Detection of Mycobacterium avium Sub sp. paratuberculosis in Pasteurized Milk Samples in Northeast of Iran by Culture, Direct Nested PCR and PCR Methods

Mycobacterium avium Subsp. paratuberculosis (MAP) is a gram-positive, small, acid-fast bacillus with high environmental resistance. In animals, especially ruminants, it leads to Paratuberculosis (PTB) or Johne's disease, which is chronic granulomatous enteritis. This bacterium as the main causative agent of Crohn's disease can be a serious threat to human health. This study aimed to detect MAP in pasteurized milk samples produced in Khorasan Razavi province, Iran, using Direct Nested PCR, PCR, and culture methods. In this study, ۵۴۴ milk samples from Pasteurized Milk Production Companies were selected randomly during the ۳-month period. DNA was extracted from milk fat after centrifugation. In order to identify the bacteria, Direct Nested PCR and PCR tests were applied using IS۹۰۰ and f۵۷, respectively. Furthermore, to detect viable MAP, positive samples resulted from Direct Nested PCR assays were cultured on Herrold's egg medium. For identification of mycobacterial isolates, all colonies were processed by PCR based on f۵۷. A total of ۵۴۴ pasteurized milk samples were assayed, and Mycobacterium paratuberculosis was detected in ۳۹% of them by IS۹۰۰ Nested PCR, and only ۴.۹% of samples were positive in the culture method. All the colonies were positive for the f۵۷using PCR. The results of this study indirectly indicated a high level of contamination of pasteurized milk to Mycobacterium paratuberculosis which is due to the large number of affected animals in livestock farms in Khorasan Razavi province. However, in comparison with the other researches, the low percentage of viable bacteria in pasteurized milk can be due to changes in temperature and time in pasteurizing systems of milk production companies in Khorasan Razavi province, Northeast of Iran.

کلیدواژه ها:

Mycobacterium avium ، Paratuberculosis ، IS۹۰۰ ، Nested PCR ، f۵۷