Genomes of cressdnaviruses, microviruses, a gyrovirus, and a caudovirus identified in the fecal samples of a mallard and double-crested cormorant

ABSTRACT Mallards and double-crested cormorants have a broad distribution across North America. In the fecal sample from two individual mallard and double-crested cormorant, we determined the genomes of a caudovirus, microviruses (n = 6), cressdnaviruses (n = 35), and a gyrovirus (chicken anemia virus, CAV). Here, we report double-crested cormorant as a CAV host.

M allards (Anas platyrhynchos) and double-crested cormorants (Nannopterum auritum) are widely distributed across North America.Although viruses in 23 families have been identified as circulating in mallards, relatively less is known about those in double-crested cormorants (1).To date, only viruses in the Flaviviridae, Orthomyxoviridae, and Paramyxoviridae have been reported in double-crested cormor ants (1).
As part of our effort to identify viruses in migratory birds that concentrate around bodies of freshwater (2,3) in the southwestern USA (part of the Pacific Flyway for migratory birds), we collected two fecal samples, one from a mallard and one from a double-crested cormorant at Kiwanis Park, Tempe (AZ, USA) on 12 January 2021.The samples were collected in 2 mL tubes following a visual defecation observation.The fecal samples were homogenized in 2 mL of SM buffer, centrifuged at 10,000 × g for 10 min, and the supernatant from each sample was filtered through 0.45 and 0.2 µm syringe filters.Viral DNA was extracted from 200 µL of the filtrate with the High Pure Viral Nucleic Acid Kit (Roche, USA).Circular viral DNA was enriched using rolling circle amplification (RCA) with the Templiphi 100 Amplification Kit (GE Healthcare, USA).Illumina sequencing libraries were generated from the RCA products using the DNA TrueSeq Nano Kit and sequenced on an Illumina HiSeq4000.Trimmomatic v0.39 (4) was used to trim the raw paired-end reads (2 × 150 nt) and were then de novo assembled using MEGAHIT v1.2.9 (5).Contigs > 1,000 nt in length were analyzed using BLASTx (6) against a viral RefSeq protein sequence database (release 207) to identify viral-like sequences.The viral-like sequences were checked for terminal redundancy to identify circular genomes.All contigs were annotated with Cenote-taker 2 (7) with manual checks to identify spliced coding regions (Fig. 1).Read depth for viral contigs was determined using BBMap (8).All bioinformatic tools were run with default parameters.
In the mallard sample, we identified a caudovirus, 6 microviruses, 3 genomoviruses, and 15 unclassified cressdnaviruses (Fig. 1).A gyrovirus and 17 cressdnaviruses (Fig. 1) were identified in the double-crested cormorant sample.A BLASTn (9) analysis of the virus genomes identified in this study against the non-redundant nucleotide database shows that 37 of the 43 genomes have low similarity (65.88%-89.09%)over a low genome coverage (3%-58%) with E-values ranging from 0 to 2 × 10 −5 (Table 1).The   viruses with accessions OR915625, OR915629, OR915630, and OR915634 from mallard and OR915600 and PP473079 from double-crested cormorant have >70% genome coverage with 73.77%-99.16%identity and E-values of 0 in the BLASTn analysis (Table 1).Other than chicken anemia virus (CAV), which infects avian species, we are unable to determine the true hosts of the caudovirus, microviruses, and cressdnaviruses.CAV has mainly been identified in chickens (Gallus gallus) where it has a negative impact on chicken health.Although CAV has been identified in two other avian hosts, including waterfowl (10) and neognath (11), here we report its identification in double-crested cormorant as a host.

TABLE 1
Summary of the top BLASTn hit to the genomes of the viruses identified in this study

TABLE 1
Summary of the top BLASTn hit to the genomes of the viruses identified in this study(Continued) July 2024 Volume 13 Issue 7 10.1128/mra.00332-244