted acetic acid to methane . This well-defined mixed consortium used furfural as its sole source of carbon and converted it to methane and CO(2) . In the mixed culture, when a methanogen inhibitor was used in the culture medium, furfural was converted to acetic acid by the Desulfovibrio strain B, but acetic acid did not undergo further metabolism . On the other hand, when the growth of Desulfovibrio in the consortium was suppressed with a specific SRB inhibitor, namely molybdate, furfural was not degraded . Thus, the metabolic activities of both Desulfovibrio strain B and M . barkeri 227 were essential for the complete degradation of furfural.

Biosci Biotechnol Biochem, 2002 Feb, 66(2), 471 - 5
Molecular cloning of the gene encoding a novel beta-N-acetylhexosaminidase from a marine bacterium, Alteromonas sp . strain O-7, and characterization of the cloned enzyme; Tsujibo H et al.; We have reported that the chitinolytic system of Alteromonas sp . strain O-7 consists of chitinases (ChiA, ChiB, and ChiC), a chitinase-like enzyme (ChiD), beta-N-acetylglucosaminidases (GlcNAcasesA, GlcNAcaseB, and GlcNAcaseC), and a novel transglycosylative enzyme (Hex99) . The gene encoding a beta-hexosaminidase with an unusual substrate specificity (hex86), located upstream of the hex99 gene, was cloned and sequenced . The gene encoded a protein of 761 amino acids with a calculated molecular mass of 86,758 Da . The deduced amino acid sequence of Hex86 showed sequence similarity with beta-hexosaminidases belonging to family 20 . The hex86 gene was expressed in Escherichia coli, and the recombinant enzyme was purified to homogeneity . The enzyme rapidly cleaved p-nitrophenyl-beta-N-acetyl-D-glucosaminide and slowly cleaved p-nitrophenyl-beta-N-acetyl-D-galactosaminide . Unexpectedly, the enzyme did not hydrolyzed chitin oligosaccharides under the assay conditions for synthetic glycosides . However, after prolonged incubation with excessive quantities of the enzyme, Hex86 hydrolyzed chitin oligosaccharides . These results indicate that Hex86 is a novel enzyme that prefers p-nitrophenyl-beta-N-acetyl-D-glucosaminide to chitin oligosaccharides as a substrate.

Biosci Biotechnol Biochem, 2002 Feb, 66(2), 416 - 21
Isolation and characterization of the genes encoding two metalloproteases (MprI and MprII) from a marine bacterium, Alteromonas sp . strain O-7; Miyamoto K et al.; An extracellular alkaline metalloprotease (MprI) from Alteromonas sp . strain O-7 was purified and characterized . The molecular mass of the purified enzyme was estimated to be 56 kDa by SDS-PAGE . The optimum pH and temperature were pH 10.0 and 60 degrees C, respectively . The gene (mprI) encoding MprI was cloned and its nucleotide sequence was analyzed . The deduced amino acid sequence of MprI showed significant similarity to metalloproteases classified into the thermolysin family . Furthermore, sequence analysis showed that another metalloprotease (MprII)-encoding gene was located downstream from mprI . The deduced amino acid sequence of MprII showed high similarity to metalloproteases of the aminopeptidase family . Similar repeated C-terminal extensions were found in both MprI and MprII.

Wien Med Wochenschr, 2002, 152(5-6), 135 - 40
{Significance of Helicobacter pylori infection for stomach lymphoma and stomach carcinoma}; Dragosics B; The discovery of Helicobacter pylori in 1983 revolutionised pathogenetic hypotheses of almost all gastric diseases and markedly enhanced research especially in the field of malignant tumours of this organ . Based on epidemiological studies indicating an association of H . pylori infection and malignant gastric tumours the WHO classified this bacterium as "class I carcinogen" already in 1994 . Although the high prevalence of this germ worldwide is sharply contrasting the low incidence of gastric carcinomas and lymphomas its role as independent risk factor in the carcinogenesis of these tumours today is reasonably evidence-based . Thus, epidemiologists are calculating a reduction of 80% of all MALT-type lymphomas and of about 60% of gastric "non cardia" carcinomas in the scenario of an "H . pylori-free" world . However, complete remission of 80% of early lymphomas of MALT-type confined to mucosa and submucosa, only, after antibiotic eradication of the bacterium is well established in literature and follow-up data are confirming sustained response after years . The strength of impact of an H . pylori infection on gastric carcinogenesis will be figured out by prospective studies on a non infected population in the future.

Carbohydr Res, 2002 Apr 30, 337(9), 869 - 72
Structure of the O-specific polysaccharide of the lipopolysaccharide of Azospirillum brasilense Sp245; Fedonenko YP et al.; An O-specific polysaccharide was isolated from the lipopolysaccharide of a plant-growth-promoting bacterium Azospirillum brasilense Sp245 and studied by sugar analyses along with one- and two-dimensional 1H and 13C NMR spectroscopy, including NOESY . The polysaccharide was found to be a new rhamnan with a pentasaccharide repeating unit having the following structure:-->2)-beta-D-Rhap-(1-->3)-alpha-D-Rhap-(1-->3)-alpha-D-Rhap-(1-->2)-alpha-D-Rhap-(1-->2)-alpha-D-Rhap-(1-->

Nat Rev Mol Cell Biol, 2002 Mar, 3(3), 167 - 76
Dynamic localization of proteins and DNA during a bacterial cell cycle; Jensen RB et al.; A cellular differentiation programme that culminates in an asymmetric cell division is an integral part of the cell cycle in the bacterium Caulobacter crescentus . Recent work has uncovered mechanisms that ensure the execution of many events at different times during the cell cycle and at specific places in the cell . Surprisingly, in this one-micron bacterial cell, the dynamic spatial disposition of regulatory proteins, structural proteins and specific regions of the chromosome are important components of both cell-cycle progression and the generation of daughter cells with different cell fates.

Mol Microbiol, 2002 May, 44(3), 819 - 28
Pyruvate kinase from Chlamydia trachomatis is activated by fructose-2,6-bisphosphate; Iliffe-Lee ER et al.; Pyruvate kinase is the final regulatory point in the catabolic Embden-Meyerhoff-Parnas pathway, which controls the carbon flux of glycolytic intermediates and regulates the level of ATP in the cell . In a previous study, we identified, cloned and sequenced pyruvate kinase from the obligate intracellular bacterium Chlamydia trachomatis and demonstrated that the enzyme was active in crude extract . Here, we report the kinetic properties of highly purified C . trachomatis pyruvate kinase . The results indicate that C . trachomatis pyruvate kinase is 53.5 kDa with a pH optima of 7.3 . Kinetic studies show that C . trachomatis pyruvate kinase requires both K+ and Mg2+ ions for activity, exhibits sigmoidal kinetics with respect to phosphoenolpyruvate and Michaelis-Menten kinetics with respect to ADP . In addition, C . trachomatis pyruvate kinase is able to use alternative nucleoside diphosphates as phosphate acceptors, although it shows the greatest activity with ADP . In contrast to other bacterial pyruvate kinases that are activated by AMP, our data show that AMP, in addition to ATP and GTP, inhibits C . trachomatis pyruvate kinase . Surprisingly, unlike any other known bacterial pyruvate kinase, C . trachomatis pyruvate kinase was allosterically activated by fructose-2,6-bisphosphate, an important regulatory metabolite that has only been reported in eukaryotes.

Bioorg Med Chem Lett, 2002 May 20, 12(10), 1339 - 42
A novel type of nonsteroidal estrone sulfatase inhibitors; Jutten P et al.; Madurahydroxylactone (MHL) is a secondary metabolite produced by the soil bacterium Nonomuria rubra and belongs to the family of benzo{a}naphthacenequinones . We report the initial results and structure-activity relationships of our study into a series of thiosemicarbazone derivatives of madurahydroxylactone as potential nonsteroidal inhibitors of the enzyme estrone sulfatase . The most active compound, the cyclohexylthiosemicarbazone, was shown to be a non-competitive inhibitor with a K(i) of 0.35microM . This compound is devoid of estrogenic properties and showed low acute toxicity in the hen's fertile egg screening test.

Dig Dis Sci, 2002 Apr, 47(4), 823 - 30
Cellular immune responses in Helicobacter heilmannii infection: evaluation of the role of the host and the bacterium; Cinque SM et al.; We evaluated some aspects of the immune response to Helicobacter heilmannii in two mouse strains . Gastritis that was more severe in infected C57BL/6 mice . A proliferative response to H . pylori antigens was observed in splenocytes from H . heilmannii-positive and -negative mice, similar in the positive- and negative-BALB/c mice, but lower in the positive- than in the negative-C57BL/6 animals . A decrease in B cells and an increase in CD4+ cells after stimulation with type I H . pylori antigen and an increase in CD8+ cells after stimulation with type I and II antigens was observed in infected C57BL/6 mice . Conversely, the percentage of CD4+, CD8+, and B cells was similar in positive- and negative-BALB/c mice . These results demonstrated that the immune response is similar in H . heilmannii and H . pylori infection and strengthened the importance of host and bacterial virulence markers in the immune response to gastric Helicobacter infections.

Scand J Gastroenterol, 2002 Apr, 37(4), 409 - 13
Role of Helicobacter pylori CagA+ infection in determining oxidative DNA damage in gastric mucosa; Papa A et al.; BACKGROUND: Although Helicobacter pylori is a risk factor for gastric cancer, the role of the bacterium in the development of this malignancy is not defined precisely . Reactive oxygen species (ROS) could play an important role in carcinogenesis by inducing DNA damage . The aims of the present study were: 1) to assess the production of ROS and 8-hydroxy-2'-deoxyguanosine (8-OHdG), a sensitive marker of oxidative DNA injury, in gastric mucosa, according to H . pylori status and cytotoxic associated gene product A (CagA); 2) to determine the relationship between ROS generation and amount of 8-OHdG . METHODS: Gastric biopsy specimens were obtained from 60 consecutive patients . ROS generation was measured by luminol enhanced chemiluminescence . 8-OHdG detection was performed by an immunoperoxidase method, using a specific anti 8-OHdG monoclonal antibody . RESULTS: 40/60 patients (67%) were H . pylori-positive . ROS generation was significantly higher in patients positive for H . pylori infection as compared to negative . 8-OHdG detection was performed in 30 patients in which CagA presence was also investigated . High expression of 8-OHdG was detected in 14/20 (70%) H . pylori-positive patients (13 CagA+ and 1 CagA-) and in 2/10 (20%) H . pylori-negative patients . A significant correlation was found between ROS production and 8-OHdG content . CONCLUSION: H . pylori infection by a CagA+ strain is associated with the highest production of ROS to which a severe oxidative DNA damage corresponds . This sequence of events could support the hypothesis that the oxygen-free radicals-mediated damage due to H . pylori cytotoxic strains could be a driving force that leads from chronic gastritis to gastric carcinoma.

Microbiology, 2002 May, 148(Pt 5), 1439 - 46
An essential virulence protein of Brucella abortus, VirB4, requires an intact nucleoside-triphosphate-binding domain; Watarai M et al.; Brucella abortus is a facultative intracellular bacterium capable of surviving inside macrophages . The VirB complex, which is highly similar to conjugative DNA transfer apparatuses, is required for intracellular replication . A conserved NTP-binding domain in VirB4 suggests that one or both proteins couple energy by NTP hydrolysis to transport of putative effector molecule(s) . Here it is shown that a mutant strain of B . abortus that contains an in-frame deletion in virB4 is unable to replicate in macrophages and survives in mice . Intracellular replication and virulence in mice are fully restored by expressing virB4 in trans, indicating that VirB4 is essential for intracellular replication and virulence in mice . An alteration within the NTP-binding region of VirB4 by site-directed mutagenesis abolished complementation of a virB4 mutant, demonstrating that an intact NTP-binding domain is critical for VirB4 function . Intracellular replication was inhibited in wild-type B . abortus after introducing a plasmid expressing a mutant VirB4 altered in the NTP-binding region . The dominant negative phenotype suggests that VirB4 either functions as a multimer or interacts with some other component(s) necessary for intracellular replication . Wild-type B . abortus-containing phagosomes lack the glycoprotein LAMP-1, which is an indicator of the normal endocytic pathway . Mutant strains were found in phagosomes that co-localized with LAMP-1, indicating that VirB4 containing the intact NTP-binding region is essential for evasion of fusion with lysosomes.

J Gastroenterol Hepatol, 2002 Jan, 17(1), 27 - 31
Reinfection rate following effective therapy against Helicobacter pylori infection in Japan; Adachi M et al.; BACKGROUND AND AIM: In developed countries, reinfection of Helicobacter pylori (H . pylori) after eradication of the bacterium is unusual, while the reinfection rate in developing countries is variable . In this study, we determined the reinfection rate after successful H . pylori eradication in Japan, a country with a high prevalence of H . pylori infection . METHODS: After successful eradication, 377 patients were followed up by endoscopy and urea breath test annually . In reinfected patients, H . pylori strains isolated initially and after reinfection were compared by using random amplification of polymorphic DNA fingerprinting . RESULTS: H . pylori became positive in four of 337 patients (1.2) 1 year after eradication and in two of 133 patients (1.5) 2 years after eradication . One patient experienced an ulcer relapse 2 years after eradication therapy . Random amplification of polymorphic DNA fingerprinting of the isolated strains from four of the six patients showed two had identical strains (at 1 year) while the other two had different strains (one at 1 year and one at 2 years) . When infection in the two patients reinfected with identical strains is considered a recrudescence, the true reinfection rate is < 0.8 per patient year . CONCLUSIONS: The reinfection rate after eradication of H . pylori is low in Japan despite the country's high prevalence of H . pylori infection.

Proc Natl Acad Sci U S A, 2002 May 14, 99(10), 7078 - 83 Epub 2002 Apr 30.
Repression of photosynthesis gene expression by formation of a disulfide bond in CrtJ; Masuda S et al.; Many species of purple photosynthetic bacteria repress synthesis of their photosystem in the presence of molecular oxygen . The bacterium Rhodobacter capsulatus mediates this process by repressing expression of bacteriochlorophyll, carotenoid, and light-harvesting genes via the aerobic repressor, CrtJ . In this study, we demonstrate that CrtJ forms an intramolecular disulfide bond in vitro and in vivo when exposed to oxygen . Mutational and sulfhydryl-specific chemical modification studies indicate that formation of a disulfide bond is critical for CrtJ binding to its target promoters . Analysis of the redox states of aerobically and anaerobically grown cells indicates that they have similar redox states of approximately -200 mV, thereby demonstrating that a change in midpoint potential is not responsible for disulfide bond formation . In vivo and in vitro analyses indicate that disulfide bond formation in CrtJ is insensitive to the addition of hydrogen peroxide but is sensitive to molecular oxygen . These results suggest that disulfide bond formation in CrtJ may differ from the mechanism of disulfide bond formation used by OxyR.

J Gastroenterol Hepatol, 2002 Apr, 17(4), 495 - 502
Gastric cancer: laboratory bench to clinic; Houghton J et al.; Gastric cancer is the second most common cause of cancer-related mortality worldwide and the 14th overall cause of death . Detection of disease usually occurs at an advanced stage and overall survival rates for gastric cancer are poor . Our current model for gastric cancer progression clearly maintains Helicobacter infection as the primary inducer of gastric metaplastic and neoplastic disease . Helicobacter pylori is a ubiquitous organism, infecting more than half the world's population . It has been suggested that this infection directly contributes to the formation of gastric cancer in up to 80% of cases; however, gastric malignancy develops in only a subset (< 1%) of infected patients . Therefore, predisposition to Helicobacter-associated gastric cancer is most likely multifactorial, including the interaction of bacterial, host and environmental components . Our understanding of how the organism interacts with the gastric mucosa and synergizes with dietary and other environmental factors to induce malignant mucosal changes is evolving . Indeed, H . pylori has direct effects on the gastric mucosa, but the major factor in disease progression appears to be a robust host Th1 immune response in the setting of a permissive environment . In combination, these factors predispose to the formation of atrophy, metaplasia and gastric cancer . Understanding the interaction of the bacterium with the host and the environment can potentially identify patients most at risk . Identifying potentially removable factors (in addition to H . pylori infection) in the acquisition and progression of neoplastic disease may provide targets for early intervention and prevention strategies .

J Basic Microbiol, 2002, 42(2), 127 - 31
Characterization of JP-7 jet fuel degradation by the bacterium Nocardioides luteus strain BAFB; Jung CM et al.; In the fall of 1996, numerous bacteria capable of degrading JP-7 jet fuel were isolated from soil collected at Beale Air Force Base in northern California . The most prevalent organism, identified as Nocardioides luteus by16s rRNA sequencing (MIDI Labs, Inc.), was selected for further analysis . Analysis of JP-7 following inoculation with N . luteus demonstrated degradation of the C(11) alkane component of the fuel . Growth rates of N . luteus were determined with alkanes of various lengths as the sole carbon and energy source . The organism grew best on shorter length alkanes (C(8) and C(10)) . Growth was measurably slower on C(11), and minimal on C(12), C(13), and C(14).

Graefes Arch Clin Exp Ophthalmol, 2002 Apr, 240(4), 291 - 5 Epub 2002 Mar 12.
Outbreak of Empedobacter brevis endophthalmitis after cataract extraction; Janknecht P et al.; BACKGROUND: We present a series of patients who were operated upon on the same day by the same surgeon . All of those patients developed postoperative endophthalmitis due to Empedobacter brevis - a bacterium hitherto unknown in ophthalmologic literature . METHODS: Twelve patients were referred because of endophthalmitis after cataract extraction . The patients' files were studied and the intraoperative and postoperative outcome was analysed . RESULTS: Twelve patients (five male, seven female, mean age 75 years) presented 1-6 days after uncomplicated cataract extraction . Although some suffered from medical or ophthalmological diseases there was no association with the severity of the endophthalmitis . Eleven patients required vitrectomy, seven as primary procedure, one primary with extraction of the lens and three secondary after anterior chamber lavage and intravitreal antibiotics . In three cases vitrectomy had to be repeated together with extraction of the intraocular lens . There were two postoperative retinal detachments that required silicon oil and in one case an encircling band . Mean visual acuity rose from 0.02 to 0.47 by 9 months after operation . Empedobacter brevis was found in the anterior chamber and in the vitreous in all except one patient . CONCLUSION: In high-volume cataract surgery endemic endophthalmitis is always possible . Sources of infection may be anything from the lens to the sterilisation process, the latter being the primary suspect in our series . Prompt, adequate and (if necessary) aggressive treatment by vitreous surgery may lead to favourable results.

J Bacteriol, 2002 May, 184(10), 2748 - 54
Biological properties and cell tropism of Chp2, a bacteriophage of the obligate intracellular bacterium Chlamydophila abortus; Everson JS et al.; A number of bacteriophages belonging to the Microviridae have been described infecting chlamydiae . Phylogenetic studies divide the Chlamydiaceae into two distinct genera, Chlamydia and Chlamydophila, containing three and six different species, respectively . In this work we investigated the biological properties and host range of the recently described bacteriophage Chp2 that was originally discovered in Chlamydophila abortus . The obligate intracellular development cycle of chlamydiae has precluded the development of quantitative approaches to assay bacteriophage infectivity . Thus, we prepared hybridomas secreting monoclonal antibodies (monoclonal antibodies 40 and 55) that were specific for Chp2 . We demonstrated that Chp2 binds both C . abortus elementary bodies and reticulate bodies in an enzyme-linked immunosorbent assay . Monoclonal antibodies 40 and 55 also detected bacteriophage Chp2 antigens in chlamydia-infected eukaryotic cells . We used these monoclonal antibodies to monitor the ability of Chp2 to infect all nine species of chlamydiae . Chp2 does not infect members of the genus Chlamydia (C . trachomatis, C . suis, or C . muridarum) . Chp2 can infect C . abortus, C . felis, and C . pecorum but is unable to infect other members of this genus, including C . caviae and C . pneumoniae, despite the fact that these chlamydial species support the replication of very closely related bacteriophages.

J Bacteriol, 2002 May, 184(10), 2719 - 27
Rescue of social motility lost during evolution of Myxococcus xanthus in an asocial environment; Velicer GJ et al.; Replicate populations of the social bacterium Myxococcus xanthus underwent extensive evolutionary adaptation to an asocial selective environment (liquid batch culture) . All 12 populations showed partial or complete loss of their social (S) motility function after 1,000 generations of evolution . Mutations in the pil gene cluster (responsible for type IV pilus biogenesis and function) were found to be at least partially responsible for the loss of S motility in the majority of evolved lines . Restoration (partial or complete) of S motility in the evolved lines by genetic complementation with wild-type pil genes positively affected their fruiting body development and sporulation while negatively affecting their competitive fitness in the asocial regime . This genetic tradeoff indicates that mutations in the pil region were adaptive in the asocial selective environment . This finding was confirmed by experiments showing that defined deletions of pil gene regions conferred a competitive advantage under asocial conditions . Moreover, an amino acid substitution in an evolved genotype was located in a region predicted by genetic complementation analysis to bear an adaptive mutation.

Mol Microbiol, 2002 Apr, 44(2), 417 - 30
StoPK-1, a serine/threonine protein kinase from the glycopeptide antibiotic producer Streptomyces toyocaensis NRRL 15009, affects oxidative stress response; Neu JM et al.; The glycopeptide antibiotic-producing bacterium, Streptomyces toyocaensis NRRL 15009, has proteins phosphorylated on Ser, Thr, Tyr and His, implying the presence of a battery of associated kinases . We have identified the Ser/Thr protein kinase gene fragments stoPK-1, stoPK-2, stoPK-3 and stoPK-4 from S . toyocaensis NRRL 15009 by a polymerase chain reaction (PCR) strategy using oligonucleotide primers based on eukaryotic Ser/Thr and Tyr kinase sequences . One of these (stoPK-1) was subsequently cloned in its entirety from a 3.2 kb genomic BamHI fragment . stoPK-1 encodes a 642-amino-acid protein with a predicted N-terminal Ser/Thr kinase domain and a C-terminal coiled-coil region divided by a membrane-spanning region . Expression of StoPK-1 in Escherichia coli yielded a protein confined to the membrane fraction, which was found to be phosphorylated exclusively on Thr residues and could transfer phosphate to the model substrates myelin basic protein and histone H1 . Both autophosphorylation and phosphoryl transfer could be inhibited by the flavanoid apigenin . Disruption of stoPK-1 with the apramycin resistance gene in the S . toyo-caensis chromosome resulted in changes in mycelial morphology and an increased sensitivity to the redox cycling agents paraquat and nitrofurantoin on glucose-containing media . Supplying stoPK-1 or the S . coelicolor homologue pkaF in trans could reverse this sensitivity, whereas a catalytically inactive mutant of stoPK-1 could not, indicating that kinase activity is essential for this phenotype . This suggests a link between this membrane-bound protein kinase in signalling pathways sensitive to oxidative stress and/or glucose metabolism . These results broaden the roles of Ser/Thr protein kinases in bacteria and underscore the diversity of signal transduction mechanisms available to respond to various stimuli.

J Appl Microbiol, 2002, 92(5), 976 - 82
The enrichment of a ruminal bacterium (Megasphaera elsdenii YJ-4) that produces the trans-10, cis-12 isomer of conjugated linoleic acid; Kim YJ et al.; AIMS: To isolate predominant ruminal bacteria that produce trans-10, cis-12 conjugated linoleic acid (CLA) from linoleic acid (LA) . METHODS AND RESULTS: Mixed bacteria from ruminal contents of a cow fed grain were enriched with DL-lactate and trypticase . They produced more trans-10, cis-12 CLA than those that were not enriched (7 vs 2 microg mg protein(-1), P < 0.05) . Enrichments had an abundance of large cocci that produced trans-10, cis-12 CLA from LA . Strain YJ-4 produced the most trans-10, cis-12 CLA (approx . 7 microg mg protein(-1)) and 16S rDNA sequencing indicated that YJ-4 was a strain of Megasphaera elsdenii . Megasphaera elsdenii T81 produced approx . 4 microg trans-10, cis-12 CLA mg protein(-1) while strains B159, AW106 and JL1 produced < 0.5 microg mg protein(-1) . The trans-10, cis-12 CLA production of YJ-4 was first order with respect to cell concentration (0-800 microg protein ml(-1)), but kinetics were not first order with respect to substrate concentration . CONCLUSIONS: Some M . elsdenii strains produce significant amounts of trans-10, cis-12 CLA . SIGNIFICANCE AND IMPACT OF THE STUDY: Trans-10, cis-12 CLA appears to cause milk fat depression in cattle fed diets supplemented with grain and polyunsaturated fatty acids, but predominant ruminal bacteria that produced trans-10, cis-12 CLA from LA had not previously been isolated.

J Appl Microbiol, 2002, 92(5), 903 - 11
Comparison of adhesion of the food spoilage bacterium Shewanella putrefaciens to stainless steel and silver surfaces; Hjelm M et al.; AIMS: To compare the number of attached Shewanella putrefaciens on stainless steel with different silver surfaces, thus evaluating whether silver surfaces could contribute to a higher hygienic status in the food industry . METHODS AND RESULTS: Bacterial adhesion to three types of silver surface (new silver, tarnished silver and sulphide-treated silver) was compared with adhesion to stainless steel (AISI 316) using the Malthus indirect conductance method to estimate the number of cfu cm(-2) . The number of attached bacteria on new silver surfaces was lower than on steel for samples taken after 24 h . However, this was not statistically significant (P > 0.05) . The numbers of attached bacteria were consistently lower when tarnished silver surfaces were compared with stainless steel and some, but not all, experiments showed statistical significance (P < 0.05) . Treating new silver with sulphide to reproduce a tarnished silver surface did not result in a similar lowering of adhering cells when compared with steel (P > 0.05) . CONCLUSIONS: New or tarnished silver surfaces caused a slight reduction in numbers of attached bacteria; however, the difference was only sometimes statistically significant . SIGNIFICANCE AND IMPACT OF THE STUDY: The lack of reproducibility in differences in numbers adhering to the different surfaces and lack of statistical significance between numbers of adhered viable bacteria do not indicate that the tested silver surfaces can be used to improve hygienic characteristics of surfaces in the food industry.

Proc Natl Acad Sci U S A, 2002 Apr 30, 99(9), 6017 - 22 Epub 2002 Apr 23.
An alternative carotenoid-to-bacteriochlorophyll energy transfer pathway in photosynthetic light harvesting; Papagiannakis E et al.; Blue and green sunlight become available for photosynthetic energy conversion through the light-harvesting (LH) function of carotenoids, which involves transfer of carotenoid singlet excited states to nearby (bacterio)chlorophylls (BChls) . The excited-state manifold of carotenoids usually is described in terms of two singlet states, S(1) and S(2), of which only the latter can be populated from the ground state by the absorption of one photon . Both states are capable of energy transfer to (B)Chl . We recently showed that in the LH1 complex of the purple bacterium Rhodospirillum rubrum, which is rather inefficient in carotenoid-to-BChl energy transfer, a third additional carotenoid excited singlet state is formed . This state, which we termed S*, was found to be a precursor on an ultrafast fission reaction pathway to carotenoid triplet state formation . Here we present evidence that S* is formed with significant yield in the LH2 complex of Rhodobacter sphaeroides, which has a highly efficient carotenoid LH function . We demonstrate that S* is actively involved in the energy transfer process to BChl and thus have uncovered an alternative pathway of carotenoid-to-BChl energy transfer . In competition with energy transfer to BChl, fission occurs from S*, leading to ultrafast formation of carotenoid triplets . Analysis in terms of a kinetic model indicates that energy transfer through S* accounts for 10-15% of the total energy transfer to BChl, and that inclusion of this pathway is necessary to obtain a highly efficient LH function of carotenoids.

J Immunol, 2002 May 1, 168(9), 4692 - 700
Helicobacter pylori induces macrophage apoptosis by activation of arginase II; Gobert AP et al.; Helicobacter pylori infection induces innate immune responses in macrophages, contributing to mucosal inflammation and damage . Macrophage apoptosis is important in the pathogenesis of mucosal infections but has not been studied with H . pylori . NO derived from inducible NO synthase (iNOS) can activate macrophage apoptosis . Arginase competes with iNOS by converting L-arginine to L-ornithine . Since we reported that H . pylori induces iNOS in macrophages, we now determined whether this bacterium induces arginase and the effect of this activation on apoptosis . NF-kappa B-dependent induction of arginase II, but not arginase I, was observed in RAW 264.7 macrophages cocultured with H . pylori . The time course of apoptosis matched those of both arginase and iNOS activities . Surprisingly, apoptosis was blocked by the arginase inhibitors N(omega)-hydroxy-L-arginine or N(omega)-hydroxy-nor-L-arginine, but not by the iNOS inhibitor N-iminoethyl-L-lysine . These findings were confirmed in peritoneal macrophages from iNOS-deficient mice and were not dependent on bacterial-macrophage contact . Ornithine decarboxylase (ODC), which metabolizes L-ornithine to polyamines, was also induced in H . pylori-stimulated macrophages . Apoptosis was abolished by inhibition of ODC and was restored by the polyamines spermidine and spermine . We also demonstrate that arginase II expression is up-regulated in both murine and human H . pylori gastritis tissues, indicating the likely in vivo relevance of our findings . Therefore, we describe arginase- and ODC-dependent macrophage apoptosis, which implicates polyamines in the pathophysiology of H . pylori infection.

J Appl Microbiol, 2002, 92(4), 753 - 8
Enumeration of Megasphaera elsdenii in rumen contents by real-time Taq nuclease assay; Ouwerkerk D et al.; AIMS: To develop a real-time Taq nuclease assay (TNA) to enable the in vivo enumeration of Megasphaera elsdenii . METHODS AND RESULTS: Megasphaera elsdenii YE34 was phenotypically characteristic of the species and had 16S rDNA sequence similarity of 98% to previously described isolates . Calibration of the number of cells of M . elsdenii against the cycle threshold of fluorescent dye release gave a straight-line relationship with a correlation coefficient approximating unity . The specificity of the assay for M . elsdenii was confirmed by performing it against a panel of 24 heterogeneous, mainly ruminal bacteria . Megasphaera elsdenii was not detected in ruminal contents from a pasture-fed steer but was readily detected 2 and 50 h after the probiotic introduction of the bacterium into the rumen . CONCLUSIONS: Real-time TNA has provided a sensitive and specific means of enumerating the M . elsdenii population in rumen contents . SIGNIFICANCE AND IMPACT OF THE STUDY: Megasphaera elsdenii is an important lactate-degrading ruminal bacterium that has been selected for probiotic use to prevent acidosis and enhance starch utilization in grain-fed cattle . The assay developed in this study provides a tool for determining the ability of probiotically-introduced M . elsdenii to establish useful populations in the rumen.

