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The Genome-Sequenced Variant of Campylobacter jejuni NCTC 11168 and the Original Clonal Clinical Isolate Differ Markedly in Colonization, Gene Expression, and Virulence-Associated Phenotypes. Erin C. Gaynor, 2004.The genome sequence of the enteric bacterial pathogen Campylobacter jejuni NCTC 11168 (11168-GS) was published in 2000, providing a valuable resource for the identification of C . jejuni-specific colonization and virulence factors . Surprisingly, the 11168-GS clone was subsequently found to colonize 1-day-old chicks following oral challenge very poorly compared to other strains . In contrast, we have found that the original clinical isolate from which 11168-GS was derived, 11168-O, is an excellent colonizer of chicks . Other marked phenotypic differences were also identified: 11168-O invaded and translocated through tissue culture cells far more efficiently and rapidly than 11168-GS, was significantly more motile, and displayed a different morphology . Serotyping, multiple high-resolution molecular genotyping procedures, and subtractive hybridization did not yield observable genetic differences between the variants, suggesting that they are clonal . However, microarray transcriptional profiling of these strains under microaerobic and severely oxygen-limited conditions revealed dramatic expression differences for several gene families . Many of the differences were in respiration and metabolism genes and operons, suggesting that adaptation to different oxygen tensions may influence colonization potential . This correlates biologically with our observation that anaerobically priming 11168-GS or aerobically passaging 11168-O caused an increase or decrease, respectively, in colonization compared to the parent strain . Expression differences were also observed for several flagellar genes and other less well-characterized genes that may participate in motility . Targeted sequencing of the sigma factors revealed specific DNA differences undetected by the other genomic methods . These observations highlight the capacity of C . jejuni to adapt to multiple environmental niches, the likelihood that this adaptation involves genetic evolution, and provides the first whole-genome molecular exploration of the effect of laboratory culture and storage on colonization and virulence properties of this pathogen . Raw Cow Milk Bacterial Population Shifts Attributable to Refrigeration. Véronique Lafarge, 2004.We monitored the dynamic changes in the bacterial population in milk associated with refrigeration . Direct analyses of DNA by using temporal temperature gel electrophoresis (TTGE) and denaturing gradient gel electrophoresis (DGGE) allowed us to make accurate species assignments for bacteria with low-GC-content (low-GC%) (<55%) and medium- or high-GC% (>55%) genomes, respectively . We examined raw milk samples before and after 24-h conservation at 4°C . Bacterial identification was facilitated by comparison with an extensive bacterial reference database (  150 species) that we established with DNA fragments of pure bacterial strains . Cloning and sequencing of fragments missing from the database were used to achieve complete species identification . Considerable evolution of bacterial populations occurred during conservation at 4°C . TTGE and DGGE are shown to be a powerful tool for identifying the main bacterial species of the raw milk samples and for monitoring changes in bacterial populations during conservation at 4°C . The emergence of psychrotrophic bacteria such as Listeria spp . or Aeromonas hydrophila is demonstrated .
A New Modified ortho Cleavage Pathway of 3-Chlorocatechol Degradation by Rhodococcus opacus 1CP: Genetic and Biochemical Evidence. Olga V. Moiseeva, 2002.The 4-chloro- and 2,4-dichlorophenol-degrading strain Rhodococcus opacus 1CP has previously been shown to acquire, during prolonged adaptation, the ability to mineralize 2-chlorophenol . In addition, homogeneous chlorocatechol 1,2-dioxygenase from 2-chlorophenol-grown biomass has shown relatively high activity towards 3-chlorocatechol . Based on sequences of the N terminus and tryptic peptides of this enzyme, degenerate PCR primers were now designed and used for cloning of the respective gene from genomic DNA of strain 1CP . A 9.5-kb fragment containing nine open reading frames was obtained on pROP1 . Besides other genes, a gene cluster consisting of four chlorocatechol catabolic genes was identified . As judged by sequence similarity and correspondence of predicted N termini with those of purified enzymes, the open reading frames correspond to genes for a second chlorocatechol 1,2-dioxygenase (ClcA2), a second chloromuconate cycloisomerase (ClcB2), a second dienelactone hydrolase (ClcD2), and a muconolactone isomerase-related enzyme (ClcF) . All enzymes of this new cluster are only distantly related to the known chlorocatechol enzymes and appear to represent new evolutionary lines of these activities . UV overlay spectra as well as high-pressure liquid chromatography analyses confirmed that 2-chloro-cis,cis-muconate is transformed by ClcB2 to 5-chloromuconolactone, which during turnover by ClcF gives cis-dienelactone as the sole product . cis-Dienelactone was further hydrolyzed by ClcD2 to maleylacetate . ClcF, despite its sequence similarity to muconolactone isomerases, no longer showed muconolactone-isomerizing activity and thus represents an enzyme dedicated to its new function as a 5-chloromuconolactone dehalogenase . Thus, during 3-chlorocatechol degradation by R . opacus 1CP, dechlorination is catalyzed by a muconolactone isomerase-related enzyme rather than by a specialized chloromuconate cycloisomerase . Irp9, Encoded by the High-Pathogenicity Island of Yersinia enterocolitica, Is Able To Convert Chorismate into Salicylate, the Precursor of the Siderophore Yersiniabactin. Cosima Pelludat, 2003.The Irp9 protein of Yersinia enterocolitica participates in the synthesis of salicylate, the precursor of the siderophore yersiniabactin . In Pseudomonas species, salicylate synthesis is mediated by two enzymes: isochorismate synthase and isochorismate pyruvate-lyase . Both enzymes are required for complementation of a Yersinia irp9 mutant . However, irp9 is not able to complement Escherichia coli entC for the production of enterobactin, which requires isochorismate as a precursor . These results suggest that Irp9 directly converts chorismate into salicylate . Use of Root Organ Cultures To Investigate the Interaction between Glomus intraradices and Pratylenchus coffeae. Annemie Elsen, 2003.The interaction between Glomus intraradices and Pratylenchus coffeae on transformed carrot roots was studied in root organ culture . G . intraradices provided the roots with increased protection against P . coffeae by suppressing nematode reproduction in the roots . The internal and external mycorrhizal development was not influenced by the presence of the nematodes .
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