J Microsc, 2005 Jan, 217(Pt 1), 109 - 16 Combined AFM and confocal fluorescence microscope for applications in bio-nanotechnology; Kassies R et al.; Summary We present a custom-designed atomic force fluorescence microscope (AFFM), which can perform simultaneous optical and topographic measurements with single molecule sensitivity throughout the whole visible to near-infrared spectral region . Integration of atomic force microscopy (AFM) and confocal fluorescence microscopy combines the high-resolution topographical imaging of AFM with the reliable (bio)-chemical identification capability of optical methods . The AFFM is equipped with a spectrograph enabling combined topographic and fluorescence spectral imaging, which significantly enhances discrimination of spectroscopically distinct objects . The modular design allows easy switching between different modes of operation such as tip-scanning, sample-scanning or mechanical manipulation, all of which are combined with synchronous optical detection . We demonstrate that coupling the AFM with the fluorescence microscope does not compromise its ability to image with a high spatial resolution . Examples of several modes of operation of the AFFM are shown using two-dimensional crystals and membranes containing light-harvesting complexes from the photosynthetic bacterium Rhodobacter sphaeroides. Int J Syst Evol Microbiol, 2005 Jan, 55(Pt 1), 91 - 4 Planococcus stackebrandtii sp . nov., isolated from a cold desert of the Himalayas, India; Mayilraj S et al.; The taxonomic position of a bacterium isolated from a cold desert of the Himalayas, India, was analysed by using a polyphasic approach . The isolated strain, designated K22-03(T), had phenotypic characteristics that matched those of the genus Planococcus and it represents a novel species . The almost-complete 16S rRNA gene sequence (1464 bases) of the novel strain was compared with those of previously studied Planococcus type strains and confirmed that the strain belongs to the genus Planococcus . 16S rRNA gene sequence analysis indicated that strain K22-03(T) differs from all other species of Planococcus by at least 2.5 % . DNA-DNA hybridization showed that it had low genomic relatedness with Planomicrobium mcmeekinii (MTCC 3704(T), 23 %), Planococcus psychrophilus (MTCC 3812(T), 61 %), Planococcus antarcticus (MTCC 3854(T), 45 %) and Planomicrobium okeanokoites (MTCC 3703(T), 51 %), the four species with which it was most closely related based on 16S rRNA gene sequence analysis (97-97.5 % similarity) . Therefore, strain K22-03(T) should be recognized as a novel species, for which the name Planococcus stackebrandtii sp . nov . is proposed . The type strain is K22-03(T) (=MTCC 6226(T)=DSM 16419(T)=JCM 12481(T)). Science, 2005 Jan 14, 307(5707), 214 - 5 Microbiology . TB--a new target, a new drug; Cole ST et al.; Genetique Moleculaire Bacterienne and Biochimie Structurale Units, Institut Pasteur, Paris 75724 Cedex 15, France . stcole@pasteur.fr Chembiochem . 2005 Jan 13; {Epub ahead of print} A New Type of Metalloprotein: The Mo Storage Protein from Azotobacter vinelandii Contains a Polynuclear Molybdenum-Oxide Cluster; Fenske D et al.; Azotobacter vinelandii is a diazotrophic bacterium characterized by the outstanding capability of storing Mo in a special storage protein, which guarantees Mo-dependent nitrogen fixation even under growth conditions of extreme Mo starvation . The Mo storage protein is constitutively synthesized with respect to the nitrogen source and is regulated by molybdenum at an extremely low concentration level (0-50 nM) . This protein was isolated as an alpha(4)beta(4) octamer with a total molecular mass of about 240 kg mol(-1) and its shape was determined by small-angle X-ray scattering . The genes of the alpha and beta subunits were unequivocally identified; the amino acid sequences thereby determined reveal that the Mo storage protein is not related to any other known molybdoprotein . Each protein molecule can store at least 90 Mo atoms . Extended X-ray absorption fine-structure spectroscopy identified a metal-oxygen cluster bound to the Mo storage protein . The binding of Mo (biosynthesis and incorporation of the cluster) is dependent on adenosine triphosphate (ATP); Mo release is ATP-independent but pH-regulated, occurring only above pH 7.1 . This Mo storage protein is the only known noniron metal storage system in the biosphere containing a metal-oxygen cluster. J Biol Inorg Chem . 2005 Jan 14; {Epub ahead of print} Isoprenoid biosynthesis in chloroplasts via the methylerythritol phosphate pathway: the (E)-4-hydroxy-3-methylbut-2-enyl diphosphate synthase (GcpE) from Arabidopsis thaliana is a {4Fe-4S} protein; Seemann M et al.; The mevalonate-independent methylerythritol phosphate pathway is widespread in bacteria . It is also present in the chloroplasts of all phototrophic organisms . Whereas the first steps, are rather well known, GcpE and LytB, the enzymes catalyzing the last two steps have been much less investigated . 2-C-Methyl-D: -erythritol 2,4-cyclodiphosphate is transformed by GcpE into 4-hydroxy-3-methylbut-2-enyl diphosphate, which is converted by LytB into isopentenyl diphosphate or dimethylallyl diphosphate . Only the bacterial GcpE and LytB enzymes have been investigated to some extent, but nothing is known about the corresponding plant enzymes . In this contribution, the prosthetic group of GcpE from the plant Arabidopsis thaliana and the bacterium Escherichia coli has been fully characterized by Mossbauer spectroscopy after reconstitution with (57)FeCl(3), Na(2)S and dithiothreitol . It corresponds to a {4Fe-4S} cluster, suggesting that both plant and bacterial enzymes catalyze the reduction of 2-C-methyl-D: -erythritol 2,4-cyclodiphosphate into (E)-4-hydroxy-3-methylbut-2-enyl diphosphate via two consecutive one-electron transfers . In contrast to the bacterial enzyme, which utilizes NADPH/flavodoxin/flavodoxin reductase as a reducing shuttle system, the plant enzyme could not use this reduction system . Enzymatic activity was only detected in the presence of the 5-deazaflavin semiquinone radical. J Antimicrob Chemother . 2005 Jan 13; {Epub ahead of print} Molecular evaluation of antibiotic susceptibility of Tropheryma whipplei in axenic medium; Boulos A et al.; Whipple's disease is a rare multisystem chronic infection, involving the intestinal tract as well as various other organs . Tropheryma whipplei is a slow-growing facultative intracellular bacterium that remains poorly understood . In vitro antibiotic susceptibility testing has previously been assessed in cells using a real-time quantitative PCR assay . In this study, we have evaluated the antibiotic susceptibility of three strains of T . whipplei grown in axenic medium using the same assay . The active compounds in axenic medium were doxycycline, macrolide compounds, penicillin G, streptomycin, rifampicin, chloramphenicol, thiamphenicol, teicoplanin, vancomycin, amoxicillin, gentamicin, aztreonam, levofloxacin and ceftriaxone, with MICs in the range 0.06-1 mg/L . Cefalothin was less active, with MICs in the range 2-4 mg/L . We found that co-trimoxazole was active with MICs in the range 0.5-1 mg/L, and sulfamethoxazole alone was active with MICs in the range 0.5-1 mg/L . MICs of trimethoprim varied from 64-128 mg/L . Co-trimoxazole was effective in vitro, but this activity was due to sulfamethoxazole alone . These results were in accordance with the fact that T . whipplei does not contain the encoding gene for dihydrofolate reductase, the target for trimethoprim. Biochem Biophys Res Commun, 2005 Feb 18, 327(3), 668 - 74 Successful recombinant production of Allochromatium vinosum cytochrome c' requires coexpression of cmm genes in heme-rich Escherichia coli JCB712; Evers TH et al.; Cytochrome c' from the purple photosynthetic bacterium Allochromatium vinosum (CCP) displays a unique, reversible dimer-to-monomer transition upon binding of NO, CO, and CN(-) . This small, four helix bundle protein represents an attractive model for the study of other heme protein biosensors, provided a recombinant expression system is available . Here we report the development of an efficient expression system for CCP that makes use of a maltose binding protein fusion strategy to enhance periplasmic expression and allow easy purification by affinity chromatography . Coexpression of cytochrome c maturase genes and the use of a heme-rich Escherichia coli strain were found to be necessary to obtain reasonable yields of cytochrome c' . Characterization using circular dichroism, UV-vis spectroscopy, and size-exclusion chromatography confirms the native-like properties of the recombinant protein, including its ligand-induced monomerization. Environ Microbiol, 2005 Jan, 7(1), 22 - 33 Isolation and properties of methanesulfonate-degrading Afipia felis from Antarctica and comparison with other strains of A . felis; Moosvi SA et al.; Summary Three novel strains of methylotrophic Afipia felis were isolated from several locations on Signy Island, Antarctica, and a fourth from estuary sediment from the River Douro, Portugal . They were identified as strains of the alpha-2 proteobacterium A . felis by 16S rRNA gene sequence analysis . Two strains tested were shown to contain the fdxA gene, diagnostic for A . felis . All strains grew with methanesulfonate (and two strains with dimethylsulfone) as sole carbon substrate . Growth on methanesulfonate required methanesulfonate monooxygenase (MSAMO), using NADH as the reductant and stimulated by reduced flavin nucleotides and Fe(II) . Polymerase chain reaction amplification of DNA from an Antarctic strain showed a typical msmA gene for the alpha-hydroxylase of MSAMO, and both Antarctic and Portuguese strains contained mxaF, the methanol dehydrogenase large subunit gene . This is the first report of methanesulfonate-degrading bacteria from the Antarctic and of methylotrophy in Afipia, and the first description of any bacterium able to use both methanesulfonate and dimethylsulfone . In contrast, the type strain of A . felis DSM 7326(T) was not methylotrophic, but grew in defined mineral medium with a wide range of single simple organic substrates . Free-living Afipia strains occurring widely in the natural environment may be significant as methylotrophs, degrading C(1)-sulfur compounds, including the recalcitrant organosulfur compound methanesulfonate. Appl Environ Microbiol, 2005 Jan, 71(1), 417 - 22 Contribution of Folate Biosynthesis to Ralstonia solanacearum Proliferation in Intercellular Spaces; Shinohara R et al.; The vigorous proliferation of Ralstonia solanacearum OE1-1 in host intercellular spaces after the invasion of host plants is necessary for the virulence of this bacterium . A folate auxotroph, RM, in which a mini-Tn5 transposon was inserted into pabB encoding para-aminobenzoate synthase component I, lost its ability to vigorously proliferate in intercellular spaces along with its systemic infectivity and virulence after inoculation into roots and infiltration into leaves of tobacco plants . Complementation of RM with the pabB gene allowed the mutant to multiply in intercellular spaces and to cause disease . In tobacco plants that were pretreated with folate, RM was able to vigorously proliferate in the intercellular spaces and cause disease . Interestingly, when it was inoculated through cut stems, the mutant multiplied in the plants and was virulent . Moreover, the mutant multiplied well in stem fluids but not in intercellular fluids, suggesting that the folate concentration within intercellular spaces may be a limiting factor for bacterial proliferation . Therefore, folate biosynthesis contributes to the vigorous proliferation of bacteria in intercellular spaces and leads to systemic infectivity resulting in virulence. Appl Environ Microbiol, 2005 Jan, 71(1), 123 - 30 Seasonal Change in Bacterial Flora and Biomass in Mountain Snow from the Tateyama Mountains, Japan, Analyzed by 16S rRNA Gene Sequencing and Real-Time PCR; Segawa T et al.; The bacterial flora and biomass in mountain snow from the Tateyama Mountains, Toyama Prefecture, Japan, one of the heaviest snowfall regions in the world, were analyzed by amplified ribosomal DNA restriction analysis followed by 16S rRNA gene sequencing and DNA quantification by real-time PCR . Samples of surface snow collected in various months during the melting season contained a psychrophilic bacterium, Cryobacterium psychrophilum, and two psychrotrophic bacteria, Variovorax paradoxus and Janthinobacterium lividum . Bacterial colonies that developed in an in situ meltwater medium at 4 degrees C were revealed to be V . paradoxus . The biomasses of C . psychrophilum, J . lividum, and V . paradoxus, as estimated by real-time PCR, showed large increases during the melting season from March to October (2.0 x 10(5)-fold, 1.5 x 10(5)-fold, and 1.0 x 10(4)-fold increases, respectively), suggesting their rapid growth in the surface snow . The biomasses of C . psychrophilum and J . lividum increased significantly from March to April, reached a maximum in August, and dropped at the end of the melting season . In contrast, the biomass of V . paradoxus did not increase as rapidly during the early melting season but continued to increase from June until October . The differences in development observed among these bacterial species suggest that their growth was promoted by different nutrients and/or environmental conditions in the snow . Since these three types of bacteria have also been reported to be present in a glacier in Antarctica and a Greenland ice core, they seem to be specialized members of the snow biota that are distributed in snow and ice environments in various parts of the world. Science, 2005 Jan 7, 307(5706), 105 - 8 Genome sequence of the PCE-dechlorinating bacterium Dehalococcoides ethenogenes; Seshadri R et al.; Dehalococcoides ethenogenes is the only bacterium known to reductively dechlorinate the groundwater pollutants, tetrachloroethene (PCE) and trichloroethene, to