Microbe-seq: high-throughput, single-microbe genomics with strain resolution, applied to a human gut microbiome

Abstract

We present Microbe-seq, a high-throughput single-microbe method that yields strain-resolved genomes from complex microbial communities. We encapsulate individual microbes into droplets with microfluidics and liberate their DNA, which we amplify, tag with droplet-specific barcodes, and sequence. We use Microbe-seq to explore the human gut microbiome; we collect stool samples from a single individual, sequence over 20,000 microbes, and reconstruct nearly-complete genomes of almost 100 bacterial species, including several with multiple subspecies strains. We use these genomes to probe genomic signatures of microbial interactions: we reconstruct the horizontal gene transfer (HGT) network within the individual and observe far greater exchange within the same bacterial phylum than between different phyla. We probe bacteria-virus interactions; unexpectedly, we identify a significant in vivo association between crAssphage, an abundant bacteriophage, and a single strain of Bacteroides vulgatus. Microbe-seq contributes high-throughput culture-free capabilities to investigate genomic blueprints of complex microbial communities with single-microbe resolution.

Ecological Stability Emerges at the Level of Strains in the Human Gut Microbiome

“…The ambiguity surrounding the bacterial species concept is well known [34] and reasonable alternatives have been proposed [19], but operationally species are, nonetheless, the predominant focus of attention in microbial ecology. This focus is reasonable, as within-host strain structure is a comparatively recent discovery [10, 28, 54] and 16S rRNA sequencing provides an inexpensive high-throughput means to examine community dynamics through OTUs. Regardless, the recapitulation of species-level macroecological dynamics at the level of strains calls into question the disproportionate focus on species as the primary locus of attention in characterizing community structure and dynamics.…”

“…The ambiguity surrounding the bacterial species concept is well known [34] and reasonable alternatives have been proposed [19], but operationally species are, nonetheless, the predominant focus of attention in microbial ecology. This focus is reasonable, as within-host strain structure is a comparatively recent discovery [10,28,54] and 16S rRNA sequencing provides an inexpensive high-throughput means to examine community dynamics through OTUs.…”

“…While a typical gut microbial species harbors considerable genetic variation both within and across hosts, the ecological and functional consequences of this diversity remain largely unknown. Although recent efforts have begun to characterize how genotypic diversity changes within healthy hosts over months to years, these trends are not, at present, quantified on the short time frames most relevant for microbial ecology – that is, over periods of days [5, 10, 15, 24, 25, 28, 30]. Understanding the typical scale of daily fluctuations in genetic variation is critical to assessing both the long-term stability of the genetic composition of the gut microbiome, as well the effects of occasional large perturbations resulting from changes in host diet, medication, travel, illness, and other factors.…”

Comprehensive Strain-Level Analysis of the Gut Microbe Faecalibacterium prausnitzii in Patients with Liver Cirrhosis

“…Nonetheless, our results suggest that strain heterogeneity should receive more attention. Recently developed single-cell microbial sequencing technologies seems to be more promising for metagenomic analysis ( 30 , 31 ), especially at the strain level. With the rapid development of sequencing technologies and experimental approaches, an increasing number of metagenomic studies will involve strain-level analysis.…”

Comparative Population Genetics in the Human Gut Microbiome

“…Over the course of just a few months, new genotypes can emerge and recombine over short time scales, and ultimately become lost or fixed over extended time scales. Indeed, it recently has been observed that adaptation and recombination can occur in the human gut over a matter of months (Zheng et al 2020;Yaffe & Relman 2020;Poyet et al 2019;Garud et al 2019;Zhao et al 2019;Lin & Kussell 2017). However, the necessary temporal resolution of sampling remains subject to the researcher's question, as short-term adaptation in response to a temporary environmental perturbation (e.g., a host consuming antibiotics over a few weeks) may require denser sampling than evolution in a relatively unchanging environment.…”