Multiplex polymerase chain reaction for identification of bovine mastitis associated pathogens

Abstract

The present study was focused on the development of a sensitive multiplex polymerase chain reaction (mPCR) for rapid detection of mastitis associated pathogens directly from milk samples. A total of 40 mastitic milk samples were processed for isolation and identification of pathogens which were mentioned below by conventional method and compared it with mPCR. The prevalence of bacterial isolates in conventional detection were found to be 20 (50%) Staphylococcus aureus, 10 (25%), Streptococcus agalactiae, 9 (22.5%) Klebsiella pneumoniae and 8 (20%) Escherichia coli, respectively. In mPCR all culture-positive samples were detected for the corresponding bacteria but additional numbers of (i.e. 5 S. aureus, 7 S. agalactiae, 2 E.coli, and 4 K. Pneumoniae) isolates were also detected by mPCR which were culture‐negative. The target sequence for primer designing were the different gene i.e. vicK gene (S. aureus), atr gene (S. agalactiae), barA gene (K. pneumoniae) and uidA gene (E. coli). It was concluded that, Staphylococcus aureus appeared as the most prevalent organism responsible for causing mastitis in cattle. Furthur, the results suggested that mPCR was significantly more sensitive than culture detection. A multiplex PCR could be used as an efficient tool for detecting pathogens of mastitis with high accuracy, sensitivity and in a short period of time.