ethene . Its 1,469,720-base pair chromosome contains large dynamic duplicated regions and integrated elements . Genes encoding 17 putative reductive dehalogenases, nearly all of which were adjacent to genes for transcription regulators, and five hydrogenase complexes were identified . These findings, plus a limited repertoire of other metabolic modes, indicate that D . ethenogenes is highly evolved to utilize halogenated organic compounds and H2 . Diversification of reductive dehalogenase functions appears to have been mediated by recent genetic exchange and amplification . Genome analysis provides insights into the organism's complex nutrient requirements and suggests that an ancestor was a nitrogen-fixing autotroph. J Clin Microbiol, 2005 Jan, 43(1), 393 - 401 Latency-related gene encoded by bovine herpesvirus 1 promotes virus growth and reactivation from latency in tonsils of infected calves; Perez S et al.; Infection of calves with bovine herpesvirus 1 (BHV-1) results in transient immunosuppression that may lead to bacterium-induced pneumonia and, occasionally, death . Although sensory neurons in the trigeminal ganglia (TG) are the primary site of BHV-1 latency, viral genomes are detected in the tonsils of latently infected calves . Dexamethasone (DEX) consistently induces reactivation from latency, and viral gene expression is detected in TG and tonsils . In sensory neurons of latently infected calves, the latency-related (LR) gene is abundantly expressed and is required for reactivation from latency . In the present study, we compared the abilities of wild-type (wt) BHV-1 and a strain with a mutation in the LR gene (the LR mutant strain) to grow in the tonsils of infected calves and reactivate from latency . Lower levels of the LR mutant virus were detected in the tonsils of acutely infected calves . LR mutant viral DNA was consistently detected by PCR in the tonsils of latently infected calves, suggesting that the establishment of a latent or persistent infection occurred . Although the LR mutant did not reactivate from latency in vivo after DEX treatment, explantation of tonsil tissue from calves latently infected with the LR mutant yielded infectious virus . Relative to wt BHV-1, the LR mutant did not induce explant-induced reactivation as efficiently . These studies indicate that the LR gene promotes virus shedding from tonsil tissue during acute infection and reactivation from latency in tonsil tissue in vivo . We suggest that incorporation of the LR gene mutation into existing modified live vaccines would prevent reactivation from latency in neural and nonneural sites and would thus prevent transmission to other animals. J Clin Microbiol, 2005 Jan, 43(1), 41 - 8 Multispacer typing technique for sequence-based typing of Bartonella quintana; Foucault C et al.; Bartonella quintana is a worldwide fastidious bacterium of the Alphaproteobacteria responsible for bacillary angiomatosis, trench fever, chronic lymphadenopathy, and culture-negative endocarditis . The recent genome sequencing of a B . quintana isolate allowed us to propose a genome-wide sequence-based typing method . To ensure sequence discrimination based on highly polymorphic areas, we amplified and sequenced 34 spacers in a large collection of B . quintana isolates . Six of these exhibited polymorphisms and allowed the characterization of 4 genotypes . However, the strain variants suggested by the noncoding sequences did not correlate with the results of pulsed-field gel electrophoresis (PFGE), which suggested a higher degree of variability . Modification of the PFGE profile of one isolate after nine subcultures confirmed that rearrangement frequencies are high in this species, making PFGE unreliable for epidemiological purposes . The low extent of sequence heterogeneity in the species suggests a recent emergence of this bacterium as a human pathogen . Direct typing of natural samples allowed the identification of a fifth genotype in the DNA extracted from a human body louse collected in Burundi . We have named the typing technique herein described multispacer typing. Proc Natl Acad Sci U S A . 2005 Jan 4; {Epub ahead of print} Brucella coopts the small GTPase Sar1 for intracellular replication; Celli J et al.; The pathogen Brucella abortus resides inside macrophages within a unique, replication-permissive organelle that is derived from the endoplasmic reticulum (ER) . Although dependent on the Brucella type IV secretion system VirB, the mechanisms governing the biogenesis of this compartment remain elusive . Here, we investigated a putative role of the early secretory pathway in ER membrane accretion by the Brucella-containing vacuoles (BCVs) . We show that BCVs interact with ER exit sites (ERES), and blockade of Sar1 activity, which disrupts ERES, prevents intracellular replication of Brucella . In cells expressing the dominant interfering form Sar1{T39N}, BCVs do not acquire ER membranes, suggesting that they are unable to mature into replicative organelles . By contrast, treatments that block subsequent secretory events do not affect bacterial replication . We propose that Sar1-dependent ERES functions, but not subsequent secretory events, are essential for the biogenesis of the Brucella replicative compartment and, thus, bacterial replication . These results assign an essential role for Sar1 in pathogenesis of an intracellular bacterium. J Biol Chem . 2005 Jan 4; {Epub ahead of print} Functional consequences of the organization of the photosynthetic apparatus in R . sphaeroides: 1 . Quinone domains and excitation transfer in chromatophores and reaction center-antenna complexes; Comayras F et al.; The purpose of this study was to gain information on functional consequences of the supramolecular organization of the photosynthetic apparatus in the bacterium Rhodobacter sphaeroides . Isolated complexes of the reaction center (RC) with its core antenna ring (LH1) were studied, in their dimeric (native) form or as monomers, with respect to excitation transfer and distribution of the quinone pool . Similar issues were examined in chromatophore membranes . The relationship between the fluorescence yield and the amount of closed centers is indicative of a very efficient excitation transfer between the two monomers in isolated dimeric complexes . A similar dependence was observed in chromatophores, suggesting that excitation transfer in vivo from a closed RC-LH1 unit is also essentially directed to its partner in the dimer . The isolated complexes were found to retain 25-30 % of the endogenous quinone acceptor pool and the distribution of this pool among the complexes suggests a cooperative character for the association of quinones to the protein complexes . In chromatophores, the decrease in the amount of photoreducible quinones when inhibiting a fraction of the centers implies a confinement of the quinone pool over small domains, including 1-6 reaction centers . We suggest that the crowding of membrane proteins may not be the sole reason for quinone confinement and that a quinone-rich region is formed around the RC-LH1 complexes. J Biol Chem . 2005 Jan 4; {Epub ahead of print} Functional consequences of the organization of the photosynthetic apparatus in R . sphaeroides: 2 . A study of PufX- membranes; Comayras F et al.; In the bacterium R . sphaeroides, polypeptide PufX is indispensable for photosynthetic growth . Its deletion is known to have important consequences on the organization of the photosynthetic apparatus . In the WT, the RC-LH1 complexes (reaction center and antenna) are associated in dimers and the LH1 does not fully encircle the RC . In the absence of PufX, the complexes become monomeric and the LH1 ring closes around the RC . We analyze functional consequences of PufX deletion . Some effects can be ascribed to the monomerization of the RC-LH1 complexes: the number of RCs that share a common antenna for excitation transfer, or a common quinone pool, both become smaller . We examined the kinetic effects of the closed LH1 ring on quinone turnover: the diffusion across the LH1 entails a delay of about 1 ms and the barrier appears located directly against the quinone binding (QB) pocket . The diffusion of ubiquinol from the RC to the cytochrome bc1 complex is about two-fold slower in the mutant, suggesting an increased distance between the two complexes . The properties of the QB pocket (binding of inhibitors, stabilization of QB- and rate of QBH2 formation) appeared modified in the mutant . Another specificity of PufX- is the accumulation of closed centers in the QA- state as the secondary acceptor pool becomes reduced, which is probably the origin of the photosynthetic incompetence . We suggest that this is related to the QB pocket alterations . The malfunction of the reaction center is probably due to a faulty association with LH1 that is prevented in the PufX-containing structure. J Biol Chem . 2005 Jan 4; {Epub ahead of print} How an enzyme binds the C1-carrier tetrahydromethanopterin: Structure of the tetrahydromethanopterin dependent formaldehyde-activating enzyme (Fae) from methylobacterium extorquens AM1; Acharya P et al.; Tetrahydromethanopterin (H4MPT) is a tetrahydrofolate (H4F) analogue involved as C1-carrier in the metabolism of various groups of microorgansims . How H4MPT is bound to the respective C1-unit converting enzymes remained elusive . We describe here the structure of the homopentameric formaldehyde-activating enzyme (Fae) from Methylobacterium extorquens AM1 established at 2.0 A without and at 1.9 A with methylene-H4MPT bound . Methylene-H4MPT is bound in a 'S'-shaped conformation into the cleft formed between two adjacent subunits . Coenzyme binding is accompanied by side chain rearrangements up to 5 A and leads to a rigidification of the C-terminal arm, a formation of a new hydrophobic cluster and an inversion of the amide side chain of Gln88 . Methylene-H4MPT in Fae shows a characteristic kink between the tetrahydropyrazine and the imidazolidine rings of 70 degrees that is more pronounced than that reported for free methylene-H4MPT in solution (50 degrees ) . Fae is an essential enzyme for energy metabolism and formaldehyde detoxification of this bacterium and catalyses the formation of methylene-H4MPT from H4MPT and formaldehyde . The molecular mechanism of this reaction involving His22 as acid catalyst is discussed. Plasmid, 2005 Jan, 53(1), 1 - 13 Epub 2004 Dec 16. The plasmids of Borrelia burgdorferi: essential genetic elements of a pathogen; Stewart PE et al.; The spirochete Borrelia burgdorferi, the causative agent of Lyme disease, has an unusual genome comprised of a linear chromosome and the largest plasmid complement of any characterized bacterium . Certain plasmid-encoded elements are required for virulence and viability, both in vitro and in vivo . The genetic tools to manipulate B . burgdorferi are sufficiently developed for precise molecular genetic investigations . B . burgdorferi now represents a prime system with which to address basic questions of plasmid biology and plasmid contributions to bacterial virulence and disease pathogenesis. J Bacteriol, 2005 Jan, 187(2), 534 - 43 Analysis of the Precursor rRNA Fractions of Rapidly Growing Mycobacteria: Quantification by Methods That Include the Use of a Promoter (rrnA P1) as a Novel Standard; Menendez Mdel C et al.; Mycobacterial species are able to control rRNA production through variations in the number and strength of promoters controlling their rrn operons . Mycobacterium chelonae and M . fortuitum are members of the rapidly growing mycobacterial group . They carry a total of five promoters each, encoded, respectively, by one and two rrn operons per genome . Quantification of precursor rrn transcriptional products (pre-rrn) has allowed detection of different promoter usage during cell growth . Bacteria growing in several culture media with different nutrient contents were compared . Balanced to stationary phases were analyzed . Most promoters were found to be used at different levels depending on the stage of bacterial growth and the nutrient content of the culture medium . Some biological implications are discussed . Sequences of the several promoters showed motifs that could be correlated to their particular level of usage . A product corresponding to the first rrnA promoter in both species, namely, rrnA P1, was found to contribute at a low and near-constant level to pre-rRNA synthesis, regardless of the culture medium used and the stage of growth analyzed . This product was used as a standard to quantitate rRNA gene expression by real-time PCR when M . fortuitum infected macrophages . It was shown that this bacterium actively synthesizes rRNA during the course of infection and that one of its rrn operons is preferentially used under such conditions. Cell Motil Cytoskeleton, 2005 Feb, 60(2), 104 - 20 Actin and alpha-actinin dynamics in the adhesion and motility of EPEC and EHEC on host cells; Shaner NC et al.; Two pathogenic Escherichia coli, Enteropathogenic E . coli (EPEC) and Enterohemorrhagic E . coli (EHEC), adhere to the outside of host cells and induce cytoskeletal rearrangements leading to the formation of membrane-encased pedestals comprised of actin filaments and other associated proteins beneath the bacteria . The structure of the pedestals induced by the two pathogens appears similar, although those induced by EHEC are shorter in length . Fluorescence Recovery After Photobleaching (FRAP) was used to determine potential differences of actin polymerization in EPEC and EHEC induced pedestals in cultured PtK2 cells expressing either Green or Yellow Fluorescent Protein (GFP or YFP) fused to actin or alpha-actinin . When all the fluorescent actin in a pedestal on EPEC-infected cells was photobleached, fluorescence recovery first occurred directly beneath the bacterium in a band that grew wider at a rate of one micron/minute . Consistently observed in all EPEC-induced pedestals, whether they were stationary, lengthening, or translocating, the rate of