Microbiome Assembly in Fermented Foods.

For thousands of years, humans have enjoyed the novel flavors, increased shelf-life, and nutritional benefits that microbes provide in fermented foods and beverages. Recent sequencing surveys of ferments have mapped patterns of microbial diversity across space, time, and production practices. But a mechanistic understanding of how fermented food microbiomes assemble has only recently begun to emerge. Using three foods as case studies (surface-ripened cheese, sourdough starters, and fermented vegetables), we use an ecological and evolutionary framework to identify how microbial communities assemble in ferments. By combining in situ sequencing surveys with in vitro models, we are beginning to understand how dispersal, selection, diversification, and drift generate the diversity of fermented food communities. Most food producers are unaware of the ecological processes occurring in their production environments, but the theory and models of ecology and evolution can provide new approaches for managing fermented food microbiomes, from farm to ferment.

Genomic, transcriptomic, and ecological diversity of Penicillium species in cheese rind microbiomes.

Although Penicillium molds can have significant impacts on agricultural, industrial, and biomedical systems, the ecological roles of Penicillium species in many microbiomes are not well characterized. Here we utilized a collection of 35 Penicillium strains isolated from cheese rinds to broadly investigate the genomic potential for secondary metabolism in cheese-associated Penicillium species, the impact of Penicillium on bacterial community assembly, and mechanisms of Penicillium-bacteria interactions. Using antiSMASH, we identified 1558 biosynthetic gene clusters, 406 of which were mapped to known pathways, including several mycotoxins and antimicrobial compounds. By measuring bacterial abundance and fungal mRNA expression when culturing representative Penicillium strains with a cheese rind bacterial community, we observed divergent impacts of different Penicillium strains, from strong inhibitors of bacterial growth to those with no impact on bacterial growth or community composition. Through differential mRNA expression analyses, Penicillium strains demonstrated limited differential gene expression in response to the bacterial community. We identified a few shared responses between the eight tested Penicillium strains, primarily upregulation of nutrient metabolic pathways, but we did not identify a conserved fungal response to growth in a multispecies community. These results in tandem suggest high variation among cheese-associated Penicillium species in their ability to shape bacterial community development and highlight important ecological diversity within this iconic genus.

Penicillium molds impact the transcriptome and evolution of the cheese bacterium Staphylococcus equorum.

The observation that Penicillium molds can inhibit the growth of Staphylococcus was a catalyst for the antibiotic revolution. Considerable attention has been paid to purified Penicillium metabolites that inhibit bacteria, but little is known about how Penicillium species impact the ecology and evolution of bacteria in multispecies microbial communities. Here, we investigated how four different species of Penicillium can impact global transcription and evolution of a widespread Staphylococcus species (S. equorum) using the cheese rind model microbiome. Through RNA sequencing, we identified a core transcriptional response of S. equorum against all five tested Penicillium strains, including upregulation of thiamine biosynthesis, fatty acid degradation, and amino acid metabolism as well as downregulation of genes involved in the transport of siderophores. In a 12-week evolution experiment where we co-cultured S. equorum with the same Penicillium strains, we observed surprisingly few non-synonymous mutations across S. equorum populations evolved with the Penicillium species. A mutation in a putative DHH family phosphoesterase gene only occurred in populations evolved without Penicillium and decreased the fitness of S. equorum when co-cultured with an antagonistic Penicillium strain. Our results highlight the potential for conserved mechanisms of Staphylococcus-Penicillium interactions and demonstrate how fungal biotic environments may constrain the evolution of bacterial species.IMPORTANCEFungi and bacteria are commonly found co-occurring both in natural and synthetic microbiomes, but our understanding of fungal-bacterial interactions is limited to a handful of species. Conserved mechanisms of interactions and evolutionary consequences of fungal-bacterial interactions are largely unknown. Our RNA sequencing and experimental evolution data with Penicillium species and the bacterium S. equorum demonstrate that divergent fungal species can elicit conserved transcriptional and genomic responses in co-occurring bacteria. Penicillium molds are integral to the discovery of novel antibiotics and production of certain foods. By understanding how Penicillium species affect bacteria, our work can further efforts to design and manage Penicillium-dominated microbial communities in industry and food production.