[1]杨晓宇,徐雷,殷鑫欢,等.猪非典型性瘟病毒与猪流行性腹泻病毒双重PCR方法的建立与应用[J].江苏农业学报,2018,(05):1081-1086.[doi:doi:10.3969/j.issn.1000-4440.2018.05.016]
　YANG Xiao-yu,XU Lei,YIN Xin-huan,et al.Establishment and application of double PCR method between atypical porcine pestivirus and porcine epidemic diarrhea virus[J].,2018,(05):1081-1086.[doi:doi:10.3969/j.issn.1000-4440.2018.05.016]
点击复制

猪非典型性瘟病毒与猪流行性腹泻病毒双重PCR方法的建立与应用()
分享到：

江苏农业学报[ISSN:1006-6977/CN:61-1281/TN]

卷:
期数:
2018年05期
页码:
1081-1086
栏目:
畜牧兽医·水产养殖
出版日期:
2018-10-25

文章信息/Info

Title:
Establishment and application of double PCR method between atypical porcine pestivirus and porcine epidemic diarrhea virus
作者:
杨晓宇1; 徐雷1; 殷鑫欢1; 张继宗1; 徐志文1; 2; 朱玲1; 2
（1.四川农业大学动物医学院，四川成都611130；2.四川农业大学动物疫病与人类健康四川省重点实验室，四川成都611130）
Author(s):
YANG Xiao-yu1; XU Lei1; YIN Xin-huan1; ZHANG Ji-zong1; XU Zhi-wen1; 2; ZHU Ling1; 2
(1.College of Veterinary Medicine, Sichuan Agricultural University, Chengdu 611130, China;2.Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Chengdu 611130, China)
关键词:
猪非典型性瘟病毒; 猪流行性腹泻病毒; 双重RT-PCR
Keywords:
atypical porcine pestivirus; porcine epidemic diarrhea virus; double RT-PCR
分类号:
S852.65+1
DOI:
doi:10.3969/j.issn.1000-4440.2018.05.016
文献标志码:
A
摘要:
为了快速准确地检测出猪非典型性瘟病毒（APPV）与猪流行性腹泻病毒（PEDV），根据 GenBank 中发布的PEDV S2基因和APPV NS3基因的序列，分别选取PEDV S2基因和APPV NS3基因的保守序列，设计、合成了1对特异性引物。通过对体系进行优化，分别扩增出190 bp和500 bp的特异性片段。建立的双重RT-PCR方法对猪繁殖与呼吸综合征病毒（PRRSV）、猪瘟病毒（CSFV）、猪轮状病毒（RV）的检测均为阴性。对PEDV和APPV的最低检出量分别为1 μl 2.65×105和1 μl 3.56×104，对四川遂宁、眉山等地采集到的腹泻、肌肉震颤的病料进行检测，PEDV、APPV阳性病料检出率分别为41.18%和11.76%，经分别与特异性检测APPV和PEDV的单一RT-PCR法检测结果进行比较分析，符合率均为100%。与单一RT-PCR检测方法具有相同特异性和重复性，可用于临床检测。
Abstract:
In order to detect the atypical porcine pestivirus (APPV) and porcine epidemic diarrhea virus (PEDV) rapidly and accurately, according to the conservative sequences of PEDV S2 gene and APPV NS3 gene deposited in GenBank, a pair of specific primer was designed and synthesized. By optimizing the system, specific fragments of 190 bp and 500 bp were amplified, respectively. The detection results of porcine reproductive and respiratory syndrome virus (PRRSV), classical swine fever virus (CSFV) and porcine rotavirus (RV) using this double RT-PCR method were negative. The minimum detectable amount of PEDV and APPV were 1 μl 2.65×105 and 1 μl 3.56×104, respectively. The detection rates for diarrhea and muscular tremor collected from Suining and Meishan in Sichuan province were 41.18% and 11.76%, respectively. The coincidence rates of the results detected by double RT-PCR method and the specific RT-PCR method for APPV and PEDV were all 100%. The double RT-PCR method has the same sensitivity, specificity and repeatability as the single RT-PCR assay and can be used for clinical detection.

参考文献/References:

