Genetic relatedness within the genus Campylobacter inferred from rpoB sequences

Abstract

The genus Campylobacter comprises 17 species, some of which are important animal and human pathogens. To gain more insight into the genetic relatedness of this genus and to improve the molecular tools available for diagnosis, a universal sequencing approach was established for the gene encoding the beta-subunit of RNA polymerase (rpoB) for the genus Campylobacter. A total of 59 strains, including the type strains of currently recognized species as well as field isolates, were investigated in the study. A primer set specific for Campylobacter species enabled straightforward amplification and sequencing of a 530 bp fragment of the rpoB gene. The 16S rRNA gene sequences of all of the strains were determined in parallel. A good congruence was obtained between 16S rRNA and rpoB gene sequence-based trees within the genus Campylobacter. The branching of the rpoB tree was similar to that of the 16S rRNA gene tree, even though a few discrepancies were observed for certain species. The resolution of the rpoB gene within the genus Campylobacter was generally much higher than that of the 16S rRNA gene sequence, resulting in a clear separation of most species and even some subspecies. The universally applicable amplification and sequencing approach for partial rpoB gene sequence determination provides a powerful tool for DNA sequence-based discrimination of Campylobacter species.The genus Campylobacter was first proposed by M. Sebald and M. Véron in 1963 and included only the type species Campylobacter fetus and Campylobacter bubulus, now known as Campylobacter sputorum (Sebald & Véron, 1963). These taxa had formerly been classified as Vibrio species. In 1973, M. Véron and R. Chatelain included several misclassified Vibrio in the distinct Campylobacter genus based on serological, biochemical and DNA-DNA hybridization investigations (On, 2001). Since then, the taxonomy of the genus has changed dramatically. At present, it comprises 17 species with validly published names and six recognized subspecies (On, 2001;Foster et al., 2004). In addition, strains belonging to C. sputorum are divided into three biovars, sputorum, fecalis and paraureolyticus, on the basis of their ability to produce catalase or urease Vandamme & On, 2001).In general, members of the genus Campylobacter colonize the mucosal surfaces of the intestinal tract, oral cavity or urogenital tract of healthy, as well as diseased, humans and animals, especially birds. Several species may act as pathogens, causing disease in both human and animal hosts. Twelve of the 17 Campylobacter species are associated with human diseases. Campylobacter jejuni and Campylobacter coli are particularly frequent causative agents of human bacterial intestinal disorders worldwide (Skirrow, 1994). There have also been reported cases of diarrhoea in man caused by Campylobacter upsaliensis and Campylobacter lari, but the frequency of these infections is very low (Bourke et al., 1998;Van Doorn et al., 1998). Occasionally, Campylobacter species are implicated as causative agents of pericard...

Multiplex Strategy for Multilocus Sequence Typing, fla Typing, and Genetic Determination of Antimicrobial Resistance of Campylobacter jejuni and Campylobacter coli Isolates Collected in Switzerland

“…Genetic identification and phylogenetic investigation of the C. jejuni and C. coli strains were performed by sequence analysis of the 16S rRNA and rpoB genes. Phylogenetic analysis of the 16S rRNA gene fragments of all 186 C. jejuni and 143 C. coli strains confirmed previous studies showing that the resolution of this target is not high enough to separate these closely related species (10,15). The 16S rRNA gene sequences of C. jejuni and C. coli match up to 100%.…”

“…Cluster I was closely related to C. jejuni NCTC 11351 T (AF372097), cluster II to C. jejuni 81-176 (CP000538), cluster III to C. jejuni NCTC 11168 (AL111168), and cluster IV to C. jejuni CCUG 12066 (DQ174200). Most C. coli isolates (93%) showed very high ropB sequence similarity (the highest sequence difference was 1 base out of 487) to the type strain of C. coli LMG 6440 T (AF372098) and formed cluster V. A very distinct cluster, VI, was found for some C. coli isolates that were observed earlier (15). This group is mainly formed by multiple isolates of the previously known ST1147 and ST1426, as well as single isolates of the newly determined ST2914, ST2915, and ST3345.…”

“…Primer sets were designed for both the flaA and flaB genes, covering the "short variable regions," and for the amplification of the gyrA fragment. Previously published primers were used to generate the fragments from the 23S rRNA gene (34), the 16S rRNA gene (16), and the rpoB gene (15). The primers and their locations, as well as the resulting fragment sizes, are listed in Table 1.…”

Arcobacter mytili sp. nov., an indoxyl acetate-hydrolysis-negative bacterium isolated from mussels

“…Sequence assembly was performed with AUTO ASSEMBLER (Applied Biosystems). In addition, the rpoB genes of the four novel isolates and 11 Arcobacter strains [two strains for each recognized species of the genus Arcobacter, with the exception of A. halophilus (see Supplementary Table S1)], were amplified using the PCR primers CamrpoB-L and RpoB-R and the conditions that have been established for use with the genus Campylobacter (Korczak et al, 2006). The expected PCR product size (524 bp) was obtained for all strains, with additional unexpected bands in some cases.…”

Melting of the Ribosomal RNA Gene Reveals Bacterial Species Identity: A Step toward a New Rapid Test in Clinical Microbiology

“…Moreover, it is well known that certain bacterial species share almost identical 16S rDNA sequences (e.g., E. coli, Salmonella and Shigella). In such cases, more discriminatory regions are necessary (18 ), or a battery of species-specific assays can be performed.…”