actin polymerization that occurred at the pedestal tip was approximately 1 mum/min . Overall, a much slower rate of actin polymerization was measured in long EHEC-induced pedestals . In contrast to the dynamics of GFP-actin, recovery of GFP-alpha-actinin fluorescence was not polarized, with the actin cross-linking protein exchanging all the length of the EPEC/EHEC induced pedestals . Surprisingly, the depolymerization and retrograde flow of pedestal actin, as well as pedestal translocations, were inhibited reversibly by either 2,3-butanedione monoxime (BDM) or by a combination of sodium azide and 2-deoxy D-glucose, leading to an increase in the lengths of the pedestals . A simple physical model was developed to describe elongation and translocation of EPEC/EHEC pedestals in terms of actin polymerization and depolymerization dynamics . Cell Motil . Cytoskeleton 60:104-120, 2005 . (c) 2004 Wiley-Liss, Inc. Rev Esp Cardiol, 2002 Jul, 55(Supl.1), 2 - 9 Mycoplasma Pneumoniae and Chlamydia Pneumoniae are associated to inflammation and rupture of the atherosclerotic coronary plaques; F Ramires JA et al.; In this review we report recent findings of our lab showing that Mycoplasma pneumoniae and Chlamydia pneumoniae are present in higher amount, associated with adventitial inflammation and positive vessel remodeling, in thrombosed coronary artery segments (CAS) of patients who died due to acute myocardial infarction . CD8T cell was the predominant lymphocytes in the plaque and CD24(B) cell in the adventitia . The mean numbers of lymphocytes were significantly higher in adventitia than in the plaque . Vulnerable plaques were usually associated with focal positive vessel remodeling and large lipidic atheromas . Mycoplasma is the only bacterium that needs cholesterol for proliferation . We hypothesized that the association of Mycoplasma pneumoniae and Chlamydia pneumoniae increases virulence of both bacteria, inducing inflammation and rupture of the plaque . The search of CMV and Helicobacter pylori resulted negative. Hum Fertil (Camb), 2004 Dec, 7(4), 271 - 6 Chlamydia trachomatis and male fertility; Pacey A et al.; There is increasing evidence that the function of human spermatozoa can be significantly affected by direct exposure to the bacterium Chlamydia trachomatis . This may contribute to sub-fertility in infected individuals by a route that is independent of any damage to the reproductive epithelium . In addition, if a C . trachomatis infection is undiagnosed it could contribute to poor outcomes in assisted conception techniques such as in vitro fertilization . The antibiotics routinely used in IVF culture systems are largely ineffective against chlamydia, emphasizing the importance of screening patients prior to treatment . Moreover, given the many thousands of semen samples provided for analysis by men in primary care (many of which will never undergo assisted conception treatment), it is suggested that this may represent a wasted opportunity to provide screening (and treatment) for the infection using an appropriate test specimen and without the need for additional hospital visits. J Inorg Biochem, 2005 Feb, 99(2), 415 - 23 Single crystal EPR study of electronic structure and exchange interactions for copper(II)(l-arginine)(2)(SO(4)).(H(2)O)(6): a model system to study exchange interactions between unpaired spins in proteins; Santana RC et al.; We report EPR measurements at 9.77 and 34.1 GHz in powder and single crystal samples of the ternary copper amino acid complex Cu(l-arginine)(2)(SO(4)).(H(2)O)(6) . The single crystal Electron Paramagnetic Resonance spectra display a single resonance for all magnetic field orientations in the ca and cb crystal planes . In the ab plane they display two resonances for most orientations of the magnetic field, and only one resonance for orientations close to the crystal axes . This behavior is a result of the selective collapse of the resonances corresponding to the four copper sites in the unit cell produced by the exchange interactions between copper ions . From the characteristics of the collapse and the angular dependences of the position and width of the resonances we evaluate the g-tensors of the copper molecules and estimate exchange interactions |J(1)/k(B)|=0.9 K and |J(2)/k(B)|=0.009 K between copper neighbors at 5.908 A and at 15.684 A, respectively . J(1) is assigned to a syn-anti equatorial-apical carboxylate bridge with a total bond length of 7.133 A . J(2) is assigned to a long bridge of 12 atoms with a total bond length of 19.789 A, that includes two hydrogen bonds . The results are discussed in terms of the crystal and electronic structure of Cu(l-arginine)(2)(SO(4)).(H(2)O)(6) . We show that J(2) is in excellent agreement with the observed magnetic interaction between the reduced quinone acceptors in the photosynthetic reaction center protein of the bacterium Rb . sphaeroides, which is transmitted along a similar chemical path containing two hydrogen bonds . Our findings indicate that it is valid to estimate values for the exchange interactions between redox centers in proteins transmitted along long chemical paths containing sigma and H-bonds, from data obtained in model systems, and emphasize the importance of measuring exchange interactions in biologically relevant model systems. Carbohydr Res, 2005 Jan 17, 340(1), 69 - 74 Structure of an acidic polysaccharide from the agar-decomposing marine bacterium Pseudoalteromonas atlantica strain IAM 14165 containing 5,7-diacetamido-3,5,7,9-tetradeoxy-l-glycero-l-manno-non-2-ulosonic acid; Perepelov AV et al.; The structure of an acidic polysaccharide from Pseudoalteromonas atlantica strain 14165 containing 5,7-diacetamido-3,5,7,9-tetradeoxy-l-glycero-l-manno-non-2-ulosonic acid (di-N-acetylpseudaminic acid, Pse5Ac7Ac) has been elucidated . The polysaccharide was studied by (1)H and (13)C NMR spectroscopy, including 2D experiments, along with sugar and methylation analyses . After a selective hydrolysis a modified polysaccharide devoid of its side chain could be isolated . It was found that the polysaccharide has pentasaccharide repeating units with following structure: Biosci Biotechnol Biochem, 2004 Dec, 68(12), 2519 - 28 Involvement of Sulfide:Quinone Oxidoreductase in Sulfur Oxidation of an Acidophilic Iron-Oxidizing Bacterium, Acidithiobacillus ferrooxidans NASF-1; Wakai S et al.; The effects of cyanide, azide, and 2-n-Heptyl-4-hydroxy-quinoline-N-oxide (HQNO) on the oxidation of ferrous ion or elemental sulfur with Acidithiobacillus ferrooxidans NASF-1 cells grown in iron- or sulfur-medium were examined . The iron oxidation of both iron- and sulfur-grown cells was strongly inhibited by cyanide and azide, but not by HQNO . Sulfur oxidation was relatively resistant to cyanide and azide, and inhibited by HQNO . Higher sulfide oxidation, ubiquinol dehydrogenase activity, and sulfide:quinone oxidoreductase (SQR) activity were observed in sulfur-grown cells more than in iron-grown cells . Sulfide oxidation in the presence of ubiquinone with the membrane fraction was inhibited by HQNO, but not by cyanide, azide, antimycin A, and myxothiazol . The transcription of three genes, encoding an aa(3)-type cytochrome c oxidase (coxB), a bd-type ubiquinol oxidase (cydA), and an sqr, were measured by real-time reverse transcription polymerase chain reaction . The transcriptional levels of coxB and cydA genes were similar in sulfur- and iron-grown cells, but that of sqr was 3-fold higher in sulfur-grown cells than in iron-grown cells . A model is proposed for the oxidation of reduced inorganic sulfur compounds in A . ferrooxidans NASF-1 cells. Infect Immun, 2005 Jan, 73(1), 155 - 65 NF-kappaB activation during Rickettsia rickettsii infection of endothelial cells involves the activation of catalytic IkappaB kinases IKKalpha and IKKbeta and phosphorylation-proteolysis of the inhibitor protein IkappaBalpha; Clifton DR et al.; Rocky Mountain spotted fever, a systemic tick-borne illness caused by the obligate intracellular bacterium Rickettsia rickettsii, is associated with widespread infection of the vascular endothelium . R . rickettsii infection induces a biphasic pattern of the nuclear factor-kappaB (NF-kappaB) activation in cultured human endothelial cells (ECs), characterized by an early transient phase at 3 h and a late sustained phase evident at 18 to 24 h . To elucidate the underlying mechanisms, we investigated the expression of NF-kappaB subunits, p65 and p50, and IkappaB proteins, IkappaBalpha and IkappaBbeta . The transcript and protein levels of p50, p65, and IkappaBbeta remained relatively unchanged during the course of infection, but Ser-32 phosphorylation of IkappaBalpha at 3 h was significantly increased over the basal level in uninfected cells concomitant with a significant increase in the expression of IkappaBalpha mRNA . The level of IkappaBalpha mRNA gradually returned toward baseline, whereas that of total IkappaBalpha protein remained lower than the corresponding controls . The activities of IKKalpha and IKKbeta, the catalytic subunits of IkappaB kinase (IKK) complex, as measured by in vitro kinase assays with immunoprecipitates from uninfected and R . rickettsii-infected ECs, revealed significant increases at 2 h after infection . The activation of IKK and early phase of NF-kappaB response were inhibited by heat treatment and completely abolished by formalin fixation of rickettsiae . The IKK inhibitors parthenolide and aspirin blocked the activities of infection-induced IKKalpha and IKKbeta, leading to attenuation of nuclear translocation of NF-kappaB . Also, increased activity of IKKalpha was evident later during the infection, coinciding with the late phase of NF-kappaB activation . Thus, activation of catalytic components of the IKK complex represents an important upstream signaling event in the pathway for R . rickettsii-induced NF-kappaB activation . Since NF-kappaB is a critical regulator of inflammatory genes and prevents host cell death during infection via antiapoptotic functions, selective inhibition of IKK may provide a potential target for enhanced clearance of rickettsiae and an effective strategy to reduce inflammatory damage to the host during rickettsial infections. Cell Microbiol, 2005 Jan, 7(1), 29 - 38 Anaplasma phagocytophilum inhibits human neutrophil apoptosis via upregulation of bfl-1, maintenance of mitochondrial membrane potential and prevention of caspase 3 activation; Ge Y et al.; Summary The inhibition of neutrophil apoptosis plays a central role in human granulocytic anaplasmosis . Intracellular signalling pathways through which the obligatory intracellular bacterium Anaplasma phagocytophilum inhibits the spontaneous apoptosis of human peripheral blood neutrophils were investigated . bfl-1 mRNA levels in uninfected neutrophils after 12 h in culture were reduced to approximately 5-25% of 0 h levels, but remained high in infected neutrophils . The eukaryotic RNA synthesis inhibitor, actinomycin D, prevented the maintenance of bfl-1 mRNA levels by A . phagocytophilum . Differences in mcl-1, bax, bcl-w, bad or bak mRNA levels in infected versus uninfected neutrophils were not remarkable . By using mitochondrial fluorescent dyes, Mitotracker Red and JC-1, it was found that most uninfected neutrophils lost mitochondrial membrane potential after 10-12 h incubation, whereas A . phagocytophilum-infected neutrophils maintained high membrane potential . Caspase 3 activity and the degree of apoptosis were lower in dose-dependent manner in A . phagocytophilum-infected neutrophils at 16 h post infection, as compared to uninfected neutrophils . Anti-active caspase 3 antibody labelling showed less positively stained population in infected neutrophils compared to those in uninfected neutrophils after 12 h incubation . These results suggest that A . phagocytophilum inhibits human neutrophil apoptosis via transcriptional upregulation of bfl-1 and inhibition of mitochondria-mediated activation of caspase 3. Acta Crystallogr D Biol Crystallogr, 2004 Dec, 60(Pt 12 Pt 2), 2361 - 3 Crystallization and preliminary X-ray characterization of the atypical glutaminyl-tRNA synthetase from Deinococcus radiodurans; Deniziak MA et al.; The glutaminyl-tRNA synthetase (GlnRS) from the radiation-resistant bacterium Deinococcus radiodurans differs from known GlnRSs and other tRNA synthetases by the presence of an additional C-terminal domain resembling the C-terminal region of the GatB subunit of tRNA-dependent amidotransferase (AdT) . This atypical synthetase was overexpressed in Escherichia coli, purified and crystallized in the presence of PEG 3350 . Orthorhombic crystals were obtained that belong to space group P2(1)2(1)2(1) and diffract to 2.3 A resolution . The crystal structure was solved by molecular replacement using the structure of E . coli GlnRS as a search model. J Infect Chemother, 2004 Dec, 10(6), 316 - 25 Animal models for the study of Helicobacter-induced gastric carcinoma; Kodama M et al.; Helicobacter pylori is considered to have a close association with gastric cancer . Many epidemiological studies have shown a strong association between chronic H . pylori infection and subsequent development of gastric carcinoma in humans . To clarify this link more clearly, it is necessary to use this bacterium in experimental studies to develop gastric carcinoma in suitable experimental animals . Persistent H . pylori infection was seen in the Japanese monkey model, and has recently been achieved in the Mongolian gerbil model . In these models, the sequential histopathological changes in the gastric mucosa are very similar to those in humans . The Japanese monkey model showed advances in atrophic change and p53 point mutations in the gastric mucosa during long-term observation . The Mongolian gerbil model demonstrated that H . pylori infection enhances gastric carcinogenesis in combination with known carcinogens such as N-methyl-N-nitrosourea (MNU) and N-methyl-N-nitro-N'-nitrosoguanidine (MNNG), and also showed that H . pylori infection alone can result in the development of gastric carcinoma . These important results provide a starting point for further studies to clarify the mechanism of gastric carcinogenesis as a result of H . pylori infection and assist in the planning of eradication therapy to prevent gastric carcinoma. Biophys J . 