Original Articles Mycobacterium avium subsp. paratuberculosis Detection in Individual and Bulk Tank Milk Samples from Bovine Herds and Caprine Flocks Lucı ´a C. Favila-Humara, 1 Gilberto G. Cha ´ vez-Gris, 2 Erika M. Carrillo-Casas, 3 and Rigoberto Herna ´ ndez-Castro 3 Abstract Paratuberculosis, or Johne’s disease, is caused by Mycobacterium avium subsp. paratuberculosis (Map), and it generates great economic losses for the dairy industry worldwide. In humans, Map has been associated with Crohn’s disease. Mexico has unknown paratuberculosis prevalence, and yet, control programs have not been applied. This study aimed to determine the presence of Map in milk samples from seropositive goats and cows and bulk tank milk samples from herds previously designated Map-infected using indirect enzyme-linked immunosorbent assay. Map DNA was detected in 100% of the bulk tank milk samples of 14 bovine herds and 3 caprine flocks using a modified insertion sequence 900 polymerase chain reaction (PCR). Additionally, Map DNA was detected in 100% of the individual milk samples from 10 cows and 8 goats. Further, based on the findings of the experimental insertion sequence 900 PCR assessment, evaluation of bulk tank and individual milk samples through a type-specific PCR was performed, which confirmed our previous findings and revealed that 56.25% cow and 63.63% goat milk had concurrent infections of the C, I, and S types. Out of 14 bulk tank milk samples, 10 had viable mycobacteria. Paratuberculosis was detected at a high frequency in cow and goat milk, which suggests that raw milk ingestion represents a potential risk of Map infection. Introduction P aratuberculosis, or Johne’s disease, is a chronic, progressive granulomatous enteritis that can affect cattle, sheep, goats (Collins et al., 2002), and other nonruminant wildlife species. This infectious disease is caused by Myco- bacterium avium subsp. paratuberculosis (Map) and is charac- terized by diarrhea and weight loss, leading to cachexia and death (Manning and Collins, 2001). As the link between Map and Crohn’s disease pathogenesis in humans remains unclear (Chamberlin et al., 2001), and considering the potential health risk, the potential routes of human exposure to MAP should be investigated (Eltholth et al., 2009). Accordingly, many ex- perimental studies have investigated Map’s resistance to the current standard pasteurization processes when high colony numbers are present (Grant et al., 2002a; Grant, 2006). More- over, its survival in cheese and raw milk has also been ad- dressed (Corti and Stephan, 2002; Grant et al., 2002b; Ayele et al., 2005; Ellingson et al., 2005; Stephan et al., 2007). Ad- ditionally, paratuberculosis is recognized as one of the main diseases in ovine flocks, and it is common in caprine and bovine herds (Cha ´ vez-Gris et al., 2004; Este ´vez-Denaives et al., 2007). Raw milk ingestion is still a common practice in Me ´xico, and fresh cheese is frequently made with unpas- teurized milk. The true disease prevalence in this country remains unknown, and yet, no paratuberculosis control pro- grams have been currently conducted. A variety of DNA probes have been developed and evaluated based on insertion sequence 900 (IS900), an insertion element regarded as spe- cies-specific for Map, which has been useful for the early de- tection of Johne’s disease (Englund et al., 2002). Culture differences and genetic polymorphism studies classify Map strains into two principal types: cattle as ‘‘C’’ and sheep as ‘‘S,’’ and also an intermediate type has been described (Pavlı ´k et al., 1995). Each type can infect a wide range of hosts; the C type has been recognized in the majority of studies, including all human isolations (Pavlı ´k et al., 1995; Cha ´ vez-Gris et al., 2004). This study aimed to identify the presence of Map and 1 National Center of Disciplinary Investigations in Animal Microbiology, National Institute of Forest, Agricultural, and Cattle Investigations, Mexico City, Mexico. 2 National Autonomous University of Mexico, Faculty of Veterinary Medicine, Diagnostic and Verification Services Unity, Teaching Investigation and Extension Center for Animal Production in the Altiplane, Tequisquiapan, Mexico. 3 Direction of Education and Investigation, Department of Ecology and Pathogenic Agents, Department of Molecular Biology and His- tocompatibility, Dr. Manuel Gea Gonzalez General Hospital, Mexico City, Mexico. FOODBORNE PATHOGENS AND DISEASE Volume 7, Number 4, 2010 ª Mary Ann Liebert, Inc. DOI: 10.1089=fpd.2009.0374 351