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Simple Summary  Mastitis is the most prevalent disease of dairy cattle that causes significant economic losses. Different agents cause mastitis which leads to increased somatic cell count (SCC) and low milk quality. Treating mastitis with antimicrobials is essential to reduce SCC and improve milk quality. Excessive use or misuse of antimicrobials in dairy farms leads to the development of antimicrobial resistant bacteria. The objectives of this study were (1) to isolate and identify the causative agent of mastitis and (2) determine antimicrobial resistance profiles of bacterial isolates. A total of 174 quarter milk samples from 151 cows with high SCC and clinical mastitis from 34 dairy farms in Tennessee, Kentucky, and Mississippi were collected. Bacterial causative agents were determined by bacteriological and biochemical tests. Antimicrobial resistance of bacterial isolates against 10 commonly used antimicrobials was tested. A total of 193 bacteria consisting of six bacterial species, which include  Staphylococcus aureus ,  Streptococcus uberis ,  Streptococcus dysgalactiae ,  Escherichia coli ,  Klebsiella oxytoca  and  Klebsiella pneumoniae  were isolated.  Staphylococcus aureus  was the predominant isolate. The proportion of resistant isolates was relatively higher in Gram-negatives than Gram-positives. Continuous antimicrobial resistance testing and identification of reservoirs of resistance traits in dairy farms are essential to implement proper mitigation measures.    Abstract  Mastitis is the most prevalent and economically important disease caused by different etiological agents, which leads to increased somatic cell count (SCC) and low milk quality. Treating mastitis cases with antimicrobials is essential to reduce SCC and improve milk quality. Non-prudent use of antimicrobials in dairy farms increased the development of antimicrobial resistant bacteria. This study’s objectives were (1) to isolate and identify etiological agents of mastitis and (2) to determine antimicrobial resistance profiles of bacterial isolates. A total of 174 quarter milk samples from 151 cows with high SCC and clinical mastitis from 34 dairy farms in Tennessee, Kentucky, and Mississippi were collected. Bacterial causative agents were determined by bacteriological and biochemical tests. The antimicrobial resistance of bacterial isolates against 10 commonly used antimicrobials was tested. A total of 193 bacteria consisting of six bacterial species, which include  Staphylococcus aureus ,  Streptococcus uberis ,  Streptococcus dysgalactiae ,  Escherichia coli ,  Klebsiella oxytoca  and  Klebsiella pneumoniae  were isolated.  Staphylococcus aureus  was the predominant isolate followed by  Strep.  spp.,  E. coli , and  Klebsiella  spp. Results of this study showed that Gram-negatives ( E. coli  and  Klebsiella  spp.) were more resistant than Gram-positives ( Staph. aureus  and  Streptococcus  spp.). Continuous antimicrobial resistance testing and identification of reservoirs of resistance traits in dairy farms are essential to implement proper mitigation measures.

Antimicrobial Resistance of Major Bacterial Pathogens from Dairy Cows with High Somatic Cell Count and Clinical Mastitis