A total of 765 samples were collected from beef carcasses, knives, cutting table surfaces, beef, hands, air, and water from four cattle slaughterhouses of Kerala, South India, to determine the occurrence and antibiotic susceptibility of Listeria species and Staphylococcus aureus. Listeria spp. were isolated from beef carcasses (2.0%), knives (3.7%), cutting table surfaces (1.9%), beef (0.7%), and water (1.3%). The identified species were Listeria monocytogenes (0.1%), Listeria innocua (0.9%), and Listeria ivanovii (0.4%). Most of the Listeria spp. were susceptible to majority of the antibiotics tested. The virulence genes were not detected in Listeria spp. However, all the L. innocua isolates were found to harbor the iap gene. The overall occurrence of S. aureus in slaughterhouses was 50.8%. The highest occurrence was observed on hands of abattoir workers (79.6%) and beef carcasses (59.9%). The isolates were commonly resistant to penicillin (38.0%), followed by ceftriaxone (31.9%), ampicillin (29.0%), amoxicillin (28.8%), tetracycline (24.4%), and chloramphenicol (23.9%). Overall, 53.0% of S. aureus isolates were resistant to three or more antibiotics. Vancomycin and methicillin resistance were observed in 8.5% and 5.4% of S. aureus isolates, respectively. Eight methicillin-resistant S. aureus isolates were found to harbor the mecA gene. In conclusion, Listeria spp. was only rarely found in the slaughterhouse environment and on beef. Nevertheless, the recovery of L. monocytogenes from a water reservoir containing sea water that was used to wash carcasses indicates the potential risk of contamination of the carcasses with L. monocytogenes when using sea water. S. aureus was frequently isolated from abattoir workers and beef carcasses, and the occurrence of S. aureus differed significantly between slaughterhouses. The high occurrence of S. aureus, which were often resistant toward different antibiotics, represents a significant public health concern.
