Rapid differentiation of Mycobacterium avium and M-paratuberculosis by PCR and restriction enzyme analysis

Mycobacterium avium subsp. avium (M. avium) and M. avium subsp. paratuberculosis (M. paratuberculosis), intracellular bacteria that can cause chronic granulomatous enteritis in cattle, are difficult to distinguish on the basis of growth and biochemical characteristics. We report the development of a PCR-based strategy for the rapid differentiation of isolates of M. paratuberculosis. Restriction fragment length polymorphism was identified by PCR amplification and subsequent restriction enzyme digestion with PstI of a 960-bp fragment of the 65-kDa heat shock protein (hsp65) from 21 clinical isolates of M. paratuberculosis and 14 isolates of M. avium. These results indicate that a restriction fragment length polymorphism in the hsp65 gene can be used for the rapid differentiation of clinical isolates of M. paratuberculosis and M. avium.