2004 Dec 21; {Epub ahead of print} Multiple scattering X-ray absorption studies of Zn2+ binding sites in bacterial photosynthetic reaction centers; Giachini L et al.; Binding of transition metal ions to the reaction center (RC) protein of the photosynthetic bacterium Rhodobacter sphaeroides has been previously shown to slow light-induced electron and proton transfer to the secondary quinone acceptor molecule, QB . On the basis of X-ray diffraction (XRD) at 2.5 A resolution a site, formed by AspH124, HisH126 and HisH128, has been identified at the protein surface which binds Cd(2+) or Zn(2+) . Using Zn K-edge X-ray absorption fine structure (XAFS) spectroscopy we report here on the local structure of Zn(2+) ions bound to purified RC complexes embedded into polyvinyl alcohol films . XAFS data were analysed by combining ab-initio simulations and multiparameter fitting; structural contributions up to the fourth coordination shell and multiple scattering paths (involving three atoms) have been included . Results for complexes characterized by a Zn to RC stoichiometry close to 1 indicate that Zn(2+) binds two O and two N atoms in the first coordination shell . Higher shell contributions are consistent with a binding cluster formed by two His, one Asp residue and a water molecule . Analysis of complexes characterized by approximately 2 Zn ions per RC reveals a second structurally distinct binding site, involving one O and three N atoms, not belonging to a His residue . The local structure obtained for the higher affinity site nicely fits the coordination geometry proposed on the basis of XRD data, but detects a significant contraction of the first shell . Two possible locations of the second new binding site at the cytoplasmic surface of the RC are proposed. Syst Appl Microbiol, 2004 Nov, 27(6), 653 - 60 Mycobacterium fluoranthenivorans sp . nov., a fluoranthene and aflatoxin B1 degrading bacterium from contaminated soil of a former coal gas plant; Hormisch D et al.; Mycobacterium strain FA4T was isolated with fluoranthene as the single carbon source from soil of a former coal gas plant, polluted with polycyclic aromatic hydrocarbons . The physiological properties, fatty acid pattern, and the 16S ribosomal RNA gene sequence indicated membership to the genus Mycobacterium, but were different from all type strains of Mycobacterium species . Based on comparative 16S rRNA gene sequence analyses strain FA4T could be assigned to the Mycobacterium neoaurum taxon showing 98% sequence similarity to M . diernhoferi as its closest neighbour . The occurrence of epoxymycolate in the cell wall differentiates FA4 from all members of this taxon which synthesize wax-ester mycolates in addition to alpha-mycolates . Strain FA4T is able to degrade aflatoxin B1 . This biological attribute might be useful in biological detoxification processes of foods and feeds . From the investigated characteristics it is concluded that strain FA4T represents a new species, for which we propose the name Mycobacterium fluoranthenivorans sp . nov . The type strain of Mycobacterium fluoranthenivorans is FA4T (DSM 44556T = CIP 108203T). Biofizika, 2004 Nov-Dec, 49(6), 1069 - 74 {A study of the content of pigments in the light-harvesting antenna of the green bacterium from the new family Oscillochloridaceae} {Physicochemical properties of two-component polyhydroxyalkanoates {poly(3HB/3HV)}} {No authors listed} A series of two-component polyhydroxyalkanoates consisting of hydroxybutyrate and hydroxyvalerate monomer at different ratios were synthesized using the bacterium Ralstonia eutropha B5786 . The properties of polyhydroxyalkanoates were compared with those of the homopolymer of hydroxybutyric acid by X-ray structure analysis, IR spectroscopy, differential scanning calorimetry, and viscosimetry . With an increase in the molar fraction of hydroxyvalerate, an equalization of the ratio of the crystalline and amorphous phases in the copolymer was observed . The degree of crystallinity of the polymer decreased from 70-80 to 45-50%; in the range of an increase in the hydroxyvalerate molar fraction from several to 25-30 mol%, the dependence was linear . The temperature characteristics, the melting temperature (T(m)), and the degradation temperature (T(d)) were lower in polyhydroxyalkanoates than in polyhydroxybutyrate, for which T(m) and T(d) were 168-170 and 260-265 degrees C, respectively . In the copolymer, as the molar fraction of hydroxyvalerate grew, both parameters decreased . In the range of variation of monomer ratio studied, they decreased to 150-160 and 200-220 degrees C, respectively . No distinct correlation between the composition of the polymer and its molecular mass was found. Dis Aquat Organ, 2004 Nov 4, 61(3), 227 - 33 Experimental infection of Pacific white shrimp Litopenaeus vannamei with Necrotizing Heptopancreatitis (NHP) bacterium by per os exposure; Vincent AG et al.; Necrotizing Hepatopancreatitis Bacterium (NHPB), which causes Necrotizing Hepatopancreatitis, was successfully transmitted in individually isolated Kona stock Litopenaeus vannamei through per os exposure . Animals (140) were individually exposed orally to a 0.05 g piece of an NHPB-infected hepatopancreas and 120 control animals were each exposed to a 0.05 g piece of NHPB-negative hepatopancreas . Shrimp were maintained in Sterilite containers with approximately 41 of artificial seawater at 30 per thousand salinity and 30 degrees C for 60 d . Mortality of infected shrimp was observed from Day 16 to Day 51 post-exposure . Infected animals sustained reduced feeding activity and displayed empty guts . Some infected animals developed a pale hepatopancreas noticeable through the carapace . Survival probabilities fit a Weibull distribution and parametric survival analysis revealed lowered survival due to NHPB infection . Median survival time of NHPB-infected animals was 34.5 d . After correcting for background daily mortality in the controls, mean acute daily mortality of NHPB was estimated at 0.09, a value much lower than that estimated for other diseases in Kona stock L . vannamei such as White Spot Syndrome Virus (0.40) and Taura Syndrome Virus (0.30) . A chronic, or carrier, state was not demonstrated in NHPB epizootics because all NHPB-positive animals experienced mortality and no animals surviving to 60 d post-exposure were diagnosed NHPB-positive through PCR or histology. Eur Biophys J . 2004 Dec 18; {Epub ahead of print} Sizing-up finite fluorescent particles with nanometer-scale precision by convolution and correlation image analysis; Gennerich A et al.; Determining the positions, shapes and sizes of finite living particles such as bacteria, mitochondria or vesicles is of interest in many biological processes . In fluorescence microscopy, algorithms that can simultaneously localize such particles as a function of time and determine the parameters of their shapes and sizes at the nanometer scale are not yet available . Here we develop two such algorithms based on convolution and correlation image analysis that take into account the position, orientation, shape and size of the object being tracked, and we compare the precision of the two algorithms using computer simulations . We show that the precision of both algorithms strongly depends on the object's size . In cases where the diameter of the object is larger than about four to five times the beam waist radius, the convolution algorithm gives a better precision than the correlation algorithm (it leads to more precise parameters), while for smaller object diameters, the correlation algorithm gives superior precision . We apply the convolution algorithm to sequences of confocal laser scanning micrographs of immobile Escherichia coli bacteria, and show that the centroid, the front end, the rear end, the left border and the right border of a bacterium can be determined with a signal-to-noise-dependent precision down to ~5 nm. J Physiol Pharmacol, 2004 Jul, 55 Suppl 2, 5 - 17 Duodenal mucosal protection by bicarbonate secretion and its mechanisms; Konturek SJ et al.; Proximal portion of duodenum is exposed to intermittent pulses of gastric H(+) discharged by the stomach . This review summarizes the mechanisms of duodenal mucosal integrity, mainly the role of mucus-alkaline secretion and the mucous barrier protecting surface epithelium against gastric H(+) . The mucous barrier protects the leaky duodenal epithelium against each pulse of gastric H(+), which penetrates this barrier and diffuses into duodenocytes, but fails to damage them due to; a) an enhanced expression of cyclooxygenase-1 (COX-1), with release of protective prostaglandins (PG) and of nitric oxide (NO) synthase (NOS) with, however, production of NO, stimulating duodenal HCO(3)(-) secretion and b) the release of several neurotransmitters also stimulating HCO(3)(-) secretion such as vasoactive intestimal peptide (VIP), pituitary adenylate-cyclase activating polypeptide (PACAP), acetylcholine, melatonin, leptin and ghrelin released by enteric nerves and mucosal cells . At the apical duodenocyte membrane at least two HCO(3)(-)/Cl(-) anion exchangers operate in response to luminal H(+) to provide adequate extrusion of HCO(3)(-) into duodenal lumen . In the basolateral portion of duodenocyte membrane, both non-electrogenic (NBC) and electrogenic (NBC(n)) Na(+) HCO(3)(-) cotransporters are activated by the exposure to duodenal acidification, causing inward movement of HCO(3)(-) from extracellular fluid to duodenocytes . There are also at least three Na(+)/H(+) (NHE1-3) amiloride-sensitive exchangers, eliminating H(+)which diffused into these cells . The Helicobacter pylori (Hp) infection and gastric metaplasia in the duodenum with bacterium inoculating metaplastic mucosa and inhibiting HCO(3)(-) secretion by its endogenous inhibitor, asymetric dimethyl arginine (ADMA), may result in duodenal ulcerogenesis. Biochem Biophys Res Commun, 2005 Jan 28, 326(4), 777 - 81 Cytokine induction by a bacterial DNA-specific modified base; Tsuchiya H et al.; Unmethylated CpG dinucleotides in DNA contribute to a rapid inflammatory response in mammals . Here we show that N(6)-methyladenine (N(6)-MeA), a bacterium-specific modified base, also causes cytokine production . An oligodeoxyribonucleotide (ODN) containing N(6)-MeA induced cytokines when injected into mice . Co-injection of N(6)-MeA and CpG ODNs enhanced cytokines 2- to 3-fold, as compared with the injection of a CpG ODN alone . Plasmid DNA containing N(6)-MeA, complexed with cationic lipids, induced IL-12 . These results indicate that the bacterium-specific base, in addition to the unmethylated CpG motif, triggers the mammalian immune response, and suggest that N(6)-MeA-containing DNA could be useful for cellular immunotherapy and DNA vaccine. Biotechnol Lett, 2004 Oct, 26(19), 1511 - 5 Expression and characterization of a thermostable beta-xylosidase from the hyperthermophile, Thermotoga maritima; Xue Y et al.; A thermostable beta-xylosidase from a hyperthermophilic bacterium, Thermotoga maritima, was over-expressed in Escherichia coli using the T7 polymerase expression system . The expressed beta-xylosidase was purified in two steps, heat treatment and immobilized metal affinity chromatography, and gave a single band on SDS-PAGE . The maximum activity on p-nitrophenyl beta-D-xylopyranoside was at 90 degrees C and pH 6.1 . The purified enzyme had a half-life of over 22-min at 95 degrees C, and retained over 57% of its activity after holding a pH ranging from 5.4 to 8.5 for 1 h at 80 degrees C . Among all tested substrates, the purified enzyme had specific activities of 275, 50 and 29 U mg(-1) on pNPX, pNPAF, and pNPG, respectively . The apparent Michaelis constant of the beta-xylosidase was 0.13 mM for p NPX with a V (max) of 280 U mg(-1) . When the purified beta-xylosidase was added to xylanase, corncob xylan was hydrolized completely to xylose. Proc Natl Acad Sci U S A, 2004 Dec 28, 101(52), 17994 - 9 Epub 2004 Dec 15. The long-range organization of a native photosynthetic membrane; Frese RN et al.; Photosynthesis relies on the delicate interplay between a specific set of membrane-bound pigment-protein complexes that harvest and transport solar energy, execute charge separation, and conserve the energy . We have investigated the organization of the light-harvesting (LH) and reaction-center (RC) complexes in native bacterial photosynthetic membranes of the purple bacterium Rhodobacter sphaeroides by using polarized light spectroscopy, linear dichroism (LD) on oriented membranes . These LD measurements show that in native membranes, which contain LH2 as the major energy absorber, the RC-LH1-PufX complexes are highly organized in a way similar to that which we found previously for a mutant lacking LH2 . The relative contribution of LH1 and LH2 light-harvesting complexes to the LD spectrum shows that LH2 preferentially resides in highly curved parts of the membrane . Combining the spectroscopic data with our recent atomic force microscopy (AFM) results, we propose an organization for this photosynthetic membrane that features domains containing linear arrays of RC-LH1-PufX complexes interspersed with LH2 complexes and some LH2 found in separate domains . The study described here allows the simultaneous assessment of both global and local structural information on the organization of intact, untreated membranes. J Bacteriol, 2005 Jan, 187(1), 257 - 65 The elastic properties of the caulobacter crescentus adhesive holdfast are dependent on oligomers of N-acetylglucosamine; Li G et al.; The aquatic bacterium Caulobacter crescentus attaches to solid surfaces through an adhesive holdfast located at the tip of its polar stalk, a thin cylindrical