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ABSTRACT Omnivorous animals, including humans, harbor diverse, species-rich gut communities that impact their growth, development, and homeostasis. Model invertebrates are broadly accessible experimental platforms that enable linking specific species or species groups to host phenotypes, yet often their specialized diets and distinct gut microbiota make them less comparable to human and other mammalian and gut communities. The omnivorous cockroach Periplaneta americana harbors ∼4 × 102 bacterial genera within its digestive tract and is enriched with taxa commonly found in omnivorous mammals (i.e., Proteobacteria, Bacteroidetes, and Firmicutes). These features make P. americana a valuable platform for identifying microbe-mediated host phenotypes with potential translations to mammals. Rearing P. americana insects under germfree conditions resulted in prolonging development time by ∼30% and an up to ∼8% reduction in body size along three dimensions. Germfree rearing resulted in downregulation of gene networks involved in growth, energy homeostasis, and nutrient availability. Reintroduction of a defined microbiota comprised of a subset of P. americana commensals to germfree insects did not recover normal growth and developmental phenotypes or transcriptional profiles observed in conventionally reared insects. These results are in contrast with specialist-feeding model insects (e.g., Drosophila), where introduction of a single endemic bacterial species to germfree condition-reared specimens recovered normal host phenotypes. These data suggest that understanding microbe-mediated host outcomes in animals with species-rich communities should include models that typically maintain similarly diverse microbiomes. The dramatic transcriptional, developmental, and morphological phenotypes linked to gut microbiome status in this study illustrates how microbes are key players in animal growth and evolution. IMPORTANCE Broadly accessible model organisms are essential for illustrating how microbes are engaged in the growth, development, and evolution of animals. We report that germfree rearing of omnivorous Periplaneta americana cockroaches resulted in growth defects and severely disrupted gene networks that regulate development, which highlights the importance of gut microbiota in these host processes. Absence of gut microbiota elicited a starvation-like transcriptional response in which growth and development were inhibited while nutrient scavenging was enhanced. Additionally, reintroduction of a subset of cockroach gut bacterial commensals did not broadly recover normal expression patterns, illustrating that a particular microbiome composition may be necessary for normal host development. Invertebrate microbiota model systems that enable disentangling complex, species-rich communities are essential for linking microbial taxa to specific host phenotypes.

ABSTRACT Disrupted interactions between host and intestinal bacteria are implicated in colorectal cancer (CRC) development. However, activities derived from these bacteria and their interplay with the host are unclear. Here, we examine this interplay by performing mouse and microbiota RNA sequencing on colon tissues and 16S and small RNA sequencing on stools from germfree (GF) and gnotobiotic ApcMinΔ850/+;Il10−/− mice associated with microbes from biofilm-positive human CRC tumor (BF+T) and biofilm-negative healthy (BF-bx) tissues. The bacteria in BF+T mice differentially expressed (DE) >2,900 genes, including genes related to bacterial secretion, virulence, and biofilms but affected only 62 host genes. Small RNA sequencing of stools from these cohorts revealed eight significant DE host microRNAs (miRNAs) based on biofilm status and several miRNAs that correlated with bacterial taxon abundances. Additionally, computational predictions suggest that some miRNAs preferentially target bacterial genes while others primarily target mouse genes. 16S rRNA sequencing of mice that were reassociated with mucosa-associated communities from the initial association revealed a set of 13 bacterial genera associated with cancer that were maintained regardless of whether the reassociation inoculums were initially obtained from murine proximal or distal colon tissues. Our findings suggest that complex interactions within bacterial communities affect host-derived miRNA, bacterial composition, and CRC development. IMPORTANCE Bacteria and bacterial biofilms have been implicated in colorectal cancer (CRC), but it is still unclear what genes these microbial communities express and how they influence the host. MicroRNAs regulate host gene expression and have been explored as potential biomarkers for CRC. An emerging area of research is the ability of microRNAs to impact growth and gene expression of members of the intestinal microbiota. This study examined the bacteria and bacterial transcriptome associated with microbes derived from biofilm-positive human cancers that promoted tumorigenesis in a murine model of CRC. The murine response to different microbial communities (derived from CRC patients or healthy people) was evaluated through RNA and microRNA sequencing. We identified a complex interplay between biofilm-associated bacteria and the host during CRC in mice. These findings may lead to the development of new biomarkers and therapeutics for identifying and treating biofilm-associated CRCs.