J Appl Microbiol, 2002, 92(4), 706 - 16
Stimulation of Alexandrium fundyense growth by bacterial assemblages from the Bay of Fundy; Ferrier M et al.; AIMS: To evaluate the influence of marine bacterial assemblages from the Bay of Fundy on Alexandrium fundyense str . CB301 growth . METHODS AND RESULTS: Bacterial assemblages were collected from the Bay of Fundy during an Alexandrium spp . bloom, serially diluted to extinction and inoculated into axenic CB301 cultures . Bacterial assemblages dramatically enhanced CB301 growth . Retrieval and analysis of 16S rDNA fragments revealed an Alteromonas sp . strain to be the only detectable bacterium in all assemblages that promoted A . fundyense and the sole bacterium found in the most dilute inoculum that promoted A . fundyense . While this bacterium has not yet been isolated, other isolates obtained from the assemblages did not stimulate A . fundyense, indicating that the observed stimulation was not a general effect of marine bacteria . CONCLUSIONS: Bay of Fundy marine bacterial assemblages dominated by a member of the family Alteromonadaceae were found to dramatically stimulate growth of A . fundyense . SIGNIFICANCE AND IMPACT OF THE STUDY: These results show that native bacteria have the potential to dramatically promote the growth of A . fundyense and may play an important role in influencing A . fundyense dynamics in the Bay of Fundy.

J Mol Evol, 2002 May, 54(5), 563 - 8
Trends in codon and amino acid usage in Thermotoga maritima; Zavala A et al.; The usage of synonymous codons and the frequencies of amino acids were investigated in the complete genome of the bacterium Thermotoga maritima using a multivariate statistical approach . The GC3 content of each gene was the most prominent source of variation of codon usage . Surprisingly the usage of UGU and UGC (synonymous triplets coding for Cys, the least frequent amino acid in this species) was detected as the second most prominent source of variation . However, this result is probably an artifact due to the very low frequency of Cys together with the nonbiased composition of this genome . The third trend was related to the preferential usage of a subset of codons among highly expressed genes, and these triplets are presumed to be translationally optimal . Concerning the amino acid usage, the hydropathy level of each protein (and therefore the frequency of charged residues) was the main trend, while the second factor was related to the frequency of usage of the smaller residues, suggesting that the cell economy strongly influences the architecture of the proteins . The third axis of the analysis discriminated the usage of Phe, Tyr, Trp (aromatic residues) plus Cys, Met, and His . These six residues have in common the property of being the preferential targets of reactive oxygen species, and therefore the anaerobic condition of T . maritima is an important factor for the amino acid frequencies . Finally, the Cys content of each protein was the fourth trend.

Curr Opin Infect Dis, 2001 Jun, 14(3), 315 - 21
The role of Helicobacter pylori in paediatrics; Vandenplas Y; Helicobacter pylori is a bacterium that colonizes the human stomach, especially during childhood . Although it has been studied intensively during the past decade, many controversies still exist on many important issues, including the clinical relevance of virulence factors, indications for treatment, recommended procedures for diagnosis and the typical symptom profile, among others . The lack of double-blind, placebo-controlled studies in children illustrates the paucity of hard scientific data, and is the major reason for differences in opinion . Therefore, the present review of literature published during the past 2 years raises many questions.

Proc Natl Acad Sci U S A, 2002 Apr 16, 99(8), 5732 - 7
Arabidopsis disrupted in SQD2 encoding sulfolipid synthase is impaired in phosphate-limited growth; Yu B et al.; The sulfolipid sulfoquinovosyldiacylglycerol is one of the three nonphosphorous glycolipids that provide the bulk of the structural lipids in photosynthetic membranes of seed plants . Unlike the galactolipids, sulfolipid is anionic at physiological pH because of its 6-deoxy-6-sulfonate-glucose (sulfoquinovose) head group . The biosynthesis of this lipid proceeds in two steps: first, the assembly of UDP-sulfoquinovose from UDP-glucose and sulfite, and second, the transfer of the sulfoquinovose moiety from UDP-sulfoquinovose to diacylglycerol . The first reaction is catalyzed by the SQD1 protein in Arabidopsis . Here we describe the identification of the SQD2 gene of Arabidopsis . We propose that this gene encodes the sulfoquinovosyltransferase catalyzing the second step of sulfolipid biosynthesis . Expression of SQD1 and SQD2 in Escherichia coli reconstituted plant sulfolipid biosynthesis in this bacterium . Insertion of a transfer DNA into this gene in Arabidopsis led to complete lack of sulfolipid in the respective sqd2 mutant . This mutant showed reduced growth under phosphate-limited growth conditions . The results support the hypothesis that sulfolipid can function as a substitute of anionic phospholipids under phosphate-limited growth conditions . Along with phosphatidylglycerol, sulfolipid contributes to maintaining a negatively charged lipid-water interface, which presumably is required for proper function of photosynthetic membranes.

Antimicrob Agents Chemother, 2002 May, 46(5), 1281 - 7
Widespread distribution of a tet W determinant among tetracycline-resistant isolates of the animal pathogen Arcanobacterium pyogenes; Billington SJ et al.; Tetracycline resistance is common among isolates of the animal commensal and opportunistic pathogen Arcanobacterium pyogenes . The tetracycline resistance determinant cloned from two bovine isolates of A . pyogenes was highly similar at the DNA level (92% identity) to the tet(W) gene, encoding a ribosomal protection tetracycline resistance protein, from the rumen bacterium Butyrivibrio fibrisolvens . The tet(W) gene was found in all 20 tetracycline-resistant isolates tested, indicating that it is a widely distributed determinant of tetracycline resistance in this organism . In 25% of tetracycline-resistant isolates, the tet(W) gene was associated with a mob gene, encoding a functional mobilization protein, and an origin of transfer, suggesting that the determinant may be transferable to other bacteria . In fact, low-frequency transfer of tet(W) was detected from mob+ A . pyogenes isolates to a tetracycline-sensitive A . pyogenes recipient . The mobile nature of this determinant and the presence of A . pyogenes in the gastrointestinal tract of cattle and pigs suggest that A . pyogenes may have inherited this determinant within the gastrointestinal tracts of these animals.

J Biol Chem, 2002 Jun 21, 277(25), 22209 - 14 Epub 2002 Apr 15.
H+-pyrophosphatase of Rhodospirillum rubrum . High yield expression in Escherichia coli and identification of the Cys residues responsible for inactivation my mersalyl; Belogurov GA et al.; H(+)-translocating pyrophosphatase (H(+)-PPase) of the photosynthetic bacterium Rhodospirillum rubrum was expressed in Escherichia coli C43(DE3) cells . Recombinant H(+)-PPase was observed in inner membrane vesicles, where it catalyzed both PP(i) hydrolysis coupled with H(+) transport into the vesicles and PP(i) synthesis . The hydrolytic activity of H(+)-PPase in E . coli vesicles was eight times greater than that in R . rubrum chromatophores but exhibited similar sensitivity to the H(+)-PPase inhibitor, aminomethylenediphosphonate, and insensitivity to the soluble PPase inhibitor, fluoride . Using this expression system, we showed that substitution of Cys(185), Cys(222), or Cys(573) with aliphatic residues had no effect on the activity of H(+)-PPase but decreased its sensitivity to the sulfhydryl modifying reagent, mersalyl . H(+)-PPase lacking all three Cys residues was completely resistant to the effects of mersalyl . Mg(2+) and MgPP(i) protected Cys(185) and Cys(573) from modification by this agent but not Cys(222) . Phylogenetic analyses of 23 nonredundant H(+)-PPase sequences led to classification into two subfamilies . One subfamily invariably contains Cys(222) and includes all known K(+)-independent H(+)-PPases, whereas the other incorporates a conserved Cys(573) but lacks Cys(222) and includes all known K(+)-dependent H(+)-PPases . These data suggest a specific link between the incidence of Cys at positions 222 and 573 and the K(+) dependence of H(+)-PPase.

Infect Immun, 2002 May, 70(5), 2591 - 7
Enhanced disease severity in Helicobacter pylori-infected mice deficient in Fas signaling; Jones NL et al.; Recent evidence suggests that immune-mediated gastric epithelial cell apoptosis through Fas-Fas ligand interactions participates in Helicobacter pylori disease pathogenesis . To define the role of Fas signaling in vivo, H . pylori strain SS1 infection in C57BL/6 mice was compared to that in mice deficient in the Fas ligand (gld) . gld mice had a degree of gastritis similar to that of C57BL/6 mice after 6 weeks (gastritis score, 5.2 +/- 0.6 {mean +/- standard error} versus 3.5 +/- 0.8) and 12 weeks (4.0 +/- 0.7 versus 3.4 +/- 0.5) of infection . Bacterial colonization was comparable in each group of mice at 12 weeks of infection (2.1 +/- 0.3 versus 1.6 +/- 0.3 for gld and C57BL/6, respectively; the difference is not significant) . Sixty-seven percent of H . pylori-infected gld mice displayed atrophic changes in the gastric mucosa, compared with 37% of infected C57BL/6 mice, at 12 weeks . In addition, atrophic changes were more severe in H . pylori-infected gld mice (P < 0.05) . Splenocytes isolated from H . pylori-infected C57BL/6 mice had a twofold increase in production of the Th1 cytokine gamma interferon (IFN-gamma) in response to H . pylori antigens at both 6 and 12 weeks compared to controls (143 +/- 65 versus 69 +/-26 pg/ml and 336 +/- 73 versus 172 +/- 60, respectively) . In contrast, there was a lack of detectable IFN-gamma in gld mice infected with the bacterium . H . pylori-infected C57BL/6 mice had increased epithelial cell apoptosis compared with sham-infected C57BL/6 mice (35.0 +/- 8.9 versus 12.3 +/- 6.9; P < 0.05) . Epithelial cell apoptosis did not differ between H . pylori-infected and control gld mice (5.2 +/- 1.6 versus 6.5 +/- 2.9 {not significant}) . These data demonstrate that mice with mutations in the Fas ligand develop more severe premalignant mucosal changes in response to infection with H . pylori in association with both an impaired gastric epithelial cell apoptotic response and IFN-gamma production . The Fas death pathway modulates disease pathophysiology following murine infection with H . pylori . Deregulation of the Fas pathway could be involved in the transition from gastritis to gastric cancers during H . pylori infection.

Infect Immun, 2002 May, 70(5), 2512 - 8
Hydrolysis of epithelial junctional proteins by Porphyromonas gingivalis gingipains; Katz J et al.; Porphyromonas gingivalis has been implicated as an etiologic agent of adult periodontitis . We have previously shown that P . gingivalis can degrade the epithelial cell-cell junction complexes, thus suggesting that this bacterium can invade the underlying connective tissues via a paracellular pathway . However, the precise mechanism(s) involved in this process has not been elucidated . The purpose of this study was to determine if the arginine- and lysine-specific gingipains of P . gingivalis (i.e., HRgpA and RgpB, and Kgp, respectively) were responsible for the degradation of E-cadherin, the cell-cell adhesion protein in the adherens junctions . In addition, we compared the degradative abilities of the whole gingipains HRgpA and Kgp to those of their catalytic domains alone . In these studies, immunoprecipitated E-cadherin as well as monolayers of polarized Madin-Darby canine kidney (MDCK) epithelial cell cultures were incubated with the gingipains and hydrolysis of E-cadherin was assessed by Western blot analysis . Incubation of P . gingivalis cells with immunoprecipitated E-cadherin resulted in degradation, whereas prior exposure of P . gingivalis cells to leupeptin and especially acetyl-Leu-Val-Lys-aldehyde (which are arginine- and lysine-specific inhibitors, respectively) reduced this activity . Furthermore, incubation of E-cadherin immunoprecipitates with the different gingipains resulted in an effective and similar hydrolysis of the protein . However, when monolayers of MDCK cells were exposed to the gingipains, Kgp was most effective in hydrolyzing the E-cadherin molecules in the adherens junction . Kgp was more effective than its catalytic domain in degrading E-cadherin at 500 nM but not at a lower concentration (250 nM) . These results suggest that the hemagglutinin domain of Kgp plays a role in degradation and that there is a critical threshold concentration for this activity . Taken together, these results provide evidence that the gingipains, especially Kgp, are involved in the degradation of the adherens junction of epithelial cells, which may be important in the invasion of periodontal connective tissue by P . gingivalis.

Infect Immun, 2002 May, 70(5), 2480 - 6
Immunization with the RgpA-Kgp proteinase-adhesin complexes of Porphyromonas gingivalis protects against periodontal bone loss in the rat periodontitis model; Rajapakse PS et al.; A major virulence factor of Porphyromonas gingivalis is the extracellular noncovalently associated complexes of Arg-X- and Lys-X-specific cysteine proteinases and adhesins designated the RgpA-Kgp complexes . In this study we investigated the ability of RgpA-Kgp as an immunogen to protect against P . gingivalis-induced periodontal bone loss in the rat . Specific-pathogen-free Sprague-Dawley rats were immunized with either formalin-killed whole P . gingivalis ATCC 33277 cells with incomplete Freund's adjuvant, RgpA-Kgp with incomplete Freund's adjuvant, or incomplete Freund's adjuvant alone . The animals were then challenged by oral inoculation with live P . gingivalis ATCC 33277 cells . Marked periodontal bone loss was observed in animals immunized with incomplete Freund's adjuvant alone; this bone loss was significantly (P < 0.05) greater than that detected in animals immunized with formalin-killed whole cells or RgpA-Kgp or in unchallenged animals . There was no significant difference in periodontal bone loss between animals immunized with formalin-killed whole cells and those immunized with RgpA-Kgp . The bone loss in these animals was also not significantly different from that in unchallenged animals . DNA probe analysis of subgingival plaque samples showed that 100% of the animals immunized with incomplete Freund's adjuvant alone and challenged with P . gingivalis ATCC 33277 were positive for the bacterium . However, P . gingivalis ATCC 33277 could not be detected in subgingival plaque samples from animals immunized with formalin-killed whole cells or with RgpA-Kgp . Immunization with formalin-killed whole cells or RgpA-Kgp induced a high-titer serum immunoglobulin G2a response . Western blot analysis of RgpA-Kgp using pooled protective antisera taken from rats immunized with RgpA-Kgp revealed immunodominant bands at 44, 39, and 27 kDa . In conclusion, immunization with RgpA-Kgp restricted colonization by P . gingivalis and periodontal bone loss in the rat.

Eur J Biochem, 2002 Apr, 269(7), 1984 - 92
A point mutation in the ATP synthase of Rhodobacter capsulatus results in differential contributions of Delta(pH) and Delta(phi) in driving the ATP synthesis reaction; Turina P et al.; The interface between the c-subunit oligomer and the a subunit in the F0 sector of the ATP synthase is believed to form the core of the rotating motor powered by the protonic flow . Besides the essential cAsp61 and aArg210 residues (Escherichia coli numbering), a few other residues at this interface, although nonessential, show a high degree of conservation, among these aGlu219 . The homologous residue aGlu210 in the ATP synthase of the photosynthetic bacterium Rhodobacter capsulatus has been substituted by a lysine . Inner membranes prepared from the mutant strain showed approximately half of the ATP synthesis activity when driven both by light and by acid-base transitions . As estimated with the ACMA assay, proton pumping rates in the inner membranes were also reduced to a similar extent in the mutant . The most striking impairment of ATP synthesis in the mutant, a decrease as low as 12 times as compared to the wild-type, was observed in the absence of a transmembrane electrical membrane potential (Delta(phi)) at low transmembrane pH difference (Delta(pH)) . Therefore, the mutation seems to affect both the mechanism responsible for coupling F1 with proton translocation by F0, and the mechanism determining the relative contribution of Delta(pH) and Delta(phi) in driving ATP synthesis.

Eur J Biochem, 2002 Apr, 269(7), 1926 - 31
Characterization of isocitrate dehydrogenase from the green sulfur bacterium Chlorobium limicola . A carbon dioxide-fixing enzyme in the reductive tricarboxylic acid cycle; Kanao T et al.; Isocitrate dehydrogenase (IDH) catalyzes the reversible conversion between isocitrate and 2-oxoglutarate accompanied by decarboxylation/carboxylation and oxidoreduction of NAD(P)+ cofactor . While this enzyme has been well studied as a catabolic enzyme in the tricarboxylic acid (TCA) cycle, here we have characterized NADP-dependent IDH from Chlorobium limicola, a green sulfur bacterium that fixes CO2 through the reductive tricarboxylic acid (RTCA) cycle, focusing on the CO2-fixation ability of the enzyme . The gene encoding Cl-IDH consisted of 2226 bp, corresponding to a polypeptide of 742 amino acid residues . The primary structure and the size of the recombinant protein indicated that Cl-IDH was a monomeric enzyme of 80 kDa distinct from the dimeric NADP-dependent IDHs predominantly found in bacteria or eukaryotic mitochondria . Apparent Michaelis constants for isocitrate (45 +/- 13 microm) and NADP+ (27 +/- 10 microm) were much smaller than those for 2-oxoglutarate (1.1 +/- 0.5 mm) and CO2 (1.3 +/- 0.3 mm) . No significant differences in kinetic properties were observed between Cl-IDH and the dimeric, NADP-dependent IDH from Saccharomyces cerevisiae (Sc-IDH) at the optimum pH of each enzyme . However, in contrast to the 20% activity of Sc-IDH toward carboxylation as compared with that toward decarboxylation at pH 7.0, the activities of Cl-IDH for both directions were almost equivalent at this pH, suggesting a more favorable property of Cl-IDH than Sc-IDH as a CO2-fixation enzyme under physiological pH . Furthermore, we found that among various intermediates, oxaloacetate was a competitive inhibitor (K(i) = 0.35 +/- 0.04 mm) for 2-oxoglutarate in the carboxylation reaction by Cl-IDH, a feature not found in Sc-IDH.

Eur J Biochem, 2002 Apr, 269(7), 1877 - 85
Role of the N- and C-terminal regions of the PufX protein in the structural organization of the photosynthetic core complex of Rhodobacter sphaeroides; Francia F et al.; The core complex of Rhodobacter sphaeroides is formed by the association of the light-harvesting antenna 1 (LH1) and the reaction center (RC) . The PufX protein is essential for photosynthetic growth; it is located within the core in a 1 : 1 stoichiometry with the RC . PufX is required for a fast ubiquinol exchange between the Q(B) site of the RC and the Qo site of the cytochrome bc1 complex . In vivo the LH1-PufX-RC complex is assembled in a dimeric form, where PufX is involved as a structural organizer . We have modified the PufX protein at the N and the C-terminus with progressive deletions . The nine mutants obtained have been characterized for their ability for photosynthetic growth, the insertion of PufX in the core LH1-RC complex, the stability of the dimers and the kinetics of flash-induced reduction of cytochrome b561 of the cytochrome bc1 complex . Deletion of 18 residues at the N-terminus destabilizes the dimer in vitro without preventing photosynthetic growth . The dimer (or a stable dimer) does not seem to be a necessary requisite for the photosynthetic phenotype . Partial C-terminal deletions impede the insertion of PufX, while the complete absence of the C-terminus leads to the insertion of a PufX protein composed of only its first 53 residues and does not affect the photosynthetic growth of the bacterium . Overall, the results point to a complex role of the N and C domains in the structural organization of the core complex; the N-terminus is suggested to be responsible mainly for dimerization, while the C-terminus is thought to be involved mainly in PufX assembly.

J Bacteriol, 2002 May, 184(9), 2529 - 32
Identification of an akinete marker gene in Anabaena variabilis; Zhou R et al.; Cyanobacteria that form akinetes as well as heterocysts present a rare opportunity to investigate the relationships between alternative differentiation processes and pattern formation processes in a single bacterium . Because no akinete marker gene has been identified, akinete formation has been little studied genetically . We report the first identification of an akinete marker gene.

Mikrobiol Z, 2001 Nov-Dec, 63(6), 25 - 31
{Effect of bacterial physiological state and heavy metals on the hyperactive reaction of tobacco leaf}; Gvozdiak RI et al.; It has been shown that the initiation and proceeding of hypersensitive reaction on the tobacco leaf under the bacterial effect are influenced by the infectious loading of the pathogen, physiological state of bacteria, their viability, temperature of the environment and heavy metal salts . It has been established that the hypersensitive reaction is most distinctly manifested when 1-3 days bacterium cultures are used . Their activity decreases essentially with age, and 10-14 days old cultures are not able to evoke the hypersensitive reaction . The bacterial cells being inactivated with temperature and UV-irradiation lose the ability to evoke the hypersensitive reaction . The reaction of hypersensitive is inhibited by salts of heavy metals in concentrations 0.1-1% and by high temperature of the environment (30 degrees C).

Lipids, 2002 Mar, 37(3), 305 - 8
Novel methoxylated FA from the Caribbean sponge Spheciospongia cuspidifera; Carballeira NM et al.; The delta6 monoenoic methoxylated FA (6Z)-2-methoxy-6-heptadecenoic acid and (6Z)-2-methoxy-6-octadecenoic acid were identified for the first time in nature in the phospholipids from the uncommon Caribbean sponge Spheciospongia cuspidifera . These findings expand the occurrence of A6 2-methoxylated FA to C17-C18 chain lengths and establish a new FA biosynthetic possibility for these marine organisms . The novel methoxylated FA also could have originated from the phospholipids of a bacterium in symbiosis with the sponge.

Lett Appl Microbiol, 2002, 34(4), 263 - 8
Development of a simple and sensitive fluorimetric method for isolation of coumaphos-hydrolysing bacteria; Harcourt RL et al.; AIMS: To develop a simple, rapid and sensitive fluorimetric assay to detect, isolate and characterize a soil bacterium capable of degrading the organophosphorus pesticide, coumaphos . METHODS AND RESULTS: A high throughput microtitre plate-based method was used to quantify coumaphos hydrolysis by the bacterium . The fluorescent hydrolysis product of coumaphos, chlorferon, was detected at levels as low as 10 nmol l(-1) . Incorporation of coumaphos into agar plates allowed the rapid detection of coumaphos-hydrolysing bacteria when exposed to an excitation wavelength of approximately 340 nm . The coumaphos-hydrolysing enzyme could be visualized when bacterial cell extracts were separated on SDS-PAGE, incubated with coumaphos and exposed to an excitation source as above . CONCLUSIONS: This method is 100-fold more sensitive than the currently used spectrophotometric method for coumaphos . SIGNIFICANCE AND IMPACT OF THE STUDY: This is a unique and versatile tool to screen for bacteria possessing phosphotriesterase activity.

Appl Microbiol Biotechnol, 2002 Mar, 58(3), 313 - 21 Epub 2002 Jan 12.
Beta-galactosidase from a cold-adapted bacterium: purification, characterization and application for lactose hydrolysis; Fernandes S et al.; The enzyme beta-galactosidase was purified from a cold-adapted organism isolated from Antarctica . The organism was identified as a psychotrophic Pseudoalteromonas sp . The enzyme was purified with high yields by a rapid purification scheme involving extraction in an aqueous two-phase system followed by hydrophobic interaction chromatography and ultrafiltration . The beta-galactosidase was optimally active at pH 9 and at 26 degrees C when assayed with o-nitrophenyl-beta-D-galactopyranoside as substrate for 2 min . The enzyme activity was highly sensitive to temperature above 30 degrees C and was undetectable at 40 degrees C . The cations Na+, K+, Mg2+ and Mn2+ activated the enzyme while Ca2+, Hg2+, Cu2+ and Zn2+ inhibited activity . The shelf life of the pure enzyme at 4 degrees C was significantly enhanced in the presence of 0.1% (w/v) polyethyleneimine . The pure beta-galactosidase was also evaluated for lactose hydrolysis . More than 50% lactose hydrolysis was achieved in 8 h in buffer at an enzyme concentration of 1 U/ml, and was increased to 70% in the presence of 0.1% (w/v) polyethyleneimine . The extent of lactose hydrolysis was 40-50% in milk . The enzyme could be immobilized to Sepharose via different chemistries with 60-70% retention of activity . The immobilized enzyme was more stable and its ability to hydrolyze lactose was similar to that of the soluble enzyme.

J Biol Chem, 2002 Jun 14, 277(24), 21397 - 404 Epub 2002 Apr 04.
Iron-sulfur cluster biosynthesis . Thermatoga maritima IscU is a structured iron-sulfur cluster assembly protein; Mansy SS et al.; Genetic evidence has indicated that Isc proteins play an important role in iron-sulfur cluster biogenesis . In particular, IscU is believed to serve as a scaffold for the assembly of a nascent iron-sulfur cluster that is subsequently delivered to target iron-sulfur apoproteins . We report the characterization of an IscU from Thermatoga maritima, an evolutionarily ancient hyperthermophilic bacterium . The stabilizing influence of a D40A substitution allowed characterization of the holoprotein . Mossbauer (delta = 0.29 +/- 0.03 mm/s, DeltaE(Q) = 0.58 +/- 0.03 mm/s), UV-visible absorption, and circular dichroism studies of the D40A protein show that T . maritima IscU coordinates a {2Fe-2S}2+ cluster . Thermal denaturation experiments demonstrate that T . maritima IscU is a thermally stable protein with a thermally unstable cluster . This is also the first IscU type domain that is demonstrated to possess a high degree of secondary and tertiary structure . CD spectra indicate 36.7% alpha-helix, 13.1% antiparallel beta-sheet, 11.3% parallel beta-sheet, 20.2% beta-turn, and 19.1% other at 20 degrees C, with negligible spectral change observed at 70 degrees C . Cluster coordination also has no effect on the secondary structure of the protein . The dispersion of signals in 1H-15N heteronuclear single quantum correlation NMR spectra of wild type and D40A IscU supports the presence of significant tertiary structure for the apoprotein, consistent with a scaffolding role, and is in marked contrast to other low molecular weight Fe-S proteins where cofactor coordination is found to be necessary for proper protein folding . Consistent with the observed sequence homology and proposed conservation of function for IscU-type proteins, we demonstrate T . maritima IscU-mediated reconstitution of human apoferredoxin.

J Mol Microbiol Biotechnol, 2002 May, 4(3), 243 - 8
Regulation of nitrogen fixation in the phototrophic purple bacterium Rhodobacter capsulatus; Masepohl B et al.; In R . capsulatus synthesis and activity of the molybdenum and the alternative nitrogenase is controlled at three levels by the environmental factors ammonium, molybdenum, light, and oxygen . At the first level, transcription of the nifA1, nifA2, and anfA genes--which encode the transcriptional activators of all other nif and anf genes, respectively--is controlled by the Ntr system in dependence on ammonium availability . Mutations in ginB (coding for the signal transduction protein PII) result in significant expression of nifA and anfA in the presence of ammonium . In contrast to GlnB, the PII-paralogue GlnK is not involved in the Ntr signal transduction mechanism . In addition to ammonium control, transcription of anfA is inhibited by traces of molybdenum via the molybdate-dependent repressor proteins MopA and MopB . At the second level of regulation, activity of NifA1, NifA2, and AnfA is inhibited by ammonium in an NtrC-independent manner . This post-translational ammonium control of NifA activity is partially released in the absence of GlnK, and completely abolished in a glnB/glnK double mutant . In contrast, AnfA activity is still inhibited by ammonium in the glnB/glnK mutant background . At the third level of regulation, both GlnB and GlnK as well as the (methyl)-ammonium transporter AmtB are involved in ammonium control of the DraT/DraG system, which mediates reversible ADP-ribosylation of both nitrogenase reductases (NifH and AnfH) in response to changes in ammonium availability or light intensity . Most remarkably, in a glnB/glnK double mutant ammonium control of the molybdenum (but not of the alternative) nitrogenase is completely relieved, leading to synthesis of active nitrogenase in the presence of high concentrations of ammonium.

Mol Microbiol, 2002 Feb, 43(3), 597 - 615
Mutations in FlbD that relieve the dependency on flagellum assembly alter the temporal and spatial pattern of developmental transcription in Caulobacter crescentus; Muir RE et al.; The transcription factor FlbD regulates the temporal and spatial transcription of flagellar genes in the bacterium Caulobacter crescentus . Activation of FlbD requires cell cycle progression and the assembly of an early (class II) flagellum structure . In this report, we identify 20 independent gain-of-function mutations in flbD that relieve regulation by flagellar assembly . One of these, flbD-1204, contained a mutation in the receiver domain (V17M) and another, flbD-1231, in the DNA binding domain (V451G) . Both of these mutations resulted in an aberrant pattern of cell cycle transcription . The presence of the FlbD-1204 allele also resulted in a loss of swarmer-pole-specific transcription . These results indicate that temporal and spatial transcription is influenced by the assembly of the nascent flagellar structure . The trans-acting positive and negative regulatory factor, FliX, couples flagellar assembly to the activation of FlbD and, as we show here, also influences temporal transcription . Furthermore, we show that FliX can suppress the activity of FlbD mutants that cannot be phosphorylated, and that FliX is required for FlbD stability, and vice versa . These results indicate that FliX may interact directly with FlbD to regulate its activity.

Curr Microbiol, 2002 May, 44(5), 363 - 7
Inhibition of biosynthesis and activity of nitrogenase in Azospirillum brasilense Sp7 under salinity stress; Tripathi AK et al.; Azospirillum brasilense is a microaerophilic, plant growth-promoting bacterium, whose nitrogenase activity has been shown to be sensitive to salinity stress . Growth of A . brasilense in semi-solid medium showed that diazotrophic growth in N-free medium was relatively less sensitive to high NaCl concentrations (200-400 mM) than that in presence of NH4+ . Increase in salinity stress to diazotrophic A . brasilense in the semi-solid medium led to the migration of the pellicle to deeper anaerobic zones . Assays of acetylene reduction and nifH- lacZ and nifA- lacZ fusions indicated that salinity stress inhibited nitrogenase biosynthesis more strongly than nitrogenase activity . Under salt stress, the amount of dinitrogenase reductase inactivated by ADP-ribosylation was strongly reduced, indicating that the dinitrogenase reductase ADP ribosyl transferase (DRAT) activity was also inhibited by increased NaCl concentrations . Movement of the pellicle to the anaerobic zone and inhibition of DRAT might be adaptive responses of A . brasilense to salinity stress under diazotrophic conditions . Supplementation of glycine betaine, which alleviates salt stress, partially reversed both responses.

Int J Cancer, 2002 Mar 20, 98(3), 446 - 9
Risk of gastric cancer among smokers infected with Helicobacter pylori; Brenner H et al.; Infection with the gastric bacterium Helicobacter pylori (in particular infection with CagA-positive strains) and smoking have been identified as risk factors for the development of gastric cancer . Both risk factors are typically acquired early in life and prevail over decades if not for life . We assessed the individual and joint impact of both risk factors on gastric cancer risk in a population-based case-control study from Germany including 71 patients with histologically verified gastric cancer and 363 patients with colorectal cancer who served as controls . Information on smoking and potential confounding factors was collected by standardized interviews . H . pylori infection was measured serologically by immunoglobulin G antibody titers against H . pylori . In addition, antibodies against the CagA antigen were determined by Western blot . Twenty-seven percent of cases compared with 15% of controls were smokers, and 43% of cases compared with 23% of controls were infected with CagA-positive H . pylori strains . After control for potential confounders, the relative risk of gastric cancer was 2.6 (95% CI 1.2-5.7) for nonsmoking subjects with CagA-positive H . pylori infections and 7.2 (95% CI 2.2-23.6) for smoking subjects with CagA-positive H . pylori infections compared with subjects without these risk factors . The corresponding relative risks for noncardia gastric cancer were 6.1 ( 95% CI 2.3-16.5) and 16.6 (95% CI 4.3-64.2) . We conclude that smoking subjects with CagA-positive H . pylori infections have a strongly increased risk of gastric cancer and may be an important group for targeting efforts of prevention and early detection .