extension of the cell membrane . In this paper, the elastic properties of the C . crescentus stalk and holdfast assembly were studied by using video light microscopy . In particular, the contribution of oligomers of N-acetylglucosamine (GlcNAc) to the elasticity of holdfast was examined by lysozyme digestion . C . crescentus cells attached to a surface undergo Brownian motion while confined effectively in a harmonic potential . Mathematical analysis of such motion enabled us to determine the force constant of the stalk-holdfast assembly, which quantifies its elastic properties . The measured force constant exhibits no dependence on stalk length, consistent with the theoretical estimate showing that the stalk can be treated as a rigid rod with respect to fluctuations of the attached cells . Therefore, the force constant of the stalk-holdfast assembly can be attributed to the elasticity of the holdfast . Motions of cells in a rosette were found to be correlated, consistent with the elastic characteristics of the holdfast . Atomic force microscopy analysis indicates that the height of a dried (in air) holdfast is approximately one-third of that of a wet (in water) holdfast, consistent with the gel-like nature of the holdfast . Lysozyme, which cleaves oligomers of GlcNAc, reduced the force constant to less than 10% of its original value, consistent with the polysaccharide gel-like nature of the holdfast . These results also indicate that GlcNAc polymers play an important role in the strength of the holdfast. J Bacteriol, 2005 Jan, 187(1), 92 - 8 Identification of two new genes involved in diazotrophic growth via the alternative Fe-only nitrogenase in the phototrophic purple bacterium Rhodobacter capsulatus; Sicking C et al.; Growth of Rhodobacter capsulatus with molecular dinitrogen as the sole N source via the alternative Fe-only nitrogenase requires all seven gene products of the anfHDGK-1-2-3 operon . In contrast to mutant strains carrying lesions in the structural genes of nitrogenase (anfH, anfD, anfG, and anfK), strains defective for either anf1, anf2, or anf3 are still able to reduce the artificial substrate acetylene, although with diminished activity . To obtain further information on the role of Anf1, we screened an R . capsulatus genomic library designed for use in yeast two-hybrid studies with Anf1 as bait . Two genes, which we propose to call ranR and ranT (for genes related to alternative nitrogenase), coding for products that interact with Anf1 were identified . A ranR mutant exhibited a phenotype similar to that of an anf1 mutant strain (no growth with N2 in the absence of molybdenum, but significant reduction of acetylene via the Fe-only nitrogenase), whereas a ranT mutant retained the ability to grow diazotrophically, but growth was clearly delayed compared to the parental strain . In contrast to the situation for anf1, expression of neither ranR nor ranT was regulated by ammonium or molybdenum . A putative role for Anf1, RanR, and RanT in the acquisition and/or processing of iron in connection with the Fe-only nitrogenase system is discussed. Extremophiles . 2004 Dec 15; {Epub ahead of print} Cold-active DnaK of an Antarctic psychrotroph Shewanella sp . Ac10 supporting the growth of dnaK-null mutant of Escherichia coli at cold temperatures; Yoshimune K et al.; Shewanella sp . Ac10 is a psychrotrophic bacterium isolated from the Antarctica that actively grows at such low temperatures as 0 degrees C . Immunoblot analyses showed that a heat-shock protein DnaK is inducibly formed by the bacterium at 24 degrees C, which is much lower than the temperatures causing heat shock in mesophiles such as Escherichia coli . We found that the Shewanella DnaK (SheDnaK) shows much higher ATPase activity at low temperatures than the DnaK of E . coli (EcoDnaK): a characteristic of a cold-active enzyme . The recombinant SheDnaK gene supported neither the growth of a dnaK-null mutant of E . coli at 43 degrees C nor lambda phage propagation at an even lower temperature, 30 degrees C . However, the recombinant SheDnaK gene enabled the E . coli mutant to grow at 15 degrees C . This is the first report of a DnaK supporting the growth of a dnaK-null mutant at low temperatures. J Biochem (Tokyo), 2004 Sep, 136(3), 363 - 9 Reconstitution and replacement of bacteriochlorophyll a molecules in photosynthetic reaction centers; Kobayashi M et al.; Reaction centers (RCs) of the photosynthetic bacterium Rhodobacter sphaeroides R-26 were reconstituted in liposomes after release of pigments (bacteriochlorophyll a (BChla) and bacteriopheophytin a (BPhea)) by treatment with acetone . As shown by absorption and circular dichroism spectroscopies, the reconstituted RCs had the same arrangement of pigments as the native RC and exhibited photoactivity of the special pair . The recovery yield of RCs of up to 30% was achieved by addition of 7.8-fold excess of BChla in the acetone treatment . Furthermore BChla was partially replaced with Zn-BChla by addition of the pigments during the acetone treatment . About 30% and 50% of the special pair and accessory pigments can be replaced with Zn-BChla, respectively . From this rate, an oxidation-reduction potential of 520 mV (vs . the normal hydrogen electrode NHE) was derived by the simulation of the experimental data, which is 35 mV higher than that of the native RC (484 mV vs . NHE). FEMS Microbiol Lett, 2004 Dec 15, 241(2), 151 - 6 Expression of glutathione S-transferase and peptide methionine sulphoxide reductase in Ochrobactrum anthropi is correlated to the production of reactive oxygen species caused by aromatic substrates; Tamburro A et al.; Peptide methionine sulphoxide reductase (MsrA) and glutathione S-transferases (GSTs) are considered as detoxification enzymes . In the xenobiotics-degrading bacterium Ochrobactrum anthropi the two enzymes are co-induced by toxic concentrations of aromatic substrates such as phenol and 4-chlorophenol . In aerobic organisms, degradation of aromatic substrates by mono- and dioxygenases leads to a generation of oxidative stress that causes the occurrence of reactive oxygen species (ROS) . A capillary electrophoretic method, using the intracellular conversion of dihydrorhodamine-123 into rhodamine-123, was developed to measure the content of ROS in the bacteria . The presence of toxic concentrations of the aromatic substrate 4-chlorophenol, an inducer of GST and MsrA, leads to a significant increase in the production of ROS . These results strongly suggest that GST and MsrA enzymes are part of the bacterial defence mechanism against particular oxidative stress conditions . As oxidative stress is known to be present predominantly close to the cytoplasmic membrane, we investigated the subcellular distribution of both MsrA and GST enzymes in this bacterium grown in the presence of 4-chlorophenol . By Western blotting, MsrA and GST was assayed in the cytoplasm as well as in the periplasm . Moreover, immunolocalisation by colloidal gold immunoelectron microscopy identified the two proteins associated with the cell envelope. J Inorg Biochem, 2005 Jan, 99(1), 280 - 92 CooA, a paradigm for gas sensing regulatory proteins; Roberts GP et al.; The heme-containing transcriptional factor CooA regulates the expression of genes involved in the oxidation of carbon monoxide (CO) in the bacterium Rhodospirillum rubrum . CooA is both a redox sensor and a specific CO sensor, a combination of properties that is unique among heme proteins . Extensive biochemical and genetic analyses, interpreted in the context of a crystal structure of one form of the protein, have allowed the creation of hypotheses concerning the mechanism of CooA activation by CO as well as the basis for its CO specificity . The article details the data in support of these hypotheses and indicates future lines of research. Clin Exp Med, 2004 Sep, 4(1), 39 - 43 Progress, problems, and perspectives in diagnosis and treatment of Whipple's disease; Mahnel R et al.; Whipple's disease is a rare chronic infectious disorder first described in 1907 by G.H . Whipple . The disorder is caused by the newly identified bacterium Tropheryma whipplei and there is evidence that altered immune functions play a role in the manifestation of the disease . The organ systems mostly affected are the joints and the gut, and in the further course often also the heart, lung, brain, and eyes . The intestinal involvement occurs with abdominal pain and diarrhea, which leads to weight loss, malnutrition, and anemia . In some cases the infection spreads to the central nervous system, which may lead to loss of memory, confusion, or disturbances in gait . In the last few years, several steps towards an improved diagnosis of the disease and characterization of the causative bacterium have been made . While untreated disease may be lethal, treatment is often able to eradicate the organism . At present, therapy is based on observations in small patient groups and personal experience . There are different antibiotic therapy regimens often starting with intravenous application for 2 weeks followed by oral medication for 1 year . The first clinical therapy study is ongoing. Proc Natl Acad Sci U S A, 2004 Dec 21, 101(51), 17737 - 40 Epub 2004 Dec 10. Mitochondrial cytochrome c1 is a collapsed di-heme cytochrome; Baymann F et al.; Cytochrome c(1) from mitochondrial complex III and the di-heme cytochromes c in the corresponding enzyme from epsilon-proteobacteria have so far been considered to represent unrelated cytochromes . A missing link protein discovered in the genome of the hyperthermophilic bacterium Aquifex aeolicus, however, provides evidence for a close evolutionary relationship between these two cytochromes . The mono-heme cytochrome c(1) from A . aeolicus contains stretches of strong sequence homology toward the epsilon-proteobacterial di-heme cytochromes . These di-heme cytochromes are shown to belong to the cytochrome c(4) family . Mapping cytochrome c(1) onto the di-heme sequences and structures demonstrates that cytochrome c(1) results from a mutation-induced collapse of the di-heme cytochrome structure and provides an explanation for its uncommon structural features . The appearance of cytochrome c(1) thus represents an extension of the biological protein repertoire quite different from the widespread innovation by gene duplication and subsequent diversification. Microbiology, 2004 Dec, 150(Pt 12), 4085 - 93 Characterization of a Myxococcus xanthus mutant that is defective for adventurous motility and social motility; Lancero H et al.; Myxococcus xanthus is a gliding bacterium that possesses two motility systems, the adventurous (A-motility) and social (S-motility) systems . A-motility is used for individual cell gliding, while S-motility is used for gliding in multicellular groups . Video microscopy studies showed that nla24 cells are non-motile on agar surfaces, suggesting that the nla24 gene product is absolutely required for both A-motility and S-motility under these assay conditions . S-motility requires functional type IV pili, wild-type LPS O-antigen, and an extracellular matrix of exopolysaccharide (EPS) and protein called fibrils . The results of expression studies and tethering assays indicate that the nla24 mutant has functional type IV pili . The nla24 mutant also produces wild-type LPS . However, several lines of evidence suggest that the nla24 mutant is defective for production of the EPS portion of the fibril matrix . The nla24 mutant is also defective for transcription of two genes (aglU and cglB) known to be required for A-motility, which is consistent with the idea that nla24 cells are defective for A-motility . Based on these findings, it is proposed that the putative transcriptional activator Nla24 regulates a subset of genes that are important for A-motility and S-motility in M . xanthus. J Invertebr Pathol, 2004 Oct-Nov, 87(2-3), 114 - 22 Insect cellular and chemical limitations to pathogen development: the Colorado potato beetle, the nematode Heterorhabditis marelatus, and its symbiotic bacteria; Armer CA et al.; This research examines possible factors limiting pathogen development and reproduction in a novel host insect . The nematode Heterorhabditis marelatus and its symbiotic bacterium, Photorhabdus luminescens, kill 98% of nematode-treated Colorado potato beetle (CPB) prepupae, but the nematode reproduces in only 1-6% of beetles . We examined nematode/bacterial inhibition at each step of the normal developmental pathway to determine host feature(s) limiting nematode reproduction . We found that in vivo encapsulation of nematodes occurred in only 1.6% of CPB, and in 5% of in vitro hanging drops of hemolymph . Thus, the cellular defense system did not strongly limit nematode reproduction in the CPB . The symbiotic bacterium was negatively affected by a heat-labile factor found in the CPB's hemolymph which often caused the bacterium to switch from the primary form that produces antibiotics and nutrients necessary for the nematodes' development, to a secondary form that provides only limited nutrients . A 58 kDa protein was isolated and bioassayed for activity against P . luminescens, but caused a delay in bacterial growth rather than the primary-secondary form switch . Thus, the identity of the heat-labile factor could not be confirmed as being the 58 kDa protein . The heat-labile factor did not directly affect the nematode . The addition of lipids in the form of olive oil to heated CPB hemolymph allowed nematodes to reproduce in 17% of hanging drops, in contrast to zero reproduction in hemolymph without oil . Reproductive nematodes were smaller when grown in CPB hemolymph than in hemolymph of the highly susceptible Galleria mellonella . These data suggest that both the toxic heat-labile factor and a lack of appropriate nutrients alter the CPB-bacterium-nematode interaction . These factors preclude the use of this otherwise highly effective nematode-bacterial complex in the longterm control of the CPB. J Bacteriol, 2004 Dec, 186(24), 8385 - 