Isolation and characterization of new filamentous phages active on Gram-positive bacteria in Swiss-type cheese

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Английский. In 1998, our team isolated one of the rare filamentous phages active and culturable on Gram-positive Bacteria. The phage infected Propionibacterium freudenreichii and was isolated from Swiss-type cheese. It is a single-stranded DNA phage, that was named B5. Until now, no related phage was found and little is known about B5-like phages diversity.The aim of this research is to isolate new filamentous phages from Swiss-type cheese to study their diversity and discuss, by comparison with phage B5, a possible evolution in recent years.This study was carried out using 30 pasteurized and raw milk cheese samples from different manufactures. After enrichment, each cheese sample was inoculated with mid-log phase culture of P.freudenreichii. Then, the phages plaques were detected using the soft agar layer method. The phages were subsequently isolated and studied using host range assay, transmission electron microscopy, S1 Nuclease hydrolysis, SDS page, and NGS sequencing.Seven filamentous phages that infect P.freudenreichii were isolated from raw milk cheese. Genome sequencing analysis confirmed that these phages were distinct but related. They indeed displayed a similar genome structure, and sequence identity ranged from a hundred mutations to a 9% identity difference. Did all the newly isolated phages evolve from B5? At which step of cheese making do they occur? The diversity of these filamentous phages isolated from cheeses of different geographical origins shows that B5-related phages have been able to evolve over the last few years. More, the fact that nearly ten filamentous phages were found in relatively few cheeses suggests a large diversity.To our knowledge, these 7 phages are the first filamentous phages active on Gram-positive bacteria found since the first in 1998. It would be interesting to continue this study by more precisely the newly isolated phages, by scanning a larger number of cheeses, and by investigating the origin of these filamentous phages.