Analysis Of Diarrhea Virus Diversity In Newborn Piglets In Guangdong And Isolation And Identification Of Porcine Teschovirus

Viral diarrhoea in piglets is characterised by acute diarrhoea,vomiting and severe dehydration,with high morbidity and mortality,causing serious economic losses to the global pig industry.The main viruses that cause diarrhoea in piglets are:TGEV,RVA,PEDV,PDCo V,PKV,PTV and among others.Viral diarrhoea in piglets is usually a mixed infection with multiple pathogens,and these viruses are prone to genetic mutation or recombination,while new pathogens causing piglet diarrhoea have emerged in recent years,bringing new challenges to the prevention and control of piglet diarrhoea.In this study,43 piglet diarrhoea samples from 11 pig farms in 8 regions of Guangdong Province were subjected to second-generation high-throughput sequencing and viral diversity analysis using viral-metagenomics;techniques.10.78Gb of data were measured,and the results showed that the reads were annotated to seven known viral families other than phage,distributed in ss RNA,ss DNA and ds DNA,with the coronaviridae accounting for the largest proportion（57.8%）.The viral reads were spliced and sequences of 12 viruses were detected by comparison of the spliced sequences with the non-redundant reference protein database（nr）.Seven viruses,including PEDV,PDCo V,TGEV,PKV,PAst V,PBo V and PTV,which were identified by viral-metagenomics analysis,were selected for validation by q RT-PCR,and the S1 gene was cloned and sequenced from PEDV-positive samples to analyse the genetic evolution and variation of the PEDV S1 gene.The results showed that all seven viruses were detected,with PEDV detected at 95.3%（41/43）,PAst V detected at16.2%（7/43）,PDCo V detected at 13.9%（6/43）,PKV detected at 9.3%（4/43）,PBo V detected at 5.65%（2/43）and TGEV The detection rate was 5.65%（2/43）and the detection rate of PTV was 2.32%（1/43）.Amplification,cloning and sequencing of the S1 gene in 41 PEDV-positive samples revealed that these strains were all mutant strains with one 9 amino acid site variation;3D structure comparison revealed that the9 amino acid mutations caused changes in the spatial structure of 6 regions of the NTD and COE regions of the S1 protein.The isolation of porcine teschovirus was carried out using ST cells on porcine teschovirus-positive samples.IFA identification,TCID₅₀determination and the full genome sequence of the virus were also obtained.The results show that the PTV was successfully isolated and identified,the full genome sequence was obtained and genetic evolutionary analysis showed that the strain is type 2 with a TCID₅₀=10^7.25/0.1 m L.In this study,a systematic analysis of the viruses carried by piglets with diarrhoea in Guangdong was conducted using viral-metagenomics technology,which deepened the understanding of the diversity of piglet diarrhoea viruses in Guangdong;at the same time,a 9-amino acid mutation PEDV was found to be prevalent in Guangdong,providing an early warning for the prevention of PEDV;the first PTV strain in Guangdong was isolated from piglets with diarrhoea,laying the foundation for further research on the pathogenesis and prevention and control measures.