Molecular Subtyping of Poultry-Associated Type AClostridium perfringensIsolates by Repetitive-Element PCR

Abstract

Clostridium perfringens strains (type A) isolated from an integrated poultry operation were subtyped using repetitive-element PCR with Dt primers. Isolates were obtained from fecal, egg shell, fluff, and carcass rinse samples as part of a previously reported temporally linked epidemiological survey. A total of 48 isolates of C. perfringens were obtained from different stages of the broiler chicken production chain from two separate breeder farms that supplied a single hatchery that in turn provided chicks to a single grow-out farm whose flocks were processed at a single plant. All 48 isolates were typeable (100% typeability) by repetitive-element PCR with Dt primers. This subtyping method was highly reproducible and discriminatory. By repetitive-element PCR with Dt primers, isolates were classified into four major branches with 12 subgroups or clades. The Simpson's index of discrimination was calculated to be 0.96 for groupings of >95% correlation. Toxin gene profiles of the isolates indicated that all of the isolates were C. perfringens alpha-toxin gene positive and 46 of 48 isolates were beta2-toxin gene positive. All strains were negative for beta-and epsilon-toxin genes. Repetitive sequence-based PCR was found to be a technically practical and reproducible means of subtyping C. perfringens libraries from specific epidemiological or production environment settings.

Molecular detection and characterization of cpb2 gene in Clostridium perfringens isolates from healthy and diseased chickens

“…The extent to which the cpb2 gene has been found in isolates from healthy flocks has also varied. Both high (46 out of 48) and low prevalences (4 out of 27) of the cpb2 gene were previously shown in isolates from healthy flocks (21,22). In the present study, we found 42 out of 43 isolates positive for the cpb2 gene.…”

“…This finding may be explained under the assumption that these isolates were taken from two different flocks originating in geographically separate areas of Iran. It appears that the type of cpb2 alleles may differ in distinct geographical locations (22,23). Siragusa et al (22) reported the presence of the atypical cpb2 allele among North American C. perfringens isolates, whereas Johansson et al (23) found that the European C. perfringens isolates carried the consensus cpb2 allele.…”

Multilocus Sequence Typing Subtypes of Poultry Clostridium perfringens Isolates Demonstrate Disease Niche Partitioning

“…Genetic relationships between C. perfringens strains of disparate origins, including human and livestock host species, have been investigated (3,10,16,32,34). Such phylogenetic studies have elucidated distinct lineages associated with specific disease syndromes attributed to this pathogen.…”

Comparison and Utilization of Repetitive-Element PCR Techniques for Typing Lactobacillus Isolates from the Chicken Gastrointestinal Tract

“…In contrast, repetitive-element PCR (Rep-PCR) has been developed for genotypically fingerprinting bacteria and is a fast and reliable high-throughput genotyping system (66,67). Rep-PCR has been used successfully to identify strains of a variety of genera (33,44,48,55). Several Rep-PCR primers, including the repetitive extragenic palindromic (REP) primers (6,9,16,65), the enterobacterial repetitive intergenic consensus (ERIC) primers (6,65), and the (GTG) 5 primer (24,40,62), have been used in the typing of lactobacilli in studies which have generally focused on species important to the dairy industry and food fermentations.…”