Biotechnol Bioeng, 2002 May 5, 78(3), 296 - 312
Stoichiometric model for evaluating the metabolic capabilities of the facultative methylotroph Methylobacterium extorquens AM1, with application to reconstruction of C(3) and C(4) metabolism; Van Dien SJ et al.; A stoichiometric model of central metabolism was developed based on new information regarding metabolism in this bacterium to evaluate the steady-state growth capabilities of the serine cycle facultative methylotroph Methylobacterium extorquens AM1 during growth on methanol, succinate, and pyruvate . The model incorporates 20 reversible and 47 irreversible reactions, 65 intracellular metabolites, and experimentally-determined biomass composition . The flux space for this underdetermined system of equations was defined by finding the elementary modes, and constraints based on experimental observations were applied to determine which of these elementary modes give a reasonable description of the flux distribution for each growth substrate . The predicted biomass yield, on a carbon atom basis, is 49.8%, which agrees well with the range of published experimental yield measurements (37-50%) . The model predicts the cell to be limited by reduced pyridine nucleotide availability during methylotrophic growth, but energy-limited when growing on multicarbon substrates . Mutation and phenotypic analysis was used to explore a previously unknown region of the metabolic map and to confirm the stoichiometry of the pathways in this region used in the metabolic model . Based on genome sequence data and simulation results, three enzymes involved in C(3)-C(4) interconversion pathways were predicted to be mutually redundant: malic enzyme, phosphoenolpyruvate carboxykinase, and phosphoenolpyruvate synthase . Insertion mutations in the genes predicted to encode these enzymes were made and these mutants were capable of growing on all substrates tested, confirming the redundancy of these pathways . Likewise, pathway analysis suggests that the TCA cycle enzymes citrate synthase and succinate dehydrogenase are essential for all growth substrates . In keeping with these predictions, null mutants could not be obtained in these genes . Finally, a similar model was developed for the ribulose monophosphate pathway obligate methylotroph Methylobacillus flagellatum KT to compare the efficiency of carbon utilization in the two types of methylotrophic carbon utilization pathways . The predicted yield for this organism on methanol is 65.9% .

Heredity, 2002 Mar, 88(3), 166 - 71
Wolbachia infection associated with all-female broods in Hypolimnas bolina (Lepidoptera: Nymphalidae): evidence for horizontal transmission of a butterfly male killer; Dyson EA et al.; Inherited bacteria that kill male hosts during embryogenesis infect a wide range of insect species . In order to ascertain if there are patterns to host infection, with particular male killing bacteria specialising on particular taxa, we investigated the male killing trait in the butterfly Hypolimnas bolina . All-female broods were first reported in this species in the 1920s . Investigation of this system in the Fiji Islands revealed the causal agent of sex ratio distortion in H . bolina to be a male killing Wolbachia bacterium . This bacterium is identical in wsp and ftsZ sequence to a male killer in the butterfly Acraea encedon in Tanzania, suggesting it has moved between host species, yet retained its phenotype . The prevalence of the Wolbachia was calculated for three different island groups of Fiji, and found to vary significantly across the country . Antibiotics failed to cure either the male killing trait or the Wolbachia infection . The implications of these results are discussed.

Platelets, 2002 Feb, 13(1), 21 - 30
Aggregation of human platelets by gingipain-R from Porphyromonas gingivalis cells and membrane vesicles; Pham K et al.; The hypothesis that there is an association between periodontitis and cardiovascular disease suggests new lines of research on the mechanism whereby oral bacteria might exert systemic effects . This study was conducted to ascertain and quantitate the effect of Porphyromonas gingivalis on human platelets in vitro . A second related objective was to purify and identify the aggregating vector . Aggregation was measured by platelet turbidometry and gingipain-R was purified from P . gingivalis membrane vesicles by Sepharose 2B and hydroxyapatite chromatography . The in vitro aggregation of platelets requires that at least 1.0 x 10(4) cells be stirred with 1.35 x 10(8) platelets . The specific activity is substantially increased in the membrane vesicles that are shed by this bacterium . Aggregability was due to gingipain-R activity, a potent cysteine protease that was found to be highly concentrated in the membrane vesicle fraction . The enzyme was purified 18-fold in high yield from the membrane vesicles, and consists of two noncovalently linked proteins that migrate at 49 and 44 kDa on SDS-PAGE . Aggregation of platelets by gingipain-R was shown to be dose-dependent, and inhibited by leupeptin and arginine, but not by anti-thrombin III . This is the first report enumerating the specific number of cells and lowest concentration of membrane vesicles necessary to evoke a full human platelet response, and the first report to assign this activity to gingipain-R.

Appl Environ Microbiol, 2002 Apr, 68(4), 2031 - 5
Rapid assessment of the physiological status of the polychlorinated biphenyl degrader Comamonas testosteroni TK102 by flow cytometry; Hiraoka Y et al.; The viability of the polychlorinated biphenyl-degrading bacterium Comamonas testosteroni TK102 was assessed by flow cytometry (FCM) with the fluorogenic ester Calcein-AM (CAM) and the nucleic acid dye propidium iodide (PI) . CAM stained live cells, whereas PI stained dead cells . When double staining with CAM and PI was performed, three physiological states, i.e., live (calcein positive, PI negative), dead (calcein negative, PI positive), and permeabilized (calcein positive, PI positive), were detected . To evaluate the reliability of this double-staining method, suspensions of live and dead cells were mixed in various proportions and analyzed by FCM . The proportion of dead cells measured by FCM directly correlated with the proportion of dead cells in the sample (y = 0.9872 x + 0.18; R(2) = 0.9971) . In addition, the proportion of live cells measured by FCM inversely correlated with the proportion of dead cells in the sample (y = -0.9776 x + 98.36; R(2) = 0.9962) . The proportion of permeabilized cells was consistently less than 2% . These results indicate that FCM in combination with CAM and PI staining is rapid (<or=1 h) and distinguishes correctly among live, dead, and permeabilized cells.

Proc R Soc Lond B Biol Sci, 2002 Mar 22, 269(1491), 623 - 9
Deleterious Wolbachia in the ant Formica truncorum; Wenseleers T et al.; Wolbachia is a maternally inherited bacterium that may manipulate the reproduction of its arthropod hosts . In insects, it is known to lead to inviable matings, cause asexual reproduction or kill male offspring, all to its own benefit, but to the detriment of its host . In social Hymenoptera, Wolbachia occurs widely, but little is known about its fitness effects . We report on a Wolbachia infection in the wood ant Formica truncorum, and evaluate whether it influences reproductive patterns . All 33 colonies of the study population were infected, suggesting that Wolbachia infection is at, or close to, fixation . Interestingly, in colonies with fewer infected workers, significantly more sexuals are produced, indicating that Wolbachia has deleterious effects in this species . In addition, adult workers are shown to have significantly lower infection rates (45%) than worker pupae (87%) or virgin queens (94%), suggesting that workers lose their infection over life . Clearance of Wolbachia infection has, to our knowledge, never been shown in any other natural system, but we argue that it may, in this case, represent an adaptive strategy to reduce colony load . The cause of fixation requires further study, but our data strongly suggest that Wolbachia has no influence on the sex ratio in this species.

J Bacteriol, 2002 Apr, 184(8), 2215 - 24
Role for vitamin B(12) in light induction of gene expression in the bacterium Myxococcus xanthus; Cervantes M et al.; A light-inducible promoter (P(B)) drives the carB operon (carotenoid genes) of the bacterium Myxococcus xanthus . A gene encoding a regulator of carotenoid biosynthesis was identified by studying mutant strains carrying a transcriptional fusion to P(B) and deletions in three candidate genes . Our results prove that the identified gene, named carA, codes for a repressor of the P(B) promoter in the dark . They also show that the carA gene product does not participate in the light activation of two other promoters connected with carotenoid synthesis or its regulation in M . xanthus . CarA is a novel protein consisting of a DNA-binding domain of the family of MerR helix-turn-helix transcriptional regulators, directly joined to a cobalamin-binding domain . In support of this, we report here that the presence of vitamin B(12) or some other cobalamin derivatives is absolutely required for activation of the P(B) promoter by light.

J Bacteriol, 2002 Apr, 184(8), 2081 - 7
Anaerobic respiration using Fe(3+), S(0), and H(2) in the chemolithoautotrophic bacterium Acidithiobacillus ferrooxidans; Ohmura N et al.; The chemolithoautotrophic bacterium Acidithiobacillus ferrooxidans has been known as an aerobe that respires on iron and sulfur . Here we show that the bacterium could chemolithoautotrophically grow not only on H(2)/O(2) under aerobic conditions but also on H(2)/Fe(3+), H(2)/S(0), or S(0)/Fe(3+) under anaerobic conditions . Anaerobic respiration using Fe(3+) or S(0) as an electron acceptor and H(2) or S(0) as an electron donor serves as a primary energy source of the bacterium . Anaerobic respiration based on reduction of Fe(3+) induced the bacterium to synthesize significant amounts of a c-type cytochrome that was purified as an acid-stable and soluble 28-kDa monomer . The purified cytochrome in the oxidized form was reduced in the presence of the crude extract, and the reduced cytochrome was reoxidized by Fe(3+) . Respiration based on reduction of Fe(3+) coupled to oxidation of a c-type cytochrome may be involved in the primary mechanism of energy production in the bacterium on anaerobic iron respiration.

Biochemistry, 2002 Apr 2, 41(13), 4358 - 70
Subcellular localization of chlorosome proteins in Chlorobium tepidum and characterization of three new chlorosome proteins: CsmF, CsmH, and CsmX; Vassilieva EV et al.; Chlorosomes are unique light-harvesting structures found in two families of photosynthetic bacteria . In this study, three chlorosome proteins (CsmF, CsmH, and CsmX) of the green sulfur bacterium Chlorobium tepidum were characterized by cloning and sequencing the genes which encode them, by overproducing the respective proteins in Escherichia coli, and by raising polyclonal antisera to the purified proteins . Three other proteins (AtpF, CT1970, and CT2144) which were identified in chlorosome fractions have similarly been characterized . The antisera were used to establish the distribution of each protein in various cellular fractions . Ten chlorosome proteins (CsmA, CsmB, CsmC, CsmD, CsmE, CsmF, CsmH, CsmI, CsmJ, and CsmX) copurified in a constant proportion together with bacteriochlorophyll c, and none of these 10 proteins was found in substantial amounts in other subcellular fractions . An antiserum to CsmH was highly effective in agglutinating chlorosomes, and antisera to CsmI, CsmJ, CsmX, and CsmA also immunoprecipitated chlorosomes to varying extents . However, an antiserum to CsmF did not agglutinate chlorosomes . The sequences of chlorosome proteins generally are not significantly similar to the sequences of other proteins in the databases . However, the N-terminal domains of three chlorosome proteins, CsmI, CsmJ, and CsmX, are related to adrenodoxin-type ferredoxins that ligate {2Fe-2S} clusters {Vassilieva, E . V., Antonkine, M . L., Zybailov, B . L., Yang, F., Jakobs, C . U., Golbeck, J . H., and Bryant, D . A . (2001) Biochemistry 40, 464-473} . The sequences of the C-terminal domains of these three proteins appear to be distantly related to CsmA and CsmE . The remaining chlorosome proteins can be divided into two additional structural families, CsmB/F and CsmC/D . CsmH is recovered in water-soluble form after overproduction in E . coli . Interestingly, this protein contains an N-terminal domain that is similar to CsmB/D, while its C-terminal domain is related to CsmC/D . The sequence relationships indicate that, although the protein composition of Chlorobium-type chlorosomes is superficially more complex than that of the chlorosomes of Chloroflexus aurantiacus, this heterogeneity is mostly produced by gene duplication and divergence among a small number of protein types.

Tuberculosis (Edinb), 2002, 82(1), 1 - 6
A variable number of tandem repeats result in polymorphic alpha -isopropylmalate synthase in Mycobacterium tuberculosis; Chanchaem W et al.; A locus of variable number of the tandem repeat, VNTR4155, resides in the putative leuA gene, encoding for alpha -isopropylmalate synthase (alpha -IPMS) of Mycobacterium tuberculosis, a repeat that is unique to the bacterium . The objective was to determine whether the polymorphic VNTR4155 was translated and resulted in a polymorphic protein . The putative leuA gene of the M . tuberculosis H37Rv strain was cloned by PCR and expressed in a His-tagged form in Escherichia coli . The enzymatic properties of the purified protein were studied . The protein was used as an antigen to immunize rabbits . Soluble proteins of several strains of M . tuberculosis were examined by Western blot analysis . The polymorphism of VNTR4155 was due to the presence of different copy number of the 57-bp tandem repeat . The putative alpha -IPMS of various strains of M . tuberculosis had different sizes, varying directly with the length of their VNTR4155 .

Biotechniques, 2002 Mar, 32(3), 551 - 2, 554, 556 passim
Bacterium-based heavy metal biosorbents: enhanced uptake of cadmium by E . coli expressing a metallothionein fused to beta-galactosidase; Yoshida N et al.; We investigated the potential utility of a recombinant E . coli that expresses the human metallothionein II gene as a fusion protein with beta-galactosidase as a heavy metal biosorbent . E . coli cells expressing the metallothionein fusion demonstrated enhanced binding of Cd2+ compared to cells that lack the metallothionein . It was shown that the metallothionein fusion was capable of efficiently removing Cd2+ from solutions . Approximately 40% of the Cd2+ accumulated by the recombinant cells free in suspension was associated with the outer cell membrane, and 60% of that was present in the cytoplasm.

Int J Clin Pharmacol Ther, 2002 Mar, 40(3), 126 - 9
Treatment behavior of doctors regarding Helicobacter pylori infections; Perez E et al.; OBJECTIVE: Since the detection of the gastric acid resistant bacterium Helicobacter pylori in the year 1982 there has been a fundamental change regarding the therapy of ulcers . According to expert opinion these infections should be treated and eradicated, whereby the so-called triple-therapies are considered to be the most effective ones . Whether such recommendations to eradicate Helicobacter pylori can be put to use in daily practice is an important question that is frequently asked . METHODS: All analyses described in the study here were done using mediplus, a longitudinal patient database with anonymous access to a representative and valid panel of physicians and patients within Germany . A total of more than 1,000 medical practices and over 75 million prescriptions can be analyzed in a cross- and/or longitudinal section . The longest time period per patient is more than 10 years starting in 1989 with monthly updates . RESULTS: With regard to existing recommendations, doctors overestimate their own compliance with the recommendations because only a fraction of traceable Helicobacter pylori infections are actually eradicated . Within the period of observation the therapy behavior has changed significantly in favor of the triple-therapies, but there are relevant differences between practitioners and internal specialists . DISCUSSION: Only a fraction of traceable Helicobacter pylori infections are adequately treated by doctors . The results show a very alarming situation due to the gap between "state of the art" and what is being achieved and carried out in daily practice . The potential for possible cost reductions and saving on resources is probably high . CONCLUSION: The results lead to the conclusion that in the treatment of Helicobacter pylori infections there is a potential for cost saving which is unused from the pharmacoeconomical point of view.

Protein Sci, 2002 Apr, 11(4), 903 - 11
Alkaline phosphatase from the hyperthermophilic bacterium T . maritima requires cobalt for activity; Wojciechowski CL et al.; The hyperthermophilic bacterium Thermotoga maritima encodes a gene sharing sequence similarities with several known genes for alkaline phosphatase (AP) . The putative gene was isolated and the corresponding protein expressed in Escherichia coli, with and without a predicted signal sequence . The recombinant protein showed phosphatase activity toward the substrate p-nitrophenyl-phosphate with a k(cat) of 16 s(-1) and a K(m) of 175 microM at a pH optimum of 8.0 when assayed at 25 degrees C . T . maritima phosphatase activity increased at high temperatures, reaching a maximum k(cat) of 100 s(-1), with a K(m) of 93 microM at 65 degrees C . Activity was stable at 65 degrees C for >24 h and at 90 degrees C for 5 h . Phosphatase activity was dependent on divalent metal ions, specifically Co(II) and Mg(II) . Circular dichroism spectra showed that the enzyme gains secondary structure on addition of these metals . Zinc, the most common divalent metal ion required for activity in known APs, was shown to inhibit the T . maritima phosphatase enzyme at concentrations above 0.3 moles Zn: 1 mole monomer . All activity was abolished in the presence of 0.1 mM EDTA . The T . maritima AP primary sequence is 28% identical when compared with E . coli AP . Based on a structural model, the active sites are superimposable except for two residues near the E . coli AP Mg binding site, D153 and K328 (E . coli numbering) corresponding to histidine and tryptophan in T . maritima AP, respectively . Sucrose-density gradient sedimentation experiments showed that the protein exists in several quaternary forms predominated by an octamer.

J Antimicrob Chemother, 2002 Apr, 49(4), 601 - 9
Helicobacter pylori susceptibility testing by disc diffusion; McNulty C et al.; The bacterium Helicobacter pylori is found in c . 40% of the population and is responsible for the development of duodenal disease . Triple treatment with a proton-pump inhibitor or bismuth salt plus two antibiotics is now commonplace in all patients diagnosed . As antibiotic resistance reduces treatment efficacy, it is time to consider routine susceptibility testing to guide individual patient treatment and surveillance of antibiotic resistance . There are no published nationally agreed standards for disc diffusion testing of H . pylori . After reviewing the literature, we recommend the following method for disc diffusion tests . A suspension of cultures < or = 4 days old equivalent to McFarland Standard no . 4 (10(8) cfu/mL) should be used on Mueller-Hinton or Columbia agar base with 5-10% blood, using a metronidazole disc strength of 5 Ig and a clarithromycin disc strength of 2 microg . Anaerobic pre-incubation of plates is unnecessary . A H . pylori control susceptible to metronidazole (e.g . NCTC 12822) should be used . Zone sizes with the Mueller-Hinton agar base for metronidazole testing are <16 mm resistant, 16-21 mm intermediate and >21 mm susceptible . We suggest that isolates in the intermediate zone should be re-tested by Etest . Zone sizes with the Columbia agar base for metronidazole testing are <10 mm resistant and > or = 10 mm susceptible . Co-infection with two strains, which may be a mixture of isolates susceptible and resistant to metronidazole leading to conflicting susceptibility results, occurs in 5-10% of patients . Zone sizes with Mueller-Hinton agar and Columbia blood agar for clarithromycin testing are resistant no zone and susceptible any zone.

Nature . 2002 Mar 21;416(6878):279.
Biochemistry: biosynthesis of an organofluorine molecule; O'Hagan D et al.; Although fluorine in the form of fluoride minerals is the most abundant halogen in the Earth's crust, only 12 naturally occurring organofluorine compounds have so far been found, and how these are biosynthesized remains a mystery . Here we describe an enzymatic reaction that occurs in the bacterium Streptomyces cattleya and which catalyses the conversion of fluoride ion and S-adenosylmethionine (SAM) to 5'-fluoro-5'-deoxyfluoroadenosine (5'-FDA) . To our knowledge, this is the first fluorinase enzyme to be identified, a discovery that opens up a new biotechnological opportunity for the preparation of organofluorine compounds.

J Immunol, 2002 Apr 1, 168(7), 3451 - 7
The NKp46 receptor contributes to NK cell lysis of mononuclear phagocytes infected with an intracellular bacterium; Vankayalapati R et al.; We used human tuberculosis as a model to investigate the role of NK cytotoxic mechanisms in the immune response to intracellular infection . Freshly isolated NK cells and NK cell lines from healthy donors lysed Mycobacterium tuberculosis-infected monocytes to a greater extent than uninfected monocytes . Lysis of infected monocytes was associated with increased expression of mRNA for the NKp46 receptor, but not the NKp44 receptor . Antisera to NKp46 markedly inhibited lysis of infected monocytes . NK cell-mediated lysis was not due to reduced expression of MHC class I molecules on the surface of infected monocytes or to enhanced production of IL-18 or IFN-gamma . NK cell lytic activity against M . tuberculosis-infected monocytes and NKp46 mRNA expression were reduced in tuberculosis patients with ineffective immunity to M . tuberculosis compared with findings in healthy donors . These observations suggest that 1) the NKp46 receptor participates in NK cell-mediated lysis of cells infected with an intracellular pathogen, and 2) the reduced functional capacity of NK cells is associated with severe manifestations of infectious disease.

Proc Natl Acad Sci U S A, 2002 Mar 19, 99(6), 3938 - 43
Chronic intracellular infection of alfalfa nodules by Sinorhizobium meliloti requires correct lipopolysaccharide core; Campbell GR et al.; Our analyses of lipopolysaccharide mutants of Sinorhizobium meliloti offer insights into how this bacterium establishes the chronic intracellular infection of plant cells that is necessary for its nitrogen-fixing symbiosis with alfalfa . Derivatives of S . meliloti strain Rm1021 carrying an lpsB mutation are capable of colonizing curled root hairs and forming infection threads in alfalfa in a manner similar to a wild-type strain . However, developmental abnormalities occur in the bacterium and the plant at the stage when the bacteria invade the plant nodule cells . Loss-of-function lpsB mutations, which eliminate a protein of the glycosyltransferase I family, cause striking changes in the carbohydrate core of the lipopolysaccharide, including the absence of uronic acids and a 40-fold relative increase in xylose . We also found that lpsB mutants were sensitive to the cationic peptides melittin, polymyxin B, and poly-l-lysine, in a manner that paralleled that of Brucella abortus lipopolysaccharide mutants . Sensitivity to components of the plant's innate immune system may be part of the reason that this mutant is unable to properly sustain a chronic infection within the cells of its host-plant alfalfa.

Lancet Oncol, 2002 Feb, 3(2), 97 - 104
Gastric MALT lymphoma: from aetiology to treatment; Du MQ et al.; The development of gastric mucosa-associated lymphoid tissue (MALT) lymphoma is dependent on Helicobacter pylori infection . Bacterial colonisation of the gastric mucosa triggers lymphoid infiltration and the formation of acquired MALT . The bacterial infection induces and sustains an actively proliferating B-cell population through direct (autoantigen) and indirect (intratumoral T cells specific for H . pylori) immunological stimulation . Moreover, the bacterial infection provokes a neutrophilic response, which causes the release of oxygen free radicals . These reactive species may promote the acquisition of genetic abnormalities and malignant transformation of reactive B cells . A transformed clone carrying the translocation t(1;18)(q21;q21) forms a MALT lymphoma, the growth of which is independent of H . pylori and will not respond to bacterial eradication . Malignant clones without t(11;18)(q21;q21), but with other genetic abnormalities, such as trisomy 3 or microsatellite instability, depend critically on immune stimulation mediated by H . pylori for their clonal expansion . In the early stages, the tumour can be successfully treated by eradication of the bacterium, whereas at later stages the tumour may escape its growth dependency through acquisition of additional genetic abnormalities such as t(1;14)(p22;q32) and t(1;2)(p22,p12) involving the BCL-10 gene . Finally, further genetic abnormalities, such as inactivation of the tumour suppressor genes, p53 and p16, can lead to high-grade transformation . Detection of these abnormalities may help with the clinical management of patients with gastric MALT lymphoma.

Expert Rev Mol Diagn, 2001 Sep, 1(3), 299 - 309
Molecular techniques in Whipple's disease; Fenollar F et al.; Whipple's disease is a systemic infection, caused by the bacterium Tropheryma whipplei, with protean clinical manifestations characterized by fever, weight loss, diarrhea, polyarthritis, skin hyperpigmentation and adenopathy . For a long time, due to the inability to culture the causative organism, diagnosis was based on histologic examination of infected tissues, usually duodenal biopsies, which revealed diastase-resistant periodic acid-Schiff-positive staining . Now, PCR of various tissues or fluid is emerging as a way to diagnose Whipple's disease . However, the presence of T . whipplei DNA in saliva, gastric juice or duodenal biopsies of healthy individuals has led to questions regarding the specificity of the molecular techniques involved . After a series of failures, stable culture was achieved in 2000 . Subsequently, the generation of rabbit polyclonal antibodies has led to the detection of the bacterium in tissues by immunohistology . However, culture and immunohistology are very recent techniques and are not yet widely used . Propagation of the bacterium will lead to extensive molecular knowledge of T . whipplei, which will help in the diagnosis and understanding of the epidemiology and pathogenicity of Whipple's disease.

Expert Rev Mol Diagn, 2001 Sep, 1(3), 290 - 8
Detection of Helicobacter pylori virulence-associated genes; van Doorn LJ; Helicobacter pylori is an important human pathogen and persistent colonization of the human gastric mucosa can cause severe gastrointestinal diseases . The bacterium should not be considered as a uniform organism, but as a population of closely related and yet genetically diverse bacteria . Several genes of H . pylori (such as vacA and cagA) have been identified as being virulence-associated and may have important clinical and epidemiological implications . Assessment of virulence-associated genes of H . pylori should be included in clinical and epidemiological studies as well as therapeutic trials, in order to stratify between patient groups, harboring H . pylori strains with particular virulence genotypes . Molecular determination of antibiotic resistance will be especially useful for treatment studies . Together with our increasing knowledge about the human genome, typing of H . pylori will facilitate the management of gastroenterological pathologies.

Genetika, 2002 Feb, 38(2), 182 - 9
{The use of fragments of the 85- and 120-MDa plasmids of Azospirillum brasilense Sp245 to study the plasmid rearrangement in this bacterium and to search for homologous sequences in plasmids of Azospirillum brasilense Sp7}; Katsy EI et al.; A spontaneous loss of the 85- (p85) and 120-MDa (p120) replicons and simultaneous generation of a plasmid of more than 300 MDa were associated with defects in synthesis of O-specific and Calcofluor-binding polysaccharides and had no effect on flagellation and motility of the Azospirillum brasilense Sp245.5 mutant . The plasmid rearrangement was studied by hybridization of DNAs from the wild-type Sp245 strain and the Sp245.5 mutant with p85 and p120 fragments that contained loci involved in formation of the polar (fla) and lateral (laf) flagella, synthesis of O-specific and Calcofluor-binding polysaccharides (lps/cal), swimming (mot), and swarming (swa) of bacteria . Hybridization with the p120 fragments revealed incorporation of the intact fla/swa loci and the altered lps/cal loci into a new megaplasmid . Two EcoRI fragments homologous to the fla/laf/mot/swa loci of p85 were found in A . brasilense Sp245 DNA, whereas only one copy was preserved in the Sp245.5 mutant . Hybridization of the p120 and p85 fragments of Sp245 to the A . brasilense Sp7 DNA for the first time revealed regions of substantial homology to these fragments in the 90- and 115-MDa Sp7 plasmids, respectively.

Lab Invest, 2002 Mar, 82(3), 303 - 11
Kinetics of CD11b/CD18 up-regulation during infection with the agent of human granulocytic ehrlichiosis in mice; Borjesson DL et al.; The agent of human granulocytic ehrlichiosis (aoHGE) is a tick-borne, obligate intracellular, granulocytotropic bacterium able to infect numerous host species . Given its unique niche and the leukopenia often noted with infection, we investigated the effect of acute aoHGE infection on neutrophil activation by evaluating surface expression of the beta 2 integrin CD11b/CD18 in a mouse model using FACS analysis . Infection resulted in neutrophil activation with up-regulation of CD11b/CD18 in multiple strains of mice, however, hematologic analysis showed no apparent role for CD11b/CD18 in mediating peripheral leukopenia . Because IFN-gamma is an important cytokine during granulocytic ehrlichiosis and is known to activate leukocytes, we investigated the potential role of IFN-gamma in CD11b/CD18 up-regulation . Neutrophils from IFN-gamma knock-out mice became activated during aoHGE infection, however, the kinetics of activation differed from wild-type mice . In addition, activation correlated directly with the presence of bacteria because neutrophils with large intracytoplasmic morula also expressed higher levels of CD11b/CD18 . CD11b/CD18 seemed to be critical to early bacterial clearance and killing in vivo because infection of mice with targeted genetic disruption of CD11b/CD18 resulted in an initial increase in bacterial burden compared with wild-type mice . Similarly, in vitro culture of neutrophils from infected CD11b/CD18 knock-out mice resulted in a marked increase in bacterial proliferation compared with congenic controls . The data support crucial roles of CD11b/CD18 and IFN-gamma-mediated cell activation as mechanisms that limit bacterial replication.

Infect Immun, 2002 Apr, 70(4), 1936 - 48
In vivo clearance of an intracellular bacterium, Francisella tularensis LVS, is dependent on the p40 subunit of interleukin-12 (IL-12) but not on IL-12 p70; Elkins KL et al.; To determine the role of interleukin-12 (IL-12) in primary and secondary immunity to a model intracellular bacterium, we have comprehensively evaluated infection with Francisella tularensis LVS in three murine models of IL-12 deficiency . Mice lacking the p40 protein of IL-12 (p40 knockout {KO} mice) and mice treated in vivo with neutralizing anti-IL-12 antibodies survived large doses of primary and secondary LVS infection but never cleared bacteria and exhibited a chronic infection . In dramatic contrast, mice lacking the p35 protein (p35 KO mice) of heterodimeric IL-12 readily survived large doses of primary sublethal LVS infection as well as maximal secondary lethal challenge, with only a slight delay in clearance of bacteria . LVS-immune wild-type (WT) lymphocytes produced large amounts of gamma interferon (IFN-gamma), but p35 KO and p40 KO lymphocytes produced much less; nonetheless, similar amounts of NO were found in all cultures containing immune lymphocytes, and all immune lymphocytes were equally capable of controlling intracellular growth of LVS in vitro . Purified CD4(+) and CD8(+) T cells from both WT and p40 KO mice controlled intracellular growth, even though T cells from WT mice produced much more IFN-gamma than those from p40 KO mice, and p40 KO T cells did not adopt a Th2 phenotype . Thus, while IL-12 p70 stimulation of IFN-gamma production may be important for bacteriostasis, IL-12 p70 is not necessary for appropriate development of LVS-immune T cells that are capable of controlling intracellular bacterial growth and for clearance of primary or secondary LVS infection . Instead, an additional mechanism dependent on the IL-12 p40 protein, either alone or in another complex such as the newly discovered heterodimer IL-23, appears to be responsible for actual clearance of this intracellular bacterium.

