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Bacterial-fungal interactions revealed by genome-wide analysis of bacterial mutant fitness.

ABSTRACT: Microbial interactions are expected to be major determinants of microbiome structure and function. Although fungi are found in diverse microbiomes, their interactions with bacteria remain largely uncharacterized. In this work, we characterize interactions in 16 different bacterial-fungal pairs, examining the impacts of 8 different fungi isolated from cheese rind microbiomes on 2 bacteria (Escherichia coli and a cheese-isolated Pseudomonas psychrophila). Using random barcode transposon-site sequencing with an analysis pipeline that allows statistical comparisons between different conditions, we observed that fungal partners caused widespread changes in the fitness of bacterial mutants compared to growth alone. We found that all fungal species modulated the availability of iron and biotin to bacterial species, which suggests that these may be conserved drivers of bacterial-fungal interactions. Species-specific interactions were also uncovered, a subset of which suggested fungal antibiotic production. Changes in both conserved and species-specific interactions resulted from the deletion of a global regulator of fungal specialized metabolite production. This work highlights the potential for broad impacts of fungi on bacterial species within microbiomes.

SUBMITTER: Pierce EC

PROVIDER: S-EPMC8515420 | biostudies-literature | 2021 Jan

REPOSITORIES: biostudies-literature

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Bacterial-fungal interactions revealed by genome-wide analysis of bacterial mutant fitness.

Pierce Emily C EC Morin Manon M Little Jessica C JC Liu Roland B RB Tannous Joanna J Keller Nancy P NP Pogliano Kit K Wolfe Benjamin E BE Sanchez Laura M LM Dutton Rachel J RJ

Nature microbiology 20201102 1

Microbial interactions are expected to be major determinants of microbiome structure and function. Although fungi are found in diverse microbiomes, their interactions with bacteria remain largely uncharacterized. In this work, we characterize interactions in 16 different bacterial-fungal pairs, examining the impacts of 8 different fungi isolated from cheese rind microbiomes on 2 bacteria (Escherichia coli and a cheese-isolated Pseudomonas psychrophila). Using random barcode transposon-site seque ...[more]

PMID: 33139882

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Project description:Microbial interactions abound in natural ecosystems and shape community structure and function. Substantial attention has been given to cataloging mechanisms by which microbes interact, but there is a limited understanding of the genetic landscapes that promote or hinder microbial interactions. We previously developed a mutualistic coculture pairing Escherichia coli and Rhodopseudomonas palustris, wherein E. coli provides carbon to R. palustris in the form of glucose fermentation products and R. palustris fixes N2 gas and provides nitrogen to E. coli in the form of NH4+ The stable coexistence and reproducible trends exhibited by this coculture make it ideal for interrogating the genetic underpinnings of a cross-feeding mutualism. Here, we used random barcode transposon sequencing (RB-TnSeq) to conduct a genome-wide search for E. coli genes that influence fitness during cooperative growth with R. palustris RB-TnSeq revealed hundreds of genes that increased or decreased E. coli fitness in a mutualism-dependent manner. Some identified genes were involved in nitrogen sensing and assimilation, as expected given the coculture design. The other identified genes were involved in diverse cellular processes, including energy production and cell wall and membrane biogenesis. In addition, we discovered unexpected purine cross-feeding from R. palustris to E. coli, with coculture rescuing growth of an E. coli purine auxotroph. Our data provide insight into the genes and gene networks that can influence a cross-feeding mutualism and underscore that microbial interactions are not necessarily predictable a prioriIMPORTANCE Microbial communities impact life on Earth in profound ways, including driving global nutrient cycles and influencing human health and disease. These community functions depend on the interactions that resident microbes have with the environment and each other. Thus, identifying genes that influence these interactions will aid the management of natural communities and the use of microbial consortia as biotechnology. Here, we identified genes that influenced Escherichia coli fitness during cooperative growth with a mutualistic partner, Rhodopseudomonas palustris Although this mutualism centers on the bidirectional exchange of essential carbon and nitrogen, E. coli fitness was positively and negatively affected by genes involved in diverse cellular processes. Furthermore, we discovered an unexpected purine cross-feeding interaction. These results contribute knowledge on the genetic foundation of a microbial cross-feeding interaction and highlight that unanticipated interactions can occur even within engineered microbial communities.

Project description:Rapid growth in deep tissue is essential to the high virulence of Yersinia pestis, causative agent of plague. To better understand the mechanisms underlying this unusual ability, we used transposon mutagenesis and high-throughput sequencing (Tn-seq) to systematically probe the Y. pestis genome for elements contributing to fitness during infection. More than a million independent insertion mutants representing nearly 200,000 unique genotypes were generated in fully virulent Y. pestis. Each mutant in the library was assayed for its ability to proliferate in vitro on rich medium and in mice following intravenous injection. Virtually all genes previously established to contribute to virulence following intravenous infection showed significant fitness defects, with the exception of genes for yersiniabactin biosynthesis, which were masked by strong intercellular complementation effects. We also identified more than 30 genes with roles in nutrient acquisition and metabolism as experiencing strong selection during infection. Many of these genes had not previously been implicated in Y. pestis virulence. We further examined the fitness defects of strains carrying mutations in two such genes-encoding a branched-chain amino acid importer (brnQ) and a glucose importer (ptsG)-both in vivo and in a novel defined synthetic growth medium with nutrient concentrations matching those in serum. Our findings suggest that diverse nutrient limitations in deep tissue play a more important role in controlling bacterial infection than has heretofore been appreciated. Because much is known about Y. pestis pathogenesis, this study also serves as a test case that assesses the ability of Tn-seq to detect virulence genes. Our understanding of the functions required by bacteria to grow in deep tissues is limited, in part because most growth studies of pathogenic bacteria are conducted on laboratory media that do not reflect conditions prevailing in infected animal tissues. Improving our knowledge of this aspect of bacterial biology is important as a potential pathway to the development of novel therapeutics. Yersinia pestis, the plague bacterium, is highly virulent due to its rapid dissemination and growth in deep tissues, making it a good model for discovering bacterial adaptations that promote rapid growth during infection. Using Tn-seq, a genome-wide fitness profiling technique, we identified several functions required for fitness of Y. pestis in vivo that were not previously known to be important. Most of these functions are needed to acquire or synthesize nutrients. Interference with these critical nutrient acquisition pathways may be an effective strategy for designing novel antibiotics and vaccines.

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Bacterial-fungal interactions revealed by genome-wide analysis of bacterial mutant fitness.

Publications

Bacterial-fungal interactions revealed by genome-wide analysis of bacterial mutant fitness.

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