DNA fingerprinting and RAPD-PCR analysis of S. aureus strains isolated from bovine origin

Bovine mastitis is the greatest source of economic loss in the dairy industry. Highly reproducible and definitive detection of causative agent is very important for treatment and control of the disease. The aim of this study was to compare culture and polymerase chain reaction (PCR) techniques for diagnosis of agents in subclinical bovine mastitis. All the mastitic milk samples were diagnosed as solely infected by Staphylococcus spp. using the classical microbiological method following culturing. The objectives of the present study were to study the PCR-based fingerprinting of Staphylococcus aureus isolated from bovine mastitis. California Mastitis Test (CMT)-positive milk samples were received from mastitis cases. Milk samples scored positive for CMT were subjected to cultural isolation. Preliminary identification and characterisation of the isolates recovered were carried out by conventional phenotypic test, that is various biochemical tests and coagulase test. Out of 245 animals screened, 93 were found positive for subclinical mastitis. From the investigation of 190 quarter samples, a total of 216 isolates were obtained. One hundred sixty eight were identified as Staphylococcus spp., and out of these, 70 were distinguished as Coagulase-positive Staphylococci (CPS), and further 56 isolates were characterised as S. aureus phenotypically. PCR-based fingerprinting of 56 S. aureus recovered from different regions was carried out using randomly amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR). The OEP-4 primer generated multiple amplicons with size ranging from 200 bp to 3000 bp in 56 isolates. In most of the isolates, amplicon of 1, 100 bp was common. On the basis of amplicon size, dendrogram was generated. The RAPD-PCR analysis types all 56 S. aureus into 15 clads (C1-C15). Location-wise study revealed that S. aureus isolates were categorised into four major clusters. The results demonstrate the genetic heterogeneity in the strains studied and the dissemination of some clones through different regions. The present findings allow the epidemiological monitoring of S. aureus causing mastitis. The DNA fingerprinting as defined for each strain of S. aureus could be useful in epidemiology studies and the identification of new strains and their origins and selection of predominant strain among CPS for development of preventive biologicals.