Curr Allergy Asthma Rep, 2001 Nov, 1(6), 566 - 71
The diagnosis and treatment of Whipple's disease; Marth T; Whipple's disease is a rare, chronic, and systemic infectious disease caused by the ubiquitously occurring bacterium Tropheryma whippelii . For two reasons, the disease represents a good example for documenting the input of modern molecular-based techniques into pathogenetic, diagnostic, and therapeutic concepts in clinical medicine . First, the unidentified and uncultivable causative organism has been characterized by novel molecular-genetic techniques . Second, in contrast to other chronic inflammatory disorders, clinical manifestations of T . whippelii infection seem to be based on reduced T-cell helper type 1 (TH1) activity . These findings have led to an improved pathophysiologic understanding of the disease and to new aspects in treatment strategies that are discussed in this paper.

Arch Microbiol, 2002 Apr, 177(4), 299 - 303 Epub 2002 Jan 31.
Purification and characterization of the methylene tetrahydromethanopterin dehydrogenase MtdB and the methylene tetrahydrofolate dehydrogenase FolD from Hyphomicrobium zavarzinii ZV580; Goenrich M et al.; Recently, it has been shown that heterotrophic methylotrophic Proteobacteria contain tetrahydrofolate (H(4)F)- and tetrahydromethanopterin (H(4)MPT)-dependent enzymes . Here we report on the purification of two methylene tetrahydropterin dehydrogenases from the methylotroph Hyphomicrobium zavarzinii ZV580 . Both dehydrogenases are composed of one type of subunit of 31 kDa . One of the dehydrogenases is NAD(P)-dependent and specific for methylene H(4)MPT (specific activity: 680 U/mg) . Its N-terminal amino acid sequence showed sequence identity to NAD(P)-dependent methylene H(4)MPT dehydrogenase MtdB from Methylobacterium extorquens AM1 . The second dehydrogenase is specific for NADP and methylene H(4)F (specific activity: 180 U/mg) and also exhibits methenyl H(4)F cyclohydrolase activity . Via N-terminal amino acid sequencing this dehydrogenase was identified as belonging to the classical bifunctional methylene H(4)F dehydrogenases/cyclohydrolases (FolD) found in many bacteria and eukarya . Apparently, the occurrence of methylene tetrahydrofolate and methylene tetrahydromethanopterin dehydrogenases is not uniform among different methylotrophic alpha-Proteobacteria . For example, FolD was not found in M . extorquens AM1, and the NADP-dependent methylene H(4)MPT dehydrogenase MtdA was present in the bacterium that also shows H(4)F activity.

Biosens Bioelectron, 2002 May, 17(5), 427 - 32
Enhancement in the sensitivity of a gas biosensor by using an advanced immobilization of a recombinant bioluminescent bacterium; Gil GC et al.; A genetically engineered bioluminescent bacterium (lac::luxCDABE) was immobilized to develop a whole cell biosensor for the detection of toxic gaseous chemicals . The toxicity of chemicals can be evaluated through the bioluminescent reaction as it reduces in intensity when the cells experience toxic or lethal conditions . This whole cell biosensor was fabricated, using an immobilization technique utilizing solid agar medium, for the measurement of toxicity through direct contact of the cells with the gas . To enhance the sensitivity of the biosenor, glass beads were used and the thickness of the agar layer was reduced . The bioluminescent response was measured using a fiber optic probe connected between the biosensor kit and a luminometer . As sample gaseous toxic chemicals, BTEX (Benzene, Toluene, Ethylbenzene, and Xylene) gases were selected and their vapors were produced by a gas generation system . The concentrations of the gaseous chemicals injected into the chamber were controlled by the time of exposure and were measured using a portable gas chromatograph (Allstech., USA) . Additions of glass beads facilitated gas diffusion through the solid medium, making the biosensor more sensitive . In addition, a thinner matrix layer was more advantageous for the detection of gas toxicity.

Microbiology, 2002 Mar, 148(Pt 3), 853 - 60
Bioenergetics of the alkaliphilic sulfate-reducing bacterium Desulfonatronovibrio hydrogenovorans; Sydow U et al.; Energy metabolism of the alkaliphilic sulfate-reducing bacterium Desulfonatronovibrio hydrogenovorans strain Z-7935 was investigated in continuous culture and in physiological experiments on washed cells . When grown in chemostats with H2 as electron donor, the cells had extrapolated growth yields {Y(max), g dry cell mass (mol electron acceptor)(-1)} of 5.5 with sulfate and 12.8 with thiosulfate . The maintenance energy coefficients were 1.9 and 1.3 mmol (g dry mass)(-1) x h(-1), and the minimum doubling times were 27 and 20 h with sulfate and thiosulfate, respectively . Cell suspensions reduced sulfate, thiosulfate, sulfite, elemental sulfur and molecular oxygen in the presence of H2 . In the absence of H2, sulfite, thiosulfate and sulfur were dismutated to sulfide and sulfate . Sulfate and sulfite were only reduced in the presence of sodium ions, whereas sulfur was reduced also in the absence of Na+ . Plasmolysis experiments showed that sulfate entered the cells via an electroneutral symport with Na+ ions . The presence of an electrogenic Na+-H+ antiporter was demonstrated in experiments applying monensin (an artificial electroneutral Na+-H+ antiporter) and propylbenzylylcholine mustard.HCl (a specific inhibitor of Na+-H+ antiporters) . Sulfate reduction was sensitive to uncouplers (protonophores), whereas sulfite reduction was not affected . Changes in pH upon lysis of washed cells with butanol indicated that the intracellular pH was lower than the optimum pH for growth (pH 9.5) . Pulses of NaCl (0.52 M) to cells incubated in the absence of Na+ did not result in ATP formation, whereas HCl pulses (shifting the pH from 9.2 to 7.0) did . Small oxygen pulses, which were reduced within a few seconds, caused a transient alkalinization . The results of preliminary experiments with chemiosmotic inhibitors provided further evidence that the alkalinization was caused by sodium--proton antiport following a primary electron-transport-driven sodium ion translocation . It is concluded that energy conservation in D . hydrogenovorans depends on a proton-translocating ATPase, whereas electron transport appears to be coupled to sodium ion translocation.

Microbiology, 2002 Mar, 148(Pt 3), 735 - 42
The growth cycle of Simkania negevensis; Kahane S et al.; Simkania negevensis, a bacterium formerly referred to as 'the micro-organism Z' or 'Simkania Z', belongs to the order Chlamydiales, assigned to the family Simkaniaceae: The purpose of this study was to investigate the production of Simkania negevensis progeny in infected cells in comparison with the well-documented Chlamydiaceae developmental cycle . It was found that replicating Simkania negevensis in Vero cells resembled the reticulate bodies of all known chlamydial species: in electron micrographs they were reticulated, homogeneously staining, and often caught in the process of binary division . These replicative forms were found in low abundance shortly after infection, but by 3 days post-infection they were the most prevalent particles in host cells . Electron-dense forms of Simkania negevensis began to appear on the third day post-infection, but quantitatively did not account for the high titre of infectivity in extracts from these host cells . These had both electron-dense and electron-lucent areas, a characteristic seen only in a few chlamydial species . Simkania negevensis infectivity did not appreciably change during the ensuing 12 days required for host cell lysis, despite an eightfold increase in the proportion of electron-dense bacteria over this time . The emergence of electron-dense bodies, increase in infectivity and host-cell lysis were not synchronized developmental events . This is a novel finding in Chlamydiales spp . and suggests that Simkania negevensis will provide new perspectives in the mechanisms of chlamydial intracellular growth.

Proc Natl Acad Sci U S A, 2002 Mar 5, 99(5), 2918 - 23
Wolbachia density and virulence attenuation after transfer into a novel host; McGraw EA et al.; The factors that control replication rate of the intracellular bacterium Wolbachia pipientis in its insect hosts are unknown and difficult to explore, given the complex interaction of symbiont and host genotypes . Using a strain of Wolbachia that is known to over-replicate and shorten the lifespan of its Drosophila melanogaster host, we have tracked the evolution of replication control in both somatic and reproductive tissues in a novel host/Wolbachia association . After transinfection (the transfer of a Wolbachia strain into a different species) of the over-replicating Wolbachia popcorn strain from D . melanogaster to Drosophila simulans, we demonstrated that initial high densities in the ovaries were in excess of what was required for perfect maternal transmission, and were likely causing reductions in reproductive fitness . Both densities and fitness costs associated with ovary infection rapidly declined in the generations after transinfection . The early death effect in D . simulans attenuated only slightly and was comparable to that induced in D . melanogaster . This study reveals a strong host involvement in Wolbachia replication rates, the independence of density control responses in different tissues, and the strength of natural selection acting on reproductive fitness.

Syst Appl Microbiol, 2001 Dec, 24(4), 618 - 22
Viable cytophaga-like bacterium in the 0.2 microm-filtrate seawater; Elsaied HE et al.; A strain of the Cytophaga-like bacterium (CLB), Nano-1, was isolated from the 0.2 microm-filtrate of natural seawater . Both cellular fatty acid and 16S rDNA sequence analyses indicated that Nano-1 is closely affiliated to the marine gliding CLB genus, Microscilla . Nano-1 was observed to undergo cyclic morphological change typical of the genus Microscilla, and sub-0.2-microm cells were formed in the late stationary phase . The sub-0.2-microm cells were repeatedly revived and subcultured . Formation of the sub-0.2-microm cells seems to be adaptive for oligotrophic growth and starvation survival.

Genome Res, 2002 Mar, 12(3), 408 - 13
Domain-level differences in microsatellite distribution and content result from different relative rates of insertion and deletion mutations; Metzgar D et al.; Microsatellites (short tandem polynucleotide repeats) are found throughout eukaryotic genomes at frequencies many orders of magnitude higher than the frequencies predicted to occur by chance . Most of these microsatellites appear to have evolved in a generally neutral manner . In contrast, microsatellites are generally absent from bacterial genomes except in locations where they provide adaptive functional variability, and these appear to have evolved under selection . We demonstrate a mutational bias towards deletion (repeat contraction) in a native chromosomal microsatellite of the bacterium Mycoplasma gallisepticum, through the collection and analysis of independent mutations in the absence of natural selection . Using this and similar existing data from two other bacterial species and four eukaryotic species, we find strong evidence that deletion biases resulting in repeat contraction are common in bacteria, while eukaryotic microsatellites generally experience unbiased mutation or a bias towards insertion (repeat expansion) . This difference in mutational bias suggests that eukaryotic microsatellites should generally expand wherever selection does not exclude them, whereas bacterial microsatellites should be driven to extinction by mutational pressure wherever they are not maintained by selection . This is consistent with observed bacterial and eukaryotic microsatellite distributions . Hence, mutational biases that differ between eukaryotes and bacteria can account for many of the observed differences in microsatellite DNA content and distribution found in these two groups of organisms.

J Bacteriol, 2002 Mar, 184(6), 1693 - 702
Regulation of nap gene expression and periplasmic nitrate reductase activity in the phototrophic bacterium Rhodobacter sphaeroides DSM158; Gavira M et al.; Bacterial periplasmic nitrate reductases (Nap) can play different physiological roles and are expressed under different conditions depending on the organism . Rhodobacter sphaeroides DSM158 has a Nap system, encoded by the napKEFDABC gene cluster, but nitrite formed is not further reduced because this strain lacks nitrite reductase . Nap activity increases in the presence of nitrate and oxygen but is unaffected by ammonium . Reverse transcription-PCR and Northern blots demonstrated that the napKEFDABC genes constitute an operon transcribed as a single 5.5-kb product . Northern blots and nap-lacZ fusions revealed that nap expression is threefold higher under aerobic conditions but is regulated by neither nitrate nor ammonium, although it is weakly induced by nitrite . On the other hand, nitrate but not nitrite causes a rapid enzyme activation, explaining the higher Nap activity found in nitrate-grown cells . Translational nap'-'lacZ fusions reveal that the napK and napD genes are not efficiently translated, probably due to mRNA secondary structures occluding the translation initiation sites of these genes . Neither butyrate nor caproate increases nap expression, although cells growing phototrophically on these reduced substrates show a very high Nap activity in vivo (nitrite accumulation is sevenfold higher than in medium with malate) . Phototrophic growth on butyrate or caproate medium is severely reduced in the NapA(-) mutants . Taken together, these results indicate that nitrate reduction in R . sphaeroides is mainly regulated at the level of enzyme activity by both nitrate and electron supply and confirm that the Nap system is involved in redox balancing using nitrate as an ancillary oxidant to dissipate excess reductant.

J Bacteriol, 2002 Mar, 184(6), 1649 - 60
RecA Protein from the extremely radioresistant bacterium Deinococcus radiodurans: expression, purification, and characterization; Kim JI et al.; The RecA protein of Deinococcus radiodurans (RecA(Dr)) is essential for the extreme radiation resistance of this organism . The RecA(Dr) protein has been cloned and expressed in Escherichia coli and purified from this host . In some respects, the RecA(Dr) protein and the E . coli RecA (RecA(Ec)) proteins are close functional homologues . RecA(Dr) forms filaments on single-stranded DNA (ssDNA) that are similar to those formed by the RecA(Ec) . The RecA(Dr) protein hydrolyzes ATP and dATP and promotes DNA strand exchange reactions . DNA strand exchange is greatly facilitated by the E . coli SSB protein . As is the case with the E . coli RecA protein, the use of dATP as a cofactor permits more facile displacement of bound SSB protein from ssDNA . However, there are important differences as well . The RecA(Dr) protein promotes ATP- and dATP-dependent reactions with distinctly different pH profiles . Although dATP is hydrolyzed at approximately the same rate at pHs 7.5 and 8.1, dATP supports an efficient DNA strand exchange only at pH 8.1 . At both pHs, ATP supports efficient DNA strand exchange through heterologous insertions but dATP does not . Thus, dATP enhances the binding of RecA(Dr) protein to ssDNA and the displacement of ssDNA binding protein, but the hydrolysis of dATP is poorly coupled to DNA strand exchange . The RecA(Dr) protein thus may offer new insights into the role of ATP hydrolysis in the DNA strand exchange reactions promoted by the bacterial RecA proteins . In addition, the RecA(Dr) protein binds much better to duplex DNA than the RecA(Ec) protein, binding preferentially to double-stranded DNA (dsDNA) even when ssDNA is present in the solutions . This may be of significance in the pathways for dsDNA break repair in Deinococcus.

J Bacteriol, 2002 Mar, 184(6), 1578 - 86
Rhodospirillum rubrum possesses a variant of the bchP gene, encoding geranylgeranyl-bacteriopheophytin reductase; Addlesee HA et al.; The bchP gene product of Rhodobacter sphaeroides is responsible for the reduction of the isoprenoid moiety of bacteriochlorophyll (Bchl) from geranylgeraniol (GG) to phytol; here, we show that this enzyme also catalyzes the reduction of the isoprenoid moiety of bacteriopheophytin (Bphe) . In contrast, we demonstrate that a newly identified homolog of this gene in Rhodospirillum rubrum encodes an enzyme, GG-Bphe reductase, capable of reducing the isoprenoid moiety of Bphe only . We propose that Rhodospirillum rubrum is a naturally occurring bchP mutant and that an insertion mutation may have been the initial cause of a partial loss of function . Normal BchP function can be restored to Rhodospirillum rubrum, creating a new transconjugant strain possessing Bchl esterified with phytol . We speculate on the requirement of Rhodospirillum rubrum for phytylated Bphe and on a potential link between the absence of LH2 and of phytylated Bchl from the wild-type bacterium . The identification of a second role for the fully functional BchP in catalyzing the synthesis of phytylated Bphe strongly suggests that homologs of this enzyme may be similarly responsible for the synthesis of phytylated pheophytin in organisms possessing photosystem 2 . In addition to bchP, other members of a photosynthesis gene cluster were identified in Rhodospirillum rubrum, including a bchG gene, demonstrated to encode a functional Bchl synthetase by complementation of a Rhodobacter sphaeroides mutant.

Appl Environ Microbiol, 2002 Mar, 68(3), 1468 - 72
Higher abundance of bacteria than of viruses in deep Mediterranean sediments; Danovaro R et al.; The interactions between viral abundance and bacterial density, biomass, and production were investigated along a longitudinal transect consisting of nine deep-sea stations encompassing the entire Mediterranean basin . The numbers of viruses were very low (range, 3.6 x 10(7) to 12.0 x 10(7) viruses g(-1)) and decreased eastward . The virus-to-bacterium ratio was always < 1.0, indicating that the deep-sea sediments of the Mediterranean Sea are the first example of a marine ecosystem not numerically dominated by viruses . The lowest virus numbers were found where the lowest bacterial metabolism and turnover rates and the largest cell size were observed, suggesting that bacterial doubling time might play an important role in benthic virus development.

Environ Sci Technol, 2002 Feb 1, 36(3), 421 - 5
Effect of oxide formation mechanisms on lead adsorption by biogenic manganese (hydr)oxides, iron (hydr)oxides, and their mixtures; Nelson YM et al.; The effects of iron and manganese (hydr)oxide formation processes on the trace metal adsorption properties of these metal (hydr)oxides and their mixtures was investigated by measuring lead adsorption by iron and manganese (hydr)oxides prepared by a variety of methods . Amorphous iron (hydr)oxide formed by fast precipitation at pH 7.5 exhibited greater Pb adsorption (gamma(max) = 50 mmol of Pb/mol of Fe at pH 6.0) than iron (hydr)oxide formed by slow, diffusion-controlled oxidation of Fe(II) at pH 4.5-7.0 or goethite . Biogenic manganese(III/IV) (hydr)oxide prepared by enzymatic oxidation of Mn(II) by the bacterium Leptothrix discophora SS-1 adsorbed five times more Pb (per mole of Mn) than an abiotic manganese (hydr)oxide prepared by oxidation of Mn(II) with permanganate, and 500-5000 times more Pb than pyrolusite oxides (betaMnO2) . X-ray crystallography indicated that biogenic manganese (hydr)oxide and iron (hydr)oxide were predominantly amorphous or poorly crystalline and their X-ray diffraction patterns were not significantly affected by the presence of the other (hydr)oxide during formation . When iron and manganese (hydr)oxides were mixed after formation, or for Mn biologically oxidized with iron(III) (hydr)oxide present, observed Pb adsorption was similar to that expected for the mixture based on Langmuir parameters for the individual (hydr)oxides . These results indicate that interactions in iron/manganese (hydr)oxide mixtures related to the formation process and sequence of formation such as site masking, alterations in specific surface area, or changes in crystalline structure either did not occur or had a negligible effect on Pb adsorption by the mixtures.

BMC Microbiol . 2002;2(1):3 . Epub 2002 Feb 13.
Serovar distribution of a DNA sequence involved in the antigenic relationship between Leptospira and equine cornea; Lucchesi PM et al.; BACKGROUND: Horses infected with Leptospira present several clinical disorders, one of them being recurrent uveitis . A common endpoint of equine recurrent uveitis is blindness . Serovar pomona has often been incriminated, although others have also been reported . An antigenic relationship between this bacterium and equine cornea has been described in previous studies . A leptospiral DNA fragment that encodes cross-reacting epitopes was previously cloned and expressed in Escherichia coli . RESULTS: A region of that DNA fragment was subcloned and sequenced . Samples of leptospiral DNA from several sources were analysed by PCR with two primer pairs designed to amplify that region . Reference strains from serovars canicola, icterohaemorrhagiae, pomona, pyrogenes, wolffi, bataviae, sentot, hebdomadis and hardjo rendered products of the expected sizes with both pairs of primers . The specific DNA region was also amplified from isolates from Argentina belonging to serogroups Canicola and Pomona . Both L . biflexa serovar patoc and L . borgpetersenii serovar tarassovi rendered a negative result . CONCLUSIONS: The DNA sequence related to the antigen mimicry with equine cornea was not exclusively found in serovar pomona as it was also detected in several strains of Leptospira belonging to different serovars . The results obtained with L . biflexa serovar patoc strain Patoc I and L . borgpetersenii serovar tarassovi strain Perepelicin suggest that this sequence is not present in these strains, which belong to different genomospecies than those which gave positive results . This is an interesting finding since L . biflexa comprises nonpathogenic strains and serovar tarassovi has not been associated clinically with equine uveitis.

Comp Biochem Physiol A Mol Integr Physiol, 2002 Mar, 131(3), 559 - 67
The Antarctic Psychrobacter sp . TAD1 has two cold-active glutamate dehydrogenases with different cofactor specificities . Characterisation of the NAD+-dependent enzyme; Camardella L et al.; Psychrobacter sp . TAD1 is a psychrotolerant bacterium from Antarctic frozen continental water that grows from 2 to 25 degrees C with optimal growth rate at 20 degrees C . The new isolate contains two glutamate dehydrogenases (GDH), differing in their cofactor specificities, subunit sizes and arrangements, and thermal properties . NADP+-dependent GDH is a hexamer of 47 kDa subunits and it is comparable to other hexameric GDHs of family-I from bacteria and lower eukaria . The NAD+-dependent enzyme, described in this communication, has a subunit weight of 160 kDa and belongs to the novel class of GDHs with large size subunits . The enzyme is a dimer; this oligomeric arrangement has not been reported previously for GDH . Both enzymes have an apparent optimum temperature for activity of approximately 20 degrees C, but their cold activities and thermal labilities are different . The NAD+-dependent enzyme is more cold active: at 10 C it retains 50% of its maximal activity, compared with 10% for the NADP+-dependent enzyme . The NADP+-dependent enzyme is more heat stable, losing only 10% activity after heating for 30 min, compared with 95% for the NAD+-dependent enzyme . It is concluded that in Psychrobacter sp . TAD1 not only does NAD+-dependent GDH have a novel subunit molecular weight and arrangement, but that its polypeptide chains are folded differently from those of NADP+-dependent GDH, providing different cold-active properties to the two enzymes.

Biomacromolecules, 2002 Jan-Feb, 3(1), 208 - 13
Formation of short chain length/medium chain length polyhydroxyalkanoate copolymers by fatty acid beta-oxidation inhibited Ralstonia eutropha; Green PR et al.; Ralstonia eutropha has been considered as a bacterium, incorporating hydroxyalkanoates of less than six carbons only into polyhydroxyalkanoates (PHAs) . Cells of the wild type cultivated with sodium octanoate as the carbon source in the presence of the fatty acid beta-oxidation inhibitor sodium acrylate synthesized PHAs composed of the medium chain length hydroxyalkanoates (3HA(MCL)) 3-hydroxyhexanoate (3HHx) and 3-hydroxyoctanoate (3HO) as well as of 3-hydroxybutyrate and 3-hydroxyproprionate as revealed by gas chromatography, (1)H NMR spectroscopy, and mass spectroscopy . The characterization of the polymer as a tetrapolymer was confirmed by differential solvent extraction and measurement of melting and glass transition temperature depression in the purified polymer compared to PHB . These data suggested that the R . eutropha PHA synthase is capable of incorporating longer chain substrates than suggested by previous in vitro studies . Furthermore, expression of the class II PHA synthase gene phaC1 from P . aeruginosa in R . eutropha resulted in the accumulation of PHAs consisting of 3HA(MCL) contributing about 3-5% to cellular dry weight . These PHAs were composed of nearly equal molar fractions of 3HO and 3-hydroxydecanoate (3HD) with traces of 3HHx . These data indicated that 3HA(MCL)-CoA thioesters were diverted from the fatty acid beta-oxidation pathway towards PHA biosynthesis in recombinant R . eutropha.

Respir Med, 2002 Jan, 96(1), 59 - 60
A world-wide internet survey of public knowledge about tuberculosis; Corless JA et al.; Four simple multiple-choice questions about tuberculosis (TB) were posted on a non-medical internet site for a 2-month period . A total of 564 responses were received . Sixty-two were excluded as individuals had made multiple attempts at the questions . Sixty-five per cent of responses were from North America, 14.5% from Europe and 12% from Australia and New Zealand, with only a small number of responses from Africa, the Indian subcontinent and South America . Of the respondents 49.5% correctly answered that cough is the commonest symptom of TB, 45% knew that TB was transmitted mainly by air-borne droplets, 37.8% knew that TB was caused by a bacterium . Only 19.5% knew that the most important risk factor for developing TB was HIV infection and only 4% answered all questions correctly . This survey suggests that knowledge about tuberculosis is limited in computer-literate individuals throughout the world.

Biochemistry, 2002 Mar 5, 41(9), 3096 - 108
A hyperthermophilic plant-type {2Fe-2S} ferredoxin from Aquifex aeolicus is stabilized by a disulfide bond; Meyer J et al.; A {2Fe-2S} ferredoxin (Fd1) from the hyperthermophilic bacterium Aquifex aeolicus has been obtained by heterologous expression of the encoding gene in Escherichia coli . Sequence comparisons show that this protein belongs to the extended family of plant- and mammalian-type {2Fe-2S} ferredoxins but also indicate that it is not closely similar to either the plant-type or mammalian-type subfamilies . Instead, it appears to bear some similarity to novel members of this family, in particular the Isc-type ferredoxins involved in the assembly of iron-sulfur clusters in vivo . The two redox levels of the {2Fe-2S}(2+/+) metal site of A . aeolicus ferredoxin have been studied by UV-visible, resonance Raman, EPR, variable temperature magnetic circular dichroism, and Mossbauer spectroscopies . A full-spin Hamiltonian analysis is given for the Mossbauer spectra . In aggregate, the spectroscopic data reveal differences with both the plant-type and mammalian-type ferredoxins, in keeping with the sequence comparisons . The midpoint potential of the {2Fe-2S}(2+/+) couple, at -375 mV versus the normal hydrogen electrode, is more negative than those of mammalian-type ferredoxins and at the upper end of the range covered by plant-type ferredoxins . A . aeolicus ferredoxin contains two cysteines in addition to the four that are committed as ligands of the {2Fe-2S} cluster . These two residues have been shown by chemical modification and site-directed mutagenesis to form a disulfide bridge in the native protein . While that cystine unit plays a significant role in the exceptional thermostability of A . aeolicus ferredoxin (T(m) = 121 degrees C at pH 7 versus T(m) = 113 degrees C in a molecular variant where the disulfide bridge has been removed), it does not bear on the properties of the {2Fe-2S}(2+/+) chromophore . This observation is consistent with the large distance (ca . 20 A) that is predicted to separate the iron-sulfur chromophore from the disulfide bridge.

Biochemistry, 2002 Mar 5, 41(9), 3081 - 8
High yield of B-branch electron transfer in a quadruple reaction center mutant of the photosynthetic bacterium Rhodobacter sphaeroides; de Boer AL et al.; A new reaction center (RC) quadruple mutant, called LDHW, of Rhodobacter sphaeroides is described . This mutant was constructed to obtain a high yield of B-branch electron transfer and to study P(+)Q(B)(-) formation via the B-branch . The A-branch of the mutant RC contains two monomer bacteriochlorophylls, B(A) and beta, as a result of the H mutation L(M214)H . The latter bacteriochlorophyll replaces bacteriopheophytin H(A) of wild-type RCs . As a result of the W mutation A(M260)W, the A-branch does not contain the ubiquinone Q(A); this facilitates the study of P(+)Q(B)(-) formation . Furthermore, the D mutation G(M203)D introduces an aspartic acid residue near B(A) . Together these mutations impede electron transfer through the A-branch . The B-branch contains two bacteriopheophytins, Phi(B) and H(B), and a ubiquinone, Q(B.) Phi(B) replaces the monomer bacteriochlorophyll B(B) as a result of the L mutation H(M182)L . In the LDHW mutant we find 35-45% B-branch electron transfer, the highest yield reported so far . Transient absorption spectroscopy at 10 K, where the absorption bands due to the Q(X) transitions of Phi(B) and H(B) are well resolved, shows simultaneous bleachings of both absorption bands . Although photoreduction of the bacteriopheophytins occurs with a high yield, no significant (approximately 1%) P(+)Q(B)(-) formation was found.

Biochemistry, 2002 Mar 5, 41(9), 2932 - 45
Identification of a thiosulfate utilization gene cluster from the green phototrophic bacterium Chlorobium limicola; Verte F et al.; Chlorobium is an autotrophic, green phototrophic bacterium which uses reduced sulfur compounds to fix carbon dioxide in the light . The pathways for the oxidation of sulfide, sulfur, and thiosulfate have not been characterized with certainty for any species of bacteria . However, soluble cytochrome c-551 and flavocytochrome c (FCSD) have previously been implicated in the oxidation of thiosulfate and sulfide on the basis of enzyme assays in Chlorobium . We have now made a number of observations relating to the oxidation of reduced sulfur compounds . (1) Western analysis shows that soluble cytochrome c-551 in Chlorobium limicola is regulated by thiosulfate, consistent with a role in the utilization of thiosulfate . (2) A membrane-bound flavocytochrome c-sulfide dehydrogenase (which is normally a soluble protein in other species) is constitutive and not regulated by sulfide as expected for an obligately autotrophic species dependent upon sulfide . (3) We have cloned the cytochrome c-551 gene from C . limicola and have found seven other genes, which are also presumably involved in sulfur metabolism and located near that for cytochrome c-551 (SoxA) . These include genes for a flavocytochrome c flavoprotein homologue (SoxF2), a nucleotidase homologue (SoxB), four small proteins (including SoxX, SoxY, and SoxZ), and a thiol-disulfide interchange protein homologue (SoxW) . (4) We have established that the constitutively expressed FCSD genes (soxEF1) are located elsewhere in the genome . (5) Through a database search, we have found that the eight thiosulfate utilization genes are clustered in the same order in the Chlorobium tepidum genome . Similar thiosulfate utilization gene clusters occur in at least six other bacterial species but may additionally include genes for rhodanese and sulfite dehydrogenase.

J Med Microbiol, 2002 Feb, 51(2), 159 - 68
Induction of apoptosis of human macrophages in vitro by Legionella longbeachae through activation of the caspase pathway; Arakaki N et al.; The cytotoxicity of the facultative intracellular bacterium, Legionella longbeachae, an important cause of legionellosis, was characterised . Apoptosis was induced in HL-60 cells, a human macrophage-like cell line, during the early stages of infection and induction of apoptosis correlated with cytotoxicity . Apoptosis was confirmed by agarose gel electrophoresis of fragmented DNA, surface exposure of phosphatidylserine and propidium iodide labelling of host cell nuclei . The involvement of macrophage infectivity potentiator (Mip) protein, a known virulence factor of L . longbeachae, was also examined . A mip mutant of L . longbeachae induced apoptosis of HL-60 cells but failed to multiply intracellularly, suggesting that intracellular replication of L . longbeachae is not essential for the induction of apoptosis of HL-60 cells . Furthermore, induction of apoptosis of L . longbeachae-infected macrophages was mediated by activation of the caspase pathway but might be independent of tumour necrosis factor-alpha- and Fas-mediated signal transduction pathways.

