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. 2024 Feb 6;10(4):e25616.
doi: 10.1016/j.heliyon.2024.e25616. eCollection 2024 Feb 29.

Metagenomic survey of viral diversity obtained from feces of piglets with diarrhea

Lingling Qian  1 , Zi Zhuang  2 , Juan Lu  1 , Huiying Wang  3 , Xiaochun Wang  1 , Shixing Yang  1 , Likai Ji  1 , Quan Shen  1 , Wen Zhang  1 , Tongling Shan  4
Affiliations

Metagenomic survey of viral diversity obtained from feces of piglets with diarrhea

Lingling Qian et al. Heliyon. .

Abstract

Pigs are natural host to various zoonotic pathogens including viruses. In this study, we analyzed the viral communities in the feces of 89 piglets with diarrhea under one month old which were collected from six farms in Jiangsu Province of the Eastern China, using the unbiased virus metagenomic method. A total of 89 libraries were constructed, and 46937894 unique sequence reads were generated by Illumina sequencing. Overall, the family Picornaviridae accounted for the majority of the total reads of putative mammalian viruses. Ten novel virus genomes from different family members were discovered, including Parvoviridae (n = 2), Picobirnaviridae (n = 4) and CRESS DNA viruses (n = 4). A large number of phages were identified, which mainly belonged to the order Caudovirales and the family Microviridae. Moreover, some identified viruses were closely related to viruses found in non-porcine hosts, highlighting the potential for cross-species virus dissemination. This study increased our understanding of the fecal virus communities of diarrhea piglets and provided valuable information for virus monitoring and preventing.

Keywords: Diarrhea piglets; Feces; Metagenomic; Viral communities; Virus genome.

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Conflict of interest statement

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Figures

Fig. 1
Fig. 1
Viral taxonomy analyses at the family level. (A) Heatmap representing the read number of each viral family in each library on log2 scale. The sample groups are shown with corresponding colors (see color legend). (B) Heatmap representing the read number of each phage virus family in each farm on log2 scale. The sample groups are shown with corresponding colors (see color legend). (C) The pie chart shows the composition of fecal virus groups of diarrhea piglets and displays the percentage of virus sequences in different virus families.
Fig. 2
Fig. 2
Diversity analysis of virus difference in six farms. (A) Principal coordinate analysis (PcoA) based on Bray-Curtis of six farms at the family level. (B)The P-value is calculated by ANOSIM. (C) Box plots were used to compare the magnitude of differences in the Alpha diversity index between farm groups using the Simpson and Shannon indexes. Each color box represented a subgroup, and the p-value above the box indicated whether or not that diversity index differed significantly across all subgroups. (D) The Upset diagram effectively represented the farms' shared and unique viruses. The horizontal bar on the left side of the figure depicted the number of virus species in each farm, and the lines connecting the dots represented the intersection of different farms, with individual dots indicating unique to that farm and nodes connected by lines indicating common. The vertical bars represented the number of corresponding intersection elements.
Fig. 3
Fig. 3
Phylogenies of Parvoviruses identified in the diarrhea piglets. (A) The genomic organization of the six Parvoviruses identified in diarrhea piglets. (B) Bayesian inference tree established based on amino acid sequences of NS protein of the genus Bocaparvovirus. (C) Bayesian inference tree established based on amino acid sequences of NS protein of the genus Chaphamaparvovirus. The viruses identified in this study are labeled with red branches and leave names. Different taxonomic clusters were represented by rectangles filled with different colors, and taxon names are indicated on the right. Tree scales indicate the amino acid substitutions per site.
Fig. 4
Fig. 4
Phylogenies of the genus Mamastrovirus and the family Caliciviriade identified in diarrhea piglets. (A) Bayesian inference tree established based on amino acid sequences of Cap protein of the genus Mamastrovirus. (B) Bayesian inference tree established based on amino acid sequences of ORF1 protein of the family Caliciviriade. The viruses identified in this study are labeled with red branches or red dots and leave names. Different taxonomic clusters were represented by rectangles filled with different colors, and taxon names are indicated on the right. Tree scales indicate the amino acid substitutions per site.
Fig. 5
Fig. 5
Phylogenies of the family Picobirnaviridae and the family Coronaviridae identified in diarrhea piglets. (A) Bayesian inference tree established based on amino acid sequences of RdRp protein of picobirnaviruses. (B) Bayesian inference tree established based on amino acid sequences of RdRp protein of coronaviruses. The viruses identified in this study are labeled with red branches or red dots and leave names. Different taxonomic clusters were represented by rectangles filled with different colors, and taxon names of coronaviruses are indicated on the right. Tree scales indicate the amino acid substitutions per site.
Fig. 6
Fig. 6
Phylogenies of the Picornaviruses identified in diarrhea piglets. (A) Bayesian inference tree established based on amino acid sequences of ORF1 protein of the genus Sapelovirus. (B) Bayesian inference tree established based on amino acid sequences of ORF1 protein of the genus Teschovirus. (C) Bayesian inference tree established based on amino acid sequences of ORF1 protein of the genus Enterovirus. (D) Bayesian inference tree established based on amino acid sequences of ORF1 protein of the genus Kobuvirus. The viruses identified in this study are labeled with red branches and leave names. Different viral groups are marked with color coding shown by the key in the upper right. Tree scales indicate the amino acid substitutions per site.
Fig. 7
Fig. 7
Phylogenies of CRESS DNA viruses identified in diarrhea piglets. (A) Bayesian inference tree established based on amino acid sequences of Rep protein of circoviruses. (B) The genomic organization of the four Genomoviruses identified in diarrhea piglets. (C) Bayesian inference tree established based on amino acid sequences of Rep protein of the family Genomoviridae (D) Bayesian inference tree established based on amino acid sequences of Rep protein of the genus Porprismacovirus. The viruses identified in this study are labeled with red dots or red branches and leave names. Different taxonomic clusters of circoviriuses and genomoviriuses were represented by rectangles filled with different colors, and taxon names are indicated on the right. Tree scales indicate the amino acid substitutions per site.
Fig. 8
Fig. 8
Phylogenies of the family Microviridae and the order Caudovirales identified in diarrhea piglets. (A) Bayesian inference tree established based on amino acid sequences of MCP protein of the family Microvirdae. The Unclassified Microvirinae were tentatively divided into ten clades artificially. (B) Bayesian inference tree established based on amino acid sequences of TERL protein of the order caudovirales. Representative strains of different subfamilies were included and marked with the color coding in the key on the top right. The viruses identified in this study are labeled with red branches and leave names. The size of the dots on nodes is positively correlated with the corresponding bootstrap score. Tree scales indicate the amino acid substitutions per site.
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