2008 A P P L I E D A N D N A T U R A L S C I E N C E F O U N D A T I O N   Journal of Applied and Natural Science 6 (2): 356-361 (2014) A study on Methicillin resistant Staphylococcus aureus mastitis in dairy cows D. Chandrasekaran 1* , P. Venkatesan 2 , K. G. Tirumurugaan 2 , A. P. Nambi 1 , P. S. Thirunavukkarasu 1 , K. Kumanan 2 and S.Vairamuthu 3 1 Department of Veterinary Clinical Medicine, Ethics and Jurisprudence, Madras Veterinary College, Tamil Nadu Veterinary and Animal Sciences University, Chennai, INDIA 2 Department of Animal Biotechnology, Madras Veterinary College, Tamil Nadu Veterinary and Animal Sciences University, Chennai, INDIA 3 Centralized Clinical Laboratory, Madras Veterinary College, Tamil Nadu Veterinary and Animal Sciences University, Chennai, INDIA *Corresponding author. E-mail: drchandrus@yahoo.com Received: June 29, 2014; Revised received: August 22, 2014 ; Accepted: September 18, 2014 Abstract: Methicillin-resistant Staphylococcus aureus (MRSA) poses a serious problem in dairy animals suffering from mastitis. The study was carried out to evaluate the incidence of Methicillin resistant S. aureus from clinical mastitis milk samples and their antibiotic resistance profile and characterised with respect to the molecular features that contributed to the resistance in these pathogens. Isolation and identification of Methicillin resistant S. aureus were performed from acute clinical mastitis samples. The isolates were tested using agar disc diffusion method for their antimicrobial susceptibility and modified resazurin assay micro dilution technique for MIC to 8 different antimicrobial drugs. A total of 235 clinical mastitis milk samples from dairy cows were cultured for incidence of S. aureus. Methicillin resistant S. aureus was isolated from a total of 12 (44.25%) of the 116 S. aureus samples. Based on the antimicrobial sensitivity and MIC results, MRSA isolates were found sensitive to gentamicin, enrofloxcain, amoxicillin+sulbactam, ceftriaxone and resistant to amoxicillin, oxytetracycline, penicillin G and oxacillin. Most of MRSA isolates were found to be multi-drug resistant. MRSA alert kit test and mecA and blaZ target gene PCR were found to be useful in the confirmation of MRSA. Keywords: Antibiotic susceptibility, Antimicrobial drug resistance, Bovine mastitis, MIC, mecA, MRSA, PCR blaZ, PVL gene INTRODUCTION Bovine mastitis, a multi-factorial disease, is characterized by physical, chemical and microbiological changes in the milk and pathological changes in the glandular tissue of udder (Radostits et al., 2008). Among the various pathogens that affect the bovine mammary gland Staphylococcus aureus is the most frequently (45-60 per cent) isolated pathogens and often results clinical or subclinical or chronic bovine mastitis. S. aureus is capable of producing several exo and endotoxins e.g coagulase and is also endowed with a capsule that makes it difficult to treat. In the control of mastitis, the improper use of antimicrobial agents on dairy farm animals is a major concern as it leads to the emergence of resistant zoonotic bacterial pathogens (Piddock, 1996). The antibiotic-resistance of prevalent S.aureus strains is another serious concern besides the pathogenicity. The emergence of antibiotic-resistance in S. aureus from mastitic dairy animals has been shown in recent years. Beta–lactam antibiotics are frequently used in mastitis therapy and the resistance is due to the production of ISSN : 0974-9411 (Print), 2231-5209 (Online) All Rights Reserved © Applied and Natural Science Foundation www.ansfoundation.org beta-lactamases and low –affinity penicillin-binding protein, PBP2A (Olsen et al., 2006) Several strains of S.aureus isolated from mastitis case have been reported to show resistance against multiple antimicrobials such as penicillin-G, gentamicin, streptomycin, ampicillin, ciprofloxacin, oxytetracycline (Kumar et al., 2011). β-lactamase resistant penicillins such as methicillin and oxacillin are not used in dairy cows except for cloxacillin that is used in products for intramammary administration (Turutoglu et al., 2006). Among the resistance to commonly used antibiotics, resistance to this bacterium, methicillin (hence termed methicillin resistance S. aureus- MRSA) is a serious cause of concern in both human and animals (Witte et al., 2007). Bovine and human MRSA strains are epidemiologically related, which indicates transmission from either cow to human or human to cow possible (Juhász- Kaszanyitzky et al., 2007). This strain is negative for the Panton Valentine Leucocidin (PVL) genes, which differentiates it from community associated MRSA ST 1, which is positive for PVL genes (Vandenesch et al., 2003). PVL genes have been associated with community-acquired MRSA (CA-