Comparative performance of different PCR assays for the identification of Campylobacter jejuni and Campylobacter coli.

The sensitivity and specificity of 7 PCR assays described for the identification of Campylobacter jejuni and Campylobacter coli were examined using alkaline cell lysates from a collection of 100 well characterized reference strains of C. jejuni, C. coli, Campylobacter lari and related Campylobacter, Helicobacter and Arcobacter species. Based on a preliminary evaluation, one multiplex test was excluded from further evaluation. The various assays differed considerably in sensitivity and specificity towards their target species. For C. coli, 4 of the 5 assays were 100% specific and sensitive, but for C. jejuni, none of the 5 assays were found to be 100% specific or sensitive. Subsequently, a statistically valid sample (n=263) was taken from a Belgian collection of 1906 human Campylobacter field isolates. This second collection was used to further evaluate two selected multiplex PCR assays. The present study indicates that PCR-based identification using each of the two selected multiplex PCR assays was highly reliable. The R-mPCR assay, followed by species-specific PCR assays or the ceu-oxr mPCR assay if necessary, is our current strategy of choice for the molecular identification of C. jejuni and C. coli. Results presented here should aid researchers in selecting a PCR assay suitable for their specific needs.Comparative StudyEvaluation StudiesJournal ArticleResearch Support, Non-U.S. Gov'tinfo:eu-repo/semantics/publishe

An rpoD gene sequence based evaluation of cultured Pseudomonas diversity on different growth media

The last decade has shown an increased interest in the utilization of bacteria for applications ranging from bioremediation to wastewater purification and promotion of plant growth. In order to extend the current number of micro-organism mediated applications, a continued quest for new agents is required. This study focused on the genus Pseudomonas, which is known to harbour strains with a very diverse set of interesting properties. The aim was to identify growth media that allow retrieval of a high Pseudomonas diversity, as such increasing the chance of isolating isolates with beneficial properties. Three cultivation media: trypticase soy agar (TSA), potato dextrose agar (PDA) and Pseudomonas isolation agar (PIA) were evaluated for their abilities to grow Pseudomonas strains. TSA and PDA were found to generate the largest Pseudomonas diversity. However, communities obtained with both media overlapped. Communities obtained with PIA, on the other hand, were unique. This indicated that the largest diversity is obtained by sampling from either PDA or TSA and from PIA in parallel. To evaluate biodiversity of the isolated Pseudomonas members on the media, an appropriate biomarker had to be identified. Hence, an introductory investigation of the taxonomic resolution of the 16S rRNA, rpoD, gyrB and rpoB genes was performed. The rpoD gene sequences not only had a high phylogenetic content and the highest taxonomic resolution amongst the genes investigated, it also had a gene phylogeny that related well with that of the 16S rRNA gene

Alcaligenes aquatilis sp. nov.: a novel bacterium from sediments of the Weser Estuary, Germany and a salt marsh Shem Creek in Charleston Harbour, USA

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A novel strategy for the isolation and identification of environmental Burkholderia cepacia complex bacteria.

The purpose of this study was to develop a novel strategy for the isolation and identification of Burkholderia cepacia complex bacteria from the home environment of cystic fibrosis (CF) patients. Water and soil samples were enriched in a broth containing 0.1% l-arabinose, 0.1% l-threonine, and a mixture of selective agents including 1 microgml(-1) C-390, 600U ml(-1) polymyxin B sulfate, 10 microgml(-1) gentamycin, 2 microgml(-1) vancomycin and 10 microgml(-1) cycloheximide. On selective media (consisting of the same components as above plus 1.8% agar), several dilutions of the enrichment broth were inoculated and incubated for 5 days at 28 degrees C. Isolates with different randomly amplified polymorphic DNA patterns were inoculated in Stewart's medium. Putative B. cepacia complex bacteria were confirmed by means of recA PCR and further identified by HaeIII-recA restriction fragment length polymorphism analysis. Our results suggest that these organisms may be more widespread in the home environment than previously assumed and that plant associated soil and pond water may be reservoirs of B. cepacia complex infection in CF patients.Journal ArticleResearch Support, Non-U.S. Gov'tSCOPUS: ar.jinfo:eu-repo/semantics/publishe