Parasitology, 2002 Feb, 124(Pt 2), 127 - 36
Natural Ehrlichia phagocytophila transmission coefficients from sheep 'carriers' to Ixodes ricinus ticks vary with the numbers of feeding ticks; Ogden NH et al.; In a longitudinal study in a UK upland site, 38% of adult sheep were detected as infected with the tick-borne bacterium Ehrlichia phagocytophila by PCR of blood samples . Infection prevalence declined significantly with sheep age but varied significantly and non-linearly with the number of adult Ixodes ricinus ticks feeding per sheep . These findings suggested that under conditions of natural repeated tick-borne challenge sheep remain partially susceptible to re-infections, but the likelihood of re-infection depended on the numbers of feeding ticks . Transmission efficiency from sheep to immature ticks also varied significantly and non-linearly with the number of adult ticks feeding per sheep: transmission efficiency was almost zero in sheep with low adult tick infestations rising to 30% at certain levels of adult tick infestation . Infection intensity in infected engorged immature ticks also varied with the number of adult ticks feeding per sheep, but neither prevalence nor intensity of infection in engorged ticks were related to sheep blood PCR result . These findings suggest that variation in the numbers of ticks feeding per sheep may influence E . phagocytophila transmission by direct effects on transmission at the tick-host interface.

J Bioenerg Biomembr, 2002 Feb, 34(1), 21 - 30
The quinol:fumarate oxidoreductase from the sulphate reducing bacterium Desulfovibrio gigas: spectroscopic and redox studies; Lemos RS et al.; The membrane bound fumarate reductase (FRD) from the sulphate-reducer Desulfovibrio gigas was purified from cells grown on a fumarate/sulphate medium and extensively characterized . The FRD is isolated with three subunits of apparent molecular masses of 71, 31, and 22 kDa . The enzyme is capable of both fumarate reduction and succinate oxidation, exhibiting a higher specificity toward fumarate (Km for fumarate is 0.42 and for succinate 2 mM) and a reduction rate 30 times faster than that for oxidation . Studies by Visible and EPR spectroscopies allowed the identification of two B-type haems and the three iron-sulplur clusters usually found in FRDs and succinate dehydrogenases: {2Fe-2S}2+/1+ (S1), {4Fe-4S}2+/1+ (S2), and {3Fe-4S}1+/0 (S3) . The apparent macroscopic reduction potentials for the metal centers, at pH 7.6, were determined by redox titrations: -45 and -175 mV for the two haems, and +20 and -140 mV for the S3 and SI clusters, respectively . The reduction potentials of the haem groups are pH dependent, supporting the proposal that fumarate reduction is associated with formation of the membrane proton gradient . Furthermore, co-reconstitution in liposomes of D . gigas FRD, duroquinone, and D . gigas cytochrome bd shows that this system is capable of coupling succinate oxidation with oxygen reduction to water.

Vaccine, 2002 Feb 22, 20(11-12), 1586 - 92
Enhanced immune protection by a liposome-encapsulated recombinant respiratory syncytial virus (RSV) vaccine using immunogenic lipids from Deinococcus radiodurans; Huang Y et al.; The radiation-resistant bacterium, Deinococcus radiodurans contains a variety of phospho-, glyco- and phosphoglycolipids, the structures of which appear to be largely unique in nature . We show here that such lipids are immunogenic when administered as liposomes intranasally in mice, as evidenced by the induction of serum antibodies which recognize D . radiodurans lipids but not other lipids by thin layer chromatographic immunostaining . By modifying a liposomal vaccine against respiratory syncytial virus (RSV) we find that vaccine efficacy is markedly enhanced by the inclusion of lipids isolated from D . radiodurans . Using dioleoylphosphatidylcholine (DOPC) or D . radiodurans lipids, liposomes were prepared which encapsulated a soluble fragment of the RSV G protein (G(128-188)) fused with a portion of the bacterial thioredoxin (Trx) protein . Mice immunized intranasally with D . radiodurans liposomes showed markedly greater protection against RSV challenge over mice immunized with DOPC liposomes . Enhanced vaccine efficacy was achieved using liposomes prepared from either whole D . radiodurans lipids or from a single isolated phosphoglycolipid previously identified as alpha-galactosylphosphatidylglyceroylalkylamine (lipid 7) . Mice immunized and protected against RSV challenge were free of pulmonary eosinophilic infiltration, an undesirable consequence of many RSV vaccines . The results provide further support for liposome-based vaccines for RSV and underline the importance of lipid composition in liposome formulations.

Biotechnol Bioeng, 2002 Apr 5, 78(1), 17 - 23
Numerical simulation for electrochemical cultivation of iron oxidizing bacteria; Matsumoto N et al.; A numerical simulation model was constructed for electrochemical cultivation of iron oxidizing bacterium, Thiobacillus ferrooxidans, based on Monod's dual limitation equation . In this model, two limiting factors were examined, low supply of Fe(II) ion and dissolved oxygen, from empirical viewpoints . The simulation model was constructed taking into consideration the energy balance based on the amount of the electronic flow from the electrode to bacteria via an iron ion, and then to oxygen . The model consisted of a logarithmic bacterial growth phase during the first three days, followed by a plateau and growth limitation thereafter . The predicted results were in agreement with the actual growth under electrochemical cultivation . It was predicted the growth limiting factor would be changed from insufficient supply of Fe(II) ions to that of oxygen by decreasing the value of oxygen transfer constant K, which correlated with the aeration rate . The optimum aeration rate was determined for the ideal electrochemical cultivation . The algorithm described here can be used in any electrochemical cultivation by modifying the parameters for each system .

Acta Crystallogr D Biol Crystallogr, 2002 Mar, 58(Pt 3), 556 - 8 Epub 2002 Feb 21.
Expression, crystallization and preliminary X-ray diffraction studies on the complete choline-binding domain of the major pneumococcal autolysin; Fernandez-Tornero C et al.; The major pneumococcal autolysin (LytA), a virulence factor of this bacterium, is composed of an amino-terminal catalytic domain plus a carboxyl-terminal choline-binding domain (ChBD) . This C-terminal domain, responsible for anchorage to the cell wall, is a tandem of six imperfect 20-residue repeats whose precise ends have been difficult to establish by sequence methods . The reported crystal structure of a shortened C-terminal fragment of the protein suggested that it might contain an additional repeat and thus an additional choline-binding site (ChBS) . The complete recombinant choline-binding domain of LytA has now been overexpressed in soluble form using a secreting Escherichia coli strain which facilitates purification with a higher yield . It has been crystallized at room temperature using MPD as the main precipitant . The crystals belong to space group P2(1) and diffract to beyond 3.2 A resolution on a synchrotron-radiation source . The molecular-replacement solution indicates that a new ChBS which fits the topology of the solenoid structure is formed in the N-terminal region.

Acta Crystallogr D Biol Crystallogr, 2002 Mar, 58(Pt 3), 522 - 3 Epub 2002 Feb 21.
Crystallization and preliminary X-ray diffraction analysis of cytochrome c peroxidase from the purple phototrophic bacterium Rhodobacter capsulatus; De Smet L et al.; Bacterial cytochrome c peroxidase (BCCP) from Rhodobacter capsulatus was heterologously expressed in Escherichia coli . It was purified to homogeneity and crystallized using the hanging-drop vapour-diffusion method . Diffraction-quality crystals of the enzyme were obtained under two conditions . The first crystal belonged to space group P2(1), with unit-cell parameters a = 99.2, b = 224.7, c = 167.9 A, beta = 105 degrees, and diffracted to 3.5A resolution . The crystallographic asymmetric unit of these crystals contained ten peroxidase molecules . R . capsulatus BCCP also crystallized in space group P2(1)2(1)2(1), with unit-cell parameters a = 67.2, b = 134.4, c = 167.9 A . These crystals diffracted to 2.7 A resolution and contained four peroxidase molecules per crystallographic asymmetric unit.

Jpn J Cancer Res, 2002 Feb, 93(2), 111 - 6
Helicobacter pylori does not promote N-methyl-N-nitrosourea-induced gastric carcinogenesis in SPF C57BL/6 mice; Nakamura Y et al.; Helicobacter pylori (H . pylori) infection has been acknowledged as a promoter and an initiator for gastric carcinogenesis in experimental models using Mongolian gerbils with H . pylori strains TN2GF4 and ATCC 43504, which have + ve cagA and vacA phenotype s1 / m1 . To get more insight into the role of H . pylori in gastric carcinogenesis, we studied the effect of H . pylori SS1, which has + ve cagA and vacA phenotype s2 / m2, on N-methyl-N-nitrosourea (MNU)-induced chemical gastric carcinogenesis using SPF C57BL / 6 mice . Thus, H . pylori SS1 was inoculated 1 week after the completion of MNU treatment to examine the promoting effect of this bacterium . The incidences of polypoid lesions, differentiated adenocarcinomas, and adenomatous hyperplasias were 67% (10 / 15), 47% (7 / 15) and 80% (12 / 15), respectively, in the MNU-alone group . The corresponding figures were 31% (8 / 26), 23% (6 / 26) and 35% (9 / 26) in the MNU + H . pylori group . The incidences of polypoid lesions and adenomatous hyperplasia were significantly different between the groups . Thus, the results indicate that H . pylori SS1 infection reduced susceptibility to chemical gastric carcinogenesis in this model . The discrepancy between the present result and previous results is likely to have been caused by differences in host factors and bacterial factors . Further study of the relationship between gastric carcinogenesis and H . pylori infection is needed.

Microb Pathog, 2002 Mar, 32(3), 135 - 41
Role of immune sera in the in-vitro phagocytosis of Bordetella pertussis strains; Stefanelli P et al.; In this study, phagocytosis of Bordetella pertussis was assessed using a human monocyte-derived macrophage line (THP-1) and immune sera from children who had received primary vaccination during the Italian clinical trial on the efficacy of two acellular three-component (PT-FHA-PRN) and one whole-cell pertussis vaccines . The results demonstrate that phagocytosis of opsonized bacteria with specific immune sera is not significantly enhanced compared with that of non-opsonized bacteria or bacteria opsonized with non-immune sera . A similar result was obtained also using B . pertussis strains showing variants of the pertactin antigen suggesting that those variations do not reduce the capability of the bacterium to invade the monocytes .

Microb Pathog, 2002 Mar, 32(3), 117 - 25
L12 enhances gonococcal transcytosis of polarized Hec1B cells via the lutropin receptor; Spence JM et al.; We previously reported that gonococci convert to a more invasive phenotype (Inv(+)GC) following contact with cells expressing the lutropin receptor (LHr) and that Inv(+)GC express a novel adhesin that interacts with LHr . We propose that this adhesion allows Inv(+)GC to activate LHr and induce gonococcal transcytosis, usurping normal LHr function in fallopian and endometrial epithelium, which is to transport fetal chorionic gonadotropin (hCG) into the mother . Infected polarized Hec1B monolayers, grown on collagen-coated transwells, showed that the passage of GC across the monolayer occurred rapidly, within 30 min, and proceeded at a constant rate with Inv(+)GC passage three-fold faster than GC grown in tissue culture media alone (Inv(-)GC) . Electron microscopy found that Inv(+)GC triggered pseudopod formation around the bacterium, with GC found throughout the Hec1B targets within 30 min, while Inv(-)GC did neither . Pre-treatment of Inv(-)GC with recombinant ribosomal protein L12, a gonococcal "hCG-like" protein previously shown to increase invasion, also increased Inv(-)GC transcytosis to the rate of Inv(+)GC . This enhancement was completely abolished by addition of luteinizing hormone, a cognate ligand of LHr . This is convincing evidence that surface expressed L12 mediates gonococcal invasion and transcytosis via LHr, a mechanism that could be important in the development of invasive gonococcal disease in women .

Dan Medicinhist Arbog . 2001;:212-8.
{Ophthalmic tuberculosis, especially in Denmark}; Norn M; The contagiousity of tuberculosis was demonstrated by Carl J . Salomonsen in 1877, injecting human caseous material in rabbit eyes, causing granulomatous iritis . It was five years before Robert Koch found the bacterium: Mycobacterium tuberculosis (MT) . Marius Tscherning as professor emeritus from the University of Copenhagen, injected bovine MT and sanocrysin (Molgaard's goldderivate) into rabbit eyes, causing corneal phlyctenules, but no ophthalmic reaction developed with only MT or sanocrysin alone (1928-29), not published) . Emil Frandsen in 1959 examined 113 BCG vaccinated persons, who developed benign phlyctenules in 11%, 6-8 weeks after vaccination, and some cases of chronic iridocyclitis and chorioiditis . In 1158 non-vaccinated, some with serious tuberculosis, Frandsen found serious phlyctenules, chronic iridocyclitis, chorioiditis (especially disseminated) and periphlebitis, but not acute iridocyclitis of tuberculous origin . Phlyctenules and other TB-eye-diseases are described from Greenland 1926-76 . Ophthalmic tuberculosis is still an important issue, also in Denmark today.

Infect Immun, 2002 Mar, 70(3), 1591 - 8
Lack of fusion of azurophil granules with phagosomes during phagocytosis of Mycobacterium smegmatis by human neutrophils is not actively controlled by the bacterium; Cougoule C et al.; Biogenesis of phagolysosomes is a very rapid event in neutrophils which takes place with nascent unclosed phagosomes, leading to the release of lysosomal enzymes such as beta-glucuronidase in the extracellular medium . We have previously shown that, under nonopsonic conditions, both pathogenic and nonpathogenic mycobacteria uncouple phagocytosis from fusion of azurophil granules (specialized secretory lysosomes) with phagosomes . In the present study we questioned whether they actively act on neutrophils to block this process or use phagocytic receptors that negatively control the biogenesis of phagolysosomes . As for live unicellular Mycobacterium smegmatis, we observed that nonopsonic phagocytosis of heat-killed mycobacteria did not induce the release of beta-glucuronidase, indicating that M . smegmatis does not actively act on the fusion process in neutrophils . In contrast, phagocytosis of unicellular M . smegmatis opsonized in immune serum or that of small nonopsonized mycobacterial aggregates restored the biogenesis of phagolysosomes . Aggregates were internalized in a CR3- and cholesterol-dependent manner as unicellular mycobacteria . However, aggregates but not unicellular bacteria triggered F-actin and Hck recruitment at the phagosomes, events that have been associated with lysosome fusion . Thus, we propose that M . smegmatis does not actively control the fusion of azurophil granules at early time points postinfection and that mycobacterial aggregates recruit large clusters of receptors at the neutrophil surface which could trap proteins implicated in the biogenesis of phagolysosomes.

Infect Immun, 2002 Mar, 70(3), 1185 - 92
Modulation of gamma interferon-induced major histocompatibility complex class II gene expression by Porphyromonas gingivalis membrane vesicles; Srisatjaluk R et al.; Gamma interferon (IFN-gamma)-induced endothelial cells actively participate in initiating immune responses by interacting with CD4(+) T cells via class II major histocompatibility complex (MHC) surface glycoproteins . Previously, Porphyromonas gingivalis membrane vesicles were shown to selectively inhibit IFN-gamma-induced surface expression of HLA-DR molecules by human umbilical cord vascular endothelial cells . In this study, we demonstrated an absence of HLA-DR alpha mRNA from IFN-gamma-induced cells in the presence of P . gingivalis membrane vesicles by using reverse transcriptase-PCR and Southern blotting . Vesicles also prevented transcription of the gene encoding class II transactivator, a transactivator protein required for IFN-gamma-induced expression of MHC class II genes . In addition, the effects of vesicles on IFN-gamma signal transduction involving Jak and Stat proteins were characterized by using immunoprecipitation and Western blot analyses . Jak1 and Jak2 proteins could not be detected in endothelial cells treated with membrane vesicles . Consequently, IFN-gamma-induced phosphorylation of Jak1, Jak2, and Stat1 alpha proteins was prevented . The class II-inhibitory effect of the membrane vesicles could be eliminated by heating vesicles at 100 degrees C for 30 min or by treating them with a cysteine proteinase inhibitor . This indicates that the cysteine proteinases were most likely responsible for the absence of Jak proteins observed in vesicle-treated cells . The observed increased binding of radiolabeled IFN-gamma to vesicle-treated cells suggests that vesicles may also modulate the IFN-gamma interactions with the cell surface . However, no evidence was obtained demonstrating that vesicles affected the expression of IFN-gamma receptors . Thus, P . gingivalis membrane vesicles apparently inhibited IFN-gamma-induced MHC class II by disrupting the IFN-gamma signaling transduction pathway . Vesicle-inhibited class II expression also occurred in other IFN-gamma-inducible cells . This suggested that the ability of P . gingivalis membrane vesicles to modulate antigen presentation by key cells may be an important mechanism used by this particular bacterium to escape immunosurveillance, thereby favoring its colonization and invasion of host tissues.

Infect Immun, 2002 Mar, 70(3), 1175 - 84
Transcript heterogeneity of the p44 multigene family in a human granulocytic ehrlichiosis agent transmitted by ticks; Zhi N et al.; Human granulocytic ehrlichiosis (HGE) is an emerging tick-borne zoonosis caused by a strain of Anaplasma phagocytophila called the HGE agent, an obligatory intracellular bacterium . The agent expresses immunodominant 44-kDa outer membrane proteins (P44s) encoded by a multigene family . The present study established an experimental process for transmission of the HGE agent from infected mice (a reservoir model) to nymphal Ixodes scapularis ticks (a biological vector) and subsequently to horses (a patient model) by the adult infected ticks . Overall, a total of 20 different p44 transcripts were detected in the mammals, ticks, and cell cultures . Among them, a transcript from a p44-18 gene was major at acute stage in mice and horses but minor in ticks . Both mRNA and protein produced from the p44-18 gene were detected in the HGE agent cultivated in HL-60 cells at 37 degrees C, but their expression levels decreased in the organisms cultivated at 24 degrees C, suggesting that temperature is one of the factors that influence the expression of members of the p44 multigene family . Several additional p44 transcripts that were not detected in the mammals at the acute stage of infection were detected in ticks . Phylogenetic analysis of the 20 different p44 transcripts revealed that the major transcripts found in mammals and ticks were distinct, suggesting a difference in surface properties between populations of the HGE agent in different host environments . The present study provides new information for understanding the role of the p44 multigene family in transmission of the HGE agent between mammals and ticks.

Infect Immun, 2002 Mar, 70(3), 1097 - 105
Dendritic cells pulsed with a recombinant chlamydial major outer membrane protein antigen elicit a CD4(+) type 2 rather than type 1 immune response that is not protective; Shaw J et al.; Chlamydia trachomatis is an obligate intracellular bacterium that infects the oculogenital mucosae . C . trachomatis infection of the eye causes trachoma, the leading cause of preventable blindness . Infections of the genital mucosae are a leading cause of sexually transmitted diseases . A vaccine to prevent chlamydial infection is needed but has proven difficult to produce by using conventional vaccination approaches . Potent immunity to vaginal rechallenge in a murine model of chlamydial genital infection has been achieved only by infection or by immunization with dendritic cells (DC) pulsed ex vivo with whole inactivated organisms . Immunity generated by infection or ex vivo antigen-pulsed DC correlates with a chlamydia-specific interleukin 12 (IL-12)-dependent CD4(+) Th1 immune response . Because of the potent antichlamydial immunizing properties of DC, we hypothesized that DC could be a powerful vehicle for the delivery of individual chlamydial antigens that are thought to be targets for more conventional vaccine approaches . Here, we investigated the recombinant chlamydial major outer membrane protein (rMOMP) as a target antigen . The results demonstrate that DC pulsed with rMOMP secrete IL-12 and stimulate infection-sensitized CD4(+) T cells to proliferate and secrete gamma interferon . These immunological properties implied that rMOMP-pulsed DC would be potent inducers of MOMP-specific CD4(+) Th1 immunity in vivo; however, we observed the opposite result . DC pulsed ex vivo with rMOMP and adoptively transferred to naive mice generated a Th2 rather than a Th1 anti-MOMP immune response, and immunized mice were not protected following infectious challenge . We conclude from these studies that the immunological properties of ex vivo pulsed DC are not necessarily predictive of the immune response generated in vivo following adoptive transfer . These findings suggest that the nature of the antigen used to pulse DC ex vivo influences the Th1-Th2 balance of the immune response in vivo.

Prikl Biokhim Mikrobiol, 2002 Jan-Feb, 38(1), 63 - 7
{Consumption of organic carbon sources and biosynthesis of lactic acid by the photosynthetic bacterium Rhodobacter sp . D-4}; Paronian AKh; Nonsulfur photosynthetic purple bacteria isolated from the Dzhermuk mineral springs (Armenia) were grown on sugar-containing media and found to be capable of synthesizing L(+)-lactic acid . Various organic compounds were tested as possible sole sources of carbon and an electron donors required to support bacterial growth and biosynthesis of lactic acid under various growth conditions.

FEBS Lett, 2002 Feb 13, 512(1-3), 129 - 32
Light control over the size of an antenna unit building block as an efficient strategy for light harvesting in photosynthesis; Yakovlev AG et al.; It was shown that an increase in the bacteriochlorophyll (BChl) c antenna size observed upon lowering growth light intensities led to enhancement of the hyperchromism of the BChl c Q(y) absorption band of the green photosynthetic bacterium Chloroflexus aurantiacus . With femtosecond difference absorption spectroscopy, it was shown that the amplitude of bleaching of the oligomeric BChl c Q(y) band (as compared to that for monomeric BChl a) increased with increasing BChl c content in chlorosomes . This BChl c bleaching amplitude was about doubled as the chlorosomal antenna size was about trebled . Both sets of findings clearly show that a unit BChl c aggregate in the chlorosomal antenna is variable in size and governed by the grow light intensity, thus ensuring the high efficiency of energy transfer within the BChl c antenna regardless of its size.

FEBS Lett, 2002 Feb 13, 512(1-3), 125 - 8
Fractionation of cytochromes of phototrophically grown Chloroflexus aurantiacus . Is there a cytochrome bc complex among them?
Yanyushin MF.
The cytochrome-containing membrane complexes of the phototrophically grown green non-sulfur bacterium Chloroflexus aurantiacus were fractionated by anion exchange chromatography . Three cytochrome b and four cytochrome c peaks were observed . None of the separated complexes met the features of the cytochrome bc complex . Two main cytochrome b-containing complexes were further purified: a dimer of identical subunits with unknown function and a succinate:quinone oxidoreductase containing three subunit species . Two novel multisubunit complexes, similar to each other, with two heme c-bearing subunits were also purified.

FEBS Lett, 2002 Feb 13, 512(1-3), 80 - 4
The proton-pumping NADH:ubiquinone oxidoreductase (complex I) of Aquifex aeolicus; Scheide D et al.; The proton-pumping NADH:ubiquinone oxidoreductase, also called complex I, is the first energy-transducing complex of many respiratory chains . Homologues of complex I are present in the three domains of life . Here, we report the properties of complex I in membranes of the hyperthermophilic bacterium Aquifex aeolicus . The complex reacted with NADH but not with NADPH and F(420)H(2) as electron donors . Short-chain analogues of ubiquinone like decyl-ubiquinone and ubiquinone-2 were suitable electron acceptors . The affinities towards NADH and ubiquinone-2 were comparable to the ones obtained with the Escherichia coli complex I . The reaction was inhibited by piericidin A at the same concentration as in E . coli . The complex showed an unusual pH optimum at pH 9 and a maximal rate at 80 degrees C . We found no evidence for the presence of an alternative, single subunit NADH dehydrogenase in A . aeolicus membranes . The NADH:ferricyanide reductase activity of detergent extracts of A . aeolicus membranes sedimented as a protein with a molecular mass of approximately 550 kDa . From the data we concluded that A . aeolicus contains a NADH:ubiquinone oxidoreductase resembling complex I of mesophilic bacteria.

J Theor Biol, 2002 Feb 21, 214(4), 647 - 56
On the aperiodic locomotor behavior of Halobacterium salinarium under periodic light stimuli; Casagrandi R et al.; Two long time series of swimming intervals of a bacterium inverting its motion under periodic light pulses are analysed . The associated next-period plots reveal, through their filiform structure, that the underlying dynamics are low-dimensional . Using recently described properties of such dynamics, a simple second-order black-box model for the swimming intervals is derived and validated . The model reinforces the conjecture that this bacterium is endowed with an oscillator controlling the switching of the flagellar motor .

Zhonghua Liu Xing Bing Xue Za Zhi, 2001 Dec, 22(6), 411 - 3
{Application and time-effect analysis of antibiotics among neonatal pneumonia treatment}; Li S et al.; OBJECTIVE: To understand the results of antibiotics use in curing neonatal pneumonia and to evaluate the time-effect of different drugs . METHODS: Through stratified sampling, all the hospitalized cases of neonatal pneumonia from 5 hospitals with different levels in Hunan province in five years were retrospectively studied . Analysis of time-effect for different antibiotics was done through Kaplan-Meier . RESULTS: Twenty-six kinds of anti-bacterium drugs were used in 685 cases of neonatal pneumonia, among which penicillin and ampicillin were mostly used but cephalosporins also became one of the main drugs used in treating neonatal pneumonia . Most cases were discovered (51.1%) using two kinds of antibiotics during the course of diseases through time-effect analysis . We found the effect of penicillin was better than others in treating neonatal pneumonia when used as basic medicine . It was not desirable to use two or more medicines at the beginning of the treatment . CONCLUSION: Penicillin and ampicillin were still the main drugs used in treating neonatal pneumonia but more cephalosporins were used than ever . The blindness in applying antibiotics should be recovered . From Kaplan-Meier analysis, we could better understand and evaluate the time-effects of different drug treatments.

J Appl Microbiol, 2002, 92(1), 140 - 6
Assessment of the fructanolytic activities in the rumen bacterium Treponema saccharophilum strain S; Kasperowicz A et al.; AIMS: To characterize the fructose polymer degrading enzymes of rumen bacterium Treponema saccharophilum strain S . METHODS AND RESULTS: Conventional methods were used to examine bacterial growth and enzyme activities . Electrophoretic zymogram under native conditions, and thin layer chromatography, were applied to identify and characterize the enzymes . Treponema saccharophilum utilized Timothy grass fructan, inulin and sucrose but not free fructose . Timothy grass fructan was degraded at a significantly higher rate than sucrose and inulin . Two fructanolytic enzymes were found in the soluble, and one in the membrane fraction of bacterial cell extract . The first degraded each mentioned carbohydrate to monosaccharides . The second released oligosaccharides only from Timothy grass fructan . CONCLUSIONS: The bacterium T . saccharophilum strain S is capable of synthesizing non-specific beta-fructofuranosidases and 2,6-beta-D-fructan fructanohydrolase . The enzymes are of constitutive character . SIGNIFICANCE AND IMPACT OF THE STUDY: It has been stated for the first time that the 2,6-beta-D-fructan fructanohydrolase is synthesized by the rumen bacterium T . saccharophilum . This organism appears to be responsible for grass fructan degradation in the rumen.

Aliment Pharmacol Ther, 2002 Mar, 16 Suppl 1, 3 - 15
Review article: natural history and epidemiology of Helicobacter pylori infection; Go MF; Helicobacter pylori is a common bacterium infecting about half the world's population . It is causally linked with a diverse spectrum of gastrointestinal clinical disorders including peptic ulcer disease, gastric cancer, and gastric MALT lymphoma . The principal reservoir is the human stomach, and transmission probably occurs by person-to-person passage . Prevalence rates are generally much higher in developing countries compared to developed countries, although there are subgroups within many regions with higher H . pylori prevalence than in the general population . The prevalence of H . pylori varies by geographical location, ethnic background, socioeconomic conditions, and age . Recent studies suggest decreasing prevalence in developed countries or those with rapidly improving socioeconomic conditions . Comparative studies of the two fully sequenced H . pylori genomes are providing understanding of its large genetic diversity and bacterial virulence factors . The discovery of the type IV secretion system in H . pylori and its role in translocation of the CagA protein from the bacterial cell into the host epithelial cell provides insight into how host-bacterial interaction may lead to host disease . Cytokine promoter polymorphisms are determinants important in host gastric acid secretion status . Understanding the changing trends in H . pylori epidemiology, details of its transmission pathways, and the bacterial and host determinants leading to gastroduodenal disease remain the challenges in this area . Global epidemiological studies, advances in technology, and medical interventions have converged to help clarify the mechanisms of interaction between this ubiquitous micro-organism and its host that result in its diverse clinical manifestations.

Biochem Biophys Res Commun, 2002 Feb 22, 291(2), 425 - 30
Activity enhancement of Cel5Z from Pectobacterium chrysanthemi PY35 by removing C-terminal region; Park SR et al.; The phytopathogenic bacterium Pectobacteium chrysanthemi PY35 secretes Cel5Z endoglucanase belonging to the glycoside hydrolase family 5 of EC 3.2.1.4 . The mutation of cel5Z::Omega gene was constructed by cloning the 2.0-kb SmaI fragment containing the streptomycin/spectinomycin-resistance gene of pHP45(Omega) into the BalI site of pPY100 . The insertion of Omega fragment generated a new stop codon, removing the Ser/Thr-rich linker region and the cellulose binding domain (CBD) in the C-terminal region of cel5Z gene . By subsequent subcloning from this 4.9-kb fragment (pPY1001), a 1.0-kb (pPY1002) fragment was obtained and designated as cel5Z::Omega . The cel5Z::Omega gene had an open reading frame (ORF) of 1011 bp, encoding 336 amino acids, starting with an ATG codon and ending with a new TGA stop codon . The molecular mass of the Cel5Z::Omega protein in E . coli transformant appeared to be 32 kDa by SDS-PAGE analysis in the presence of carboxymethyl-cellulose (CMC) . The Cel5Z::Omega protein hydrolyzed CMC with 1.7-fold higher activity than the intact Cel5Z cellulase . (c)2002 Elsevier Science (USA).

J Bacteriol, 2002 Mar, 184(5), 1498 - 501
Characterization of the petI and res operons of Acidithiobacillus ferrooxidans; Levican G et al.; DNA sequence analysis and bioinformatic interpretations have identified two adjacent clusters of genes potentially involved in the formation of a bc1 complex and in the maturation of a cytochrome c-type protein in two strains (ATCC 19859 and ATCC 33020) of the acidophilic, chemolithoautotrophic bacterium Acidithiobacillus ferrooxidans (formerly Thiobacillus ferrooxidans) . Reverse transcriptase-PCR experiments suggest that the two clusters are organized as operons, and +1 start sites of transcription for the operons have been determined by primer extension experiments . Potential promoters have been identified . The presence of these operons lends support to a recent model of reverse electron flow and is consistent with previous reports of phenotypic switching in this bacterium.

