华北农学报
華北農學報
화북농학보
ACTA AGRICULTURAE BOREALI-SINICA
2014年
1期
232-238
,共7页
王小敏%何孔旺%周忠涛%茅爱华%俞正玉%汪伟%倪艳秀%温立斌%张雪寒%郭容利%李彬%于洋
王小敏%何孔旺%週忠濤%茅愛華%俞正玉%汪偉%倪豔秀%溫立斌%張雪寒%郭容利%李彬%于洋
왕소민%하공왕%주충도%모애화%유정옥%왕위%예염수%온립빈%장설한%곽용리%리빈%우양
猪繁殖与呼吸综合征病毒%分离与鉴定%遗传变异
豬繁殖與呼吸綜閤徵病毒%分離與鑒定%遺傳變異
저번식여호흡종합정병독%분리여감정%유전변이
Porcine reproductive and respiratory syndrome virus( PRRSV)%Isolation and identification%Genet-ic variation
从江苏某猪繁殖与呼吸综合征( PRRS)发病猪场采集的血清样本中分离到1株猪繁殖与呼吸综合征病毒( PRRSV)毒株,对该毒株的生物学特性进行了鉴定,并对其Nsp2和ORF5序列进行测定和序列比对分析。利用细胞传代的方法对PRRSV检测阳性,PPV和PRV检测阴性的猪血清进行病毒分离。通过RT-PCR、CPE和IFA对PRRSV分离毒株进行鉴定,并测定其TCID50。测序获得Nsp2和ORF5基因,并利用DNAStar软件进行序列同源性和进化分析。成功分离获得1株PRRSV毒株,该毒株能够在Marc-145细胞上增殖并产生细胞病变,免疫荧光检测可观察到特异性荧光,TCID50为105.5。 Nsp2序列分析表明,该分离株具有HP-PRRSV 30个氨基酸缺失的基因特征,同时495~516位也存在缺失。分离株ORF5基因与其他参考毒株氨基酸同源性为57.1%~99.5%,与NJ-1106的同源性最高,为99.5%。系统进化树分析表明,该分离株属于北美型,与WUH4、NJ-1106等HP-PRRSV毒株属于同一分支。本试验分离得到一株HP-PRRSV变异毒株,为分析我国PRRSV的遗传变异和防控奠定了基础。
從江囌某豬繁殖與呼吸綜閤徵( PRRS)髮病豬場採集的血清樣本中分離到1株豬繁殖與呼吸綜閤徵病毒( PRRSV)毒株,對該毒株的生物學特性進行瞭鑒定,併對其Nsp2和ORF5序列進行測定和序列比對分析。利用細胞傳代的方法對PRRSV檢測暘性,PPV和PRV檢測陰性的豬血清進行病毒分離。通過RT-PCR、CPE和IFA對PRRSV分離毒株進行鑒定,併測定其TCID50。測序穫得Nsp2和ORF5基因,併利用DNAStar軟件進行序列同源性和進化分析。成功分離穫得1株PRRSV毒株,該毒株能夠在Marc-145細胞上增殖併產生細胞病變,免疫熒光檢測可觀察到特異性熒光,TCID50為105.5。 Nsp2序列分析錶明,該分離株具有HP-PRRSV 30箇氨基痠缺失的基因特徵,同時495~516位也存在缺失。分離株ORF5基因與其他參攷毒株氨基痠同源性為57.1%~99.5%,與NJ-1106的同源性最高,為99.5%。繫統進化樹分析錶明,該分離株屬于北美型,與WUH4、NJ-1106等HP-PRRSV毒株屬于同一分支。本試驗分離得到一株HP-PRRSV變異毒株,為分析我國PRRSV的遺傳變異和防控奠定瞭基礎。
종강소모저번식여호흡종합정( PRRS)발병저장채집적혈청양본중분리도1주저번식여호흡종합정병독( PRRSV)독주,대해독주적생물학특성진행료감정,병대기Nsp2화ORF5서렬진행측정화서렬비대분석。이용세포전대적방법대PRRSV검측양성,PPV화PRV검측음성적저혈청진행병독분리。통과RT-PCR、CPE화IFA대PRRSV분리독주진행감정,병측정기TCID50。측서획득Nsp2화ORF5기인,병이용DNAStar연건진행서렬동원성화진화분석。성공분리획득1주PRRSV독주,해독주능구재Marc-145세포상증식병산생세포병변,면역형광검측가관찰도특이성형광,TCID50위105.5。 Nsp2서렬분석표명,해분리주구유HP-PRRSV 30개안기산결실적기인특정,동시495~516위야존재결실。분리주ORF5기인여기타삼고독주안기산동원성위57.1%~99.5%,여NJ-1106적동원성최고,위99.5%。계통진화수분석표명,해분리주속우북미형,여WUH4、NJ-1106등HP-PRRSV독주속우동일분지。본시험분리득도일주HP-PRRSV변이독주,위분석아국PRRSV적유전변이화방공전정료기출。
A porcine reproductive and respiratory syndrome virus( PRRSV) strain was isolated from the serum of a PRRSV infected pig in Jiangsu Province. The biological characteristics of this isolate was identified and the ge-nome of Nsp2 and ORF5 were determined and analyzed. The serum which were detected PRRSV positive and PPV, PRV negative was inoculated and passaged on Marc-145 cells. This isolate was identified by RT-PCR and CPE,and confirmed by immunofluorescence assay(IFA). The TCID50 of this isolate was measured and the genome of Nsp2 and ORF5 were sequenced and its homology and evolutionary relationship were analyzed by DNAStar software. The results indicated that the PRRSV strain was successfully isolated and it could induce cytopathogenic effect( CPE) on Marc-145 cells. The specific fluorescence could be observed and the TCID50 of this strain was 105. 5 . Comparison a-nalysis of the Nsp2 genome of the isolate 2010 JS02 and other strains revealed that the isolate has a discontinuous deletion of 30 amino acids and another deletion at the position of 495-516 amino acid′s site. According to the se-quences analysis of GP5,the isolate 2010JS02 showed identity of 57. 1%-99. 5% among other strains,a high iden-tity of 99 . 5% with NJ-1106 . The phylogenetic tree was combined and showed that 2010 JS02 belonged to NA and classified into HP-PRRSV like WHU4, NJ-1106 and other HP-PRRSV strains. We successfully isolated a HP-PRRSV variant and named it as 2010 JS02 . These results provided foundations for further studying the genetic varia-tion and the prevention of PRRSV.
