skip to main content
US FlagAn official website of the United States government
dot gov icon
Official websites use .gov
A .gov website belongs to an official government organization in the United States.
https lock icon
Secure .gov websites use HTTPS
A lock ( lock ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.


Title: Single cell genome sequencing of laboratory mouse microbiota improves taxonomic and functional resolution of this model microbial community
Laboratory mice are widely studied as models of mammalian biology, including the microbiota. However, much of the taxonomic and functional diversity of the mouse gut microbiome is missed in current metagenomic studies, because genome databases have not achieved a balanced representation of the diverse members of this ecosystem. Towards solving this problem, we used flow cytometry and low-coverage sequencing to capture the genomes of 764 single cells from the stool of three laboratory mice. From these, we generated 298 high-coverage microbial genome assemblies, which we annotated for open reading frames and phylogenetic placement. These genomes increase the gene catalog and phylogenetic breadth of the mouse microbiota, adding 135 novel species with the greatest increase in diversity to the Muribaculaceae and Bacteroidaceae families. This new diversity also improves the read mapping rate, taxonomic classifier performance, and gene detection rate of mouse stool metagenomes. The novel microbial functions revealed through our single-cell genomes highlight previously invisible pathways that may be important for life in the murine gastrointestinal tract.  more » « less
Award ID(s):
1826734
PAR ID:
10325468
Author(s) / Creator(s):
; ; ; ;
Editor(s):
Kuo, Chih-Horng
Date Published:
Journal Name:
PLOS ONE
Volume:
17
Issue:
4
ISSN:
1932-6203
Page Range / eLocation ID:
e0261795
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. Jansson, Janet K. (Ed.)
    ABSTRACT Soil ecosystems harbor diverse microorganisms and yet remain only partially characterized as neither single-cell sequencing nor whole-community sequencing offers a complete picture of these complex communities. Thus, the genetic and metabolic potential of this “uncultivated majority” remains underexplored. To address these challenges, we applied a pooled-cell-sorting-based mini-metagenomics approach and compared the results to bulk metagenomics. Informatic binning of these data produced 200 mini-metagenome assembled genomes (sorted-MAGs) and 29 bulk metagenome assembled genomes (MAGs). The sorted and bulk MAGs increased the known phylogenetic diversity of soil taxa by 7.2% with respect to the Joint Genome Institute IMG/M database and showed clade-specific sequence recruitment patterns across diverse terrestrial soil metagenomes. Additionally, sorted-MAGs expanded the rare biosphere not captured through MAGs from bulk sequences, exemplified through phylogenetic and functional analyses of members of the phylum Bacteroidetes . Analysis of 67 Bacteroidetes sorted-MAGs showed conserved patterns of carbon metabolism across four clades. These results indicate that mini-metagenomics enables genome-resolved investigation of predicted metabolism and demonstrates the utility of combining metagenomics methods to tap into the diversity of heterogeneous microbial assemblages. IMPORTANCE Microbial ecologists have historically used cultivation-based approaches as well as amplicon sequencing and shotgun metagenomics to characterize microbial diversity in soil. However, challenges persist in the study of microbial diversity, including the recalcitrance of the majority of microorganisms to laboratory cultivation and limited sequence assembly from highly complex samples. The uncultivated majority thus remains a reservoir of untapped genetic diversity. To address some of the challenges associated with bulk metagenomics as well as low throughput of single-cell genomics, we applied flow cytometry-enabled mini-metagenomics to capture expanded microbial diversity from forest soil and compare it to soil bulk metagenomics. Our resulting data from this pooled-cell sorting approach combined with bulk metagenomics revealed increased phylogenetic diversity through novel soil taxa and rare biosphere members. In-depth analysis of genomes within the highly represented Bacteroidetes phylum provided insights into conserved and clade-specific patterns of carbon metabolism. 
    more » « less
  2. The house mouse species complex (Mus musculus) is comprised of three primary subspecies. A large number of secondary subspecies have also been suggested on the basis of divergent morphology and molecular variation at limited numbers of markers. While the phylogenetic relationships among the primary M. musculus subspecies are well-defined, relationships among secondary subspecies and between secondary and primary subspecies remain less clear. Here, we integrate de novo genome sequencing of museum-stored specimens of house mice from one secondary subspecies (M. m. bactrianus) and publicly available genome sequences of house mice previously characterized as M. m. helgolandicus, with whole genome sequences from diverse representatives of the three primary house mouse subspecies. We show that mice assigned to the secondary M. m. bactrianus and M. m. helgolandicus subspecies are not genetically differentiated from M. m. castaneus and M. m. domesticus, respectively. Overall, our work suggests that the M. m. bactrianus and M. m. helgolandicus subspecies are not well-justified taxonomic entities, emphasizing the importance of leveraging whole-genome sequence data to inform subspecies designations. Additionally, our investigation provides tailored experimental procedures for generating whole genome sequences from air-dried mouse skins, along with key genomic resources to inform future genomic studies of wild mouse diversity. 