J Wildl Dis, 2002 Jan, 38(1), 40 - 6
Prevalence of Bordetella avium infection in selected wild and domesticated birds in the eastern USA; Raffel TR et al.; Bordetella avium is the etiologic agent of bordetellosis, a highly contagious upper respiratory disease of young poultry . Its prevalence among domesticated turkeys is well-known, but information on prevalence of this bacterium in other birds is limited . A survey of the prevalence of B . avium in wild and domesticated birds was conducted from June 1998 to January 2000, using tracheal cultures and serology . Of 237 blood samples from 61 species, 100 individuals from 41 species had antibodies against B . avium as determined with a microtiter agglutination test . Nine isolates of B . avium were cultured from 128 tracheal samples . Ribotype analysis of seven isolates from mallards (Anas platyrhynchos), one from a wild turkey (Meleagris gallopavo), and one from a Canada goose (Branta canadensis) indicated that they represent three strains, two of which were indistinguishable from clinical isolates from domesticated turkeys . Bordetella avium is present in wild bird populations of multiple species . Transmission from free-living avian populations to domesticated poultry populations may be possible and should be examined.

Int J Syst Evol Microbiol, 2002 Jan, 52(Pt 1), 215 - 21
Desulfovibrio magneticus sp . nov., a novel sulfate-reducing bacterium that produces intracellular single-domain-sized magnetite particles; Sakaguchi T et al.; A novel type of dissimilatory sulfate-reducing bacterium, designated strain RS-1T, capable of producing intracellular magnetite particles (magnetosomes) was isolated from freshwater sulfide-rich sediments . Phylogenetic analysis based on 16S rDNA sequences revealed that RS-1T is a member of the genus Desulfovibrio . Its closest known relative is Desulfovibrio burkinensis (sequence similarity of 98.7%) . Strain RS-1T contains desulfoviridin, c-type cytochromes and, unlike other Desulfovibrio spp., it possesses menaquinone MK-7(H2) instead of MK-6 or MK-6(H2) . Strain RS-1T is also unique compared with other members of Desulfovibrio in its ability to synthesize intracellular magnetite particles . A novel species, Desulfovibrio magneticus sp . nov., is proposed for RS-1T (= ATCC 700980T = DSM 13731T), a sulfate-reducing magnetotactic bacterium.

Hereditas, 2001, 134(3), 263 - 6
The Stockholm populations of Adalia bipunctata (L) (Coleoptera: Coccinellidae)--a case of extreme female-biased population sex ratio; Zakharov IA et al.; The genetic composition and sex ratio in the Stockholm populations of Adalia bipunctata have been studied . The overall frequency of melanics is 3.2%, which is significantly lower than in the populations of St . Petersburg and other large cities along the Baltic Sea . The secondary sex ratio in the Stockholm populations is female-biased 82:18 . More than half of A . bipunctata females are infected with the male-killing Spiroplasma bacterium . Beetles of the co-existing species Adalia decempunctata are infected with a different bacterium belonging to the genus Rickettsia.

Microbiology, 2002 Feb, 148(Pt 2), 605 - 14
Control of dimethylsulfoxide reductase expression in Rhodobacter capsulatus: the role of carbon metabolites and the response regulators DorR and RegA; Kappler U et al.; Regulation of the expression of dimethylsulfoxide (DMSO) reductase was investigated in the purple phototrophic bacterium Rhodobacter capsulatus . Under phototrophic, anaerobic conditions with malate as carbon source, DMSO caused an approximately 150-fold induction of DMSO reductase activity . The response regulator DorR was required for DMSO-dependent induction and also appeared to slightly repress DMSO reductase expression in the absence of substrate . Likewise, when pyruvate replaced malate as carbon source there was an induction of DMSO reductase activity in cells grown at low light intensity (16 W m(-2)) and again this induction was dependent on DorR . The level of DMSO reductase activity in aerobically grown cells was elevated when pyruvate replaced malate as carbon source . One possible explanation for this is that acetyl phosphate, produced from pyruvate, may activate expression of DMSO reductase by direct phosphorylation of DorR, leading to low levels of induction of dor gene expression in the absence of DMSO . A mutant lacking the global response regulator of photosynthesis gene expression, RegA, exhibited high levels of DMSO reductase in the absence of DMSO, when grown phototrophically with malate as carbon source . This suggests that phosphorylated RegA acts as a repressor of dor operon expression under these conditions . It has been proposed elsewhere that RegA-dependent expression is negatively regulated by the cytochrome cbb3 oxidase . A cco mutant lacking cytochrome cbb3 exhibited significantly higher levels of phi{dorA::lacZ} activity in the presence of DMSO compared to wild-type cells and this is consistent with the above model . Pyruvate restored DMSO reductase expression in the regA mutant to the same pattern as found in wild-type cells . These data suggest that R . capsulatus contains a regulator of DMSO respiration that is distinct from DorR and RegA, is activated in the presence of pyruvate, and acts as a negative regulator of DMSO reductase expression.

Nitric Oxide, 2002 Feb, 6(1), 61 - 8
Nitric oxide-mediated protection of A . actinomycetemcomitans-infected murine macrophages against apoptosis; Nakashima K et al.; We investigated apoptotic cell death in murine macrophage cell line J774.1 following Actinobacillus actinomycetemcomitans infection . Infected macrophages generally kill bacteria within phagosomes with nitric oxide (NO) . Our previous study demonstrated that DNA fragmentation in infected cells increased significantly on addition of S-Methylisothiourea (SMT), a selective inhibitor of inducible NO synthetase (iNOS) . The purpose of the present study was to determine the mechanism via which NO affects apoptosis of infected macrophages . J774.1 cells were infected with A . actinomycetemcomitans Y4 at a bacterium/cell ratio of 500:1 . The infected cells were then cultured in the presence or absence of SMT (400 microM) . Culture supernatant was removed 21 h after the infection to measure LDH activity . Additionally, cellular proteins were extracted from the infected cells and measured for histone-associated DNA fragmentation and caspase-1, -3, -5, -6, -8, -9 activities . LDH activity and DNA fragmentation were significantly elevated by the infection; moreover, levels increased further on addition of SMT . Caspase activity of infected cells, particularly caspase-3, was significantly higher than that of uninfected cells . Furthermore, caspase activity increased on addition of SMT . These findings indicate that NO protects infected J774.1 cells, at least in part, against apoptotic cell death via a decrease in caspase activity.

J Biol Chem, 2002 May 10, 277(19), 17300 - 7 Epub 2002 Feb 04.
Molecular cloning and characterization of sphingolipid ceramide N-deacylase from a marine bacterium, Shewanella alga G8; Furusato M et al.; Recently, lyso-sphingolipids have been identified as ligands for several orphan G protein-coupled receptors, although the molecular mechanism for their generation has yet to be clarified . Here, we report the molecular cloning of the enzyme, which catalyzes the generation of lyso-sphingolipids from various sphingolipids (sphingolipid ceramide N-deacylase) . The 75-kDa enzyme was purified from the marine bacterium, Shewanella alga G8, and its gene was cloned from a G8 genomic library using sequences of the purified enzyme . The cloned enzyme was composed of 992 amino acids, including a signal sequence of 35 residues, and its molecular weight was estimated to be 109,843 . Significant sequence similarities were found with an unknown protein of Streptomyces fradiae Y59 and a Lumbricus terrestris lectin but not other known functional proteins . The 106-kDa recombinant enzyme expressed in Escherichia coli hydrolyzed various glycosphingolipids and sphingomyelin, although it seems to be much less active than the native 75-kDa enzyme . In vitro translation using wheat germ extract revealed the activity of a 75-kDa deletion mutant lacking a C terminus to be much stronger than that of the full-length enzyme, suggesting that C-terminal processing is necessary for full activity.

Biosci Biotechnol Biochem, 2001 Dec, 65(12), 2780 - 4
Analysis of the molecular species of hydrogenase in the cells of an obligately chemolithoautotrophic, marine hydrogen-oxidizing bacterium, Hydrogenovibrio marinus; Nishihara H et al.; Hydrogenovibrio marinus was suggested to have only membrane-bound hydrogenase (MBH) . The change of cultivation pO2 did not affect the molecular species of hydrogenase expressed . We propose the MBH is grouped in class I {NiFe} MBH according to the subunit composition, size (Mw 38,000 and Mw 74,000 subunits) and N-terminal sequences of the subunits, and arrangement of the structural genes . Ni-requirement for the autotrophic growth on H2 also suggested the MBH is the Ni-containing type . Southern hybridization analysis using a part of the MBH gene showed a possibility of the presence of two highly homologous MBHs which were not separated by SDS-PAGE.

Electrophoresis, 2002 Jan, 23(1), 122 - 37
Proteome analysis and identification of symbiosis-related proteins from Medicago truncatula Gaertn . by two-dimensional electrophoresis and mass spectrometry; Bestel-Corre G et al.; Time-course analysis of root protein profiles was studied by two-dimensional gel electrophoresis and silver staining in the model plant Medicago truncatula, inoculated either with the arbuscular mycorrhizal fungus Glomus mosseae or with the nitrogen fixing bacterium Sinorhizobium meliloti . Protein modifications in relation to the development of both symbioses included down- and upregulations, as well as newly induced polypeptides . Matrix assisted laser desorption/ionization-time of flight-mass spectrometry after trypsin digestion clearly identified one polypeptide induced in nodulated roots as a M . truncatula leghemoglobin . Internal sequencing with a quadrupole time-of-flight mass spectrometer and database searches confirmed the induction of proteins previously described in root symbioses, and revealed the implication of other proteins . In nodulated roots, one polypeptide was identified as an elongation factor Tu from S . meliloti, while another one could not be assigned a function . In mycorrhizal roots, analyzed proteins also included a protein of unknown function, as well as a glutathione-S-transferase, a fucosidase, a myosin-like protein, a serine hydroxymethyltransferase and a cytochrome-c-oxidase . These results emphasize the usefulness of proteome analysis in identifying molecular events occurring in plant root symbioses.

Appl Environ Microbiol, 2002 Feb, 68(2), 881 - 92
Transcriptional and proteomic analysis of a ferric uptake regulator (fur) mutant of Shewanella oneidensis: possible involvement of fur in energy metabolism, transcriptional regulation, and oxidative stress; Thompson DK et al.; The iron-directed, coordinate regulation of genes depends on the fur (ferric uptake regulator) gene product, which acts as an iron-responsive, transcriptional repressor protein . To investigate the biological function of a fur homolog in the dissimilatory metal-reducing bacterium Shewanella oneidensis MR-1, a fur knockout strain (FUR1) was generated by suicide plasmid integration into this gene and characterized using phenotype assays, DNA microarrays containing 691 arrayed genes, and two-dimensional polyacrylamide gel electrophoresis . Physiological studies indicated that FUR1 was similar to the wild-type strain when they were compared for anaerobic growth and reduction of various electron acceptors . Transcription profiling, however, revealed that genes with predicted functions in electron transport, energy metabolism, transcriptional regulation, and oxidative stress protection were either repressed (ccoNQ, etrA, cytochrome b and c maturation-encoding genes, qor, yiaY, sodB, rpoH, phoB, and chvI) or induced (yggW, pdhC, prpC, aceE, fdhD, and ppc) in the fur mutant . Disruption of fur also resulted in derepression of genes (hxuC, alcC, fhuA, hemR, irgA, and ompW) putatively involved in iron uptake . This agreed with the finding that the fur mutant produced threefold-higher levels of siderophore than the wild-type strain under conditions of sufficient iron . Analysis of a subset of the FUR1 proteome (i.e., primarily soluble cytoplasmic and periplasmic proteins) indicated that 11 major protein species reproducibly showed significant (P < 0.05) differences in abundance relative to the wild type . Protein identification using mass spectrometry indicated that the expression of two of these proteins (SodB and AlcC) correlated with the microarray data . These results suggest a possible regulatory role of S . oneidensis MR-1 Fur in energy metabolism that extends the traditional model of Fur as a negative regulator of iron acquisition systems.

Appl Environ Microbiol, 2002 Feb, 68(2), 545 - 54
Regulation of endo-acting glycosyl hydrolases in the hyperthermophilic bacterium Thermotoga maritima grown on glucan- and mannan-based polysaccharides; Chhabra SR et al.; The genome sequence of the hyperthermophilic bacterium Thermotoga maritima encodes a number of glycosyl hydrolases . Many of these enzymes have been shown in vitro to degrade specific glycosides that presumably serve as carbon and energy sources for the organism . However, because of the broad substrate specificity of many glycosyl hydrolases, it is difficult to determine the physiological substrate preferences for specific enzymes from biochemical information . In this study, T . maritima was grown on a range of polysaccharides, including barley beta-glucan, carboxymethyl cellulose, carob galactomannan, konjac glucomannan, and potato starch . In all cases, significant growth was observed, and cell densities reached 10(9) cells/ml . Northern blot analyses revealed different substrate-dependent expression patterns for genes encoding the various endo-acting beta-glycosidases; these patterns ranged from strong expression to no expression under the conditions tested . For example, cel74 (TM0305), a gene encoding a putative beta-specific endoglucananse, was strongly expressed on all substrates tested, including starch, while no evidence of expression was observed on any substrate for lam16 (TM0024), xyl10A (TM0061), xyl10B (TM0070), and cel12A (TM1524), which are genes that encode a laminarinase, two xylanases, and an endoglucanase, respectively . The cel12B (TM1525) gene, which encodes an endoglucanase, was expressed only on carboxymethyl cellulose . An extracellular mannanase encoded by man5 (TM1227) was expressed on carob galactomannan and konjac glucomannan and to a lesser extent on carboxymethyl cellulose . An unexpected result was the finding that the cel5A (TM1751) and cel5B (TM1752) genes, which encode putative intracellular beta-specific endoglucanases, were induced only when T . maritima was grown on konjac glucomannan . To investigate the biochemical basis of this finding, the recombinant forms of Man5 (M(r), 76,900) and Cel5A (M(r), 37,400) were expressed in Escherichia coli and characterized . Man5, a T . maritima extracellular enzyme, had a melting temperature of 99 degrees C and an optimun temperature of 90 degrees C, compared to 90 and 80 degrees C, respectively, for the intracellular enzyme Cel5A . While Man5 hydrolyzed both galactomannan and glucomannan, no activity was detected on glucans or xylans . Cel5A, however, not only hydrolyzed barley beta-glucan, carboxymethyl cellulose, xyloglucan, and lichenin but also had activity comparable to that of Man5 on galactomannan and higher activity than Man5 on glucomannan . The biochemical characteristics of Cel5A, the fact that Cel5A was induced only when T . maritima was grown on glucomannan, and the intracellular localization of Cel5A suggest that the physiological role of this enzyme includes hydrolysis of glucomannan oligosaccharides that are transported following initial hydrolysis by extracellular glycosidases, such as Man5.

Can J Microbiol, 2001 Dec, 47(12), 1137 - 40
Peptidases affecting recombinant protein production by Streptomyces lividans; Yun SI et al.; The influence of peptidases on human interleukin-3 (rhIL-3) production by a recombinant Streptomyces lividans strain was investigated . The bacterium produced several general peptidases and tripeptidyl peptidases compromising the authenticity of rhIL-3 . The level of peptidases depended on growth morphology . Growing S . lividans as compact pellets successfully reduced peptidase activity . Maximum general peptidase activity in pellet culture was delayed after maximum rhIL-3 concentration was achieved . The activity of the tripeptidyl peptidase was product (rhIL-3) associated.

Syst Appl Microbiol, 2001 Nov, 24(3), 368 - 76
Classification of a Brevundimonas strain detectable after PCR with a Helicobacter-specific primer pair; Buczolits S et al.; In a study for the isolation of new Helicobacter strains, biopsy samples were taken from the gastric mucosa of dogs and subjected to PCR amplification using a Helicobacter-specific primer pair (H276f and H676r, directed to the 16S rDNA) to identify members of this genus in the specimens . From one Helicobacter positive sample, a bacterial strain was isolated which displayed a characteristic band after PCR amplification with the Helicobacter-specific primer pair . The isolate designated H2/98-FUNDUS was motile, oxidase-, catalase- and aminopeptidase-positive and grew only under microaerophilic conditions at 37 degrees C . The bacterium was classified by a polyphasic approach, including analysis of the isoprenoid quinones, fatty acids, polar lipids and partial 16S rDNA sequence . Analysis of the 16S rDNA sequence (1003 bases) indicated that the strain H2/98-FUNDUS is a member of the genus Brevundimonas and most closely related to Brevundimonas aurantiaca DSM 4731T (99.5% sequence similarity) . Isolate H2/98-FUNDUS contained a predominant ubiquinone Q-10 and a fatty acid profile with the major compounds C18:1 and C16:0 . In the polar lipid extract, phosphatidylglycerol, six unknown phospholipids, one unknown phosphoglycolipid, two unknown glycolipids and two unknown aminolipids were detected . All these results indicate that H2/98-FUNDUS represents a new member of the genus Brevundimonas which gives a positive signal upon PCR employing the Helicobacter-specific primer pair.

Wien Med Wochenschr, 2001, 151(24), 594 - 9
{Infection hypothesis of coronary heart disease}; Jahn J et al.; It is well accepted that coronary artery disease is linked to an inflammatory process . It is unproven however whether either infectious agents may cause or accelerate coronary artery disease or the inflammatory process is due to metabolic or toxic effects . Among the possible infectious agents Chlamydia pneumoniae is the most likely bacterium involved in atherosclerosis . The arguments in favour of Chlamydia pneumoniae originate from seroepidemiologic studies and from detection and isolation of bacteria from vascular lesions . This review summarises the present understanding of the role of bacterial infection for development or progression of coronary artery disease.

Wien Med Wochenschr, 2001, 151(24), 590 - 3
{Chlamydia pneumoniae--chronic infection and atherosclerosis}; Tiran B; Recent evidence suggests that common chronic infections may contribute to the initiation and/or progression of atherosclerosis . Infection of the vascular wall with Chlamydia pneumoniae, a gramnegative bacterium, has been linked with coronary heart disease, myocardial infarction and stroke in epidemiological studies and in pathological studies using immunohistochemistry and electron microscopy . In addition striking evidence for an active role of Chlamydia pneumoniae in atherogenesis has been provided in animal models and from preliminary data of intervention trials . Although these observations strongly indicate an involvement of Chlamydia pneumoniae in the pathogenesis of atherosclerosis, a causal relationship has not been established yet . In the last years several interesting papers have dealt with the molecular mechanisms how an infection with Chlamydia pneumoniae affects the vascular wall to initiate or facilitate vascular dysfunction.

World J Gastroenterol, 1998 Jun, 4(3), 249 - 251
Serum IgG response to differentiated antigens of Helicobacter pylori; Hua JS et al.; AIM:To detect antibodies against Helicobacter pylori spiral and coccoid antigens in human sera.METHODS:Blood samples were collected from 278 patients with gastric diseases . A 3-day-old culture of H . pylori on chocolate blood agar was used to providespiral form . Synchronous coccoids were cultured in (BHY) (brain heart infusion supplemented with 10% horse serum and 0.4% yeast extract) medium in a chemostat.Antigens from spiral and coccoid form were prepared using acid glycine extraction.Enzyme-linked immunosorbent assay (ELISA) was performed to detect serum IgG anti-bodies against spiral and coccoid forms of H . pylori.RESULTS:Seroprevalence of H . pylori infection was higher in patients with gastric ulcer (79%) and gastric cancer (83%) than those with non-ulcer dyspepsia (NUD)(44%) and other diseases (45%) (P <0.05) . IgG antibodies against spiral and coccoid antigens were detected in 50.7% (141/278) and 49.6% (138/278), respectively.CONCLUSION:The spiral and coccoid forms of H.pylori coexist in patients infected with the bacterium.

Analyst, 2001 Dec, 126(12), 2153 - 8
Sequences of E . coli O157:H7 detected by a PCR-acoustic wave sensor combination; Deisingh AK et al.; In this paper, we report on the amplification by polymerase chain reaction (PCR) of a 509 base sequence unique to E . coli O157:H7 . Immobilization of a probe for the bacterium on an acoustic wave sensor by the biotin-neutravidin interaction was employed to detect the on-line hybridization of the sequence with the sample obtained from PCR . The limit of detection was found to be 10(-8) M, which suggests that the method may be useful for detecting the organism in food, water and clinical samples . Finally, this approach lays the groundwork for incorporating the method into an integrated system for rapid PCR-based DNA analysis.

Parasitology, 2002 Jan, 124(Pt 1), 87 - 95
Does fertilization in the filarial nematode Dirofilaria immitis occur through endocytosis of spermatozoa?
Sacchi L, Corona S, Casiraghi M, Bandi C.
Information on the ultrastructural details of fertilization in filarial nematodes are still unavailable . Here we report new data on this process in Dirofilaria immits, the heartworm of dogs and cats . Electron microscopy allowed us to observe oocytes engulfing spermatozoa through an endocytosis-like process . We also observed spermatozoa inside the oocytes which still possessed their plasma membrane and which were clearly enveloped by a further membrane, likely derived from the endocytosis process . At this stage, at the interface between the sperm membrane and the endocytotic membrane (vacuolar space), we observed flocculent material in the proximity of the membranous organelles (MOs) of the sperm . In the proximity of the MOs, we also observed the enlargement of the vacuolar space . Other images showed the dissolution of the sperm membrane, and the release of nuclear masses and organelles in the egg cytoplasm . We did not observe the fusion of lysosomes to the endocytotic vacuoles . In addition, the lysis of the sperm organelles has never been observed inside the vacuoles containing the whole sperm . Thus we suggest that the degradation of the endocytotic and sperm plasma membranes is determined by material released by the MOs . Since we did not observe the entry of sperm into the oocytes by other mechanisms, we also suggest that endocytosis is the normal process used by the spermatozoon to get into the egg cytoplasm in D . immitis . Finally, during our observations of the seminal receptacle we did not observe any structure in the spermatozoa which could be interpreted as an intracellular bacterium . This is consistent with previous results indicating that the bacterium Wolbachia in filarial nematodes is not transmitted through the sperm.

Environ Sci Technol, 2002 Jan 1, 36(1), 16 - 20
Iron(II,III) hydroxycarbonate green rust formation and stabilization from lepidocrocite bioreduction; Ona-Nguema G et al.; Bioreduction of the well-crystallized ferric oxyhydroxide gamma-FeOOH lepidocrocite was investigated in batch cultures using Shewanella putrefaciens bacterium (strain CIP 8040) at initial pH 7.5 in bicarbonate buffer . The cultures were performed with formate as electron donor without phosphate, in the presence or absence of anthraquinone-2,6-disulfonate (AQDS) as electron shuttle . During lepidocrocite reduction, the iron(II,III) hydroxycarbonate green rust GR(CO32-) was characterized by X-ray diffraction, transmission electron microscopy, and transmission Mossbauer spectroscopy . The AQDS accelerated the kinetics of GR formation . GR was the major end product when bacterial reduction was not stopped by lack of electron donor, and between 55 and 86% of the iron from gamma-FeOOH precipitated in GR(CO32-) . However, when the bacterial reduction was stopped by freezing/thawing or the electron donor was exhausted, the large quantity of remaining lepidocrocite induced a transformation of GR into magnetite . This confirms that GR is metastable with respect to magnetite in the presence of gamma-FeOOH.

Med Electron Microsc, 1999 May, 32(1), 62 - 65
A new method for preparing electron microscopic specimens of Helicobacter pylori; Kai J et al.; Abstract A new method for preparing electron microscopic specimens of Helicobacter pylori was developed and used to examine the ultrastructure of this bacterium . We have also investigated the morphological changes in the bacterium when exposed to amoxicillin using our new method . Bacterial specimens for electron microscopy are usually prepared by collecting the bacteria by centrifugation during the fixation and dehydration processes . In our new method the bacteria are filtered through and adsorbed onto a filter before fixation, and the entire filter containing the adhered bacteria is fixed and dehydrated . Using this method we were able to obtain electron photomicrographs in which the external appearances or internal structures of the bacteria were well conserved . The advantages of this method are that it uses only a small amount of bacterial suspension, shortens the time required for the dehydration procedure, and keeps the artifacts to the minimum . Amoxicillin treatment resulted in coccoid form with blebs in the bacterial surface and the appearance of vacuoles, granules, and an area of low electron density in the cytoplasm at one and four minimum inhibitory concentrations . These changes were consistent with results previously reported in the literature.

Protein Eng, 2001 Dec, 14(12), 975 - 82
Heat labile ribonuclease HI from a psychrotrophic bacterium: gene cloning, characterization and site-directed mutagenesis; Ohtani N et al.; The rnhA gene encoding RNase HI from a psychrotrophic bacterium, Shewanella sp . SIB1, was cloned, sequenced and overexpressed in an rnh mutant strain of Escherichia coli . SIB1 RNase HI is composed of 157 amino acid residues and shows 63% amino acid sequence identity to E.coli RNase HI . Upon induction, the recombinant protein accumulated in the cells in an insoluble form . This protein was solubilized and purified in the presence of 7 M urea and refolded by removing urea . Determination of the enzymatic activity using M13 DNA-RNA hybrid as a substrate revealed that the enzymatic properties of SIB1 RNase HI, such as divalent cation requirement, pH optimum and cleavage mode of a substrate, are similar to those of E.coli RNase HI . However, SIB1 RNase HI was much less stable than E.coli RNase HI and the temperature (T(1/2)) at which the enzyme loses half of its activity upon incubation for 10 min was approximately 25 degrees C for SIB1 RNase HI and approximately 60 degrees C for E.coli RNase HI . The optimum temperature for the SIB1 RNase HI activity was also shifted downward by 20 degrees C compared with that of E.coli RNase HI . Nevertheless, SIB1 RNase HI was less active than E.coli RNase HI even at low temperatures . The specific activity determined at 10 degrees C was 0.29 units/mg for SIB1 RNase HI and 1.3 units/mg for E.coli RNase HI . Site-directed mutagenesis studies suggest that the amino acid substitution in the middle of the alphaI-helix (Pro52 for SIB1 RNase HI and Ala52 for E.coli RNase HI) partly accounts for the difference in the stability and activity between SIB1 and E.coli RNases HI.

Biotechnol Bioeng, 2002 Mar 20, 77(6), 614 - 8
Preparation of luciferase-bacterial magnetic particle complex by artificial integration of MagA-luciferase fusion protein into the bacterial magnetic particle membrane; Matsunaga T et al.; MagA is an iron-translocating protein found in the membranes of magnetic bacterium . Luciferase-bacterial magnetic particle (BMP) complexes were prepared by artificially inserting MagA-luciferase fusion proteins into the membranes of BMPs from Magnetospirillum magneticum strain AMB-1 . Fusion proteins were from recombinant Escherichia coli membranes . MagA-Luc fusion proteins were integrated by sonication in vitro . Successful integration of fusion proteins was confirmed by luciferase luminescence on BMPs . Maximum luminescence was obtained after sonication for 3 min with a solution containing 300 mM NaCl, and is 18 times higher compared with recombinant Luc-BMPs generated using previously reported gene fusion techniques .

Acta Crystallogr D Biol Crystallogr, 2002 Feb, 58(Pt 2), 292 - 5 Epub 2002 Jan 24.
Crystallization and preliminary X-ray diffraction analysis of a novel pectate lyase from Azospirillum irakense; Novoa de Armas H et al.; The PelA gene from the N(2)-fixing plant-associated bacterium Azospirillum irakense encodes a pectate lyase . Analysis of the corresponding amino-acid sequence revealed no homology to other bacterial, plant and fungal pectinases of known published structure, resulting in the classification of the enzyme in a new pectate lyase family . The A . irakense PelA has been crystallized using the hanging-drop vapour-diffusion method at 277 K . The crystals are hexagonal, with unit-cell parameters a = b = 85.55, c = 230.13 A, gamma = 120 degrees, and belong to space group P6(5)22 or P6(1)22, having one molecule per asymmetric unit . Diffraction data to a resolution of 1.97 A were collected at synchrotron facilities, as well as a three-wavelength MAD data set from an Hg-derivative crystal to a resolution of 2.6 A.

J Exp Bot, 2002 Feb, 53(367), 387 - 9
Expression of a bean acid phosphatase cDNA is correlated with disease resistance; Jakobek JL et al.; A cDNA clone, designated Hra28 (for hypersensitive reaction associated), was identified corresponding to an RNA transcript that accumulates in bean during the hypersensitive reaction . The Hra28 cDNA is 1084 nucleotides in length and is predicted to encode an acid phosphatase of 264 amino acids . Northern analysis demonstrated that the Hra28 transcript accumulated differentially in response to bacteria which induce a hypersensitive response (HR), a bacterium which causes disease, and a Hrp(-) mutant which does not elicit an HR or cause disease . In contrast, the Hra28 transcript did not accumulate in response to wounding . Thus, the Hra28 gene is induced by multiple stimuli and appears to be regulated in a complex manner.

J Bacteriol, 2002 Feb, 184(4), 1089 - 94
Microviridae, a family divided: isolation, characterization, and genome sequence of phiMH2K, a bacteriophage of the obligate intracellular parasitic bacterium Bdellovibrio bacteriovorus; Brentlinger KL et al.; A novel single-stranded DNA phage, phiMH2K, of Bdellovibrio bacteriovorus was isolated, characterized, and sequenced . This phage is a member of the Microviridae, a family typified by bacteriophage phiX174 . Although B . bacteriovorus and Escherichia coli are both classified as proteobacteria, phiMH2K is only distantly related to phiX174 . Instead, phiMH2K exhibits an extremely close relationship to the Microviridae of Chlamydia in both genome organization and encoded proteins . Unlike the double-stranded DNA bacteriophages, for which a wide spectrum of diversity has been observed, the single-stranded icosahedral bacteriophages appear to fall into two distinct subfamilies . These observations suggest that the mechanisms driving single-stranded DNA bacteriophage evolution are inherently different from those driving the evolution of the double-stranded bacteriophages.

J Bacteriol, 2002 Feb, 184(4), 913 - 8
A bacterial signal transduction system controls genetic exchange and motility; Lang AS et al.; The bacterium Rhodobacter capsulatus is capable of an unusual form of genetic exchange, mediated by a transducing bacteriophage-like particle called the gene transfer agent (GTA) . GTA production by R . capsulatus is controlled at the level of transcription by a cellular two-component signal transduction system that includes a putative histidine kinase (CckA) and response regulator (CtrA) . We found that, in addition to regulating genetic exchange by R . capsulatus, this signal transduction system controls motility . As with the regulation of GTA production, the control of motility by CckA and CtrA occurs through modulation of gene transcription . Disruptions of the cckA and ctrA genes resulted in a loss of class II, class III, and class IV flagellar gene transcripts, suggesting that cckA and ctrA function in motility as class I flagellar genes . We also found that, analogous to the GTA genes, transcription of R . capsulatus flagellar genes appears to be growth phase dependent: class II flagellar gene transcripts are maximal in the mid-log phase of the culture growth cycle, whereas class III gene transcripts are maximal in the late-log phase of growth . We speculate that coordinate regulation of motility and GTA-mediated genetic exchange in R . capsulatus exists because these two processes are complementary mechanisms for cells to cope with unfavorable conditions in natural environments.

