Antilisterial activity of raw sheep milk from two native Epirus breeds: Culture-dependent identification, bacteriocin gene detection and primary safety evaluation of the antagonistic LAB biota.

Raw milk from native small ruminant breeds in Epirus, Greece, is a valuable natural source of autochthonous lactic acid bacteria (LAB) strains with superior biotechnological properties. In this study, two bulk milks (RM1, RM2) from two local sheep yards, intended for traditional Kefalotyri cheese production, were preselected for bacteriocin-like antilisterial activity by in vitro tests. Their antagonistic LAB biota was quantified followed by polyphasic (16S rRNA gene sequencing; IGS for ; a multiplex-PCR for ) identification of 42 LAB (RM1/18; RM2/24) isolates further evaluated for bacteriocin encoding genes and primary safety traits. Representative isolates of the numerically dominant mesophilic LAB were (10) in both RMs, (7) in RM2, and (1) in RM1; the subdominant thermophilic LAB isolates were (8), (6), (3), (1), (1), (2), (1) and (1). Based on their and profiles, six strains (8 isolates) were atypical lying between the subspecies and whereas two strains profiled with subsp. that is first-time reported in Greek dairy food. Two RM1 strain biotypes (3 isolates) showed strong, enterocin-mediated antilisterial activity due to possession. One from RM1 possessed and , while additional nine RM2 isolates of the group processed or singly. All showed direct (cell-associated) antilisterial activity only, as also both strains from RM2 did strongly. Desirably, no LAB isolate was β-hemolyrtic, or cytolysin-positive, or possessed for vancomycin resistance, or and virulence genes. However, all three from RM2 possessed and/or virulence genes. In conclusion, all strains, the two safe, enterocin A-B-P-producing strains, and the two antilisterial strains should be validated further as potential costarter or adjunct cultures in Kefalotyri cheese. The prevalence of α-hemolytic pyogenic streptococci in raw milk, mainly in RM2, requires consideration in respect to subclinical mastitis in sheep and the farm hygiene overall.