NEW YORK (GenomeWeb News) – An international team led by investigators in the US and Australia reported online yesterday in Nature that it has sequenced draft genomes for hundreds of microbes from a range of environmental sites around the world.
As part of an effort to generate the Genomic Encyclopedia of Bacteria and Archaea "Microbial Dark Matter," or GEBA-MDM dataset, the researchers used single-cell sequencing strategies to sequence more than 200 uncultivated microbes formerly known only from metagenomic sequencing studies, 16S ribosomal RNA gene profiling, and the like.
These genomes made it possible for the study's authors to not only assign slews of existing microbial metagenome sequence reads, but also to look at the potential metabolic capabilities of the newly sequenced bacteria and archaea and their positions in the microbial tree of life.
"What we are now discovering are unexpected metabolic features that extend our understanding of biology and challenge established boundaries between the domains of life," co-corresponding author Tanja Woyke, with the US Department of Energy's Joint Genome Institute, said in a statement.
"There is still a staggering amount of diversity to explore," she added. "To try to capture 50 percent of just the currently known phylogenetic diversity, we would have to sequence 20,000 more genomes, and these would have to be selected based on being members of underrepresented branches on the tree."
Nearly 90 percent of the microbes that can be grown in the lab fall into just four bacterial phyla, Woyke and her co-authors noted, but rRNA sequencing studies and phylogenetic analysis hint that there are dozens of other bacterial and archaeal phyla that are missed by those looking only at those cultivated microbes.
Likewise, many of the microbes sequenced to date have come from a limited portion of the microbial tree, though projects such as GEBA have gone to great pains to begin remedying such omissions by sequencing microbes from the tree's poorly represented branches.
More options have become available for putting together genome sequences for uncultured microbes, too — including approaches for fishing sequences for individual candidate organisms from a mishmash of metagenomic sequences that represent many microbes in a given environmental sample.
Another method that's sometimes used instead of, or in addition to, metagenomic sequencing is single-cell sequencing, which circumvents some of the complications associated with trying to make sense of reads from a complicated DNA collection.
"In particular, natural populations that have a high degree of genomic heterogeneity will be more accessible through single-cell genomics than through metagenomics," authors of the new study noted, "as co-assembly of multiple strains is avoided."
For the current study, the researchers relied on the single-cell approach to sequence 201 individual microbial cells from nine marine, fresh, or brackish water, hydrothermal, soil, or bioreactor environments.
The sites sampled included British Columbia's Sakinaw Lake; the Homestake Mine in South Dakota; Nevada's Great Boiling Spring; the Gulf of Maine; a marine site off of Oahu's northern coast; a Mexican sludge reactor; Etoliko Lagoon in western Greece; the Tropical Gyre in the south Atlantic; and the East Pacific Rise.
After isolating individual microbial cells by flow sorting, the team amplified DNA from single microbial cells, selected with help from rRNA sequence screening to narrow in on novel organisms.
The genomic DNA from each cell was then sequenced and assembled using a digital normalization approach that dialed back reads from parts of the genome prone to bias during the amplification step.
In the process, researchers generated draft genome assemblies that were between around 10 percent and 90 percent complete. Together, the 201 newly sequenced organisms are believed to span 21 bacterial and eight archaeal lineages that were only scantily represented by DNA sequence data in the past.
The team's phylogenetic analysis of these microbes refined the relationships within and between several bacterial and archaeal phyla and super-phyla. That improved view of the microbial phylogeny proved useful for linking existing sequence reads to a given lineage or organism, too, prompting the group to assign or re-assign hundreds of millions of existing metagenomic sequence reads.
The draft genome sequences also made it possible for researchers to make predictions about the metabolic wherewithal of the uncultivated microbes — patterns that themselves offer clues about what each organism might be up to in a given environment.
There were surprises in that analysis, too, researchers noted. Within at least three archaeal organisms, for example, they identified a set of transcription initiation factors best known in bacteria.
The genome of another archaeal representative contained genes from an enzyme normally found in a slime mold, pointing to possible lateral gene transfer from that eukaryote into the archaea, while some bacterial genomes contained genes used by archaea to produce the purine compound that forms the basis for nucleotides such as adenine and guanine.
Such findings "extend our understanding of biology and challenge established boundaries between the [bacterial, archaeal, and eukaryotic] domains of life," the study's authors wrote, noting that ongoing efforts to unravel additional microbial genomes will continue to reveal information about the biology and evolution of organisms from all reaches of life.