Molecular Characterization and Toxicity Confirmation of LukM/ F0-PV Producing Staphylococcus aureus Isolated from Bovine Mastitis Samples in Mysore, India.

The widespread status of subclinical condition of
bovine mastitis is often associated with the production of leukotoxin
M/F0-PV producing Staphylococcus aureus. The
present study aims for the profiling of such leukotoxin producers
through conventional and molecularmethods 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 isolateswas 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.