Antibiotic resistance is a growing problem worldwide, with frequent transmission between pathogens and environmental organisms. Rural rivers can support high levels of recreational use by people unaware of inputs from treated wastewater, while wastewater treatment plants (WWTPs) can generate a small but significant portion of flow volume into a river surrounded by forest and agriculture.

In-person college instruction during the 2020 pandemic required effective and economical monitoring of COVID-19 prevalence. Kenyon College and the Village of Gambier conducted measurement of SARS-CoV-2 RNA from the village wastewater plant and from an on-campus sewer line. Wastewater RNA detection revealed virus prevalence leading to individual testing and case identification. Wastewater surveillance also showed when case rates had subsided, thus limiting the need for individual clinical testing. Overall, wastewater virus surveillance allows more targeted use of individual testing and increases community confidence in student population management.

Classifying taxa, including those that have not previously been identified, is a key task in characterizing the microbial communities of under-described habitats, including permanently ice-covered lakes in the dry valleys of the Antarctic. Current supervised phylogeny-based methods fall short on recognizing species assembled from metagenomic datasets from such habitats, as they are often incomplete or lack closely known relatives. Here, we report an efficient software suite, "Genome Constellation", that is capable of rapidly characterizing a large number of metagenome-assembled genomes. Genome Constellation estimates similarities between genomes based on their k-mer matches, and subsequently uses these similarities for classification, clustering, and visualization. The clusters of reference genomes formed by Genome Constellation closely resemble known phylogenetic relationships while simultaneously revealing unexpected connections. In a dataset containing 1,693 draft genomes assembled from the Antarctic lake communities where only 40% could be placed in a phylogenetic tree, Genome Constellation improves taxa assignment to 61%. The clustering-based analysis revealed several novel taxa groups, including six clusters that may represent new bacterial phyla. Remarkably, we discovered 63 new giant viruses, 3 of which could not be found by using the traditional marker-based approach. In summary, we demonstrate that Genome Constellation provides an unbiased option to rapidly analyze a large number of microbial genomes and visually explore their relatedness. The software is available under BSD license at: https://bitbucket.org/berkeleylab/jgi-genomeconstellation/.

The perennial ice-covered lakes of the Antarctic McMurdo Dry Valleys harbour oligotrophic microbial communities that are separated geographically from other aquatic systems. Their microbiomes include planktonic microbes as well as lift-off mat communities that emerge from the ice. We used the ShortBRED protein family profiler to quantify the antibiotic resistance genes (ARGs) from metagenomes of lift-off mats emerging from ice and from filtered water samples of Lake Fryxell and Lake Bonney. The overall proportion of ARG hits was similar to that found in temperate-zone rural ponds with moderate human inputs. Specific ARGs showed distinct distributions for the two lakes and for mat vs planktonic sources. Metagenomic taxa distributions showed that mat phototrophs consisted mainly of cyanobacteria or Betaproteobacteria, whereas the water column phototrophs were mainly protists. An enrichment culture of the Betaproteobacterium Rhodoferax antarcticus from a Lake Fryxell mat sample showed an unusual mat-forming phenotype not previously reported for this species. Its genome showed no ARGs associated with Betaproteobacteria but had ARGs consistent with a minor Pseudomonas component. The Antarctic lake mats and water showed specific ARGs distinctive to the mat and water sources, but overall ARG levels were similar to those of temperate water bodies with moderate human inputs.

The Antarctic Taylor Valley Lakes Fryxell and Bonney harbor oligotrophic microbial communities that are separated geographically from other aquatic systems. Their microbiomes include planktonic as well as lift-off mat communities that float to the underside of the perennial ice cover and eventually emerge at the surface. We investigated the antibiotic resistance genes (ARGs) from metagenomes of lift-off mats emerging from ice, from filtered water samples of Lake Fryxell, and from filtered water samples of Lake Bonney. ARG sequence markers were designed by ShortBRED-Identify using the Comprehensive Antibiotic Resistance Database (CARD). The overall proportion of ARG hits in the metagenomes was found to be similar to that found in temperate-zone rural water bodies with moderate human inputs (0.0002-0.0007%). The specific ARGs found showed distinct distributions for the two lakes, and for mat versus planktonic sources. An enrichment culture of Rhodoferax antarcticus from a Lake Fryxell mat sample showed a mat-forming phenotype not previously reported for this species. Its genome showed no ARGs associated with Betaproteobacteria, but had ARGs consistent with a Pseudomonas minor component. The Antarctic lake mats and water showed specific ARGs distinctive to the mat and water sources, but overall ARG levels were similar to those of temperate water bodies.