Identification Of The Intestinal Virome Of Rats Using Viral Metagenomics

In recent years, the health of human and animals was seriously threatened by emerging and re-emerging viruses, which has become a public problem. A number of human viral infectious diseases were caused by viruses from animals. Rats contact closely with human and carry a large number of pathogenic viruses such as Hemorrhagic Fever With Renal Syndrome(HFRS)caused by Hantavirus, the viruses of the family of Arenaviridae could lead the disease of human Lassa fever, acute gastroenteritis caused by Norwalk virus, hantavirus pulmonary syndrome(HPS)caused by Sin Nombre virus(SNV) and so on.Therefore, monitoring and recognizing the situation of rodents carrying virus is very important for the control, prevention and curing viral infection. Traditional technology and methods have large limitation in the discovery of new and divergent viruses which are present in emerging infectious diseases. The viral metagenomics combining high-throughput sequencing technology can analyze all the viral nucleotide in the samples directly. Using viral metagenomics, we could identify both known viruses and new viruses effectively. This method provides technical support for exploring new virus and diagnosing new viral infectious disease. The present study analyzed the composition of virus population and discovered new virus from 40 intestinal samples of rats which were collected from Taizhou city, Jiangsu Province, China.Results : 1. The composition of intestinal virus community of ratsResults show that the intestinal virus community of rats include: Adenoviridae, Anelloviridae, Bunyaviridae, Caliciviridae, Flaviviridae, Coronaviridae, Dicistroviridae, Herpesviridae, HERV, Microviridae, Papillomaviridae, Parvoviridae, Phycodnaviridae, Picornaviridae, Poxviridae, Retroviridae, Totiviridae, Virgaviridae, Circoviridae, Astroviridae, viruses without known family and other viruses. Virus community analysis indicated:(1) Retroviridae accounts more than 50% of virus population in the four pools.(2) pools no. TRat07, TRat08 and TRat09 contain Caliciviridae and Adenoviridae. Pool no. TRat07 contains more sequences than TRat08 and TRat09.(3) All the pools contain more than two viruses which can infect mammal animals.2.Novel viruses of rats’ fecal virus community(1) GemycircularvirusIn the present study, a novel gemycircularvirus was identified from the intestinal contents of rats. We designed a set of nest primers based on the Rep gene sequence of the gemycircularviruses to screen 50 blood samples from experimental rats captured in five laboratories in China, results indicated that 4 of the 50 blood samples were positive for gemycircularvirus. Sequence analysis indicated the four 361 bp sequences shared 100 % identity with each other. One of the positive samples was randomly selected to determine the complete genome, a novel gemycircularvirus named Chzjrat-01 from blood samples of experimental rats was characterized. The novel gemycircularvirus encodes two major proteins, including a capsid protein(Cap) and a replication-associated protein(Rep). Phylogenetic analysis based on the amino acid sequence of Rep indicated that Ch-zjrat-01 clusters with two gemycircularviruses discovered from bird(KF371635) and mosquito(HQ335086), sharing 48.7 and 49.4 % sequence identities with them, respectively.(2) Bufavirus In this study, a novel protoparvovirus, named Rat Bu V, was identified from the intestinal contents of rats using viral metagenomics. The near complete genome was 4643 nt encoding NS1, VP1 and VP2 proteins. Phylogenetic analysis based on the near complete genome indicated that Rat Bu V clustered with Mpulungu Bu V from shrews forming a separate clade. Sequence analysis over the amino acid sequences of NS1, VP1 and VP2 of Rat Bu V shared identities of 50.6-77.2%, 48.3-77.3%, and 47.1-78.3%, respectively with those of human Bu Vs, Mp Bu V, and WUHARV parvovirus, suggesting Rat Bu V belongs to a new species of protoparvovirus. Our epidemiologic study indicated that the prevalence rate of Rat Bu V in the cohort of 40 wild rats is 12.5%(5/40), which is higher than that of Bu V in humans in a previous study. Primers was designed to screen the 40 rat fecal samples, results indicated that 5 samples were positive with the positive rate of 12.5%. The five 379 bp sequences of the positive samples shared >98% nucleotide identities, suggesting a single strain virus was present in the rats in the area.Conclusion:(1) Using viral metagenomics method, the composition of intestinal virus community from rats is characterized.(2) The composition of intestinal virus community from rats is complicated.(3) A novel Gemycircularvirus and a novel Bufavirus were identified from the intestinal contents of rats.