The widespread status of subclinical condition of
bovine mastitis is often associated with the production of
leukotoxin M/F'-PV producing Staphylococcus aureus. The present study aims for the profiling of such
leukotoxin producers through conventional and molecular methods in parallel to their leukotoxicity. The incidence of this particular pathogen was assessed in
mastitis infected Holstein-Friesian cattle, where eight isolates of staphylococci were found to be present in 20 % of collected samples. Being intermediately resistant to
vancomycin, they showed characteristic double zone
hemolysis on 7 % sheep blood
agar and typical type II reaction for
coagulase test indicating the pathogenic attributes. Further with RAPD-PCR and 16S
rDNA-RFLP, epidemiological specificity and genotypic relatedness of isolates to S. aureus was confirmed. Subsequently, the presence of
leukotoxin (lukM) gene in native isolates was detected by
leukotoxin gene specific PCR. MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium) assay evaluated for secreted
leukotoxin in cell free supernatant was estimated to be 223 toxic units which had an LD50 cytotoxic activity on bovine neutrophil. Thus, the data acquired during study can be of prime diagnostic method for timely and accurate analysis of subclinical
mastitis samples which goes undetected at consumer level.