Comparative analysis of gut DNA viromes in wild and captive Himalayan vultures

Abstract

Introduction: Himalayan vultures (Gyps hinalayensis) are widely distributed on the Qinghai-Tibetan Plateau and play a crucial role in maintaining the ecological balance by feeding on decayed corpses of wild and domestic animals. Large-scale culture and metagenomics studies have broadened our understanding of viral diversity in animals' gastrointestinal tracts. However, despite the importance of gut viral communities in regulating bacterial diversity and performing symbiotic functions, no gut viral study has been conducted on Himalayan vultures. Furthermore, the impact of captivity on the gut virome of these vultures remains unknown.

Methods: In this study, metagenomic sequencing methods targeting DNA of virus-like particles enriched from feces were used to characterize the gut DNA viromes of wild and captive Himalayan vultures.

Results: In total, 22,938 unique viral operational taxonomic units (vOTUs) were identified and assigned to 140 viral genera in 41 viral families. These families included viruses associated with bacteria, animals, plants, insects, and archaea. Phage communities, including Siphoviridae, Microviridae, Myoviridae, Inoviridae, and Herelleviridae, dominated the gut virome of Himalayan vultures. Wild vultures exhibited higher viral richness and diversity compared with those in captivity. The functional capacity of the gut virome was characterized by identifying 93 KEGG pathways, which were significantly enriched in metabolism and genetic information processing. Abundant auxiliary metabolic genes, such as carbohydrate-active enzyme, and antibiotic resistance genes, were also found in the vultures' gut virome.

Discussion: Our findings reveal the complex and diverse viral community present in the gut virome of Himalayan vultures, which varies between wild, and captive states. The DNA virome dataset establishes a baseline for the vultures' gut virome and will serve as a reference for future virus isolation and cultivation. Understanding the impact of captivity on the gut virome contributes to our knowledge of vultures' response to captivity and aids in optimizing their rehabilitation and implementing protective measures.

Keywords: Gyps himalayensis; conservation biology; high-throughput sequencing technology; phage; scavenger; viral metagenomics; zoo.

Figure 1

Map of the Qinghai-Tibetan Plateau with sampling sites (A), and the results of the completeness of viral Operational Taxonomic Units (vOTUs) evaluated by CheckV (B), and the Venn diagram of shared and unique vOTUs in the wild and zoo groups (C).

Figure 2

Viral community structural composition and distribution on the family and genus level. (A) Composition and hierarchical clustering of gut virome at the family level. (B) Boxplot shows the differential viral families when compared between the wild and zoo groups. (C) Composition and hierarchical clustering of gut virome at the genus level. (D) Boxplot shows the differential viral general when compared between the wild and zoo groups.

Figure 3

Comparison of alpha and beta diversity in gut DNA virome between the wild and zoo Himalayan vultures. (A) Boxplot shows the richness and Shannon index that differ between the two groups. (B) NMDS analysis based on Bray-Curtis distance between the two groups. The R² values and p-values of the adonis analysis are shown.

Figure 4

Comparison of DNA viral functions between the wild and zoo Himalayan vultures. (A) Composition of viral functional categories at the KEGG pathway level. (B) Boxplot shows the KEGG pathways that differed in abundance between the two groups. (C) Number of detected CAZymes. AA, auxiliary activity; CBM, carbohydrate-binding module; CE, carbohydrate esterase; GH, glycoside hydrolase; GT glycosyltransferase; PL, polysaccharide lyase. (D) Boxplot shows the CAZymes that differed in abundance between the two groups.

Figure 5

Host predictions of the viral Operational Taxonomic Units (vOTUs). (A) Stacked bar graph of the vOTUs classification at different taxonomic ranks by their assigned homology results of host information. (B) Boxplot shows the taxonomies that differed in abundance between the two groups. Sankey diagrams shows the connections between the top five viral families and their potential hosts in the wild group (C) and the zoo group (D).

Figure 6

Maximum likelihood phylogenetic tree of Siphoviridae (A), Microviridae (B), Baculoviridae (C), Phycodnaviridae (D). Virus names are followed by the GenBank accession numbers. The viral Operational Taxonomic Units (vOTUs) identified in the present study are marked with red color. The numbers beside the branches represent statistical confidence in clades based on 1,000 bootstrap replicates, only bootstrap values ≥90% are shown.