Genetic diversity of Clostridium perfringens type A isolates from animals, food poisoning outbreaks and sludge.

BACKGROUND: Clostridium perfringens, a serious pathogen, causes enteric diseases in domestic animals and food poisoning in humans. The epidemiological relationship between C. perfringens isolates from the same source has previously been investigated chiefly by pulsed-field gel electrophoresis (PFGE). In this study the genetic diversity of C. perfringens isolated from various animals, from food poisoning outbreaks and from sludge was investigated. RESULTS: We used PFGE to examine the genetic diversity of 95 C. perfringens type A isolates from eight different sources. The isolates were also examined for the presence of the beta2 toxin gene (cpb2) and the enterotoxin gene (cpe). The cpb2 gene from the 28 cpb2-positive isolates was also partially sequenced (519 bp, corresponding to positions 188 to 706 in the consensus cpb2 sequence). The results of PFGE revealed a wide genetic diversity among the C. perfringens type A isolates. The genetic relatedness of the isolates ranged from 58 to 100% and 56 distinct PFGE types were identified. Almost all clusters with similar patterns comprised isolates with a known epidemiological correlation. Most of the isolates from pig, horse and sheep carried the cpb2 gene. All isolates originating from food poisoning outbreaks carried the cpe gene and three of these also carried cpb2. Two evolutionary different populations were identified by sequence analysis of the partially sequenced cpb2 genes from our study and cpb2 sequences previously deposited in GenBank. CONCLUSION: As revealed by PFGE, there was a wide genetic diversity among C. perfringens isolates from different sources. Epidemiologically related isolates showed a high genetic similarity, as expected, while isolates with no obvious epidemiological relationship expressed a lesser degree of genetic similarity. The wide diversity revealed by PFGE was not reflected in the 16S rRNA sequences, which had a considerable degree of sequence similarity. Sequence comparison of the partially sequenced cpb2 gene revealed two genetically different populations. This is to our knowledge the first study in which the genetic diversity of C. perfringens isolates both from different animals species, from food poisoning outbreaks and from sludge has been investigated.

Genetic relatedness and netB prevalence among environmental Clostridium perfringens strains associated with a broiler flock affected by mild necrotic enteritis.

In a previous study we investigated pulsed-field gel electrophoresis (PFGE) genotype diversity and prevalence of the netB toxin gene in Clostridium perfringens (CP) isolates recovered from a broiler flock (flock 1) affected by necrotic enteritis (NE). In this follow-up work, we examined samples collected before placement of flock 1, to see if NE during rearing could be traced back to the cleaned and empty building or the day-old chicks. Litter from the next flock in the same building (flock 2) was also examined. We detected 25 different PFGE genotypes, five of which were found only in litter from flock 2. Six genotypes which had been found in flock 1 during rearing were detected in samples collected before placement. NetB positive isolates belonging to two of these genotypes had been recovered from NE lesions during rearing, suggesting that virulent strains were transmitted from the cleaned and disinfected broiler house. NetB frequency among isolates from the empty building was 45%, indicating that netB positive strains were prevalent in a building that previously had housed a healthy flock offered in-feed narasin (flock 0). NetB frequency among isolates from litter used by flock 2 was 22%, indicating that netB positive strains were present in the environment of a 14-days-old healthy flock offered in-feed narasin. Two prevalent genotypes were consistently either netB negative or netB positive. However, the presence of genotypes represented by both negative and positive isolates may suggest that the gene can spread horizontally among different CP strains.

Genetic diversity and prevalence of netB in Clostridium perfringens isolated from a broiler flock affected by mild necrotic enteritis.

This study was undertaken to examine the genetic diversity of Clostridium perfringens isolated from a single broiler flock reared without in-feed antimicrobials (antibacterial growth promoters and anticoccidials) and affected by mild necrotic enteritis (NE). We used pulsed-field gel electrophoresis (PFGE) to investigate the genetic diversity of C. perfringens isolates from broilers of varying disease status, and from litter. The prevalence of the toxin gene netB was also investigated. Altogether 32 PFGE genotypes were found among 88 isolates. Several genotypes were detected in C. perfringens-associated organ lesions from chickens that were sampled at random and alive without clinical symptoms, suggesting that these genotypes proliferated concurrently in such lesions. More than 90% of all isolates from NE-specific organ lesions carried netB which codes for a recently described pore-forming toxin. NetB positive isolates were less predominant in non-lesion samples from broilers affected by NE, and found infrequently or not at all in healthy birds and isolates from litter. These findings show that the presence of netB in C. perfringens strains is associated with NE and suggest that mild NE differs from severe NE with regard to C. perfringens genotype diversity.