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Clostridium perfringens (C. perfringens), is a spore-forming and toxin-producing pathogenic Gram-positive rod-shaped bacterium with immense public health/zoonotic concern. Rodents are well-known reservoirs and vectors for a large number of zoonoses and strong links have been recognized between synanthropic rodents and foodborne disease outbreaks throughout the world. To date, no study has been conducted for studying the prevalence of C. perfringens in rodents and shrews. In this study, we investigated faecal samples from free-living rodents and shrews trapped in Meghalaya, a North-eastern hill state of India for the presence of virulent and antimicrobial-resistant C. perfringens. A total of 122 animals comprising six species of rodents and one species of shrews were trapped: Mus musculus (n = 15), Mus booduga (n = 7), Rattus rattus (n = 9), Rattus norvegicus (n = 3), Bandicota indica (n = 30), Bandicota bengalensis (n = 32) and Suncus murinus (n = 26). The faecal swabs were collected and processed for the isolation of C. perfringens. Toxinotyping was done using PCR. Antimicrobial susceptibility testing and biofilm forming ability testing were done using Kirby Bauer disc diffusion method and crystal violet assay. C. perfringens was isolated from 27 of the 122 faecal swabs (22.1%), from six species of rodents and shrews. Five of the host species were rodents, Bandicota bengalensis (25%), Bandicota indica (16.7%), Rattus norvegicus (33.3%), Mus musculus (13.3%), Mus booduga (42.8%) and Suncus murinus (shrew) (29.6%). The common toxinotype was type A (59.2%) followed by Type A with beta2 toxin (33.3%), Type C (3.7%) and Type C with beta2 toxin (3.7%). None of the isolates harboured cpe, etx, iap, and NetB genes and therefore none was typed as either B, D, E, F, or G. Nine isolates (33.3%) turned out to be multi-drug resistant (MDR), displaying resistance to three or more categories of antibiotics tested. Twenty-three out of twenty-seven isolates (85.2%) were forming biofilms. Globally, this is the first study to report the prevalence of C. perfringens and its virulence profile and antimicrobial resistance in free-living rodents and shrews. The rodents and shrews can potentially contaminate the food and environment and can infect humans and livestock with multi-drug resistant/virulent Type A and Type C C. perfringens. [Display omitted] • Globally, first study to describe the faecal prevalence of Clostridium perfringens in free-living rodents and shrews. • C. perfringens is 22.1% prevalent in free-living rodents and shrews. • C. perfringens type A, type A cpb2 +, type C, and type C cpb2 + strains were detected. • Multi-drug resistance was displayed by 33.3% of isolates. • Biofilm formation was identified in 85.2% of isolates. [ABSTRACT FROM AUTHOR]

Clostridium perfringens (C. perfringens), a prolific toxin-producing anaerobe is an important foodborne pathogen with a huge public health concern. Rapid and on-site detection of C. perfringens is of specific importance in developing countries. In the present study, saltatory rolling circle amplification (SRCA) assay was developed for culture-independent, rapid and visual detection of C. perfringens and evaluated in meat with pork as a model. The specificity of the SRCA assay was ascertained by using 62 C. perfringens and 18 non- C. perfringens strains. The analytical sensitivity of the developed SRCA, conventional and real-time PCR assays were 80 fg, 800 fg and 800 fg DNA per tube, respectively. The limit of detection of the SRCA assay was 80 CFU/g of pork in the absence of enrichment and 8 CFU/g after short enrichment of 6 h. The detection limits of 80 CFU/g and 8 CFU/g of pork were attained within 120 min and 8 h, respectively. Real-world or field relevancy of the developed assay was evaluated by screening 82 raw and processed pork samples. As the developed assay is simple, user-friendly, cost-effective and sophisticated-equipment free, it would be more suitable for on-site testing of C. perfringens in foods. To our information, this is the first report to apply SRCA for the detection of C. perfringens. [Display omitted] • A novel SRCA assay was developed for rapid and visual detection of Clostridium perfringens in meat. • The analytical sensitivity of the SRCA assay (80 fg) was 10 fold more than the conventional and real-time PCR. • The detection limit of 80 CFU and 8 CFU per gram of meat was attained within 120 min and 8 h, respectively. • This is the first report to use SRCA for the detection of C. perfringens. [ABSTRACT FROM AUTHOR]