Rapid Identification of Thermotolerant Campylobacter jejuni , Campylobacter coli , Campylobacter lari , and Campylobacter upsaliensis from Various Geographic Locations by a GTPase-Based PCR-Reverse Hybridization Assay

Abstract

Recently, a gene from Campylobacter jejuni encoding a putative GTPase was identified. Based on two semiconserved GTP-binding sites encoded within this gene, PCR primers were selected that allow amplification of a 153-bp fragment from C. jejuni, C. coli, C. lari, and C. upsaliensis. Sequence analysis of these PCR products revealed consistent interspecies variation, which allowed the definition of species-specific probes for each of the four thermotolerantCampylobacter species. Multiple probes were used to develop a line probe assay (LiPA) that permits analysis of PCR products by a single reverse hybridization step. A total of 320 reference strains and clinical isolates from various geographic origins were tested by the GTP-based PCR-LiPA. The PCR-LiPA is highly specific in comparison with conventional identification methods, including biochemical and whole-cell protein analyses. In conclusion, a simple method has been developed for rapid and highly specific identification of thermotolerant Campylobacter species.

Development of a novel triplex PCR assay for the detection and differentiation of thermophilic species of Campylobacter using 16S-23S rDNA internal transcribed spacer (ITS) region

“…However, these target genes have had limited impact in differentiating Campylobacters species due to insufficient variability in their sequences (Gonzalez et al. 1997; Fermer and Engvall 1999; van Doorn et al. 1999; Steinhauserova et al.…”

“…Existing PCR‐based assays for detection and differentiation in Campylobacter have used primers targeting the genes encoding 16S rRNA, 23S rRNA, GTPase and ceuE (Gonzalez et al. 1997; Hurtado and Owen 1997; Fermer and Engvall 1999; van Doorn et al. 1999; Steinhauserova et al.…”

Campylobacter Species and Neutrophilic Inflammatory Bowel Disease in Cats

“…This was screened by Langford Veterinary Services (Bristol, UK) for the presence of Salmonella, pathogenic E. coli , Campylobacter spp., and Clostridium spp. A 153‐bp DNA fragment unique to Campylobacter was amplified by PCR and sequenced to confirm the Campylobacter species present.…”

Molecular typing of micro-organisms: at the centre of diagnostics, genomics and pathogenesis of infectious diseases?

“…Hybridisation-mediated analyses of these DNA fragments con®rmed that these sub-genomic DNA molecules frequently harboured strictly species-speci®c sequences. Sequence elucidation revealed that one of the most reliable species-speci®c probes derived from a gene encoding a C. jejuni GTPase, based on which phylogenetic comparisons could be made to identify distinct species in even more detail [6]. Furthermore, the sequence heterogeneity thus documented among species could be used to improve diagnostic width and to develop and validate a reliable reversed hybridisa-tion-mediated line probe assay (Innogenetics, Ghent, Belgium).…”