J Inorg Biochem, 2002 Jan 15, 88(2), 223 - 7
Cell surface display of synthetic phytochelatins using ice nucleation protein for enhanced heavy metal bioaccumulation; Bae W et al.; Synthetic phytochelatins (ECs) composed of (Glu-Cys)nGly are protein analogs of phytochelatin that exhibit improved metal-binding capacity over metallothioneins (MTs) . Expression of EC20 on the surface of E . coli using the Lpp-OmpA anchor resulted in improved bioaccumulation of cadmium and mercury, providing a new method for treating heavy metal contamination . To further improve the whole-cell accumulation of heavy metals, EC20 was expressed on the surface of Moraxella sp., a bacterium known to survive in contaminated environments, using the truncated ice nucleation protein (INPNC) anchor . Production of EC20 was approximately three-fold higher in Moraxella sp . than E . coli . As a consequence, the mercury-binding capacity of the recombinant Moraxella sp . was increased by more than 10-fold . Owing to the very high level of surface expression, the use of Moraxella sp . and INPNC anchor may prove to be useful for the remediation of other environmental contaminants.

J Immunol, 2002 Feb 1, 168(3), 1328 - 37
Induction of TNF in human alveolar macrophages as a potential evasion mechanism of virulent Mycobacterium tuberculosis; Engele M et al.; The ability of macrophages to release cytokines is crucial to the host response to intracellular infection . In particular, macrophage-derived TNF plays an important role in the host response to infection with the intracellular pathogen Mycobacterium tuberculosis . In mice, TNF is indispensable for the formation of tuberculous granulomas, which serve to demarcate the virulent bacterium . TNF is also implicated in many of the immunopathological features of tuberculosis . To investigate the role of TNF in the local immune response, we infected human alveolar macrophages with virulent and attenuated mycobacteria . Infection with virulent strains induced the secretion of significantly higher levels of bioactive TNF than attenuated strains correlating with their ability to multiply intracellularly . Treatment of infected macrophages with neutralizing anti-TNF Abs reduced the growth rate of intracellular bacteria, whereas bacterial replication was augmented by addition of exogenous TNF . Infected and uninfected macrophages contributed to cytokine production as determined by double-staining of M . tuberculosis and intracellular TNF . The induction of TNF by human alveolar macrophages at the site of infection permits the multiplication of intracellular bacteria and may therefore present an evasion mechanism of human pathogens.

Phys Rev Lett . 2001 Dec 31;87(27 Pt 1):278102 . Epub 2001 Dec 10.
Dynamic compartmentalization of bacteria: accurate division in E . coli; Howard M et al.; Positioning of the midcell division plane within the bacterium E . coli is controlled by the min system of proteins: MinC, MinD, and MinE . These proteins coherently oscillate from end to end of the bacterium . We present a reaction-diffusion model describing the diffusion of min proteins along the bacterium and their transfer between the cytoplasmic membrane and cytoplasm . Our model spontaneously generates protein oscillations in good agreement with experiments . We explore the oscillation stability, frequency, and wavelength as a function of protein concentration and bacterial length.

Arch Microbiol, 2001 Dec, 177(1), 117 - 21 Epub 2001 Oct 24.
A novel pmoA lineage represented by the acidophilic methanotrophic bacterium Methylocapsa acidiphila {correction of acidophila} B2; Dedysh SN et al.; A fragment of the functional gene pmoA, which encodes the active-site polypeptide of particulate methane monooxygenase (pMMO), was PCR-amplified from DNA of the recently described acidophilic methanotrophic bacterium Methylocapsa acidiphila {corrected} B2 . This methanotroph was isolated from an acidic Sphagnum peat bog and possesses a novel type III arrangement of intracytoplasmic membranes . Comparative sequence analysis revealed that the inferred peptide sequence of pmoA of Methylocapsa acidiphila {corrected} B2 belongs to a novel PmoA lineage . This lineage was only distantly related to the PmoA sequence cluster of type II methanotrophs, with identity values between 69.5% and 72% . The identity values between the PmoA of Methylocapsa acidiphila {corrected} B2 and PmoA sequences of type I methanotrophs ranged from 55.5 to 68% . However, the PmoA of this acidophilic methanotroph was more closely affiliated with the inferred peptide sequences of pmoA clones retrieved from various acidic upland soils showing atmospheric methane consumption . The PmoA identity values with these clones were 79.5-81% . Nonetheless, the apparent affinity for methane exhibited by Methylocapsa acidiphila {corrected} B2 was 1-2 microM, which is similar to values measured in other methanotrophic bacteria . This finding suggests that the pMMO of Methylocapsa acidiphila {corrected} B2 is not a novel enzyme specialized to have a high affinity for methane and that apparent "high-affinity" methane uptake is either the result of particular culture conditions or is performed by another pMMO type.

Infect Immun, 2002 Feb, 70(2), 889 - 98
Rapid activation of protein tyrosine kinase and phospholipase C-gamma2 and increase in cytosolic free calcium are required by Ehrlichia chaffeensis for internalization and growth in THP-1 cells; Lin M et al.; Ehrlichia chaffeensis, a bacterium that cannot survive outside the eukaryotic cell, proliferates exclusively in human monocytes and macrophages . In this study, signaling events required for ehrlichial infection of human monocytic cell line THP-1 were characterized . Entry and proliferation of E . chaffeensis in THP-1 cells were significantly blocked by various inhibitors that can regulate calcium signaling, including 8-(diethylamino)octyl-3,4,5-trimethoxybenzoate and 2-aminoethoxydiphenyl borate (intracellular calcium mobilization inhibitors), verapamil and 1-{beta-{3-(4-methoxyphenyl)propyl}-4-methoxyphenethyl}-1H-imidazole (SKF-96365) (calcium channel inhibitors), neomycin and 1-(6-{{17beta-3-methoxyestra-1,3,5(10)-trien-17-yl}amino}hexyl)-1H-pyrrole-2,5-dione (U-73122) (phospholipase C {PLC} inhibitors), monodansylcadaverine (a transglutaminase {TGase} inhibitor), and genistein (a protein tyrosine kinase {PTK} inhibitor) . Addition of E . chaffeensis resulted in rapid increases in the level of inositol 1,4,5-trisphosphate (IP(3)) and the level of cytosolic free calcium ({Ca(2+)}(i)) in THP-1 cells, which were prevented by pretreatment of THP-1 cells with inhibitors of TGase, PTK, and PLC . E . chaffeensis induced rapid tyrosine phosphorylation of PLC-gamma2, and the presence of a PLC-gamma2 antisense oligonucleotide in THP-1 cells significantly blocked ehrlichial infection . Furthermore, tyrosine-phosphorylated proteins and PLC-gamma2 were colocalized with ehrlichial inclusions, as determined by double-immunofluorescence labeling . The heat-sensitive component of viable E . chaffeensis cells was essential for these signaling events . E . chaffeensis, therefore, can recruit interacting signal-transducing molecules and induce the following signaling events required for the establishment of infection in host cells: protein cross-linking by TGase, tyrosine phosphorylation, PLC-gamma2 activation, IP(3) production, and an increase in {Ca(2+)}(i).

Infect Immun, 2002 Feb, 70(2), 491 - 7
Differential binding of host complement inhibitor factor H by Borrelia burgdorferi Erp surface proteins: a possible mechanism underlying the expansive host range of Lyme disease spirochetes; Stevenson B et al.; The Lyme disease spirochete, Borrelia burgdorferi, is capable of infecting a wide variety of vertebrates . This broad host range implies that B . burgdorferi possesses the ability to contravene the immune defenses of many potential hosts . B . burgdorferi produces multiple different Erp proteins on its outer membrane during mammalian infection . It was reported previously that one Erp protein can bind human factor H (J . Hellwage, T . Meri, T . Heikkila, A . Alitalo, J . Panelius, P . Lahdenne, I . J . T . Seppala, and S . Meri, J . Biol . Chem . 276:8427-8435, 2001) . In this paper we report that the ability to bind the complement inhibitor factor H is a general characteristic of Erp proteins . Furthermore, each Erp protein exhibits different relative affinities for the complement inhibitors of various potential animal hosts . The data suggest that the presence of multiple Erp proteins on the surface can allow a single B . burgdorferi bacterium to resist complement-mediated killing in any of the wide range of potential hosts that it might infect . Thus, Erp proteins likely contribute to the persistence of B . burgdorferi in nature and to the ability of this bacterium to cause Lyme disease in humans and other animals.

J Am Osteopath Assoc, 2001 Dec, 101(12 Suppl Pt 1), S1 - 6
Role of infection in Alzheimer's disease; Balin BJ et al.; Alzheimer's disease (AD) is a chronic condition in which inflammation has been shown to contribute to neurodegeneration . Current thinking suggests that deposition of beta-amyloid in the brain promotes inflammation resulting in neuronal damage/death . Alternatively, our data suggest that chronic inflammation observed in late-onset sporadic AD may be stimulated by infection with the obligate, intracellular bacterium, Chlamydia pneumoniae . Our results indicate that C . pneumoniae is found in high frequency in glial cells in areas of neuropathology within the brains of patients with AD . Based on our evidence, nervous system infection with C . pneumoniae should be considered a risk factor for sporadic AD.

J Clin Lab Anal, 2001, 15(6), 301 - 7
Laboratory tests for the evaluation of Helicobacter pylori infections; Nakamura RM; Helicobacter pylori is a highly motile bacterium with multiple unipolar flagella, and it produces the urease enzyme . The flagella and urease are the virulence factors of H . pylori . H . pylori often establishes a chronic infection in the stomach that may lead to gastric and duodenal ulcers, gastric cancers, gastric lymphomas, and other gastrointestinal diseases . There are several different invasive and noninvasive clinical laboratory tests for H . pylori . Laboratory testing is not indicated in asymptomatic patients and should be considered only if treatment of H . pylori infection is planned . Invasive tests for H . pylori, such as tissue histology, culture, and rapid urease tests, are used if an endoscopy is performed on the patient . The noninvasive tests for H . pylori, such as enzyme antibody and urea breath tests, are recommended in patients whose symptoms do not warrant endoscopy . The urea breath test is very useful and is recommended to evaluate effectiveness in the eradication and treatment of H . pylori infections . Nucleic acid tests can complement other diagnostic procedures, and are useful in evaluating fixed biopsy tissue, environmental samples, gastric juices, oral secretions, and stool samples .

J Cell Sci, 2001 Dec, 114(Pt 24), 4637 - 50
How the parasitic bacterium Legionella pneumophila modifies its phagosome and transforms it into rough ER: implications for conversion of plasma membrane to the ER membrane; Tilney LG et al.; Within five minutes of macrophage infection by Legionella pneumophila, the bacterium responsible for Legionnaires' disease, elements of the rough endoplasmic reticulum (RER) and mitochondria attach to the surface of the bacteria-enclosed phagosome . Connecting these abutting membranes are tiny hairs, which are frequently periodic like the rungs of a ladder . These connections are stable and of high affinity - phagosomes from infected macrophages remain connected to the ER and mitochondria (as they were in situ) even after infected macrophages are homogenized . Thin sections through the plasma and phagosomal membranes show that the phagosomal membrane is thicker (72+/-2 A) than the ER and mitochondrial membranes (60+/-2 A), presumably owing to the lack of cholesterol, sphingolipids and glycolipids in the ER . Interestingly, within 15 minutes of infection, the phagosomal membrane changes thickness to resemble that of the attached ER vesicles . Only later (e.g . after six hours) does the ER-phagosome association become less frequent . Instead ribosomes stud the former phagosomal membrane and L . pneumophila reside directly in the rough ER . Examination of phagosomes of various L . pneumophila mutants suggests that this membrane conversion is a four-stage process used by L . pneumophila to establish itself in the RER and to survive intracellularly . But what is particularly interesting is that L . pneumophila is exploiting a poorly characterized naturally occurring cellular process.

Microbiol Immunol, 2001, 45(11), 729 - 36
Mycobacterium leprae and leprosy: a compendium; Sasaki S et al.; Leprosy is a chronic infectious disease caused by Mycobacterium leprae, which was discovered by G.H.A . Hansen in 1873 . M . leprae is an exceptional bacterium because of its long generation time and no growth in artificial media . Entire sequencing of the bacterial genome revealed numerous pseudogenes (inactive reading frames with functional counterparts in M . tuberculosis) which might be responsible for the very limited metabolic activity of M . leprae . The clinical demonstration of the disease is determined by the quality of host immune response . Th1-type immune response helps to kill the bacteria, but hosts are encroached upon when Th2-type response is predominant . The bacteria have affinity to the peripheral nerves and are likely to cause neuropathy . M . leprae/laminin-alpha2 complexes bind to alpha/beta dystroglycan complexes expressed on the Schwann cell surface . WHO recommends a chemotherapy protocol {multidrug therapy (MDT)} which effectively controls the disease and contributes to the global elimination program . Leprosy has been stigmatized throughout history, and recent topics regarding the disease in Japan are also discussed.

J Bacteriol, 2002 Feb, 184(3), 746 - 53
Rubrivivax gelatinosus acsF (previously orf358) codes for a conserved, putative binuclear-iron-cluster-containing protein involved in aerobic oxidative cyclization of Mg-protoporphyrin IX monomethylester; Pinta V et al.; This study describes the characterization of orf358, an open reading frame of previously unidentified function, in the purple bacterium Rubrivivax gelatinosus . A strain in which orf358 was disrupted exhibited a phenotype similar to the wild type under photosynthesis or low-aeration respiratory growth conditions . In contrast, under highly aerated respiratory growth conditions, the wild type still produced bacteriochlorophyll a (Bchl a), while the disrupted strain accumulated a compound that had the same absorption and fluorescence emission spectra as Mg-protoporphyrin but was less polar, suggesting that it was Mg-protoporphyrin monomethylester (MgPMe) . These data indicated a blockage in Bchl a synthesis at the oxidative cyclization stage and implied the coexistence of two different mechanisms for MgPMe cyclization in R . gelatinosus, an anaerobic mechanism active under photosynthesis or low oxygenation and an aerobic mechanism active under high-oxygenation growth conditions . Based on these results as well as on sequence analysis indicating the presence of conserved putative binuclear-iron-cluster binding motifs, the designation of orf358 as acsF (for aerobic cyclization system Fe-containing subunit) is proposed . Several homologs of AcsF were found in a wide range of photosynthetic organisms, including Chlamydonomas reinhardtii Crd1 and Pharbitis nil PNZIP, suggesting that this aerobic oxidative cyclization mechanism is conserved from bacteria to plants.

Genome Biol . 2001;2(12):REVIEWS1032 . Epub 2001 Nov 22.
Genome interdependence in insect-bacterium symbioses; Zientz E et al.; Symbioses between unicellular and multicellular organisms have contributed significantly to the evolution of life on Earth . As exemplified by several studies of bacterium-insect symbioses, modern genomic techniques are providing exciting new information about the molecular basis and the biological roles of these complex relationships, revealing for instance that symbionts have lost many genes for functions that are provided by the host, but that they can provide amino acids that the host cannot synthesize.

J Theor Biol, 2002 Jan 7, 214(1), 105 - 34
Putting intentions into cell biochemistry: an artificial intelligence perspective; Jonker CM et al.; The living cell exists by virtue of thousands of nonlinearly interacting processes . This complexity greatly impedes its understanding . The standard approach to the calculation of the behaviour of the living cell, or part thereof, integrates all the rate equations of the individual processes . If successful extremely intensive calculations often lead the calculation of coherent, apparently simple, cellular "decisions" taken in response to a signal: the complexity of the behavior of the cell is often smaller than it might have been . The "decisions" correspond to the activation of entire functional units of molecular processes, rather than individual ones . The limited complexity of signal and response suggests that there might be a simpler way to model at least some important aspects of cell function . In the field of Artificial Intelligence, such simpler modelling methods for complex systems have been developed . In this paper, it is shown how the Artificial Intelligence description method for deliberative agents functioning on the basis of beliefs, desires and intentions as known in Artificial Intelligence, can be used successfully to describe essential aspects of cellular regulation . This is demonstrated for catabolite repression and substrate induction phenomena in the bacterium Escherichia coli . The method becomes highly efficient when the computation is automated in a Prolog implementation . By defining in a qualitative way the food supply of the bacterium, the make-up of its catabolic pathways is readily calculated for cases that are sufficiently complex to make the traditional human reasoning tedious and error prone .

Avian Dis, 2001 Oct-Dec, 45(4), 828 - 43
Viral and bacterial agents associated with experimental transmission of infectious proventriculitis of broiler chickens; Huff GR et al.; Proventriculitis of broilers can be reproduced by oral inoculation of day-old chicks with a proventricular homogenate from affected 3-wk-old broilers . The objective of the following studies was to isolate from this homogenate viral and bacterial isolates that could produce proventriculitis . A monoclonal antibody to infectious bursal disease virus (IBDV) was used to precipitate virus from the homogenate . A primary chicken digestive tract cell culture system was also used to isolate virus from a 0.2-microm filtrate of the homogenate, and a bacterium was also isolated from the homogenate . In trial 1, day-old birds were orally inoculated with either proventriculus homogenate or monoclonal antibody immunoprecipitated IBDV (MAB-IBDV) . At 4, 7, 14, and 21 days postinfection (PI), 12 birds from each treatment group were subjected to necropsy . In trial 2, day-old birds were orally inoculated with either infectious proventriculus homogenate, suspect virus isolated in cell culture and propagated in embryo livers and spleens, or a bacterial isolate . Twelve birds from each treatment were subjected to necropsy at days 7, 14, 21, and 28 PI . In trial 3, treatments were maintained in negative pressure isolation chambers, and an additional treatment included virus plus bacterial isolate . Twenty-four birds from each treatment were subjected to necropsy at day 21 PI . In trial 1, infectious homogenate decreased body weight and relative gizzard weights at 4, 7, 14, and 21 days PI . Proventriculus relative weight was increased at days 7, 14, and 21 PI, and proventriculus lesion scores were increased at days 14 and 21 PI . Bursa/spleen weight ratios were decreased at day 14, and feed conversion was increased at days 4 and 21 . The MAB-IBDV treatment decreased proventriculus and gizzard relative weights at day 4 PI, increased proventriculus lesion scores and bursa/spleen weight ratios at day 14, and decreased heterophil/lymphocyte ratios at day 21 . In trial 2, all infected birds had significantly higher mean relative proventriculus weights at 21 days PI and had higher 4-wk mean proventriculus scores as compared with both control groups . In trial 3, birds treated with homogenate and birds treated with both suspect virus and the bacterial isolate had significantly higher proventriculus lesion scores; higher relative weights of proventriculus, gizzard, liver, and heart; lower body weights; and lower relative bursa weights compared with the saline control group . These studies suggest that infectious proventriculitis has a complex etiology involving both viral and bacterial infection.

Biol Sci Space, 1996 Sep, 10(2), 97 - 101
Recovery of Deinococcus radiodurans from radiation damage was enhanced under microgravity; Kobayashi Y et al.; Effect of microgravity on recovery of bacterial cells from radiation damage was examined on the IML-2 mission in 1994 using extremely radioresistant bacterium Deinococcus radiodurans . The cells were lyophilized and exposed to 60Co gamma-rays with doses 2 to 12 kGy before the space flight . At the end of the mission, the cells were mixed on board with liquid nutrient medium to allow the cells to start recovery process from the radiation damage . Afterwards the cells were stored at 4 degrees C until landing . The influence of cosmic radiation was negligible, because total absorbed dose of space radiation measured during the mission was less than 2 mGy and this bacterium does not decrease its viability after both gamma-rays and high-LET heavy charged particles irradiation with doses up to 5 kGy . The survival of the cells incubated in space increased significantly compared with the ground controls, suggesting that the recovery of this bacterium from radiation damage was enhanced under microgravity.

Mikrobiologiia, 2001 Nov-Dec, 70(6), 765 - 75
{Ultrastructural organization of cytoplasmatic membrane of Anaerobacter polyendosporus studied by electron microscopic cryofractography}; Duda VI et al.; Anaerobacter polyendosporus cells do not have typical mesosomes . However, the analysis of this anaerobic multispore bacterium by electron microscopic cryofractography showed that its cytoplasmic membrane contains specific intramembrane structures in the form of flat lamellar inverted lipid membranes tenths of nanometers to several microns in size . It was found that these structures are located in the hydrophobic interior between the outer and inner leaflets of the cytoplasmic membrane and do not contain intramembrane particles that are commonly present on freeze-fracture replicas . The flat inverted lipid membranes were revealed in bacterial cells cultivated under normal growth conditions, indicating the existence of a complex-type compartmentalization in biological membranes, which manifests itself in the formation of intramembrane compartments having the appearance of vesicles and inverted lipid membranes.

Microbiology, 2002 Jan, 148(Pt 1), 267 - 76
Quantitative speciation of sulfur in bacterial sulfur globules: X-ray absorption spectroscopy reveals at least three different species of sulfur; Prange A et al.; X-ray absorption near edge structure (XANES) spectroscopy at the sulfur K-edge was applied to probe the speciation of sulfur of metabolically different sulfur-accumulating bacteria in situ . Fitting the spectra using a least-square fitting routine XANES reveals at least three different forms of sulfur in bacterial sulfur globules . Cyclooctasulfur dominates in the sulfur globules of Beggiatoa alba and the very recently described giant bacterium Thiomargarita namibiensis . A second type of sulfur globules is present in Acidithiobacillus ferrooxidans: here the sulfur occurs as polythionates . In contrast, in purple and green sulfur bacteria the sulfur mainly consists of sulfur chains, irrespective of whether it is accumulated in globules inside or outside the cells . These results indicate that the speciation of sulfur in the sulfur globules reflects the different ecological and physiological properties of different metabolic groups of bacteria.

Microb Pathog, 2002 Jan, 32(1), 1 - 13
Outer membrane-like vesicles secreted by Actinobacillus actinomycetemcomitans are enriched in leukotoxin; Kato S et al.; Actinobacillus actinomycetemcomitans is associated with early onset periodontal diseases and secretes membranous vesicles that appear to contain several virulence-associated proteins . However, the composition of these vesicles and the process leading to their secretion are not well defined . Electron micrographs of thin sectioned bacterial cells and purified vesicle preparations showed that vesicles are spherical lipid bilayers, 50-100 nm in diameter, that appear to form by budding from the outer membrane of the bacterium . Thin layer chromatography identified the predominant lipid components of vesicles as lipopolysaccharide, phosphatidylethanolamine and cardiolipin, similar to the main lipid constituents of the outer membrane . However, vesicles also contained minor lipids that were not detected in outer membrane samples . The major protein constituents of vesicles co-migrated with proteins in outer membrane extracts of A . actinomycetemcomitans, but the outer membrane preparations possessed polypeptides that were not detected in vesicles . Three vesicle proteins were identified; the heat-modifiable OmpA homologue of A . actinomycetemcomitans, a 28 kDa lipoprotein related to the major outer membrane lipoprotein of Mannheimia haemolytica and leukotoxin . Incubation of leukotoxin-sensitive human HL60 cells with vesicles from A . actinomycetemcomitans strains JP2 and 652 resulted in cell lysis, indicating that vesicle-associated leukotoxin is biologically active . Vesicles from the highly leukotoxic strain JP2 were five- to 10-fold more toxic than vesicles from the minimally leukotoxic 652 strain . Furthermore, the specific leukotoxic activity of JP2 vesicles was approximately four- to five-fold higher than isolated outer membrane preparations from JP2, suggesting that vesicles are enriched in leukotoxin . Together, these results suggest that the formation of A . actinomycetemcomitans vesicles occurs by a process that results in the enrichment of leukotoxin .

Clin Immunol, 2002 Jan, 102(1), 28 - 36
Chlamydia trachomatis infection inhibits airway eosinophilic inflammation induced by ragweed; Bilenki L et al.; While much progress has been achieved in controlling infectious diseases, there is a startling increase in the prevalence of allergic disorders in developed countries . Previous studies using experimental murine models of asthma have demonstrated that mycobacterial infections are capable of suppressing asthma-like reactions induced by ovalbumin (OVA) . Using a different intracellular bacterium, Chlamydia trachomatis mouse pneumonitis (MoPn), we examined the effect of infection on the development of allergic responses to a common natural airborne allergen, ragweed (RW) . The data showed that airway eosinophilia induced by ragweed sensitization/challenge was significantly reduced in MoPn-infected mice . MoPn-infected mice also exhibited significantly lower levels of allergen-driven Th2 cytokine production, namely IL-4, IL-5, IL-10, and IL-13, following ragweed exposure in comparison with those treated with ragweed only . Additionally, the production of eotaxin, a C-C chemokine for eosinophil chemoattraction following RW exposure, was significantly reduced in the lungs of MoPn-infected mice . However, MoPn infection did not reduce the levels of RW-specific IgE and IgG1 production in the sera, nor did it diminish the level of total serum IgE . These data provide evidence that the suppression of the allergic airway inflammation induced by a common environmental allergen is attainable through intracellular bacterial infection . (c)2001 Elsevier Science.

RNA, 2001 Dec, 7(12), 1702 - 7
Escherichia coli RNase M is a multiply altered form of RNase I; Subbarayan PR et al.; RNase M, an enzyme previously purified to homogeneity from Escherichia coli, was suggested to be the RNase responsible for mRNA degradation in this bacterium . Although related to the endoribonuclease, RNase I, its distinct properties led to the conclusion that RNase M was a second, low molecular mass, broad specificity endoribonuclease present in E . coli . However, based on sequence analysis, southern hybridization, and enzyme activity, we show that RNase M is, in fact, a multiply altered form of RNase I . In addition to three amino acid substitutions that confer the properties of RNase M on the mutated RNase I, the protein is synthesized from an rna gene that contains a UGA nonsense codon at position 5, apparently as a result of a low level of readthrough . We also suggest that RNase M is just one of several previously described endoribonuclease activities that are actually manifestations of RNase I.

Genetics, 2001 Dec, 159(4), 1415 - 22
On the mod resc model and the evolution of Wolbachia compatibility types; Charlat S et al.; Cytoplasmic incompatibility (CI) is induced by the endocellular bacterium Wolbachia . It results in an embryonic mortality occurring when infected males mate with uninfected females . The mechanism involved is currently unknown, but the mod resc model allows interpretation of all observations made so far . It postulates the existence of two bacterial functions: modification (mod) and rescue (resc) . The mod function acts in the males' germline, before Wolbachia are shed from maturing sperm . If sperm is affected by mod, zygote development will fail unless resc is expressed in the egg . Interestingly, CI is also observed in crosses between infected males and infected females when the two partners bear different Wolbachia strains, demonstrating that mod and resc interact in a specific manner: Two Wolbachia strains are compatible with each other only if they harbor the same compatibility type . Here we focus on the evolutionary process involved in the emergence of new compatibility types from ancestral ones . We argue that new compatibility types are likely to evolve under a wider range of conditions than previously thought, through a two-step process . First, new mod variants can arise by mutation and spread by drift . This is possible because mod is expressed in males and Wolbachia is transmitted by females . Second, once such a mod variant achieves a certain frequency, it can create the conditions for the deterministic invasion of a new resc variant, allowing the invasion of a new mod resc pair . Furthermore, we show that a stable polymorphism might be maintained in natural populations, allowing the long-term existence of "suicidal" Wolbachia strains.

Front Biosci, 2002 Jan 01, 7, d1 - 11
Entry into host cells by Legionella; Samrakandi MM et al.; Many respiratory diseases are caused by extracellular bacterial pathogens; however, two very important lung infections are due to intracellular pathogens, Legionnaires' disease and tuberculosis . Legionnaires' disease remains problematic due to our inability to predict where sporadic epidemics will occur and the speed at which the bacterium debilitates its victims . The development of better methods for prevention would greatly alleviate public concern and the economic impacts of eradication efforts where infections occur . Legionella, the causative agent of Legionnaires' disease, has been shown to replicate within eukaryotic cells both during disease and in the environment . During disease these bacteria are found primarily within macrophages, though they have the ability to enter and survive within a number of different mammalian cell types . In the environment Legionella replicate within free-living protozoa . Thus, the ability to enter into host cells successfully and efficiently is critical to the ability of Legionella to survive . The process by which Legionella gains access to the intracellular environment involves a number of steps; including, finding an appropriate host cell, adherence, signal transduction, entry and initial survival . Unless Legionella accomplishes each of these steps properly, few viable bacteria will be observed intracellularly and reduced intracellular replication may occur . However, the importance of each of these individual steps in the pathogenesis of Legionella is unclear . Herein we discuss the potential mechanisms of entry by Legionella into host cells, a critical early event in the production of Legionnaires' disease.

Biochim Biophys Acta, 2001 Dec 30, 1522(3), 158 - 66
Leaving group stabilization by metal ion coordination and hydrogen bond donation is an evolutionarily conserved feature of group I introns; Kuo LY et al.; To understand the behavior of group I introns on a biologically fundamental level, we must distinguish those traits that arise as the products of natural selection (selected traits) from those that arise as the products of neutral drift (non-selected traits) . In practice, this distinction relies on comparing the similarities and differences among widely divergent introns to identify conserved traits . Here we address whether the strategies used by the eukaryotic group I intron from the Tetrahymena ciliate to stabilize the leaving group during splicing are maintained in the group I intron from the widely divergent Azoarcus bacterium . A substrate analogue containing a 3'-phosphorothiolate linkage, in which a sulfur atom replaces the bridging 3'-oxygen atom of the scissile phosphate, reacts 20-fold slower in the Azoarcus reaction than the corresponding unmodified substrate in the presence of Mg(II) as the only divalent cation . However, Mn(II) relieves this negative effect such that the 3'-S-P bond cleaves 21-fold faster than does the 3'O-P bond . Other thiophilic divalent metal ions such as Co(II), Cd(II), and Zn(II) similarly support cleavage of the S-P bond . These results indicate that a metal ion directly coordinates to the leaving group in the transition state of the Azoarcus ribozyme reaction . Additionally, the 3'-sulfur substitution eliminates the approximately 10(3)-fold contribution of the adjacent 2'-OH to transition state stabilization . Considering that sulfur accepts hydrogen bonds weakly compared to oxygen, this result suggests that the 2'-OH contributes to catalysis by donating a hydrogen bond to the 3'-oxygen leaving group in the transition state, presumably acting in conjunction with the metal ion to stabilize the developing negative charge . These same catalytic strategies of metal ion coordination and hydrogen bond donation operate in the Tetrahymena ribozyme reaction, suggesting that these features of catalysis have been conserved during evolution and thus extend to all group I introns . The two ribozymes also exhibit quantitative differences in their response to 3'-sulfur substitution . The Azoarcus ribozyme binds and cleaves the phosphorothiolate substrate more efficiently relative to the natural substrate than the Tetrahymena ribozyme under the same conditions, suggesting that the Azoarcus ribozyme better accommodates the phosphorothiolate at the active site both in the ground state and in the transition state . These differences may reflect either a less tightly knit Azoarcus structure and/or spatial deviations between backbone atoms in the two ribozymes that arise during divergent evolution, analogous to the well-documented relationship between protein sequence and structure.

