key: cord-1038022-oghe1dny
authors: Ash, K. T.; Alamilla, I.; Li, Y.; Joyner, D. C.; Williams, D. E.; McKay, P. J.; Green, B. M.; Iler, C.; DeBlander, S. E.; Kara-Murdoch, F.; Swift, C. M.; Hazen, T. C.
title: Coding-Complete Genome Sequence of a SARS-CoV-2 Variant Obtained from Raw Sewage at the University of Tennessee—Knoxville Campus
date: 2021-11-24
journal: Microbiol Resour Announc
DOI: 10.1128/mra.01049-21
sha: 4175e351440154242b30ff9f82e57811374a1312
doc_id: 1038022
cord_uid: oghe1dny

Reported here is a coding-complete genome sequence of a SARS-CoV-2 variant obtained from raw wastewater samples at the University of Tennessee—Knoxville campus. This sequence provides insight into SARS-CoV-2 variants that circulate on large college campuses but remain mostly undetected.

sequencing due to its high average quantification cycle (C q ) value (C q = 24.31) from the RT-qPCR analysis. The sample was amplified using the PCR tiling ARTIC primer pools and prepared for sequencing using the NEBNext Ultra II DNA library prep kit for Illumina (NEB catalog number E7645L) (5) . Sequencing was performed on a NovaSeq 6000 instrument at the Institute for Genome Sciences, University of Maryland. The genome assembly was produced using viral-ngs v1.25.0 utilizing viral-pipelines v2.1.19.0 from the Broad Institute (6) . The Illumina paired-end reads were aligned to the SARS-CoV-2 reference strain (GenBank accession number NC_045512.2). The total number of reads aligned to the reference was 15,317,116, with a mean length of 249.7 bp. The mean coverage depth was 132,599.4 reads, with a standard deviation of 125,391.3 reads.

This SARS-CoV-2 isolate is 29,874 bp long, with a GC content of 39.3%. A BLAST comparison, under default parameters, revealed that the UTK-840 sequence shares 99.93% nucleotide similarity with the Wuhan reference strain (GenBank accession number NC_045512.2) (7). The differences between the sequences are represented by 20 single nucleotide mutations, of which 15 are transition and 5 are transversion mutations. Half of these FIG 1 Phylogenetic tree of the UTK-840 variant in relation to the reference variant and known variants of concern. The evolutionary distance was inferred using the UPGMA method (9). The optimal tree is shown. The tree is drawn to scale, with branch lengths in the same units as those of the evolutionary distances used to infer the phylogenetic tree (10) . The evolutionary distances were computed using the Jukes-Cantor method (11) and are given in the number of base substitutions per site. This analysis involved 6 whole-genomic sequences. The codon positions included were first, second, third, and noncoding. All ambiguous positions were removed for each sequence pair (pairwise deletion option). There were a total of 29,907 positions in the final data set. Evolutionary analyses were conducted using MEGA11 (12, 13) . mutations occurred in the Orf1ab gene. Three mutations are in the nucleocapsid gene and only one in the spike gene, which is the D614G mutation that has been observed in most of the variants of concern. A sequence comparison made using the Audacity Instant program on GISAID, under default parameters, revealed the UTK-840 sequence to be closely related to lineage B.1.2 and the GH neighbor clade (8) (Fig. 1) .

Data availability. The whole-genome sequence has been deposited at GenBank under the accession number MZ265375. The raw sequencing reads have been deposited at the NCBI SRA database under the accession number SRR16202037.

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We acknowledge the University of Tennessee Office of Research and Engagement for providing funding for this project and the Maryland Institute for Genome Sciences (IGS) Microbiome Service Laboratory (MSL) (University of Maryland) for sequencing of the sample.