ABSTRACT
Staphylococcus aureus is the major contagious bovine mastitis pathogen and has no effective vaccine. Strain variation and limited knowledge of common immunogenic antigen/s are among major constraints for developing effective vaccines. S. aureus cell surface proteins that are exposed to the host immune system constitute good vaccine candidates. The objective of this study was to compare two novel S. aureus surface protein extraction methods with biotinylation method and evaluate immune-reactivity of extracted proteins. Surface proteins were extracted from nine genetically distinct S. aureus strains from cases of bovine mastitis. After extraction, bacterial cell integrity was examined by Gram staining and electron microscopy to determine if extraction methods caused damage to cells that may release non-surface proteins. The extracted proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and evaluated for immune-reactivity using western blot. Results showed that all three extraction methods provided multiple protein bands on SDS-PAGE. Western blot result showed several immunoreactive surface proteins, in which some proteins strongly (well-resolved, thick, dark, and intense band) reacted across the nine strains tested. The three methods are valid for the extraction of surface proteins and hexadecane, and cholic acid methods are more feasible than biotinylation since both are easier, cheaper, and have minor effects on the bacterial cell. Strongly immune-reactive surface proteins may serve as potential candidates for a vaccine to control S. aureus mastitis in dairy cows.
ABSTRACT
Staphylococcus aureus is a frequent and major contagious mastitis bacterial pathogen. The antibiotic treatment cure rates vary considerably from 4% to 92%. Staphylococcus aureus readily becomes resistant to antibiotics, resulting in persistent noncurable intramammary infection that usually results in culling of infected animals. Because of its notorious ability to acquire resistance to the commonly used as well as last resort antimicrobials such as methicillin and vancomycin and the development of multidrug-resistant strains, antimicrobial resistance (AMR) in S. aureus is of paramount importance in human medicine. The objective of this study was to evaluate the prevalence of AMR and genetic diversity of S. aureus isolates from milk of dairy cattle. Staphylococcus aureus isolates (n = 239) from 33 dairy farms in Tennessee were tested against 10 antimicrobials by broth microdilution method using the Sensititer system. Genetic diversity of resistant isolates was evaluated by pulsed-field gel electrophoresis (PFGE). Overall, AMR of the S. aureus isolates varied from as low as 1.3% for ceftiofur to as high as 25% for sulfadimethoxine. Out of 239 S. aureus isolates, 82 (34.3%) of them were resistant to at least 1 of the 10 antimicrobials. The AMR isolates belonged to two major PFGE types, indicating the presence of dominant clonal patterns among the resistant isolates. In general, there was a variation of prevalence of AMR within and among farms over time, with an increasing trend in tetracycline resistance. Judicious use of antimicrobials in dairy cattle farms can reduce the development of antimicrobial-resistant S. aureus.
