The aim of this study was to investigate the presence of tetracycline resistance in lactobacilli isolated from traditional Serbian white brined raw milk cheeses (Homolje, Sjenica, Zlatar). Isolation of presumptive lactobacilli was initially performed using MRS-S agar without tetracycline, or supplemented with 16 and 64 µg/mL of tetracycline. Rep-PCR (GTG)5 genotyping showed a high diversity of the isolates obtained, as examination of 233 isolates resulted in 156 different Rep-PCR fingerprints. Ninety out of 156 (57.69%) of the strains, representatives with different (GTG)5 fingerprints, were identified by MALDI-TOF MS as lactobacilli, while 66 out of 156 (42.31%) strains were identified as members of other LAB genera. All except one out of 90 Lactobacillus isolates further tested by microdilution method, demonstrated unimodal distribution of tetracycline MIC values which were equal to or lower from the breakpoint MIC values (EFSA in EFSA J 10: 1-10, 2012. 10.2903/j.efsa.2012.2740). Only one Lb. paracasei isolate showed the presence of tet(M) gene, while the other analyzed tet genes [tet(A), tet(B), tet(C) tet(K), tet(L), tet(O) and tet(W)] were not detected in any of the isolates. The results of this study indicates that lactobacilli from traditional Serbian raw milk cheeses do not present considerable tetracycline resistance reservoirs. For final conclusions about the safety of these autochthonous cheeses regarding the possible tetracycline resistance transferability, the assessment of the entire cheese microbiota is needed.
Keywords: Antibiotic resistance; Lactobacilli; MALDI TOF mass spectrometry; Raw milk cheese; Rep-PCR; Tetracycline.