Metagenomic analysis of dairy bacteriophages: extraction method and pilot study on whey samples derived from using undefined and defined mesophilic starter cultures

Despite their huge potential for characterizing the biodiversity of phages, metagenomic studies are currently not available for dairy bacteriophages, partly due to the lack of a standard procedure for phage extraction. We optimized an extraction method that allows to remove the bulk protein from whey and milk samples with losses of less than 50% of spiked phages. The protocol was applied to extract phages from whey in order to test the notion that members of Lactococcus (Lc.) lactis 936 (now Sk1virus ), P335, c2 (now C2virus ) and Leuconostoc phage groups are the most frequently encountered in the dairy environment. The relative abundance and diversity of phages in eight and four whey mixtures from dairies using undefined mesophilic mixed-strain DL-starter cultures and defined cultures, respectively, was assessed. Results obtained from transmission electron microscopy and high-throughput sequence analyses revealed the dominance of Lc. lactis 936 phages (order Caudovirales, family Siphoviridae ) in dairies using undefined DL-starter cultures and Lc. lactis c2 phages (order Caudovirales, family Siphoviridae ) in dairies using defined cultures. The 936 and Leuconostoc phages demonstrated limited diversity. Possible co-induction of temperate P335 prophages and satellite phages in one of the whey mixtures was also observed.