Tetracycline resistance phenotypes and genotypes of coagulase-negative staphylococcal isolates from bubaline mastitis in Egypt.

Aim: This study was devoted to elucidate the tetracycline resistance of coagulase-negative staphylococci (CNS) derived from normal and subclinical mastitic (SCM) buffaloes' milk in Egypt.

Materials And Methods: A total of 81 milk samples from 46 normal buffalo milk samples and 35 SCM buffalo milk samples at private dairy farms of Egypt were used in this study. CNS were identified using phenotypic and molecular methods (polymerase chain reaction [PCR]). CNS isolates were tested for tetracycline resistance using routine methods and multiplex PCR targeting tetracycline () resistance genes followed by sequencing of positive PCR products and phylogenetic analysis.

Results: Isolation and identification of 28 (34.5%) CNS from normal and SCM buffaloes' milk, namely, (39.2%), (25.0%), (10.7%), (10.7%), and 3.5% to each of , , , and . Using nested PCR, all the 28 CNS isolates revealed positive for 16srRNA gene specific for genus staphylococci and negative for thermonuclease () gene specific for species. The presence of tetracycline resistance-encoding genes (K, L, M, and O) was detected by multiplex PCR. All isolates were negative for L, M, and O genes while 14 (50%) CNS isolates were positive for K gene, namely, (100%), (100%), (66.6%), (45.4%), and (42.8%). Nucleotide sequencing of K gene followed by phylogenetic analysis showed the high homology between our CNS isolates genes of tetracycline resistance with isolates including Egyptian ones. This proves the transfer of the tetracycline resistance encoding genes between coagulase-negative and coagulase positive spp.

Conclusion: CNS isolates have distinguishingly high resistance to tetracycline. Abundant tetracycline usage for mastitis treatment leads to the spread of genetic resistance mechanisms inside CNS strains and among all spp. Consequently, tetracycline is not effective anymore.