Methicillin resistance and selective genetic determinants of Staphylococcus aureus isolates with bovine mastitis milk origin

Abstract

Background and objectives: Staphylococcus aureus is one of the major causes of bovine mastitis, which can be transmitted from animals to humans. Methicillin-resistant S. aureus (MRSA) isolates are more attentive and if not treated promptly, they can cause death. The aim of this study was to determine the prevalence of methicillin resistance and frequency of selected virulence factors of S. aureus isolates with bovine mastitis milk origin in Ahvaz, southwest of Iran.

Materials and methods: During a two-year period (2014-2015), 75 S. aureus isolates were recovered from referred clinical and sub-clinical bovine mastitis milk samples. The isolates were phenotypically investigated for resistance to cefoxitin by Kirby-Bauer method. DNA were analyzed by PCR for mecA and selected genes that encode the virulence factors.

Results: According to the results, the spa, ebpS, fnb, bbp, clfA, clfB, and cna genes were detected in 98.7, 97.3, 97.3, 86.7, 84, 84 and 65.3% of the isolates, respectively. Among the 75 isolates, only one (1.3%) isolate was methicillin-resistant. Totally, 39 isolates (50.7%) had all of these virulence factors except mecA. The results showed that 96% of the isolates had at least the fnb, ebpS and spa genes, signifying the noteworthy role of these genes in the pathogenesis of S. aureus bovine intra-mammary infection in this area.

Conclusion: In the present study, the prevalence of mecA was relatively low, possibly indicating that cows do not play a significant role in community-acquired MRSA infection in this area. According to the results, studied virulence factors were somewhat prevalent, bearing in mind the probable risk of transmission of these isolates from cows to humans, especially those that are in close contact with infected cattle. The data presented here can be used for the introduction of a protective vaccine against this infection.

Keywords: Bovine mastitis; Methicillin resistance; Staphylococcus aureus; Virulence factor.

Fig. 1.

Agarose gel electrophoresis of the PCR products for amplification of aroA in S. aureus suspected isolates, Lanes 1 and 3: negative and positive controls, respectively, lanes 4–6, aroA positive isolates and lane 2, 100bp ladder

Fig. 2.

Agarose gel electrophoresis of the PCR products for amplification of mecA in suspected MRSA Lane 1: positive control S. aureus (ATCC 33591), lane 2: MRSA isolate, lanes 3, 4 negative mecA isolates, lane 6: negative control and lane 5, 100bp ladder.

Fig. 3.

Agarose gel electrophoresis of the PCR products for detection of clfA, clfB, cna and spa gene of some S. aureus isolates, Lanes 1 and 3 negative and positive controls for clfA and clfB genes, lanes 4 and 5, clfA⁺ clfB⁺ isolates, lanes 6 and 8 negative and positive controls for cna and spa genes, lane 9 and 10, cna⁺ spa⁺ and spa⁺ isolates respectively, lanes 2 and 7, 100bp ladder.

Fig. 4.

Agarose gel electrophoresis of the PCR products for detection of fnb, bbp and ebpS gene of some S. aureus isolates, Lanes 1 and 3 negative and positive controls for fnb and bbp genes, lanes 4 and 5, fnb⁺ bbp⁺ isolates, lanes 6 and 8 negative and positive controls for ebpS gene, lane 9 and 10, ebpS⁺ and ebpS⁻ isolates, respectively. lanes 2 and 7, 100bp ladder.