Genome persistence & survival of Clostridium perfringens from ichthyofauna of dal lake Himalaya-A possible future public health hazard

Abstract

Background: Clostridium perfringens is one of the most prolific toxin-producing bacteria, an indicator of fecal contamination in aquatic habitats. This bacterium is not known to cause any disease in fish but its importance in fish is primarily due to the health hazards to humans, as C. perfringens has been found to be the causative agent of many outbreaks of food poisoning associated with the consumption of fish and its products. Aquatic resources of Kashmir are in a serious state of eutrophication-particularly cultural, more pronounced in Dal Lake, which has witnessed an increase in food borne pathogens. We aimed to investigate the presence of different toxinotypes of C. perfringens in commonly consumed fishes of Kashmir. Methods and materials: Collection of samples. Isolation and identification by culture & staining. DNA extraction and PCR using species-specific 16S rRNA gene amplification, The toxinotyping of C. perfringens isolates by multiplex-PCR targeting six toxin genes (cpa, cpb, ɛtx, ιtx, cpb2 and cpe). Phylogenetic analysis and antibiotic sensitivity tests. Results: All the 45 isolates from water and 37 isolates from fish (21 from scale-carp and 16 from snow-trout) were found to carry cpa gene alone as a major toxin gene and thus were designated as toxinotype A. None of the isolates carried cpb, ɛtx, ιtx, cpb2 or cpe genes indicating the absence of C. perfringens toxinotypes B, C, D or E in the water and fish samples of the Dal Lake. The prevalence of C. perfringens was slightly higher in scale-carps (35%) compared to snow-trouts (26.6%).Phylogenetic analysis of the cpa toxin genes of C. perfringens revealed 95% to 98% homology with corresponding GenBank published sequences and close relationship with corresponding AA sequences of C. perfringens strains reported from India, Egypt and China. Conclusion: Presence of C. perfringens toxinotype A in fish for the first time in the Dal Lake of the Kashmir valley and implied that fish may be a possible source of C. perfringens type A intoxication to humans through food chain posing a serious public health concern.The antibiogram pattern of the C. perfringens type A isolates revealed a higher antimicrobial resistance to amikacin, ceftriaxone, metronidazole, norfloxacin and tetracycline.