A Diverse Virome Is Identified in Parasitic Flatworms of Domestic Animals in Xinjiang, China

Abstract

Parasitic flatworms infect diverse vertebrates and are major threats to animal and even human health; however, little is known about the virome of these lower life forms. Using viral metagenomic sequencing, we characterized the virome of the parasitic flatworms collected from major domestic animals, including Dicrocoelium lanceatum and Taenia hydatigena, Echinococcus granulosus sensu stricto and Echinococcus multilocularis. Seven and three different viruses were discovered from D. lanceatum and T. hydatigena, respectively, and no viral sequences were found in adult tapeworms and protoscoleces of E. granulosus sensu stricto and E. multilocularis. Two out of the five parasitic flatworm species carry viruses, showing a host specificity of these viruses. These viruses belong to the Parvoviridae, Circoviridae, unclassified circular, Rep-encoding single-stranded (CRESS) DNA virus, Rhabdoviridae, Endornaviridae, and unclassified RNA viruses. The presence of multiple highly divergent RNA viruses, especially those that cluster with viruses found in marine animals, implies a deep evolutionary history of parasite-associated viruses. In addition, we found viruses with high identity to common pathogens in dogs, including canine circovirus and canine parvovirus 2. The presence of these viruses in the parasites implies that they may infect parasitic flatworms but does not completely exclude the possibility of contamination from host intestinal contents. Furthermore, we demonstrated that certain viruses, such as CRESS DNA virus may integrate into the genome of their host. Our results expand the knowledge of viral diversity in parasites of important domestic animals, highlighting the need for further investigations of their prevalence among other parasites of key animals. IMPORTANCE Characterizing the virome of parasites is important for unveiling the viral diversity, evolution, and ecology and will help to understand the "Russian doll" pattern among viruses, parasites, and host animals. Our data indicate that diverse viruses are present in specific parasitic flatworms, including viruses that may have an ancient evolutionary history and viruses currently circulating in parasite-infected host animals. These data also raise the question of whether parasitic flatworms acquire and/or carry some viruses that may have transmission potential to animals. In addition, through the study of virus-parasite-host interactions, including the influence of viral infection on the life cycle of the parasite, as well as its fitness and pathogenicity to the host, we could find new strategies to prevent and control parasitic diseases.

Keywords: metagenomics; parasitic flatworms; viral metagenomics; virome; virus discovery; virus evolution.

FIG 1

Overview of all the viruses in the different sample pools of parasites. (A) Pictures of D. lanceatum and E. granulosus isolated from the same sheep liver. (B) Pictures of T. hydatigena isolated from dog intestine. The relative abundance of different eukaryotic viruses in adult flukeworms of D. lanceatum from sheep liver (A) and T. hydatigena from dog intestines (B) is shown. No viral sequences were detected in the sample pools of E. granulosus from wither sheep liver and dog intestine or from sample pools of E. multilocularis from dog intestine.

FIG 2

The genomic structures and evolutionary relationships of the newly discovered Rhabdo-like virus (DRLV1/2) with representative viruses from the Rhabdoviridae. (A) The genomic structures of DRLV1/2 and its closely related homolog. Hypothetical ORFs, the L protein including the RdRp region, and genetic identities of the RdRp region between DRLV1/2 and its homolog are shown. (B) Phylogenetic trees were generated using the full-length coding sequences of RdRp. MEGA 7 was used for phylogeny inference using the maximum-likelihood (ML) method based on the LG+G model, and the analyses were conducted with 1,000 bootstrap replicates. Representative viruses from different genera of Rhabdoviridae and the most closely related viruses from the database were included.

FIG 3

The genomic structures and evolutionary relationships of the newly discovered Alphaendornavirus (DiEV) with representative viruses from the Endornaviridae. (A) The genomic structures of DiEV and its closely related homolog. Hypothetical ORF and genetic identities of ORFs between DiEV and its homolog are shown. (B) Phylogenetic trees were generated using the full-length coding sequences of ORFs. MEGA 7 was used for phylogeny inference using the maximum-likelihood (ML) method based on the GTR+G+I model, and the analyses were conducted with 1,000 bootstrap replicates. Representative viruses from different genera of Endornaviridae and the most closely related viruses from the database were included.

FIG 4

The genomic structures and evolutionary relationships of the newly discovered unclassified RNA viruses with other representative RNA viruses. (A) The genomic structures of unclassified RNA viruses and their closely related homolog. Hypothetical ORFs, the hypothetical protein including the RdRp region, and genetic identities of the RdRp region between unclassified RNA viruses and its homolog are shown. (B) Phylogenetic trees were generated using the full-length coding sequences of the RdRp. MEGA 7 was used for phylogeny inference using the maximum-likelihood (ML) method based on the LG+G model, and the analyses were conducted with 1,000 bootstrap replicates. Representative viruses from different genera of unclassified RNA virus families and the most closely related viruses from the database were included.

FIG 5

The genomic structure and evolutionary relationships of parasite-associated CRESS DNA virus. (A) The circular genomic structure of DiCV with a length of 2,364 bp; a typical stem-loop (TAGATTAC) is present between Rep and Cap. (B) Evolutionary relationships of DiCV with other representative viruses from CRESS DNA virus. Using the Rep protein, the phylogenetic tree was built using the maximum-likelihood (ML) method under the GTR+G+I model. (C) Evolutionary relationships of the cap protein of DiCV and its most closely related bat circovirus with the genomic sequences of different parasitic flatworms. Phylogenetic analyses were conducted with 1,000 bootstrap replicates.

FIG 6

Evolutionary relationships of all cycloviruses and circoviruses found in this study with representative viruses from the Circoviridae. Phylogenetic trees were generated using the coding sequences of Rep in the new cyclovirus. MEGA 7 was used for phylogeny inference using the maximum-likelihood (ML) method based on the GTR+G+I model, and the analyses were conducted with 1,000 bootstrap replicates. Representative viruses from different genera of Circoviridae and the most closely related viruses from the database were included.

FIG 7

Evolutionary relationships of the parvovirus found in this study with representative viruses from the Parvoviridae. Phylogenetic trees were generated based on the full-length coding sequences of the NS and VP regions. MEGA 7 was used for phylogeny inference using the maximum-likelihood (ML) method based on the GTR+G+I model. Analyses were conducted with 1,000 bootstrap replicates. Representative viruses from different genera of Parvoviridae and the most closely related viruses from the database were included.