400 Transcriptomic and proteomic characterization of the Fur modulon in the metal-reducing bacterium Shewanella oneidensis; Wan XF et al.; The availability of the complete genome sequence for Shewanella oneidensis MR-1 has permitted a comprehensive characterization of the ferric uptake regulator (Fur) modulon in this dissimilatory metal-reducing bacterium . We have employed targeted gene mutagenesis, DNA microarrays, proteomic analysis using liquid chromatography-mass spectrometry, and computational motif discovery tools to define the S . oneidensis Fur regulon . Using this integrated approach, we identified nine probable operons (containing 24 genes) and 15 individual open reading frames (ORFs), either with unknown functions or encoding products annotated as transport or binding proteins, that are predicted to be direct targets of Fur-mediated repression . This study suggested, for the first time, possible roles for four operons and eight ORFs with unknown functions in iron metabolism or iron transport-related functions . Proteomic analysis clearly identified a number of transporters, binding proteins, and receptors related to iron uptake that were up-regulated in response to a fur deletion and verified the expression of nine genes originally annotated as pseudogenes . Comparison of the transcriptome and proteome data revealed strong correlation for genes shown to be undergoing large changes at the transcript level . A number of genes encoding components of the electron transport system were also differentially expressed in a fur deletion mutant . The gene omcA (SO1779), which encodes a decaheme cytochrome c, exhibited significant decreases in both mRNA and protein abundance in the fur mutant and possessed a strong candidate Fur-binding site in its upstream region, thus suggesting that omcA may be a direct target of Fur activation. J Bacteriol, 2004 Dec, 186(24), 8221 - 8 HMW1 is required for stability and localization of HMW2 to the attachment organelle of Mycoplasma pneumoniae; Willby MJ et al.; The cytoskeletal proteins HMW1 and HMW2 are components of the terminal organelle of the cell wall-less bacterium Mycoplasma pneumoniae . HMW1 is required for a tapered, filamentous morphology but exhibits accelerated turnover in the absence of HMW2 . Here, we report that a reciprocal dependency exists between HMW1 and HMW2, with HMW2 subject to accelerated turnover with the loss of HMW1 . Furthermore, the instability of HMW2 correlated with its failure to localize to the attachment organelle . The C-terminal domain of HMW1 is essential for both function and its accelerated turnover in the absence of HMW2 . We constructed HMW1 deletion derivatives lacking portions of this domain and examined each for stability and function . The C-terminal 41 residues were particularly important for proper localization and function in cell morphology and P1 localization, but the entire C-terminal domain was required to stabilize HMW2 . The significance of these findings in the context of attachment organelle assembly is considered. Ying Yong Sheng Tai Xue Bao, 2004 Aug, 15(8), 1459 - 62 {Effect of organophosphorous insecticides on Chinese chive insect pests and their degradation by pesticide-degrading bacterium}; Jiang J et al.; 3.00 kg(a . i) x hm(-2) phoxin and 2.63 kg(a . i) x hm(-2) methyl parathion were respectively applied to control the Taeniothrips alliorum on Chinese chive . Compared to no pesticide treatment, the decline rate of the insect density was 98.28% and 98.39% at the 3rd day after spraying pesticides, and 89.94% and 94.04% at the 20th day after spraying pesticides, respectively . At the 3rd day after spraying 15.00, 18.00 and 21.00 kg(a . i) x hm(-2) phoxin, the insect density of Bradysia odoriphaga decreased 80.77%, 93.10% and 96.98%, and at the 35th day after spraying, it decreased 92.44%, 95.05% and 96.81%, respectively . The application of pesticide-degrading bacterium had not any effect on controlling insect pests, but could markedly degrade pesticide . At the 3rd day after spraying 45.00 L x hm(-2) pesticide-degrading bacterium to control Taeniothrips alliorum, the degradion rate of phoxin and methyl parathion was 99.52% and 98.83%, and at the 3rd after spraying 75.00 L x hm(-2) pesticide-degrading bacterium to control Bradysia odoriphaga, the degradation rate of three concentrations of phoxin was 100%, 100% and 99.69%, respectively. J Nat Prod, 2004 Nov, 67(11), 1897 - 9 Petrobactin sulfonate, a new siderophore produced by the marine bacterium Marinobacter hydrocarbonoclasticus; Hickford SJ et al.; Culture of the oil-degrading marine bacterium Marinobacter hydrocarbonoclasticus gave the known siderophore petrobactin (1) and the new metabolite petrobactin sulfonate (2), the first marine siderophore containing a sulfonated 3,4-dihydroxy aromatic ring . The structure of petrobactin sulfonate was elucidated from spectral data, resulting in a revision of the NMR assignments of petrobactin. Nat Prod Rep, 2004 Dec, 21(6), 773 - 84 Epub 2004 Dec. Fluorometabolite biosynthesis and the fluorinase from Streptomyces cattleya; Deng H et al.; This review outlines the recent developments in uncovering the enzymes and intermediates involved in fluorometabolite biosyntheses in the bacterium Streptomyces cattleya . A particular emphasis is placed on the purification and characterisation of the fluorinase, the C-F bond forming enzyme which initiates the biosynthesis . Nature has hardly developed a biochemistry around fluorine, yet fluorinated organics are important commercial entities, therefore a biotransformation from inorganic to organic fluorine is novel and of contemporary interest. Analyst, 2004 Dec, 129(12), 1234 - 7 Epub 2004 Dec. Monitoring viral DNA release with capillary electrophoresis; Krylova SM et al.; Viral DNA injection into host cells is one of the primary mechanisms of viral propagation . Drug development that targets viral propagation requires fast and sensitive methods for monitoring the release of viral DNA in vitro . Here we demonstrate the use of capillary electrophoresis (CE) for monitoring DNA release from virus particles . As a model for this study, we used T5 bacteriophages that infect the bacterium Escherichia coli K-12 by binding to the outer membrane FhuA receptor and then injecting DNA . DNA release from the T5 phages in vitro was induced by either elevated temperature or by interaction with the purified FhuA receptor . After DNA release, the viral samples were stained with the high affinity fluorescent dye YOYO-1, injected into the capillary and subjected to electrophoresis . YOYO-1-stained DNA generated a well-defined peak, allowing reliable detection of viral DNA from as few as 10(5) viral particles . The staining to track T5 phage DNA release exemplifies the great versatility that CE offers in studying viral systems . This CE-based method can be used to study molecular mechanisms of viral infections and to evaluate anti-viral drug candidates. Vet Microbiol, 2004 Dec 9, 104(3-4), 219 - 25 The high prevalence of Helicobacter sp . in porcine pyloric mucosa and its histopathological and molecular characteristics; Park JH et al.; This study examined the prevalence of Helicobacter infection in the pyloric mucosa of pigs and its histopathological and molecular characteristics . Forty porcine pyloric samples were examined for Helicobacter infection by silver staining and PCR assay . The PCR product (376 bp) was digested with NdeII to differentiate between Helicobacter heilmannii and Helicobacter pylori . Another PCR assay run to produce an 1157 bp fragment was performed using a primer set designed from the 16S rRNA gene of Candidatus H . suis, and its product was cloned and sequenced . Infection rates were 62.5% (25/40) and 95.0% (38/40) as determined by silver staining and the PCR assay, respectively . On histopathological examination, lymphoid follicle aggregation in the pyloric mucosa and granulocytic migration into the lumen of pyloric glands were observed in 24 (60.0%) and 33 (82.5%) gastric samples, respectively . All PCR products, except that of H . pylori, were cut into two fragments of 147 and 229 bp by enzymatic digestion with NdeII . Sequencing of the 16S rRNA gene showed that the bacterium had 99.57% (1152 bp/1157 bp) homology to the 16S rRNA gene of Candidatus H . suis. Heredity . 2004 Nov 24; {Epub ahead of print} Multiple infections and diversity of cytoplasmic incompatibility in a haplodiploid species; Mouton L et al.; Cytoplasmic incompatibility (CI) is a sperm-egg incompatibility commonly induced by the intracellular endosymbiont bacterium Wolbachia that, in diploid species, results in embryo mortality . In haplodiploid species, two types of CI exist depending on whether the incompatible fertilized eggs develop into males (male development (MD)) or abort (female mortality (FM)) . CI allows multiple infections to be maintained in host populations, and thus allows interactions to occur between co-infecting strains . In Leptopilina heterotoma, three Wolbachia strains coexist naturally (wLhet1, wLhet2, wLhet3) . When these three strains are all present, they induce a CI of FM type, whereas wLhet1 alone expresses a CI phenotype intermediate between MD and FM . Here, we compare CI effects in crosses involving insect lines sharing the same nuclear background, but harboring different mixtures of strains . Mating experiments showed that: (i) wLhet2 and wLhet3 also induce an intermediate CI when acting alone, and show a bidirectional incompatibility; (ii) there is no interaction between the co-infecting strains in CI expression; (iii) the diversity of Wolbachia present within a male host influences the expression of CI: an increase in the number of strains is correlated with a decrease in the proportion of the MD type, which is also correlated with an increase in bacterial density . All these data suggest that the CI of FM type results from a stronger effect than the MD type, which conflicts with the conventional hypotheses used to explain CI diversity in haplodiploids, and could provide some new information about CI mechanisms in insects.Heredity advance online publication, 24 November 2004; doi:10.1038/sj.hdy.6800596. Antimicrob Agents Chemother, 2004 Dec, 48(12), 4703 - 12 Chalcomycin biosynthesis gene cluster from Streptomyces bikiniensis: novel features of an unusual ketolide produced through expression of the chm polyketide synthase in Streptomyces fradiae; Ward SL et al.; Chalcomycin, a 16-membered macrolide antibiotic made by the bacterium Streptomyces bikiniensis, contains a 2,3-trans double bond and the neutral sugar D-chalcose in place of the amino sugar mycaminose found in most other 16-membered macrolides . Degenerate polyketide synthase (PKS)-specific primers were used to amplify DNA fragments from S . bikiniensis with very high identity to a unique ketosynthase domain of the tylosin PKS . The resulting amplimers were used to identify two overlapping cosmids encompassing the chm PKS . Sequencing revealed a contiguous segment of >60 kb carrying 25 putative genes for biosynthesis of the polyketide backbone, the two deoxysugars, and enzymes involved in modification of precursors of chalcomycin or resistance to it . The chm PKS lacks the ketoreductase and dehydratase domains in the seventh module expected to produce the 2,3-double bond in chalcomycin . Expression of PKS in the heterologous host Streptomyces fradiae, from which the tyl genes encoding the PKS had been removed, resulted in production of at least one novel compound, characterized as a 3-keto 16-membered macrolactone in equilibrium with its 3-trans enol tautomer and containing the sugar mycaminose at the C-5 position, in agreement with the structure predicted on the basis of the domain organization of the chm PKS . The production of a 3-keto macrolide from the chm PKS indicates that a discrete set of enzymes is responsible for the introduction of the 2,3-trans double bond in chalcomycin . From comparisons of the open reading frames to sequences in databases, a pathway for the synthesis of nucleoside diphosphate-D-chalcose was proposed. Mycorrhiza . 