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Effect of Erythromycin on Chronic Respiratory Infection Caused by Pseudomonas aeruginosa with Biofilm Formation in an Experimental Murine Model. Towako Nagata, 2004.Diffuse panbronchiolitis (DPB) is a chronic lower respiratory tract infection commonly associated with persistent late-stage Pseudomonas aeruginosa infection . However, low-dose long-term therapy with certain macrolides is effective in most patients with DPB . The present study was designed to examine the effects of long-term erythromycin (ERY) therapy by using our established murine model of chronic respiratory P . aeruginosa infection . ERY or saline was administered from day 80 after intubation with a P . aeruginosa-precoated tube for the subsequent 10, 20, 40, and 80 days . Bacteriologic and histologic analyses of the murine lungs and electron microscopy of the intubated tube were performed . In the murine model, treatment with ERY for 80 days significantly reduced the number of viable P . aeruginosa organisms in the lungs (P < 0.05) . The biofilm formed in situ by P . aeruginosa on the inner wall of the inoculation tube placed into the murine bronchus became significantly thinner after 80 days of ERY treatment . We conclude that the clinical efficacy of macrolides in DPB may be due at least in part to the reduction in P . aeruginosa biofilm formation . Isolation, Characterization, and U(VI)-Reducing Potential of a Facultatively Anaerobic, Acid-Resistant Bacterium from Low-pH, Nitrate- and U(VI)-Contaminated Subsurface Sediment and Description of Salmonella subterranea sp . nov.. Evgenya S. Shelobolina, 2004.A facultatively anaerobic, acid-resistant bacterium, designated strain FRCl, was isolated from a low-pH, nitrate- and U(VI)-contaminated subsurface sediment at site FW-024 at the Natural and Accelerated Bioremediation Research Field Research Center in Oak Ridge, Tenn . Strain FRCl was enriched at pH 4.5 in minimal medium with nitrate as the electron acceptor, hydrogen as the electron donor, and acetate as the carbon source . Clones with 16S ribosomal DNA (rDNA) sequences identical to the sequence of strain FRCl were also detected in a U(VI)-reducing enrichment culture derived from the same sediment . Cells of strain FRCl were gram-negative motile regular rods 2.0 to 3.4 µm long and 0.7 to 0.9 µm in diameter . Strain FRCl was positive for indole production, by the methyl red test, and for ornithine decarboxylase; it was negative by the Voges-Proskauer test (for acetylmethylcarbinol production), for urea hydrolysis, for arginine dihydrolase, for lysine decarboxylase, for phenylalanine deaminase, for H2S production, and for gelatin hydrolysis . Strain FRCl was capable of using O2, NO3–, S2O32–, fumarate, and malate as terminal electron acceptors and of reducing U(VI) in the cell suspension . Analysis of the 16S rDNA sequence of the isolate indicated that this strain was 96.4% similar to Salmonella bongori and 96.3% similar to Enterobacter cloacae . Physiological and phylogenetic analyses suggested that strain FRCl belongs to the genus Salmonella and represents a new species, Salmonella subterranea sp . nov . Characterization of the Bacillus subtilis ywsC Gene, Involved in  -Polyglutamic Acid Production.Yuji Urushibata, 2002.The genes required for -polyglutamic acid (PGA) production were cloned from Bacillus subtilis IFO16449, a strain isolated from fermented soybeans . There were four open reading frames in the cloned 4.2-kb DNA fragment, and they were almost identical to those in the ywsC and ywtABC genes of B . subtlis 168 . Northern blot analysis showed that the four genes constitute an operon . Three genes, ywsC, ywtA, and ywtB, were disrupted to determine which gene plays a central role in PGA biosynthesis . No PGA was produced in
 ywsC and
ywtA strains, indicating that both of these genes are essential for PGA production . To clarify the function of the YwsC protein, histidine-tagged YwsC (YwsC-His) was produced in the
ywsC strain and purified from the lysozyme-treated lysate of the transformant by Ni-nitrilotriacetic acid affinity chromatography . Western blot analysis revealed that the YwsC-His protein consists of two subunits, the 44-kDa and 33-kDa proteins, which are encoded by in-phase overlapping in the ywsC gene. 14C-labeled PGA was synthesized by the purified proteins from L-[14C]-glutamate in the presence of ATP and MnCl2, through an acylphosphate intermediate, indicating that the ywsC gene encodes PGA synthetase (EC 6.3.2), a crucial enzyme in PGA biosynthesis .
Features of Rhodobacter sphaeroides CcmFH. Carlos Rios-Velazquez, 2003.In this study, the in vivo function and properties of two cytochrome c maturation proteins, CcmF and CcmH from Rhodobacter sphaeroides, were analyzed . Strains lacking CcmH or both CcmF and CcmH are unable to grow under anaerobic conditions where c-type cytochromes are required, demonstrating their critical role in the assembly of these electron carriers . Consistent with this observation, strains lacking both CcmF and CcmH are deficient in c-type cytochromes when assayed under permissive growth conditions . In contrast, under permissive growth conditions, strains lacking only CcmH contain several soluble and membrane-bound c-type cytochromes, albeit at reduced levels, suggesting that this bacterium has a CcmH-independent route for their maturation . In addition, the function of CcmH that is needed to support anaerobic growth can be replaced by adding cysteine or cystine to growth media . The ability of exogenous thiol compounds to replace CcmH provides the first physiological evidence for a role of this protein in thiol chemistry during c-type cytochrome maturation . The properties of R . sphaeroides cells containing translational fusions between CcmF and CcmH and either Escherichia coli alkaline phosphatase or ß-galactosidase suggest that they are each integral cytoplasmic membrane proteins with their presumed catalytic domains facing the periplasm . Analysis of CcmH shows that it is synthesized as a higher-molecular-weight precursor protein with an N-terminal signal sequence . Mass Transport of Macromolecules within an In Vitro Model of Supragingival Plaque. Thomas Thurnheer, 2003.The aim of this study was to examine the diffusion of macromolecules through an in vitro biofilm model of supragingival plaque . Polyspecies biofilms containing Actinomyces naeslundii, Fusobacterium nucleatum, Streptococcus oralis, Streptococcus sobrinus, Veillonella dispar, and Candida albicans were formed on sintered hydroxyapatite disks and then incubated at room temperature for defined periods with fluorescent markers with molecular weights ranging from 3,000 to 900,000 . Subsequent examination by confocal laser scanning microscopy revealed that the mean square penetration depths for all tested macromolecules except immunoglobulin M increased linearly with time, diffusion coefficients being linearly proportional to the cube roots of the molecular weights of the probes (range, 10,000 to 240,000) . Compared to diffusion in bulk water, diffusion in the biofilms was markedly slower . The rate of diffusion for each probe appeared to be constant and not a function of biofilm depth . Analysis of diffusion phenomena through the biofilms suggested tortuosity as the most probable explanation for retarded diffusion . Selective binding of probes to receptors present in the biofilms could not explain the observed extent of retardation of diffusion . These results are relevant to oral health, as selective attenuated diffusion of fermentable carbohydrates and acids produced within dental plaque is thought to be essential for the development of carious lesions .
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