    more » « less
  3. Abstract Natural microbial communities are phylogenetically and metabolically diverse. In addition to underexplored organismal groups 1 , this diversity encompasses a rich discovery potential for ecologically and biotechnologically relevant enzymes and biochemical compounds 2,3 . However, studying this diversity to identify genomic pathways for the synthesis of such compounds 4 and assigning them to their respective hosts remains challenging. The biosynthetic potential of microorganisms in the open ocean remains largely uncharted owing to limitations in the analysis of genome-resolved data at the global scale. Here we investigated the diversity and novelty of biosynthetic gene clusters in the ocean by integrating around 10,000 microbial genomes from cultivated and single cells with more than 25,000 newly reconstructed draft genomes from more than 1,000 seawater samples. These efforts revealed approximately 40,000 putative mostly new biosynthetic gene clusters, several of which were found in previously unsuspected phylogenetic groups. Among these groups, we identified a lineage rich in biosynthetic gene clusters (‘ Candidatus Eudoremicrobiaceae’) that belongs to an uncultivated bacterial phylum and includes some of the most biosynthetically diverse microorganisms in this environment. From these, we characterized the phospeptin and pythonamide pathways, revealing cases of unusual bioactive compound structure and enzymology, respectively. Together, this research demonstrates how microbiomics-driven strategies can enable the investigation of previously undescribed enzymes and natural products in underexplored microbial groups and environments. 
    more » « less
  4. The flora and fauna of island systems, especially those in the Indo-Pacific, are renowned for their high diversification rates and outsized contribution to the development of evolutionary theories. The total diversity of geographic radiations of many Indo-Pacific fauna is often incompletely sampled in phylogenetic studies due to the difficulty in obtaining single island endemic forms across the Pacific and the relatively poor performance of degraded DNA when using museum specimens for inference of evolutionary relationships. New methods for production and analysis of genome-wide datasets sourced from degraded DNA are facilitating insights into the complex evolutionary histories of these influential island faunas. Here, we leverage whole genome resequencing (20X average coverage) and extensive sampling of all taxonomic diversity within Todiramphus kingfishers, a rapid radiation of largely island endemic Great Speciators. We find that whole genome datasets do not outright resolve the evolutionary relationships of this clade: four types of molecular markers (UCEs, BUSCOs, SNPs, and mtDNA) and tree building methods did not find a single well-supported and concordant species-level topology. We then uncover evidence of widespread incomplete lineage sorting and both ancient and contemporary gene flow and demonstrate how these factors contribute to conflicting evolutionary histories. Our complete taxonomic sampling allowed us to further identify a novel case of mitochondrial capture between two allopatric species, suggesting a potential historical (but since lost) hybrid zone as islands were successively colonized. Taken together, these results highlight how increased genomic and taxon sampling can reveal complex evolutionary patterns in rapid island radiations. 
    more » « less
  5. Abstract Host-microbe interactions are intimately linked to eukaryotic evolution, particularly in sap-sucking insects that often rely on obligate microbial symbionts for nutrient provisioning. Cicadas (Cicadidae: Auchenorrhyncha) specialize on xylem fluid and derive many essential amino acids and vitamins from intracellular bacteria or fungi (Hodgkinia,Sulcia, andOphiocordyceps) that are propagated via transmission from mothers to offspring. Despite the beneficial role of these non-gut symbionts in nutrient provisioning, the role of beneficial microbiota within the gut remains unclear. Here, we investigate the relative abundance and impact of host phylogeny and ecology on gut microbial diversity in cicadas using 16S ribosomal RNA gene amplicon sequencing data from 197 wild-collected cicadas and new mitochondrial genomes across 38 New Zealand cicada species, including natural hybrids between one pair of two species. We find low abundance and a lack of phylogenetic structure and hybrid effects but a significant role of elevation in explaining variation in gut microbiota. 
    more » « less