2004 Nov 19; {Epub ahead of print} Characterisation of new symbiotic Medicago truncatula (Gaertn.) mutants, and phenotypic or genotypic complementary information on previously described mutants; Morandi D et al.; From a pool of Medicago truncatula mutants-obtained by gamma-irradiation or ethyl methanesulfonate mutagenesis-impaired in symbiosis with the N-fixing bacterium Sinorhizobium meliloti, new mutants are described and genetically analysed, and for already reported mutants, complementary data are given on their phenotypic and genetic analysis . Phenotypic data relate to nodulation and mycorrhizal phenotypes . Among the five new mutants, three were classified as {Nod(+) Fix(-) Myc(+)} and the mutations were ascribed to two loci, Mtsym20 (TRV43, TRV54) and Mtsym21 (TRV49) . For the two other new mutants, one was classified as {Nod(-/+) Myc(+)} with a mutation ascribed to gene Mtsym15 (TRV48), and the other as {Nod(-) Myc(-/+)} with a mutation ascribed to gene Mtsym16 (TRV58) . Genetic analysis of three previously described mutants has shown that {Nod(-/+) Myc(+)} TR74 mutant can be ascribed to gene Mtsym14, and that {Nod(-/+) Myc(-/+)} TR89 and TRV9 mutants are ascribed to gene Mtsym2 ( dmi2) . Using a detailed analysis of mycorrhizal phenotype, we have observed a delayed typical arbuscular mycorrhizal formation on some mutants that present thick lens-shaped appressoria . This phenotype was called {Myc(-/+)} and mutants TR25, TR26, TR89, TRV9, P1 and Y6 were reclassified as {Myc(-/+)} . Mutant P1 was reclassified as {Nod(-/+)} because of a late nodulation observed on roots of this mutant. Infect Immun, 2004 Dec, 72(12), 6852 - 9 Rapid sequential changeover of expressed p44 genes during the acute phase of Anaplasma phagocytophilum infection in horses; Wang X et al.; Anaplasma phagocytophilum immunodominant polymorphic major surface protein P44s have been hypothesized to go through antigenic variation, but the within-host dynamics of p44 expression has not been demonstrated . In the present study we investigated the composition and changes of p44 transcripts in the blood during the acute phase of well-defined laboratory A . phagocytophilum infections in naive equine hosts . Three traveling waves of sequential population changeovers of the p44 transcript species were observed within a single peak of rickettsemia of less than 1 month . During the logarithmic increase, the rapid switch-off of the initial dominant transcript p44-18 occurred regardless of whether the bacterium was transmitted by ticks or by intravenous inoculation . Each of the subsequently dominant p44 transcript species was phylogenetically dissimilar from p44-18 . Development of antibody to the hypervariable region of P44-18 during the rickettsemia suggests the suppression of dominance of immuno-cross-reactive p44 populations . When A . phagocytophilum was preincubated with plasma from the infected horse and then coincubated with HL-60 cells, the dominance of the p44-18 transcript was rapidly suppressed in vitro and most of the newly emerged p44 transcript species were previously undetected in this horse . This work provides experimental evidence of within-host p44 antigenic variation . Results suggest that the rapid and synchronized switch of expression is an intrinsic property of p44s reinitiated after transmission to naive mammalian hosts and shaped upon exposure to immune plasma. FEMS Microbiol Lett, 2004 Dec 1, 241(1), 33 - 40 Nitrogen regulation in Sinorhizobium meliloti probed with whole genome arrays; Davalos M et al.; Using whole genome arrays, we systematically investigated nitrogen regulation in the plant symbiotic bacterium Sinorhizobium meliloti . The use of glutamate instead of ammonium as a nitrogen source induced nitrogen catabolic genes independently of the carbon source, including two glutamine synthetase genes, various aminoacid transporters and the glnKamtB operon . These responses depended on both the ntrC and glnB nitrogen regulators . Glutamate repressible genes included glutamate synthase and a H+-translocating pyrophosphate synthase . The smc01041-ntrBC operon was negatively autoregulated in a glnB-dependent fashion, indicating an involvement of phosphorylated NtrC . In addition to the nitrogen response, glutamate remodelled expression of carbon metabolism by inhibiting expression of the Entner-Doudoroff and pentose phosphate pathways, and by stimulating gluconeogenetic genes independently of ntrC. FEBS Lett, 2004 Nov 19, 577(3), 403 - 8 The N-terminal rhodanese domain from Azotobacter vinelandii has a stable and folded structure independently of the C-terminal domain; Melino S et al.; Sulfurtransferase are enzymes involved in the formation, conversion and transport of compounds containing sulfane-sulfur atoms . Although the three-dimensional structure of the rhodanese from the nitrogen-fixing bacterium Azotobacter vinelandii is known, the role of its two domains in the protein conformational stability is still obscure . We have evaluated the susceptibility to proteolytic degradation of the two domains of the enzyme . The two domains show different resistance to the endoproteinases and, in particular, the N-terminal domain shows to be more stable to digestion during time than the C-terminal one . Cloning and overexpression of the N-terminal domain of the protein was performed to better understand its functional and structural role . The recombinant N-terminal domain of rhodanese A . vinelandii is soluble in water solution and the spectroscopic studies by circular dichroism and heteronuclear NMR spectroscopy indicate a stable fold of the protein with the expected alpha/beta topology . The results indicate that this N-terminal domain has already got all the elements necessary for an C-terminal domain independent folding . Its solution structure by NMR, actually under course, will be a valid contribution to understand the role of this domain in the folding process of the sulfurtransferase. FEBS Lett, 2004 Nov 19, 577(3), 371 - 5 The pur6 gene of the puromycin biosynthetic gene cluster from Streptomyces alboniger encodes a tyrosinyl-aminonucleoside synthetase; Angel Rubio M et al.; The pur6 gene of the puromycin biosynthetic gene (pur) cluster from Streptomyces alboniger is shown to be essential for puromycin biosynthesis . Cell lysates from this mycelial bacterium were active in linking L-tyrosine to both 3'-amino-3'-deoxyadenosine and N6,N6-dimethyl-3'-amino-3'-deoxyadenosine with a peptide-like bond . Identical reactions were performed by cell lysates from Streptomyces lividans or Escherichia coli transformants that expressed pur6 from a variety of plasmid constructs . Physicochemical and biochemical analyses suggested that their products were tridemethyl puromycin and O-demethylpuromycin, respectively . Therefore, it appears that Pur6 is the tyrosinyl-aminonucleoside synthetase of the puromycin biosynthetic pathway. BMC Biochem . 2004 Nov 22;5(1):17. Rhodobacter capsulatus porphobilinogen synthase, a high activity metal ion independent hexamer; Bollivar DW et al.; BACKGROUND: The enzyme porphobilinogen synthase (PBGS), which is central to the biosynthesis of heme, chlorophyll and cobalamins, has long been known to use a variety of metal ions and has recently been shown able to exist in two very different quaternary forms that are related to metal ion usage . This paper reports new information on the metal ion independence and quaternary structure of PBGS from the photosynthetic bacterium Rhodobacter capsulatus . RESULTS: The gene for R . capsulatus PBGS was amplified from genomic DNA and sequencing revealed errors in the sequence database . R . capsulatus PBGS was heterologously expressed in E . coli and purified to homogeneity . Analysis of an unusual phylogenetic variation in metal ion usage by PBGS enzymes predicts that R . capsulatus PBGS does not utilize metal ions such as Zn2+, or Mg2+, which have been shown to act in other PBGS at either catalytic or allosteric sites . Studies with these ions and chelators confirm the predictions . A broad pH optimum was determined to be independent of monovalent cations, approximately 8.5, and the Km value shows an acidic pKa of approximately 6 . Because the metal ions of other PBGS affect the quaternary structure, gel permeation chromatography and analytical ultracentrifugation experiments were performed to examine the quaternary structure of metal ion independent R . capsulatus PBGS . The enzyme was found to be predominantly hexameric, in contrast with most other PBGS, which are octameric . A protein concentration dependence to the specific activity suggests that the hexameric R . capsulatus PBGS is very active and can dissociate to smaller, less active, species . A homology model of hexameric R . capsulatus PBGS is presented and discussed . CONCLUSION: The evidence presented in this paper supports the unusual position of the R . capsulatus PBGS as not requiring any metal ions for function . Unlike other wild-type PBGS, the R . capsulatus protein is a hexamer with an unusually high specific activity when compared to other octameric PBGS proteins. Cell, 2004 Nov 24, 119(5), 615 - 27 Structural basis of ligand activation in a cyclic nucleotide regulated potassium channel; Clayton GM et al.; Here we describe the initial functional characterization of a cyclic nucleotide regulated ion channel from the bacterium Mesorhizobium loti and present two structures of its cyclic nucleotide binding domain, with and without cAMP . The domains are organized as dimers with the interface formed by the linker regions that connect the nucleotide binding pocket to the pore domain . Together, structural and functional data suggest the domains form two dimers on the cytoplasmic face of the channel . We propose a model for gating in which ligand binding alters the structural relationship within a dimer, directly affecting the position of the adjacent transmembrane helices. Int J Syst Evol Microbiol, 2004 Nov, 54(Pt 6), 2313 - 7 Mycobacterium pyrenivorans sp . nov., a novel polycyclic-aromatic-hydrocarbon-degrading species; Derz K et al.; The taxonomic position of a polycyclic-aromatic-hydrocarbon-degrading bacterium, strain 17A3(T), isolated from contaminated soil was determined using a combination of phenotypic and genotypic properties . The isolate showed phenotypic properties that were diagnostic for species of the genus Mycobacterium . Comparative 16S rRNA gene sequence analysis assigned 17A3(T) to the 16S rRNA gene subgroup that contains Mycobacterium aurum, Mycobacterium austroafricanum, Mycobacterium vaccae and Mycobacterium vanbaalenii, but it could clearly be distinguished from these species using a combination of physiological, chemotaxonomic markers and internal rRNA gene spacer analyses . The data showed that strain 17A3(T) (=DSM 44605(T)=NRRL B-24244(T)) merits recognition as the type strain of a novel species of the genus Mycobacterium . The name Mycobacterium pyrenivorans sp . nov . is proposed for the species because of its ability to use pyrene as a sole source of carbon and energy. Int J Syst Evol Microbiol, 2004 Nov, 54(Pt 6), 2203 - 11 Description of 'Candidatus Helicobacter heilmannii' based on DNA sequence analysis of 16S rRNA and urease genes; O'Rourke JL et al.; While Helicobacter pylori is accepted as the major bacterial agent of gastric disease in humans, some patients and many animals are infected with a larger, tightly helical-shaped bacterium previously referred to as 'Helicobacter heilmannii' or 'Gastrospirillum hominis' . Taxonomic classification of these bacteria has been hampered by the inability to cultivate them in vitro and by the inadequate discriminatory power of 16S rRNA gene sequence analysis . This study describes the detection and phylogenetic analysis of 26 different gastrospirillum isolates from humans and animals, which incorporates sequence data based on the 16S rRNA and urease genes . Fifteen gastrospirilla detected in humans, primates and pigs clustered with 'Candidatus Helicobacter suis', thus expanding the host range for this organism . By comparison, based on 16S rRNA data, the remaining 11 gastrospirilla could not be differentiated from Helicobacter felis, Helicobacter bizzozeronii and Helicobacter salomonis . However, urease gene sequence analysis allowed for the discrimination of this latter group into four discrete clusters, three of which contained the above recognized species . The fourth cluster contained isolates from human and feline hosts, and should provisionally be considered a unique bacterial species, for which the name 'Candidatus Helicobacter heilmannii' is proposed. Int J Syst Evol Microbiol, 2004 Nov, 54(Pt 6), 1943 - 9 Shewanella profunda sp . nov., isolated from deep marine sediment of the Nankai Trough; Toffin L et al.; A novel piezotolerant, mesophilic, facultatively anaerobic, organotrophic, polarly flagellated bacterium (strain LT13a(T)) was isolated from a deep sediment layer in the Nankai Trough (Leg 190, Ocean Drilling Program) off the coast of Japan . This organism used a wide range of organic substrates as sole carbon and energy sources: pyruvate, glutamate, succinate, fumarate, lactate, citrate, peptone and tryptone . Oxygen, nitrate, fumarate, ferric iron and cystine were used as electron acceptors . Maximal growth rates were observed at a hydrostatic pressure of 10 MPa . Hydrostatic pressure for growth was in the range 0.1-50 MPa . Predominant cellular fatty acids were 16 : 1omega7c, 15 : 0 iso, 16 : 0 and 13 : 0 iso . The G+C content of the DNA was 44.9 mol% . On the basis of 16S rRNA gene sequences, strain LT13a(T) was shown to belong to the gamma-Proteobacteria, being closely related to Shewanella putrefaciens (98 %), Shewanella oneidensis (97 %) and Shewanella baltica (96 %) . Levels of DNA homology between strain LT13a(T) and S . putrefaciens, S . oneidensis and S . baltica were <20 %, indicating that strain LT13a(T) represents a novel species . Genetic evidence and phenotypic characteristics showed that isolate LT13a(T) constitutes a novel species of the genus Shewanella . Because of the deep origin of the strain, the name Shewanella profunda sp . nov . is proposed, with LT13a(T) (=DSM 15900(T)=JCM 12080(T)) as the type strain. Chem Commun (Camb), 2004 Nov 21, (22), 2590 - 1 Epub 2004 Sep 30. A mixed ladderane/n-alkyl glycerol diether membrane lipid in an anaerobic ammonium-oxidizing bacterium; Sinninghe Damste JS et al.; A novel glycerol diether containing ladderane and tetradecyl moieties has been identified in an anaerobic ammonium-oxidizing bacterium by GC/MS and high-field NMR spectroscopy. Am Nat, 2004 Nov, 164 Suppl 5, S19 - 32 The evolution of virulence when parasites cause host castration and gigantism; Ebert D; It has been suggested that the harm parasites cause to their hosts is an unavoidable consequence of parasite reproduction with costs not only for the host but also for the parasite . Castrating parasites are thought to minimize their costs by reducing host fecundity, which may minimize the chances of killing both host and parasite prematurely . We conducted a series of experiments to understand the evolution of virulence of a castrating bacterium in the planktonic crustacean Daphnia magna . By manipulating food levels during the infection of D . magna with the bacterium Pasteuria ramosa, we showed that both antagonists are resource-limited and that a negative correlation between host and parasite reproduction exists, indicating resource competition among the antagonists . Pasteuria ramosa also induces enhanced growth of its hosts (gigantism), which we found to be negatively correlated with host fecundity but positively correlated with parasite reproduction . Because infected hosts never recovered from infections, we concluded that gigantism is beneficial only for the parasite . Hosts, however, have evolved counteradaptations . We showed that infected hosts have enhanced reproduction before castration . This shift to earlier reproduction increases overall host fecundity and compromises parasite reproduction . Finally, we showed that this resource conflict is subject to genetic variation among host and parasite genotypes within a population and is therefore likely to be an important force in the coevolution of virulence in this system . A verbal model is presented and suggests that the adaptive value of gigantism is to store host resources, which are liberated after parasitic castration for later use by the growing parasite . This hypothesis assumes that infections are long lasting, that is, that they have a high life expectancy. Bioinformatics . 