Bio-safety of milk products and mycobacterium avium subspecies paratuberculosis as major microbial contaminant using multiple tests including culture and sybr green real-time assay

Abstract

Twelve types of milk products belonging to 22 market brands were purchased from local shops in South Uttar Pradesh and were screened for the presence of Mycobacterium avium subspecies paratuberculosis (MAP) as major contaminant using six tests (microscopy, Indirect Fluorescent Antibody Test, IS900 PCR, Indigenous Enzyme linked Immuno Sorbent Assay, dot_ELISA, and Latex agglutination Test). Sample positive in any one of the six tests was considered as positive. Of 276 milk products screened, the cumulative bio-presence of MAP was, 52.8% (146) and was highest in butter (75.0%), followed by curd (66.0%), buttermilk (52.9%), lassi (50.0%), cheese (40.0%) and ice-cream (28.5%). Bio-typing of MAP DNA from milk products using IS1311 PCR_REA revealed presence of ‘Indian Bison Type’ as a major biotype. Kappa (0.700 – 0.815) and two-tailed p (<0.0001–1.0) values for six tests were significant for all six tests. This study for the first time revealed large scale contamination of ‘milk products’ marketed by leading commercial brands in India, with MAP bacilli and therefore not safe for human consumption.