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The LysR-Type Transcriptional Regulator VirR Is Required for Expression of the Virulence Gene vapA of Rhodococcus equi ATCC 33701. Dean A. Russell, 2004.The virulence of the intracellular pathogen Rhodococcus equi in foals is dependent on the presence of an 81-kb virulence plasmid encoding the virulence protein VapA . Expression of this protein is induced by exposure to oxidative stress, high temperatures, and low pHs, which reflect the conditions encountered by R . equi when it enters the host environment . The aim of this study was to determine whether the LysR-type transcriptional regulator VirR, which is encoded by the virulence plasmid, is required for the expression of vapA . It was shown that the virR gene is cotranscribed with four downstream genes, one of which encodes a two-component response regulator . The expression of VapA, as monitored by Western blotting, was completely dependent on the presence of virR . Maximal expression was observed when vapA was present together with the complete virR operon, suggesting that at least one of the virR operon genes, in addition to virR, is required for the expression of vapA to wild-type levels . The transcriptional start site of vapA was determined to be a cytidine located 226 bp upstream from the vapA initiation codon . His-tagged VirR protein was expressed in Escherichia coli and purified by nickel affinity chromatography . DNA binding studies showed that purified VirR binds to a DNA fragment containing the vapA promoter . We therefore conclude that VirR is required for the activation of vapA transcription . Colonization with Extraintestinal Pathogenic Escherichia coli among Nursing Home Residents and Its Relationship to Fluoroquinolone Resistance. Joel N. Maslow, 2004.In a cross-sectional fecal prevalence survey involving 49 residents of a Veterans Affairs nursing home, 59% of subjects were colonized with extraintestinal pathogenic Escherichia coli (ExPEC), 22% were colonized with adhesin-positive E . coli, and 51% were colonized with fluoroquinolone-resistant E . coli . Among 80 unique isolates, adhesins correlated negatively and aerobactin correlated positively with fluoroquinolone resistance . Biodegradation of Chloromethane by Pseudomonas aeruginosa Strain NB1 under Nitrate-Reducing and Aerobic Conditions. David L. Freedman, 2004.Pseudomonas aeruginosa strain NB1 uses chloromethane (CM) as its sole source of carbon and energy under nitrate-reducing and aerobic conditions . The observed yield of NB1 was 0.20 (±0.06) (mean ± standard deviation) and 0.28 (±0.01) mg of total suspended solids (TSS) mg of CM–1 under anoxic and aerobic conditions, respectively . The stoichiometry of nitrate consumption was 0.75 (±0.10) electron equivalents (eeq) of NO3– per eeq of CM, which is consistent with the yield when it is expressed on an eeq basis . Nitrate was stoichiometrically converted to dinitrogen (0.51 ± 0.05 mol of N2 per mol of NO3–) . The stoichiometry of oxygen use with CM (0.85 ± 0.21 eeq of O2 per eeq of CM) was also consistent with the aerobic yield . Stoichiometric release of chloride and minimal accumulation of soluble metabolic products (measured as chemical oxygen demand) following CM consumption, under anoxic and aerobic conditions, indicated complete biodegradation of CM . Acetylene did not inhibit CM use under aerobic conditions, implying that a monooxygenase was not involved in initiating aerobic CM metabolism . Under anoxic conditions, the maximum specific CM utilization rate (k) for NB1 was 5.01 (±0.06) µmol of CM mg of TSS–1 day–1, the maximum specific growth rate (µmax) was 0.0506 day–1, and the Monod half-saturation coefficient (Ks) was 0.067 (±0.004) µM . Under aerobic conditions, the values for k, µmax, and Ks were 10.7 (±0.11) µmol of CM mg of TSS–1 day–1, 0.145 day–1, and 0.93 (±0.042) µM, respectively, indicating that NB1 used CM faster under aerobic conditions . Strain NB1 also grew on methanol, ethanol, and acetate under denitrifying and aerobic conditions, but not on methane, formate, or dichloromethane . Inactivation of Feline Calicivirus and Adenovirus Type 40 by UV Radiation. Jeanette A. Thurston-Enriquez, 2003.Little information regarding the effectiveness of UV radiation on the inactivation of caliciviruses and enteric adenoviruses is available . Analysis of human calicivirus resistance to disinfectants is hampered by the lack of animal or cell culture methods that can determine the viruses' infectivity . The inactivation kinetics of enteric adenovirus type 40 (AD40), coliphage MS-2, and feline calicivirus (FCV), closely related to the human caliciviruses based on nucleic acid organization and capsid architecture, were determined after exposure to low-pressure UV radiation in buffered demand-free (BDF) water at room temperature . In addition, UV disinfection experiments were also carried out in treated groundwater with FCV and AD40 . AD40 was more resistant than either FCV or coliphage MS-2 in both BDF water and groundwater . The doses of UV required to achieve 99% inactivation of AD40, coliphage MS-2, and FCV in BDF water were 109, 55, and 16 mJ/cm2, respectively . The doses of UV required to achieve 99% inactivation of AD40, coliphage MS-2, and FCV in groundwater were slightly lower than those in BDF water . FCV was inactivated by 99% by 13 mJ/cm2 in treated groundwater . A dose of 103 mJ/cm2 was required for 99% inactivation of AD40 in treated groundwater . The results of this study indicate that if FCV is an adequate surrogate for human caliciviruses, then their inactivation by UV radiation is similar to those of other single-stranded RNA enteric viruses, such as poliovirus . In addition, AD40 appears to be more resistant to UV disinfection than previously reported . Use of Antibiotic Resistance Analysis for Representativeness Testing of Multiwatershed Libraries. Bruce A. Wiggins, 2003.The use of antibiotic resistance analysis (ARA) for microbial source tracking requires the generation of a library of isolates collected from known sources in the watershed . The size and composition of the library are critical in determining if it represents the diversity of patterns found in the watershed . This study was performed to determine the size that an ARA library needs to be to be representative of the watersheds for which it will be used and to determine if libraries from different watersheds can be merged to create multiwatershed libraries . Fecal samples from known human, domesticated, and wild animal sources were collected from six Virginia watersheds . From these samples, enterococci were isolated and tested by ARA . Based on cross-validation discriminant analysis, only the largest of the libraries (2,931 isolates) were found to be able to classify nonlibrary isolates as well as library isolates (i.e., were representative) . Small libraries tended to have higher average rates of correct classification, but were much less able to correctly classify nonlibrary isolates . A merged multiwatershed library (6,587 isolates) was created and was found to be large enough to be representative of the isolates from the contributing watersheds . When isolates that were collected from the contributing watersheds approximately 1 year later were analyzed with the multiwatershed library, they were classified as well as the isolates in the library, suggesting that the resistance patterns are temporally stable for at least 1 year . The ability to obtain a representative, temporally stable library demonstrates that ARA can be used to identify sources of fecal pollution in natural waters .
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