2004 Nov 11; {Epub ahead of print} A parallel graph decomposition algorithm for DNA sequencing with nanopores; Bokhari SH et al.; MOTIVATION: With the potential availability of nanopore devices that can sense the bases of translocating single-stranded DNA (ssDNA), it is likely that 'reads' of length approximately 10(5) will be available in large numbers and at high speed . We address the problem of complete DNA sequencing using such reads . We assume that approximately 10(2) copies of a DNA sequence are split into single strands that break into randomly sized pieces as they translocate the nanopore in arbitrary orientations . The nanopore senses and reports each individual base that passes through, but all information about orientation and complementarity of the ssDNA subsequences is lost . Random errors (both biological and transduction) in the reads create further complications . RESULTS: We have developed an algorithm that addresses these issues . It can be considered an extreme variation of the well-known Eulerian path approach . It searches over a space of de Bruijn graphs until it finds one in which (a) the impact of errors is eliminated and (b) both possible orientations of the two ssDNA sequences can be identified separately and unambiguously . Our algorithm is able to correctly reconstruct real DNA sequences of the order of 10(6) bases (e.g . the bacterium Mycoplasma pneumoniae) from simulated erroneous reads on a modest workstation in about one hour . We describe, and give measured timings of, a parallel implementation of this algorithm on the Cray Multithreaded Architecture (MTA-2) supercomputer, whose architecture is ideally suited to this 'unstructured' problem . Our parallel implementation is crucial to the problem of rapidly sequencing long DNA sequences and also to the situation where multiple nanopores are used to obtain a high-bandwidth stream of reads. Appl Microbiol Biotechnol . 2004 Nov 6; {Epub ahead of print} Purification, cloning, and properties of alpha-galactosidase from Saccharopolyspora erythraea and its use as a reporter system; Post DA et al.; An alpha-galactosidase from the erythromycin-producing bacterium Saccharopolyspora erythraea was purified to near homogeneity . The enzyme has an apparent molecular mass of 45 kDa as determined by SDS-PAGE . The pH optimum, K(m) for p-nitrophenyl-alpha- d-glucopyranoside (pNPalphaG), K(m) for melibiose and the V(max) are similar to those of other studied alpha-galactosidase enzymes . The N-terminal amino-acid sequence of this protein was determined . PCR amplification was used to generate a 640-bp product using oligonucleotide primers based on the N-terminal amino-acid sequence and a downstream region that is conserved in other related alpha-galactosidase enzymes . This fragment was used as a probe to clone the alpha-galactosidase gene, designated melA, from a S . erythraea lambda phage chromosomal library . S . erythraea appears to possess an unique alpha-galactosidase enzyme, encoded by melA, that can utilize galactopyranosides as carbon sources . Furthermore, the ability to use the product of melA as a reporter enzyme in S . erythraea has been demonstrated . The alpha-galactosidase uses the substrates 5-bromo-4-chloro-3-indoyl-alpha- d-galactosidase (X-alpha-gal) on agar media and pNPalphaG in liquid media. J Med Entomol, 2000 May, 37(3), 349 - 56 Ehrlichia chaffeensis (Rickettsiales: Ehrlichieae) infection in Amblyomma americanum (Acari: Ixodidae) at Aberdeen Proving Ground, Maryland; Stromdahl EY et al.; Human monocytic ehrlichiosis (HME) is a sometimes fatal, emerging tick-borne disease caused by the bacterium Ehrlichia chaffeensis . It is frequently misdiagnosed because its symptoms mimic those of the flu . Current evidence indicates that Amblyomma americanum (L.), the lone star tick, is the major vector of HME . To determine if E . chaffeensis is present in ticks at Aberdeen Proving Ground, MD, questing A . americanum ticks were collected from 33 sites . Nucleic acid was extracted from 34 adult and 81 nymphal pools . Sequences diagnostic for E . chaffeensis from three different loci (16S rRNA, 120-kDa protein, and a variable-length polymerase chain reaction {PCR} target, or VLPT) were targeted for amplification by the PCR . Fifty-two percent of the collection sites yielded pools infected with E . chaffeensis, confirming the presence and widespread distribution of E . chaffeensis at Aberdeen Proving Ground . Analysis with the both the 120-kDa protein primers and the VLPT primers showed that genetic variance exists . A novel combination of variance for the two loci was detected in two tick pools . The pathogenic implications of genetic variation in E . chaffeensis are as yet unknown. Biochemistry, 2004 Nov 16, 43(45), 14379 - 84 Trapped tyrosyl radical populations in modified reaction centers from Rhodobacter sphaeroides; Narvaez AJ et al.; The photosynthetic reaction center from the purple bacterium Rhodobacter sphaeroides has been modified such that the bacteriochlorophyll dimer, when it becomes oxidized after light excitation, is capable of oxidizing tyrosine residues . One factor in this ability is a high oxidation-reduction midpoint potential for the dimer, although the location and protein environment of the tyrosine residue appear to be critical as well . These factors were tested in a series of mutants, each of which contains changes, at residues L131, M160, M197, and M210, that give rise to a bacteriochlorophyll dimer with a midpoint potential of at least 800 mV . The protein environment was altered near tyrosine residues that are either present in the wild type or introduced by mutagenesis, focusing on residues that could act as acceptors for the phenolic proton of the tyrosine upon oxidation . These mutations include Ser M190 to His, which is near Tyr L162, the combination of His M193 to Tyr and Arg M164 to His, which adds a Tyr-His pair, and the combinations of Arg L135 to Tyr with Tyr L164 to His, Arg L135 to Tyr with Tyr L144 to Glu, and Arg L135 to Tyr with Tyr L164 to Phe . Radicals were produced in the mutants by using light to initiate electron transfer . The radicals were trapped by freezing the samples, and the relative populations of the oxidized dimer and tyrosyl radicals were determined by analysis of low-temperature electron paramagnetic resonance spectra . The mutants all showed evidence of tyrosyl radical formation at high pH, and the extent of radical formation at Tyr L135 with pH differed depending on the identity of L144 and L164 . The results show that tyrosine residues within approximately 10 A of the dimer can become oxidized when provided with a suitable protein environment. Folia Microbiol (Praha), 2004, 49(4), 479 - 83 Effect of short-chain acids on the carboxymethylcellulase activity of the ruminal bacterium Ruminococcus albus; Paggi RA et al.; The addition of 100-300 mmol/L of acetic, propionic, butyric or lactic acids (short-chain acids), or of acetic, propionic, and butyric acids (volatile fatty acids, VFA) mixtures increased the degradation of carboxymethyl cellulose (CMC) by R . albus (7.5 to 46 and 6 to 39 %, respectively) . Differences among individual acids were observed at 300 mmol/L whereas VFA mixtures differed at 100 mmol/L . When assayed at the same concentration, CMCase activity was increased less by NaCl than by the short-chain acids, whereas ethylene glycol decreased the activity . Since osmolarity and/or ionic strength changes in the medium cannot completely account for the observed increases of carboxymethylcellulase (CMCase) activity, it is suggested that the anions of short-chain acids produce changes in the reaction media polarity that contribute to the effects observed . Alterations in the media could also bring about conformational changes in CMCase leading to increased rates of reaction and subsequent increases in CMC degradation . Finally, explanations for the observed phenomena based on the direct effect of the compounds tested on the cellulosome complex, its domains, and/or its component enzymes are proposed. Cell Mol Biol (Noisy-le-grand), 2004 Jun, 50(4), 311 - 6 High pressure nmr study of dihydrofolate reductase from a deep-sea bacterium Moritella profunda; Hata K et al.; We have investigated the effect of pressure and temperature on the structural and thermodynamic stability of a protein dihydrofolate reductase from a deep-sea bacterium Moritella profunda in its folate-bound form in the pressure range between 3 and 375 MPa and the temperature range between -5 and 30 degrees C . The on-line cell variable pressure 1H NMR spectroscopy has been used to analyze the chemical shift and signal intensity in one-dimensional 1H NMR spectra . Thermodynamic analysis based on signal intensities from protons in the core part indicates that the thermodynamic stability of Moritella profunda DHFR is relatively low over the temperature range between -5 and 30 degrees C (deltaG0=15.8 +/- 4.1 kJ/mol at 15 degrees C), but is well adapted to the living environment of the bacterium (2 degrees C and 28 MPa), with the maximum stability around 5 degrees C (at 0.1 MPa) and a relatively small volume change upon unfolding (deltaV= 66 +/- 19 ml/mol) . Despite the relatively low overall stability, the conformation in the core part of the folded protein remains intact up to approximately 200 MPa, showing marked stability of the core of this protein. Nat Biotechnol, 2004 Nov, 22(11), 1399 - 408 Recombinant protein folding and misfolding in Escherichia coli; Baneyx F et al.; The past 20 years have seen enormous progress in the understanding of the mechanisms used by the enteric bacterium Escherichia coli to promote protein folding, support protein translocation and handle protein misfolding . Insights from these studies have been exploited to tackle the problems of inclusion body formation, proteolytic degradation and disulfide bond generation that have long impeded the production of complex heterologous proteins in a properly folded and biologically active form . The application of this information to industrial processes, together with emerging strategies for creating designer folding modulators and performing glycosylation all but guarantee that E . coli will remain an important host for the production of both commodity and high value added proteins. Appl Environ Microbiol, 2004 Nov, 70(11), 6897 - 900 Degradation of a nonylphenol single isomer by Sphingomonas sp . strain TTNP3 leads to a hydroxylation-induced migration product; Corvini PF et al.; Sphingomonas sp . strain TTNP3 degrades 4(3',5'-dimethyl-3'-heptyl)-phenol and unidentified metabolites that were described previously . The chromatographic analyses of the synthesized reference compound and the metabolites led to their identification as 2(3',5'-dimethyl-3'-heptyl)-1,4-benzenediol . This finding indicates that the nonylphenol metabolism of this bacterium involves unconventional degradation pathways where an NIH shift mechanism occurs. Appl Environ Microbiol, 2004 Nov, 70(11), 6595 - 602 Effects of the metalloid oxyanion tellurite (TeO32-) on growth characteristics of the phototrophic bacterium Rhodobacter capsulatus; Borghese R et al.; This work examines the effects of potassium tellurite (K2TeO3) on the cell viability of the facultative phototroph Rhodobacter capsulatus . There was a growth mode-dependent response in which cultures anaerobically grown in the light tolerate the presence of up to 250 to 300 microg of tellurite (TeO3(2-)) per ml, while dark-grown aerobic cells were inhibited at tellurite levels as low as 2 microg/ml . The tellurite sensitivity of aerobic cultures was evident only for growth on minimal salt medium, whereas it was not seen during growth on complex medium . Notably, through the use of flow cytometry, we show that the cell membrane integrity was strongly affected by tellurite during the early growth phase (< or =50% viable cells); however, at the end of the growth period and in parallel with massive tellurite intracellular accumulation as elemental Te0 crystallites, recovery of cytoplasmic membrane integrity was apparent (> or =90% viable cells), which was supported by the development of a significant membrane potential (Deltapsi = 120 mV) . These data are taken as evidence that in anaerobic aquatic habitats, the facultative phototroph R . capsulatus might act as a natural scavenger of the highly soluble and toxic oxyanion tellurite. FEBS Lett, 2004 Nov 5, 577(1-2), 255 - 8 Biochemical characterization of the major adenylyl cyclase, Cya1, in the cyanobacterium Synechocystis sp . PCC 6803; Masuda S et al.; We report herein the biochemical properties of an adenylyl cyclase, Cya1, from the cyanobacterium Synechocystis sp . PCC 6803 . Heterologously expressed Cya1 catalyzed cyclic AMP formation with a Km for ATP of approximately 2.2 microM at pH 7.5 . Although cellular Cya1 activity is increased by blue light illumination {Terauchi and Ohmori, Mol . Microbiol . 52 (2004) 303}, purified Cya1 did not contain any chromophores, and the activity was light-insensitive . This suggests that an unknown blue light-responsive factor interacts with the N-terminal regulatory domain of Cya1 to control its adenylyl cyclase activity . Finally, our results show that the sensor of blue light using FAD (BLUF) protein, Slr1694, does not appear to be involved in the regulation of Cya1-mediated cAMP signal transduction in this