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Despite being potentially highly useful 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 the removal of 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 ofLactococcus lactis 936 (nowSk1virus ), P335, c2 (nowC2virus ) andLeuconostoc 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 cultures containingLactococcus lactis subsp.lactis biovar diacetylactis andLeuconostoc species (i.e., DL starter cultures) and defined cultures, respectively, were assessed. Results obtained from transmission electron microscopy and high-throughput sequence analyses revealed the dominance ofLc. lactis 936 phages (orderCaudovirales , familySiphoviridae ) in dairies using undefined DL starter cultures andLc. lactis c2 phages (orderCaudovirales , familySiphoviridae ) in dairies using defined cultures. The 936 andLeuconostoc phages demonstrated limited diversity. Possible coinduction of temperate P335 prophages and satellite phages in one of the whey mixtures was also observed.IMPORTANCE The method optimized in this study could provide an important basis for understanding the dynamics of the phage community (abundance, development, diversity, evolution, etc.) in dairies with different sizes, locations, and production strategies. It may also enable the discovery of previously unknown phages, which is crucial for the development of rapid molecular biology-based methods for phage burden surveillance systems. The dominance of only a few phage groups in the dairy environment signifies the depth of knowledge gained over the past decades, which served as the basis for designing current phage control strategies. The presence of a correlation between phages and the type of starter cultures being used in dairies might help to improve the selection and/or design of suitable, custom, and cost-efficient phage control strategies.

applied and environmental microbiology

Metagenomic Analysis of Dairy Bacteriophages: Extraction Method and Pilot Study on Whey Samples Derived from Using Undefined and Defined Mesophilic Starter Cultures