J Econ Entomol, 2001 Dec, 94(6), 1506 - 10
Seasonal flight activity of two Homalodisca species (Homoptera: Cicadellidae) that spread Xylella fastidiosa in southern California; Blua MJ et al.; Homalodisca coagulata (Say) and Homalodisca lacerta (Fowler) are vectors of a new bacterial disease of oleander in California known as oleander leaf scorch, induced by the bacterium Xylella fastidiosa . H . coagulata also has been implicated in the spread of the strain of X . fastidiosa that induces Pierce's disease of grapevines in California . We monitored the flight activity of H . coagulata and H . lacerta in oleander and citrus by using yellow sticky cards at three southern California locations where outbreaks of oleander leaf scorch have been documented, and where vector compliments are different . Areas sampled included a mesic coastal area (Irvine, CA) that supports predominantly H . coagulata and few H . lacerta, a dry inland location (Palm Desert, CA) that supports predominantly H . lacerta and few H . coagulata, and an intermediate area (Riverside, CA) supporting both Homalodisca species . From November 1996 to October 1999 peak catches of both Homalodisca species occurred during the midsummer at all locations . H . coagulata was trapped in greater numbers in citrus than in oleander at both the Riverside and the Irvine sites . Likewise, H . lacerta in Riverside was more associated with citrus than oleander, yet H . lacerta in Palm Desert was trapped in greater numbers in oleander than citrus.

J Virol, 2002 Feb, 76(3), 1285 - 92
Potent immunosuppressive activities of cytomegalovirus-encoded interleukin-10; Spencer JV et al.; Cytomegalovirus (CMV) has highly evolved mechanisms for avoiding detection by the host immune system . Recently, in the genomes of human and primate CMV, a novel gene comprising segments of noncontiguous open reading frames was identified and found to have limited predicted homology to endogenous cellular interleukin-10 (IL-10) . Here we investigate the biological activities of the CMV IL-10-like gene product and show it to possess potent immunosuppressive properties . Both purified bacterium-derived recombinant CMV IL-10 and CMV IL-10 expressed in supernatants of human cells were found to inhibit proliferation of mitogen-stimulated peripheral blood mononuclear cells (PBMCs), with specific activity comparable to that of recombinant human IL-10 . In addition, CMV IL-10 expressed from human cells inhibited cytokine synthesis, as treatment of stimulated PBMCs and monocytes with CMV IL-10 led to a marked decrease in production of proinflammatory cytokines . Finally, CMV IL-10 was observed to decrease cell surface expression of both major histocompatibility complex (MHC) class I and class II molecules, while conversely increasing expression of the nonclassical MHC allele HLA-G . These results demonstrate for the first time that CMV has a biologically active IL-10 homolog that may contribute to immune evasion during virus infection.

Appl Environ Microbiol, 2002 Jan, 68(1), 423 - 6
Characterization of a Sinorhizobium isolate and its extracellular polymer implicated in pollutant transport in soil; Janecka J et al.; A bacterium isolated from soil (designated 9702-M4) synthesizes an extracellular polymer that facilitates the transport of such hydrophobic pollutants as polynuclear aromatic hydrocarbons, as well as the toxic metals lead and cadmium in soil . Biolog analysis, growth rate determinations, and percent G+C content identify 9702-M4 as a strain of Sinorhizobium meliloti . Sequence analysis of a 16S rDNA fragment gives 9702-M4 a phylogenetic designation most closely related to Sinorhizobium fredii . The extracellular polymer of isolate 9702-M4 is composed of both an extracellular polysaccharide (EPS) and a rough lipopolysaccharide . The EPS component is composed mainly of 4-glucose linkages with monomers of galactose, mannose, and glucuronic acid and has pyruval and acetyl constituents . The lipid fraction and the negative charge associated with carbonyl groups of the exopolymer are thought to account for the binding of polynuclear aromatic hydrocarbons and cationic metals.

Appl Environ Microbiol, 2002 Jan, 68(1), 346 - 55
Molecular characterization of novel red green nonsulfur bacteria from five distinct hot spring communities in Yellowstone National Park; Boomer SM et al.; We characterized and compared five geographically isolated hot springs with distinct red-layer communities in Yellowstone National Park . Individual red-layer communities were observed to thrive in temperatures ranging from 35 to 60 degrees C and at pH 7 to 9 . All communities were dominated by red filamentous bacteria and contained bacteriochlorophyll a (Bchl a), suggesting that they represented novel green nonsulfur (GNS) bacteria . The in vivo absorption spectra of individual sites were different, with two sites showing unusual Bchl a protein absorption bands beyond 900 nm . We prepared and analyzed 16S rRNA libraries from all of these sites by using a combination of general bacterial primers and new GNS-specific primers described here . These studies confirmed the presence of novel GNS-like bacteria in all five communities . All GNS-like clones were most similar to Roseiflexus castenholzii, a red filamentous bacterium from Japan that also contains only Bchl a . Phylogenies constructed by using GNS-like clones from Yellowstone red-layer communities suggest the presence of a moderately diverse new "red" cluster within the GNS lineage . Within this cluster, at least two well-supported subclusters emerged: YRL-A was most similar to Roseiflexus and YRL-B appeared to be novel, containing no known isolates . While these patterns showed some site specificity, they did not correlate with observed Bchl a spectrum differences or obvious features of the habitat.

Appl Environ Microbiol, 2002 Jan, 68(1), 263 - 70
Identification and characterization of the gene cluster involved in chitin degradation in a marine bacterium, Alteromonas sp . strain O-7; Tsujibo H et al.; Alteromonas sp . strain O-7 secretes chitinase A (ChiA), chitinase B (ChiB), and chitinase C (ChiC) in the presence of chitin . A gene cluster involved in the chitinolytic system of the strain was cloned and sequenced upstream of and including the chiA gene . The gene cluster consisted of three different open reading frames organized in the order chiD, cbp1, and chiA . The chiD, cbp1, and chiA genes were closely linked and transcribed in the same direction . Sequence analysis indicated that Cbp1 (475 amino acids) was a chitin-binding protein composed of two discrete functional regions . ChiD (1,037 amino acids) showed sequence similarity to bacterial chitinases classified into family 18 of glycosyl hydrolases . The cbp1 and chiD genes were expressed in Escherichia coli, and the recombinant proteins were purified to homogeneity . The highest binding activities of Cbp1 and ChiD were observed when alpha-chitin was used as a substrate . Cbp1 and ChiD possessed a chitin-binding domain (ChtBD) belonging to ChtBD type 3 . ChiD rapidly hydrolyzed chitin oligosaccharides in sizes from trimers to hexamers, but not chitin . However, after prolonged incubation with large amounts of ChiD, the enzyme produced a small amount of (GlcNAc)(2) from chitin . The optimum temperature and pH of ChiD were 50 degrees C and 7.0, respectively.

J Chromatogr A, 2001 Dec 14, 938(1-2), 137 - 43
Simultaneous determination of delta-aminolevulinic acid, porphobilinogen, levulinic acid and glycine in culture broth by capillary electrophoresis; Kim JN et al.; Capillary electrophoretic simultaneous determination of a mixture containing delta-aminolevulinic acid, porphobilinogen, levulinic acid and glycine was investigated . With increases in the sodium tetraborate buffer concentration (5-70 mM), resolution of the four components was improved, but the migration time was increased . Alternatively, with increases in the applied voltage (5-22.5 kV), a shortened migration time was seen but this adversely affected resolution . The components were separated with high resolution by using a fused-silica capillary column (75 cm x 75 microm I.D.) filled with 30 mM sodium tetraborate buffer (pH 9.3-9.4) under the applied voltage of 20 kV (constant voltage mode) . When the established method was applied to the culture broth of Rhodopseudomonas sphaeroides, a photosynthetic bacterium, the four components mentioned above were separated with good resolution . Furthermore, the use of this method would provide a fast, sensitive and specific method for monitoring the administration of delta-aminolevulinic acid in photodynamic cancer therapy, for the measurement of delta-aminolevulinic acid dehydratase activity in erythrocytes, and for testing the delta-aminolevulinic acid assay and for impurities in drug formulation.

Microbiol Res, 2001, 156(4), 323 - 35
Degradation of poly (3-hydroxybutyrate) and its copolymer poly (3-hydroxybutyrate-co-3-hydroxyvalerate) by a marine Streptomyces sp . SNG9; Mabrouk MM et al.; A marine Streptomyces sp . SNG9 was characterized by its ability to utilize poly(3-hydroxybutyrate) (PHB) and its copolymer poly(3-hydroxybutyrate-co-3-hydroxyvalerate P (3HB-co-HV) . The bacterium grew efficiently in a simple mineral liquid medium enriched with 0.1 % poly(3-hydroxybutyrate) powder as the sole carbon source . Cells excreted PHB depolymerase and degraded the polymer particles to complete clarity in 4 days . The degradation activity was detectable by the formation of a clear zone around the colony (petri plates) or a clear depth under the colony (test tubes) . The expression of PHB depolymerase was repressed by the presence of simple soluble carbon sources . Bacterial degradation of the naturally occurring sheets of poly(3-hydroxybutyrate) and its copolymer poly(3-hydroxybutyrate-co-3-hydroxyvalerate) was observed by scanning electron microscopy (SEM) . Morphological alterations of the polymers sheets were evidence for bacterial hydrolysis.

Microbiol Res, 2001, 156(4), 311 - 5
Perchlorate and nitrate reductase activity in the perchlorate-respiring bacterium perclace; Giblin T et al.; The perchlorate (ClO4(-))-respiring organism, strain perclace, can grow using nitrate (NO3(-)) as a terminal electron acceptor . In resting cell suspensions, NO(-) grown cells reduced ClO4(-), and ClO4(-) grown cells reduced NO3(-) . Activity assays showed that nitrate reductase (NR) activity was 1.31 micromol min(-1) (mg protein)-1 in (ClO4)- grown cells, and perchlorate reductase (PR) activity was 4.24 micromol min(-1) (mg protein)(-1) in NO3(-) grown cells . PR activity was detected within the periplasmic space, with activities as high as 14 pmol min(-1) (mg protein)(-1) . The NR had a pH optimum of 9.0 while the PR had an optimum of 8.0 . This study suggests that separate terminal reductases are present in strain perclace to reduce NO3(-) and ClO4(-).

J Food Prot, 2001 Dec, 64(12), 2103 - 10
Crohn's disease and Mycobacterium avium subsp . paratuberculosis: current issues; Harris JE et al.; Crohn's disease is a chronic debilitating inflammatory bowel disease of unknown etiology . Proposed causes include bacterial or viral infection, diet or exposure to tobacco smoke, genetic abnormality, and immune dysfunction . The bacterium Mycobacterium avium subsp . paratuberculosis (Map) has received much research attention as a potential cause of the disease . Map causes Johne's disease in ruminants . The pathology of Johne's disease superficially resembles that of Crohn's disease in humans . Some researchers have shown evidence of Map in intestinal tissues of Crohn's disease patients . Studies are in progress to investigate the possibility that Map exists in milk from infected cows and survives pasteurization . This is a controversial subject with the potential for media attention and public outcry . We examined the current literature and concluded that insufficient evidence exists at this time to implicate any one factor, including Map in milk, as the definitive cause of Crohn's disease . The high degree of uncertainty in this issue requires regulators to recognize the need for effective risk communication as ongoing research provides additional information about the disease.

Zhonghua Jie He He Hu Xi Za Zhi, 2001 Oct, 24(10), 596 - 8
{Study of pefloxacin concentration in blood and sputum in the aged pneumonia patients with impairment of renal function}; Ma L et al.; OBJECTIVE: To study the drug concentration in blood and sputum, clinical effect and drug toxicity of pefloxacin in the aged pneumonia patients with different degree of impairment of renal function . METHODS: The patients were divided into four groups according to the impairment of renal functions pefloxacin 400 mg/12 h venous inflow, period of treatment is 10 days . Clinical manifestation and experimental index were registered; the drug concentration in blood and sputum was measured with biochemical technique, then compared and analyzed . RESULTS: The drug concentration in blood and sputum in four groups differed from the degree of the impairment of renal function . The concentration of drug in blood and sputum of the normal renal function was close to that in the low-grade impairment of renal function . Their clinical effective ratios were 83%, 80%, bacterium cleanup ratio was 86% . The difference of drug concentration in blood and sputum in the severe impairment of renal function was greater than that in the moderate renal function group, their clinical effective ratios were 66%, 53%, bacterial cleanup ratios were 57%, 36%, adverse reaction was growing along with degree of impairment of renal function . This drug has renal toxicity for moderate and severe impairment of renal function . CONCLUSION: In case of moderate impairment of renal function, prolongation of dose interval should be considered; pefloxacin should be avoided in severe impairment of renal function.

Mol Plant Microbe Interact, 2001 Dec, 14(12), 1463 - 7
ngl9: a third MADS box gene expressed in alfalfa root nodules; Zucchero JC et al.; Expression of MADS box genes has previously been localized to the infected cells of alfalfa (Medicago sativa) root nodules . These genes represent the first putative transcription factors to be identified in nodules and are hypothesized to be involved in a signal transduction pathway initiated by the intracellular bacterium . The eventual activation of specific target genes defines pertinent characteristics of this nitrogen-fixing differentiated cell . In this study, we identify a third nodule MADS box gene, ngl9, and demonstrate that the DNA-binding activity of its protein product is dependent on the presence of a second MADS box protein, NMH7 . Despite previous results to the contrary, both genes are expressed in the early stages of flower development, further strengthening the premise that nodule developmental programming may capitalize upon existing developmental cascades.

Gastroenterol Clin North Am, 2001 Dec, 30(4), 937 - 52
Helicobacter pylori and nonsteroidal anti-inflammatory drugs; Chan FK; The complex interaction between H . pylori and NSAIDs implies that it is over simplistic to conclude that their relationship is independent, synergistic, or antagonistic without considering the influence of other factors . Factors such as previous exposure to NSAIDs, a history of ulcer complication, concurrent use of acid-suppressant therapy, and the difference between NSAIDs and low-dose aspirin all affect the outcome . Several recommendations can be made with regard to the indications of H . pylori eradication for patients requiring NSAIDs . First, patients taking NSAIDs who have ulcers or previous ulcer disease should be tested for the bacterium, and it should be eradicated if present because it is impossible to determine whether the ulcers are caused by H . pylori or NSAIDs or both . Antiulcer drugs should be prescribed to prevent ulcer recurrence for patients who continue to require NSAIDs . Although the efficacy of omeprazole is enhanced by H . pylori infection, it is not justified to leave a pathogen in the stomach in exchange for a modest therapeutic gain . Second, for patients who take low-dose aspirin, eradication of H . pylori substantially reduces the risk of ulcer bleeding . It is advisable that patients taking low-dose aspirin who are at risk of ulcer bleeding should be tested for H . pylori and treated for it if the infection is found . Third, for patients who are about to start NSAIDs, screen-and-treat H . pylori has the potential of reducing the ulcer risk at an affordable incremental cost . It might be argued that any interaction between H . pylori and NSAIDs would become irrelevant in the era of COX-2-selective NSAIDs . Even among patients who are receiving a COX-2-selective NSAID, however, a large-scale study showed that the ulcer risk is significantly higher in H . pylori-positive patients than in uninfected patients . This finding suggests that the relative importance of H . pylori in ulcer development might increase with a reduced toxicity of COX-2-selective NSAIDs . With an increasing use of low-dose aspirin for cardiovascular prophylaxis, the problem of aspirin-related ulcer disease is expected to rise . Given the significant role of H . pylori in the latter condition, screen-and-treat H . pylori might be a useful strategy for the prevention of ulcer complications in high-risk patients receiving low-dose aspirin in the future.

Virchows Arch, 2001 Nov, 439(5), 653 - 60
Helicobacter pylori, N-methyl-N'-nitro-N'-nitrosoguanidine, and bile modulate gastric cell kinetics in experimental cancer; Loogna P et al.; Helicobacter pylori infection is a risk factor for gastric cancer . How the bacterium contributes to this process is still unclear . We present a new Wistar rat model that was used to evaluate the effect of H . pylori on early preneoplastic events as judged from epithelial cell turnover and histopathological changes . One hundred and four rats were colonized with H . pylori and exposed MNNG (N-methyl-N'-nitro-N'-nitrosoguanidine) and/or taurocholic acid . Inflammation, goblet cell-like metaplasia, atrophy, dysplasia, and adenocarcinoma were scored in a blinded manner . Apoptotic cells were counted after staining with terminal uridine deoxynucleotidyl nick end labeling, and epithelial cell proliferation was determined by means of the Ki-67 labeling index . No early tumor enhancement with H . pylori could be found in ordinary histology . However, H . pylori significantly enhanced the epithelial cell proliferation compared with the control group, and the combination with taurocholic acid appeared to have a synergistic effect . MNNG significantly increased the normal gastric epithelial apoptosis . This increase was reduced in antral mucosa with H . pylori infection . The findings suggest that H . pylori, especially when combined with bile . has an influence on cell kinetics, contributing to the development of gastric cancer . The reduced apoptosis of MNNG also observed in infected animals indicates a dual function of H . pylori.

J Wildl Dis, 2001 Oct, 37(4), 661 - 70
Intracranial abscessation in white-tailed deer of North America; Baumann CD et al.; From January 1996 through April 1997, the geographic distribution, etiology, demographics, seasonality, and prevalence of an intracranial abscessation/suppurative meningoencephalitits syndrome in white-tailed deer (Odocoileus virginianus) were evaluated by surveying wildlife disease diagnostic laboratories and by examining both natural mortality and hunter-harvested deer skulls from North America . Intracranial abscesses were diagnosed as the cause of death or illness in 97 of nearly 4,500 (2.2%) white-tailed deer examined from 12 states and four Canadian provinces by the diagnostic laboratories . The bacterium Arcanobacterium pyogenes was isolated from 61% of cases; 18 other genera of bacteria also were isolated . The disease was strongly gender-biased (P < 0.01) with 87% of cases occurring in males, and the overall prevalence among males was 4.9% . Cases were most common among antlered males (> or = 1 yr) with few cases among male fawns . Among antlered males, cases were seasonal, primarily occurring from September through April . Four hundred eighteen skulls from deer found dead in the field were examined from southeastern USA, and of the 119 used for further evaluation, 9% had characteristic lesions . Skulls from hunter-harvested males in the southeastern USA had a lesion prevalence of 1.4% . The similarity of disease prevalence among male deer found dead in the field (9.0%) and deer examined as southeastern diagnostic laboratory cases (8.4%) suggests that this disease accounts for slightly < 10% of the natural mortality for yearling and adult male white-tailed deer in the southeastern region . The strong bias for occurrence among males suggests this disease may affect quality deer management strategies.

Antioxid Redox Signal, 2001 Oct, 3(5), 825 - 38
Electronic structure studies of quinones and semiquinones: accurate calculation of spin densities and electron paramagnetic resonance parameters; O'Malley PJ; The application of electronic structure methods to the prediction of geometries, spin densities, and hyperfine couplings for biologically relevant quinones and semiquinones is reviewed . It is demonstrated that hybrid-type density functional methods are particularly suitable for such studies . Hydrogen bonding to the semiquinone oxygen by appropriate donors is shown to lead to a redistribution of spin density in the carbonyl group of the semiquinone . Experimental trends are well reproduced by the calculated values . Symmetric and asymmetric models of hydrogen bonding are modelled . It is shown that the symmetric models give good agreement with solution studies in vitro . The asymmetric models of hydrogen bonding give quite good agreement with values measured for in vivo semiquinones generated in the reaction centres of the purple photosynthetic bacterium, Rb sphaeroides, and also for the phyllosemiquinone free radical formed during electron transfer in Photosystem I of green plants . These recent advances in electronic structure calculations, in particular the applicability of density functional methods to the study of free radical properties, have opened up an exciting avenue for the complete characterisation of their electronic properties . In particular, the combination of experimental methods of electron paramagnetic resonance and such calculations should in future provide a clearer understanding of free radical chemistry in many areas of biology.

Int J Syst Evol Microbiol, 2001 Nov, 51(Pt 6), 2055 - 61
Desulfomusa hansenii gen . nov., sp . nov., a novel marine propionate-degrading, sulfate-reducing bacterium isolated from Zostera marina roots; Finster K et al.; The physiology and phylogeny of a novel sulfate-reducing bacterium, isolated from surface-sterilized roots of the marine macrophyte Zostera marina, are presented . The strain, designated P1T, was enriched and isolated in defined oxygen-free, bicarbonate-buffered, iron-reduced seawater medium with propionate as sole carbon source and electron donor and sulfate as electron acceptor . Strain P1T had a rod-shaped, slightly curved cell morphology and was motile by means of a single polar flagellum . Cells generally aggregated in clumps throughout the growth phase . High CaCl2 (10 mM) and MgCl2 (50 mM) concentrations were required for optimum growth . In addition to propionate, strain P1T utilized fumarate, succinate, pyruvate, ethanol, butanol and alanine . Oxidation of propionate was incomplete and acetate was formed in stoichiometric amounts . Strain P1T thus resembles members of the sulfate-reducing genera Desulfobulbus and Desulforhopalus, which both oxidize propionate incompletely and form acetate in addition to CO2 . However, sequence analysis of the small-subunit rDNA and the dissimilatory sulfite reductase gene revealed that strain P1T was unrelated to the incomplete oxidizers Desulfobulbus and Desulforhopalus and that it constitutes a novel lineage affiliated with the genera Desulfococcus, Desulfosarcina, Desulfonema and 'Desulfobotulus' . Members of this branch, with the exception of 'Desulfobotulus sapovorans', oxidize a variety of substrates completely to CO2 . Strain P1T (= DSM 12642T = ATCC 700811T) is therefore proposed as Desulfomusa hansenii gen . nov., sp . nov . Strain p1T thus illustrates the difficulty of extrapolating rRNA similarities to physiology and/or ecological function.

Biosci Biotechnol Biochem, 2001 Oct, 65(10), 2146 - 53
Regulation by two CatR proteins that differ in binding affinity to catB promoters expressing two cat gene clusters; Takashima A et al.; We isolated the two LysR-type regulatory proteins CatR1 and CatR2, which regulate the expression of cat1 and cat2 gene clusters, respectively, required for catechol degradation in the bacterium Frateuria sp . ANA-18 . In a gel mobility shift assay using CatR1 and the DNA fragment containing the catB1 promoter region, the formation of two complexes, complex 1-1 (C1-1) and complex 1-2 (C1-2), was observed in the presence of cis,cis-muconate . On the other hand, CatR2 and the DNA fragment containing the catB2 promoter region formed only complex 2-2 (C2-2) at a lower concentration of cis,cis-muconate than that at which C1-1 and C1-2 were formed . As the concentration of cis,cis-muconate decreased, the production of the muconate cycloisomerase isozyme MC II encoded by catB2 decreased as well as that of MC I encoded by catB1 . However, the amount of MC II synthesized was larger than that of MC I at low concentrations . On the basis of these results, we concluded that the catB2 promoter was activated at low concentrations of cis,cis-muconate.

Ying Yong Sheng Tai Xue Bao, 2001 Aug, 12(4), 636 - 8
{Degradation of phenanthrene and pyrene in contaminated soil by immobilized Zoogloea sp . and Fusarium sp.}; Wang X et al.; Immobilized with PVA, sodium alginate and activated carbon, both Zoogloea sp . and Fusarium sp . strains could degrade phenanthrene and pyrene efficiently . The optimal carrier was made of 100 rho.g-1 L PVA, 5 sodium alginate rho.g-1 L and 50 activated carbon rho.g-1 L . The degradation rates of phenanthrene and pyrene in 10 days were 87.48% and 75.34% by the immobilized bacterium, 37.04% and 20.85% higher than those by the free bacterium, and the rates in 15 days were 84.36% and 74.87% by the immobilized fungus, 5.35% and 11.23% higher than those by the free fungus.

J Bacteriol, 2002 Jan, 184(2), 525 - 30
Luteolin and GroESL modulate in vitro activity of NodD; Yeh KC et al.; In the early stages of symbiosis between the soil bacterium Sinorhizobium meliloti and its leguminous host plant, alfalfa, bacterial nodulation (nod) genes are controlled by NodD1, NodD2, and NodD3, members of the LysR family of transcriptional regulators, in response to flavonoid and other inducers released by alfalfa . To gain an understanding of the biochemical aspects of this action, epitope-tagged recombinant NodD1 and NodD3 were overexpressed in Escherichia coli . The DNA binding properties of the purified recombinant NodD proteins were indistinguishable from those of NodD isolated from S . meliloti . In addition, the E . coli GroEL chaperonin copurified with the recombinant NodD proteins . In this study, we showed that NodD proteins are in vitro substrates of the GroESL chaperonin system and that their DNA binding activity is modulated by GroESL . This confirmed the earlier genetic implication that the GroESL chaperonin system is essential for the function of these regulators . Increased DNA binding activity by NodD1 in the presence of luteolin confirmed that NodD1 is involved in recognizing the plant signal during the early stages of symbiosis.

FEBS Lett, 2001 Dec 14, 509(3), 350 - 4
A novel metallo-beta-lactamase, Mbl1b, produced by the environmental bacterium Caulobacter crescentus; Simm AM et al.; Caulobacter crescentus 101123 possesses a gene (Mbl1b) encoding a metallo-beta-lactamase with 32% amino acid identity to the L1 metallo-beta-lactamase from Stenotrophomonas maltophilia . The gene was cloned into an expression vector and the enzyme, Mbl1b, was expressed in Escherichia coli . Mbl1b was purified . Catalytic properties for several antibiotics were determined . The enzyme exhibits Michaelis-Menten kinetics for imipenem, meropenem and nitrocefin but substrate inhibition kinetics with cefoxitin, cefaloridine, penicillin G and ampicillin . A homology model predicts Mbl1b has the same structural fold as other metallo-beta-lactamases with a detailed structure very similar to L1 but whereas L1 is a homotetramer, Mbl1b is monomeric . The main differences between Mbl1 and L1 are in the N-terminal region.

Infect Immun, 2002 Jan, 70(1), 218 - 25
Lipopolysaccharides from periodontopathic bacteria Porphyromonas gingivalis and Capnocytophaga ochracea are antagonists for human toll-like receptor 4; Yoshimura A et al.; Toll-like receptors (TLRs) 2 and 4 have recently been identified as possible signal transducers for various bacterial ligands . To investigate the roles of TLRs in the recognition of periodontopathic bacteria by the innate immune system, a Chinese hamster ovary (CHO) nuclear factor-kappaB (NF-kappaB)-dependent reporter cell line, 7.7, which is defective in both TLR2- and TLR4-dependent signaling pathways was transfected with human CD14 and TLRs . When the transfectants were exposed to freeze-dried periodontopathic bacteria, Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Capnocytophaga ochracea, and Fusobacterium nucleatum, and a non-oral bacterium, Escherichia coli, all species of the bacteria induced NF-kappaB-dependent CD25 expression in 7.7/huTLR2 cells . Although freeze-dried A . actinomycetemcomitans, F . nucleatum, and E . coli also induced CD25 expression in 7.7/huTLR4 cells, freeze-dried P . gingivalis did not . Similarly, lipopolysaccharides (LPS) extracted from A . actinomycetemcomitans, F . nucleatum, and E . coli induced CD25 expression in 7.7/huTLR4 cells, but LPS from P . gingivalis and C . ochracea did not . Furthermore, LPS from P . gingivalis and C . ochracea attenuated CD25 expression in 7.7/huTLR4 cells induced by repurified LPS from E . coli . LPS from P . gingivalis and C . ochracea also inhibited the secretion of interleukin-6 (IL-6) from U373 cells, the secretion of IL-1beta from human peripheral blood mononuclear cells, and ICAM-1 expression in human gingival fibroblasts induced by repurified LPS from E . coli . These findings indicated that LPS from P . gingivalis and C . ochracea worked as antagonists for human TLR4 . The antagonistic activity of LPS from these periodontopathic bacteria may be associated with the etiology of periodontal diseases.

Infect Immun, 2002 Jan, 70(1), 55 - 61
Role of Bcl-2 family members in caspase-independent apoptosis during Chlamydia infection; Perfettini JL et al.; Infection with an obligate intracellular bacterium, the Chlamydia trachomatis lymphogranuloma venereum (LGV/L2) strain or the guinea pig inclusion conjunctivitis serovar of Chlamydia psittaci, leads to apoptosis of host cells . The apoptosis is not affected by a broad-spectrum caspase inhibitor, and caspase-3 is not activated in infected cells, suggesting that apoptosis mediated by these two strains of Chlamydia is independent of known caspases . Overexpression of the proapoptotic Bcl-2 family member, Bax, was previously shown to induce caspase-independent apoptosis, and we find that Bax is activated and translocates from the cytosol to the mitochondria in C . psittaci-infected cells . C . psittaci-induced apoptosis is inhibited in host cells overexpressing Bax inhibitor-1 and is inhibited through overexpression of Bcl-2, which blocks both caspase-dependent and -independent apoptosis . As Bax and mitochondria are ideally located to sense stress-related metabolic changes emanating from the interior of an infected cell, it is likely that Bax-dependent apoptosis may also be observed in cells infected with other intracellular pathogens.

Biotechnol Bioeng, 2001 Dec 5, 75(5), 497 - 503
The in vivo synthesis of plant sesquiterpenes by Escherichia coli; Martin VJ et al.; Three plant genes encoding (+)-delta-cadinene, 5-epi-aristolochene, and vetispiradiene cyclases were expressed in Escherichia coli to evaluate the potential of this bacterium to synthesize sesquiterpenes in vivo . Various growth temperatures, carbon sources, and host strains were examined to optimize terpene production . The highest levels of sesquiterpene production occurred when the enzymes were expressed in strain DH5alpha from the trc promoter (Ptrc) of the high-copy plasmidpTrc99A in M9 medium supplemented with 0.2% (v/v) glycerol at 30 degrees C for 5-epi-aristolochene and vetispiradiene and 37 degrees C for (+)-delta-cadinene . The highest concentrations of sesquiterpenes observed were 10.3 microg of (+)-delta-cadinene, 0.24 microg of 5-epi-aristolochene (measured as (+)-delta-cadinene equivalents), and 6.4 microg of vetispiradiene (measured as (+)-delta-cadinene equivalents) per liter of culture . These sesquiterpene production levels are >500-fold lower than carotenoid production, both of which are synthesized from endogenous trans-farnesyl diphosphate (FDP) in E . coli . Based on these results, we conclude that the limiting factor for sesquiterpene synthesis in E . coli is the poor expression of the cyclase enzyme and not supply of the FDP precursor . Copyright 2001 John Wiley & Sons, Inc.

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