Viromics Analysis Of Tick Borne Viruses In Northeast China

Ticks,as blood-sucking ectoparasites,can carry a variety of pathogenic microorganisms including bacteria,fungi,viruses,causing disease to the infected people and animals.Tick borne viruses(TBVs)are typical arboviruses,with over 160 typical Tick borne viruses have been found.In recent years,a variety of novel TBVs have been reported,and it has been found that these viruses can cause human infection.Therefore,increasing the diversity background investigation of TBVs and analyzing their influencing factors is of great significance for the prevention and control of TBVs.Due to its special geographical environment,the Northeast provinces of China are natural habitats for a variety of ticks,as well as important focal points for Tick-borne viral diseases such as Tick-borne encephalitis,Severe fever with thrombocytopenia syndrome,and so on.To explore the overall situation of TBVs carried by ticks in the Northeastern provinces of China,deeply excavate the TBVs that can cause zoonotic diseases,and enrich the background data of TBVs in these regions,this study collected tick samples from 15 sampling points in the three Northeastern provinces of China from May to June 2021,including 3 in Liaoning province,7 in Jilin province,and 5 in Heilongjiang province.The collection environment was forest and grassland,and the developmental stages of ticks included larvae and adult ticks.Based on their morphology and habitat conditions,it can be inferred that they all did not suck blood.And in order to analyze the ecological factors that affect the diversity of TBVs,we included ticks infested on the surface of goats at three sampling sites in Hubei province from May to June 2016 as comparative groups.Multiple displacement amplification(MDA)and Meta-transcriptomics(MTT)were used to study the tick-borne DNA virome and tick-borne RNA virome.The viromic diversity and genetic characteristics of viral sequences of ticks were studied by a series of bioinformatics and related technologies.Tick species were identified by morphology and mitochondrial gene sequencing.A total of 1,391 ticks were collected from three genera and six species which included Haemaphysalis concinna,Haemaphysalis japonicas,Haemaphysalis longicornis,Ixodes persulcatus,Dermacentor nuttalli,Dermacentor silvarum,and the ticks collected in Hubei were Haemaphysalis longicornis,with 180 in total.In this study,a total of 37 tick viromes were established,including 20 DNA viromes and 17 RNA viromes.A total of 71,577,962 reads from 37 tick viromes were annotated as Eukaryotic viruses.These Eukaryotic viruses are distributed in 19 known viral families and some viruses not classified into specific viral families,including single-stranded circular DNA(sscDNA)viruses,double stranded RNA(dsRNA)viruses,ssRNA(-)viruses,and ssRNA(+)viruses.The natural hosts of these viruses include arthropods,vertebrates,plants and fungi and etc.DNA viromic data clustering based on K-mer distance showed that there was no tick species specificity in DNA virome.Phylogenetic analyses revealed that these tick-borne CRESS DNA viruses assembled from DNA viromes were related to viruses recovered from animal feces,tissues and even environmental samples,suggesting that their presence may be largely explained by environmental factors rather than by tick species and host blood meals.Viral families with higher abundance in RNA viromes include Nairoviridae,Phenuiviridae and Rhabdoviridae.Based on the annotation results of RNA virus species,alpha and beta diversity analysis were conducted.The results showed that Haemaphysalis concinna and Ixodes persulcatus had high alpha diversity,the RNA virus communities of Ixodes persulcatus and Haemaphysalis longicornis had certain differences from other tick species,that is,the two ticks had high beta diversity,and there was no significant difference in the above analysis results.Even if the ticks in the control group were engorged ticks,compared with the free ticks in the Northeastern provinces,there were fewer RNA virus species and lower abundance.The Nairovirus,Phenuivirus and Orthomyxovirus analyzed in this study showed wide amino acid identities with the known related TBVs:(1)The L fragment of Songling virus strain HCYJ01 was amplified from Haemaphysalis concinna in Jilin province,and its amino acid identity with that of Songling virus strain YC585 in Heilongjiang province reached 99.06%,belonging to different mutant strains of the same viral species.(2)The full-length sequences of L and S fragments of Nairo-like virus IPLJ001 obtained by amplification,the amino acid identity of its L fragment with all members of Nairo-like viruses group was 58.6%-92.7%,and it belongs to a mutant strain of Nairo-like viruses group.(3)In this study,five sequences were amplified and annotated to different fragments of Orthomyxovirus,of which four were approximately full length and one was partial sequence.These sequences were 31.4%-84.2%identity with the amino acid of known Orthomyxovirus.Combined with the results of phylogenetic analysis,the viral sequences obtained in this study belong to the new virus sequences in the family Orthomyxoviridae Quaranjavirus.The above studies have showed the composition and distribution of TBVs in some regions of the Northeast provinces,analyzed the diversity of tick-borne DNA virome and RNA virome respectively.And revealed the genetic diversity of some TBVs and the situation of new viral species through phylogenetic analysis.The research results have enriched the background data of TBVs in these regions,and have important reference value for preventing and controlling the epidemic and transmission of TBVs.