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Mycobacterium avium paratuberculosis (MAP) is causative agent of Johne's disease (JD) in domestic animals and has broad host range. JD infected animals shed viable MAP in their milk, feces, blood, and tissues which get transmitted to human beings directly or indirectly by consumption of animal products, through contact, animal handling and through contaminated environment, aerosols. In this current study, we developed hydrolysis probe based TaqMan® real-time PCR assay where samples were investigated by targeting IS900 mRNA and ModD gene to differentiate live MAP shedders from inactive/dead MAP bacilli shedding animals. The IS900 mRNA and ModD gene primers were designed using discontiguous unique conserved sequences of IS900 more towards the 3' end and fibronectin attachment protein (FAP) genes, respectively. Two different reporter dyes Cy5 and TexasRed, with compatible quenchers BHQ-1 and BHQ-2, respectively, were used for probe designing of IS900 and ModD genes. Triplex PCR assay was developed by using serially diluted positive MAP culture in log

Staphylococcus aureus is one of the most prevalent pathogens, and a causative agent of a variety of infections in humans and animals. A total of 48 semen samples were collected from healthy bucks of different breeds to investigate the prevalence of S. aureus . Antimicrobial resistance and virulence of the Staphylococcus isolates were determined to assess the adverse effects of them on buck fertility. The bacterial isolates were tentatively confirmed as Staphylococcus spp. based on the Gram’s staining, growth on Mannitol salt agar and catalase test. Overall, 75% (n = 36) of the samples were positive for Staphylococcus spp. from the total 48 buck semen ejaculates from different breeds. Out of 36 staphylococcal isolates, 23 (47.92%) were coagulase negative (CoNS) and 13 (27.08%) were coagulase positive Staphylococcus (CoPS) based on the slide coagulase test. In the current study, on the basis of molecular characterization, we identified S. aureus , S. chromogenes, S. haemolyticus, S. sciuri, S. simulans and S. epidermidis amongst the staphylococcal isolates in the buck semen. This study revealed a high prevalence of Staphylococcus species in semen of the healthy bucks. The isolates exhibited varying degrees of multidrug resistance genotypically as well as phenotypically. The presence of antibiotic resistance and virulence genes may pose a potential threat to reproductive health of animals, highlighting the need for vigilant monitoring of these isolates at the time of semen cryopreservation.

Johne’s disease is major infectious disease of domestic livestock in India. Despite low per animal productivity, the country is yet to initiate plan for the survey and control of disease at National scale. A total of 81 clinical samples from Mandya breed of sheep were collected from an organised farm (Livestock Research and Information Centre, Nagmangala) and farmer’s flocks suspected for Johne’s disease on the basis of clinical symptoms (weakness, emaciation, diarrhea and alopecia). Only 10.0% samples from two farms were screened for bio-load of Mycobacterium avium subspecies paratuberculosis (MAP) using multiple tests (microscopy, indigenous ELISA, IS900 PCR and culture). Results showed that 100.0% samples were positive for MAP infection both in microscopy and “Indigenous ELISA”. IS1311 PCR-REA bio-typed 60.0% (feces) and 100.0% (blood and tissue) isolates as “Indian Bison Type”. Typing investigated for the first time that biotype was of Indian origin and present in sheep flocks of South India having wide geographical distribution and broad host range. Study reported high bio-load of MAP in Mandya breed of sheep in Nagamangala, Mandya district and Bilagi taluk of Bagalkot district in South India. Being Johne’s disease a spectral disease, multiple tests were useful for the screening of MAP infection in sheep farms. Study emphasised the need for initiation of short and long term control programs to reduce production losses and sharing of MAP strains by other domestic livestock species and prevent contamination of human food chain and reduce risk to human population.