Identification and Resolution of Microdiversity through Metagenomic Sequencing of Parallel Consortia

Abstract

cTo gain a predictive understanding of the interspecies interactions within microbial communities that govern community function, the genomic complement of every member population must be determined. Although metagenomic sequencing has enabled the de novo reconstruction of some microbial genomes from environmental communities, microdiversity confounds current genome reconstruction techniques. To overcome this issue, we performed short-read metagenomic sequencing on parallel consortia, defined as consortia cultivated under the same conditions from the same natural community with overlapping species composition. The differences in species abundance between the two consortia allowed reconstruction of near-complete (at an estimated >85% of gene complement) genome sequences for 17 of the 20 detected member species. Two Halomonas spp. indistinguishable by amplicon analysis were found to be present within the community. In addition, comparison of metagenomic reads against the consensus scaffolds revealed within-species variation for one of the Halomonas populations, one of the Rhodobacteraceae populations, and the Rhizobiales population. Genomic comparison of these representative instances of inter-and intraspecies microdiversity suggests differences in functional potential that may result in the expression of distinct roles in the community. In addition, isolation and complete genome sequence determination of six member species allowed an investigation into the sensitivity and specificity of genome reconstruction processes, demonstrating robustness across a wide range of sequence coverage (9؋ to 2,700؋) within the metagenomic data set. Microdiversity refers to the diversity of organisms that are closely related phylogenetically yet exhibit different metabolic activities and therefore occupy distinct niches. Genomic studies comparing multiple strains of the same species have revealed that while much of the genome sequence is highly conserved, significant functional variation can exist, arising from changes in gene function due to mutation, the introduction of genes by horizontal gene transfer, or changes in gene regulation due to mutation or genome rearrangement. Microbial community diversity is usually measured via sequencing of either all or a part of the 16S rRNA gene. It is well established that bacteria that have near-identical or identical 16S rRNA sequences can have significantly divergent genome complements, cell morphologies, and metabolic functions (1-4).Recent developments in sequencing technologies and analysis have enabled cultivation-independent sequencing of intact microbial communities (metagenomics) and prediction of functional potential of individual microbial populations within a community. Early metagenomic work employed the same technology used for isolate genome sequencing, the Sanger method, which provided only a limited sequencing depth. Nonetheless, initial attempts to reconstruct member genomes in very-low-diversity communities or abundant members of complex communities were successful (5, 6)....

Measurement of bacterial replication rates in microbial communities

“…Together, they make up considerably more than 15% of bacterial diversity 15,28 , yet they are known almost exclusively from genomic sampling 7,15,18,29–33 . Based on having small cells and genomes with only a few tens of ribosomes, it was inferred that these organisms grow slowly 27,34 .…”

“…4). Symbiosis has been inferred as a general life strategy for these organisms 7,15,18,29–33 , and has been demonstrated in a few cases 35–38 . Rapid growth of CPR bacteria may require rapid growth of host cells.…”

Microbial Diversity of Source and Point-of-Use Water in Rural Haiti – A Pyrosequencing-Based Metagenomic Survey

“…The advent of the next generation sequencing (NGS) based metagenomics methods enable a broad spectrum identification of different microbes along with a high-resolution visualization of the overall microbiota of a given sample without the need of culturing the bacteria [16, 17]. Studies utilizing NGS techniques to understand the overall microbial composition, ecology, and diversity of different types of environmental media have gained remarkable attention from the scientific communities which is evident by a growing number of reports in recent years [18–21]. Moreover, recent studies indicate that metagenomic tools may identify new set of microbial parameters and targets for a wider, comprehensive, and more representative risk characterization [22, 23].…”

Trade-offs between microbiome diversity and productivity in a stratified microbial mat

“…Interestingly, this cyanobacterium is known to be motile and metabolically coupled to heterotrophic bacteria that match or are very similar to the OTUs abundant near the surface of the mat, for example, Roseibacterium sp. HL7711_P4H5 (Cole et al, 2014;Nelson et al, 2016). The most abundant OTU identified in this system belonged to a cyanobacterium of the family Pseudanabaenaceae.…”

“…These niche spaces mapped to common taxa that were associated with measured variance in community structure (beta diversity shown in Figure 4). Some OTUs, belonging to the most abundant families (Supplementary Figure S5), matched species that have been previously described and sequenced (Cole et al, 2014;Nelson et al, 2016) allowing function, relating to microbial lifestyle and niche space, to be inferred. The Phormidium sp.…”