畜牧兽医学报2024，Vol.55Issue(7)：3049-3063,15.DOI:10.11843/j.issn.0366-6964.2024.07.024
一株猪流行性腹泻病毒强毒株的分离与鉴定
Isolation and Identification of a Highly Pathogenic Strain of Porcine Epidemic Diarrhea Virus
摘要
Abstract
To understand the pathogenic characteristics and pathogenicity of the latest epidemic strain of porcine epidemic diarrhea virus(PEDV),isolation and identification of PEDV were conducted this study.PEDV positive samples were inoculated into Vero cells for continuous transmission for isolating PEDV epidemic strains.Then the culture characteristics,whole genome sequence of the isolated strains were analyzed,and their pathogenicity in piglets were evaluated.Results showed that a PEDV strain(named CHN-SC2021)was isolated from the 3rd passage(P3)Vero cells of continuous inoculation of PEDV positive sample.Under the electron microscope,the diameter of the virus was about 60-110 nm,forming classic"syncytium"on the cell;The virus titer from P10 to P35 generations increased from 1047 TCID50 mL-1 to 106.43 TCID50·mL-1;The whole genome sequence of CHN-SC2021(P5)(GenBank accession number:OM505025.1)is 28 014 bp,and its nucleotide sequence homology about 99.16％-96.32％,99.01％-96.57％and 98.94％-96.33％compared with the reference strains from China,Europe and America,and other Asian countries,respectively;The phylogenetic analysis showed that the CHN-SC2021 strain is closely related to Jiangxi CH/JXJA/2017(MF375374.1),Henan CH/HNAY/2015(KT199103.1),and CH/HNKF-16(KY649107.1)strains,belonging to the G Ⅱ a genotype type.The similarity between the S gene sequence and other strains ranges from 98.7％to 93.4％.Oral infection of 6-day-old suckling piglets with 3 mL of 105 TCID50·mL-1 per pig of CHN-SC2021 virus solution resulted in loss of appetite at 12 hours,vomiting at 20 hours,and severe vomiting and yellow watery diarrhea at 24 hours;The main pathological changes during dissection are gastrointestinal distension,transparent jejunum and ileum;Histopathological changes showed that each segment of the intestine had varying degrees of intestinal villus lesions,shedding,loose connective tissue,and widened space between the muscularis mucosae and the muscle layer;Immunohistochemistry showed that viruses were present in all segments of the small intestine,with the most viruses present in the ileum.In conclusion,a genotype G Ⅱ a PEDV virulent strain was isolated,which will contribute to enriching the information of PEDV circulating strains in China and be helpful for clinical prevention and control of PED.关键词
猪流行性腹泻病毒/分离/鉴定/序列分析/哺乳仔猪/致病性Key words
porcine epidemic diarrhea virus/isolation/identification/sequence analysis/sucking piglets/pathogenicity分类
农业科技引用本文复制引用
刘维哲,罗成刚,袁蓉,廖艺杰,文艺悯,孙莹,俞恩波,曹三杰,黄小波..一株猪流行性腹泻病毒强毒株的分离与鉴定[J].畜牧兽医学报,2024,55(7):3049-3063,15.基金项目
四川省"十四五"川猪重大科技专项(2021ZDZX0010) （2021ZDZX0010）
成华猪保护基地生物危害防控经费项目(ZZSS-2024-193) （ZZSS-2024-193）
云南省重大科技专项计划(202102AE090039) （202102AE090039）
四川省产教融合示范项目"饲料工业全产业链转型升级产教融合创新示范" （）
