Culturomics of the pig tonsil microbiome identifies new species and an untapped source of novel antimicrobials

Background: In humans and pigs, altered composition of the microbiota associated with the epithelium of the palatine tonsils has been associated with bacterial or viral infection and lymphoid tissue inflammation. Tonsil lymphoid tissue is important for immunity and considered an important portal of entry for pathogens such as Streptococcus suis. Little is known about correlations between tonsil-associated microbiota, tonsillar infections, and the species that might confer colonization resistance against pathogens. Here, we describe a large collection of representative bacterial species from the tonsil surface biofilm and used genome mining and in vitro assays to assess their potential as probiotics to reduce infections by S. suis and other pathogens. Results: Data on tonsil microbiota composition from over 100 piglets from 11 farms and 3 countries revealed a core microbiota comprising Actinobacillus, Streptococcus, and Moraxella and 11 other less abundant but prevalent genera. To establish a collection of culturable core species, we plated 5 tonsil swabs taken from healthy piglets on different farms and countries on 8 different media and isolated 518 pure cultures belonging to 23 genera. To identify candidate probiotic strains, we tested for antagonistic activity against a panel of pathogens and in silico genome mining to find biosynthetic gene clusters (BGCs) in isolates that might produce antimicrobial compounds. We identified two novel species with potential probiotic activities: a Brevibacterium species and Corynebacterium species producing a heat and proteolytically stable lanthipeptide variant of flavucin, inhibiting in vitro growth of the opportunistic pathogens S. suis and Staphylococcus aureus. Conclusions: We defined the core tonsil microbiota of piglets and cultured representative single bacterial isolates for research on microbiota–host interactions in the oral cavity. Several isolates inhibiting the growth of bacterial pathogens that might be exploited as probiotics to promote colonization resistance were deposited in publicly available strain repositories.