Omnivorous animals, including humans, harbor diverse, species-rich gut communities that impact their growth, development, and homeostasis. Model invertebrates are broadly accessible experimental platforms that enable linking specific species or species groups to host phenotypes, yet often their specialized diets and distinct gut microbiota make them less comparable to human and other mammalian and gut communities. The omnivorous cockroach Periplaneta americana harbors ∼4 × 10 2 bacterial genera within its digestive tract and is enriched with taxa commonly found in omnivorous mammals (i.e., Proteobacteria, Bacteroidetes , and Firmicutes ). These features make P. americana a valuable platform for identifying microbe-mediated host phenotypes with potential translations to mammals. Rearing P. americana insects under germfree conditions resulted in prolonging development time by ∼30% and an up to ∼8% reduction in body size along three dimensions. Germfree rearing resulted in downregulation of gene networks involved in growth, energy homeostasis, and nutrient availability. Reintroduction of a defined microbiota comprised of a subset of P. americana commensals to germfree insects did not recover normal growth and developmental phenotypes or transcriptional profiles observed in conventionally reared insects. These results are in contrast with specialist-feeding model insects (e.g., Drosophila ), where introduction of a single endemic bacterial species to germfree condition-reared specimens recovered normal host phenotypes. These data suggest that understanding microbe-mediated host outcomes in animals with species-rich communities should include models that typically maintain similarly diverse microbiomes. The dramatic transcriptional, developmental, and morphological phenotypes linked to gut microbiome status in this study illustrates how microbes are key players in animal growth and evolution. IMPORTANCE Broadly accessible model organisms are essential for illustrating how microbes are engaged in the growth, development, and evolution of animals. We report that germfree rearing of omnivorous Periplaneta americana cockroaches resulted in growth defects and severely disrupted gene networks that regulate development, which highlights the importance of gut microbiota in these host processes. Absence of gut microbiota elicited a starvation-like transcriptional response in which growth and development were inhibited while nutrient scavenging was enhanced. Additionally, reintroduction of a subset of cockroach gut bacterial commensals did not broadly recover normal expression patterns, illustrating that a particular microbiome composition may be necessary for normal host development. Invertebrate microbiota model systems that enable disentangling complex, species-rich communities are essential for linking microbial taxa to specific host phenotypes.

Disrupted interactions between host and intestinal bacteria are implicated in colorectal cancer (CRC) development. However, activities derived from these bacteria and their interplay with the host are unclear. Here, we examine this interplay by performing mouse and microbiota RNA sequencing on colon tissues and 16S and small RNA sequencing on stools from germfree (GF) and gnotobiotic Apc Min Δ 850/+ ; Il10 -/- mice associated with microbes from biofilm-positive human CRC tumor (BF+T) and biofilm-negative healthy (BF-bx) tissues. The bacteria in BF+T mice differentially expressed (DE) >2,900 genes, including genes related to bacterial secretion, virulence, and biofilms but affected only 62 host genes. Small RNA sequencing of stools from these cohorts revealed eight significant DE host microRNAs (miRNAs) based on biofilm status and several miRNAs that correlated with bacterial taxon abundances. Additionally, computational predictions suggest that some miRNAs preferentially target bacterial genes while others primarily target mouse genes. 16S rRNA sequencing of mice that were reassociated with mucosa-associated communities from the initial association revealed a set of 13 bacterial genera associated with cancer that were maintained regardless of whether the reassociation inoculums were initially obtained from murine proximal or distal colon tissues. Our findings suggest that complex interactions within bacterial communities affect host-derived miRNA, bacterial composition, and CRC development. IMPORTANCE Bacteria and bacterial biofilms have been implicated in colorectal cancer (CRC), but it is still unclear what genes these microbial communities express and how they influence the host. MicroRNAs regulate host gene expression and have been explored as potential biomarkers for CRC. An emerging area of research is the ability of microRNAs to impact growth and gene expression of members of the intestinal microbiota. This study examined the bacteria and bacterial transcriptome associated with microbes derived from biofilm-positive human cancers that promoted tumorigenesis in a murine model of CRC. The murine response to different microbial communities (derived from CRC patients or healthy people) was evaluated through RNA and microRNA sequencing. We identified a complex interplay between biofilm-associated bacteria and the host during CRC in mice. These findings may lead to the development of new biomarkers and therapeutics for identifying and treating biofilm-associated CRCs., (Copyright © 2020 Tomkovich et al.)