[1]DONE J T, WOOLLEY J, UPCOTT D H, et al. Porcine congenital tremor type AII: Spinal cord morphometry[J]. British Veterinary Journal,1986, 142：145-150.
[2]DONE J T, HARDING J D. Congenital tremor in pigs (trembling disease of piglets): lesions and causes[J]. Dtsch Tierarztl Wochenschr,1967, 74：333-336.
[3]BLAKEMORE W F, HARDING J D, DONE J T. Ultrastructural observations on the spinal cord of a Landrace pig with congenital tremor type AIII[J]. Research in Veterinary Science, 1974, 17：174-178.
[4]PATTERSON D S, SWEASEY D, BRUSH P J, et al. Neurochemistry of the spinal cord in British Saddleback piglets affected with congenital tremor, type A-IV, a second form of hereditary cerebrospinal hypomyelinogenesis[J]. Journal of Neurochemistry,1973, 21：397-406.
[5]KNOX B, ASKAA J, BASSE A, et al. Congenital ataxia and tremor with cerebellar hypoplasia in piglets borne by sows treated with Neguvon vet. (metrifonate, trichlorfon) during pregnancy[J]. Nordisk Veterinaermedicin,1978, 30：538-545.
[6]DONE J T, WOOLEY J, UPCOTT D H, et al. Porcine congenital tremor type AI: Spinal cord morphometry[J]. British Veterinary Journal,1986,142 (2):145-150.
[7]HAUSE B. Discovery of a novel putative atypical porcine pestivirus in pigs in the United States[J]. Journal of General Virology,2015,96(10):2994.
[8]POSTEL A, HANSMANN F, BAECHLEIN C, et al .Presence of atypical porcine pestiviru（APPV）genomes in new-born piglets correlates with congenital tremor[J]. Sci Rep,2016,6：27735.
[9]杨春华，王强，王毅，等．猪流行性腹泻的诊断与科学防治[J]，畜牧与饲料科学，2009(7):34-36.
[10]杨云乔，郑建高，姜军华，等．复方中药治疗流行性腹泻病毒感染猪的消化道黏膜电镜观察及相关酶检测[J]. 江苏农业科学，2017,45(17):159-163.
[11]SUN R Q, CAI R J, CHEN Y Q， et al. [J]. Emerging Infectious Diseases, 2012 , 18 (1) :161-163.
[12]JUNG K, SAIF L J. Porcine epidemic diarrhea virus infection:Etiology,epidemiology，pathogenesis and immunoprophylaxis[J]. Veterinary J,2015,204(2):134-143.
[13]KLOBASA F, WERHAHN E, BUTLER J E. Composition of sow milk during lactation[J]. J Animal Sci,1987，64(5):1458-1466.
[14]WESLEY R D, LAGER K M. Increased litter survival rates，reduced clinical illness and better lactogenic immunity against TGEV in gilts that were primed as neonates with porcine respiratory coronavirus (PRCV)[J]. Veterinary Microbiol,2003,95(3):175-186.
[15]YUAN J, HAN Z, LI J, et al .Atypical porcine pestivirus as a novel type of pestivirus in pigs in China[J]. Front Microbiol，2017,8:862.
[16]SCHWARZ L, RIEDEL C, HOGLER S, et al.Congenital infection with atypical porcine pestivirus (APPV) is associated with disease and viral persistence[J]. Vet Res，2017,48(1)：1.

相似文献/References:

[1]祁光宇,刘斌,赵兴绪.猪流行性腹泻病毒HB2015分离株N基因表达及单克隆抗体的制备[J].江苏农业学报,2018,(01):76.[doi:doi:10.3969/j.issn.1000-4440.2018.01.011]
　QI Guang-yu,LIU Bin,ZHAO Xing-xu.Expression of N gene of porcine epidemic diarrhea virus（PEDV） HB2015 strain and preparation of monoclonal antibodies against PEDV[J].,2018,(05):76.[doi:doi:10.3969/j.issn.1000-4440.2018.01.011]
[2]郭佳琪,杨晓宇,冯宇,等.猪非典型性瘟病毒RT-PCR检测方法的建立及初步应用[J].江苏农业学报,2019,(02):357.[doi:doi:10.3969/j.issn.1000-4440.2019.02.016]
　GUO Jia-qi,YANG Xiao-yu,FENG Yu,et al.Establishment and preliminary application of RT-PCR detection method for atypical porcine pestivirus[J].,2019,(05):357.[doi:doi:10.3969/j.issn.1000-4440.2019.02.016]
[3]丁芳艺,宋海鑫,梁荣,等.猪流行性腹泻病毒感染对猪小肠上皮细胞I型干扰素产生通路的影响[J].江苏农业学报,2021,(01):99.[doi:doi:10.3969/j.issn.1000-4440.2021.01.013]
　DING Fang-yi,SONG Hai-xin,LIANG Rong,et al.Effects of porcine epidemic diarrhea virus infection on IFN-β production pathway in porcine intestinal epithelial cells[J].,2021,(05):99.[doi:doi:10.3969/j.issn.1000-4440.2021.01.013]

备注/Memo

备注/Memo:
收稿日期：2018-01-12 基金项目：四川省科技支撑计划项目（2017NZ0038）；四川省“十三五”育种攻关项目（2016NYZ0052）；国家“十二五”科技支撑计划项目（2015BAD12B04-2.3） 作者简介：杨晓宇（1993-），男，山东烟台人，硕士研究生，主要从事动物传染病病原分子生物学研究。（E-mail）1491618529@qq.com 通讯作者：朱玲，（Tel）08352885846；（E-mail）abtcxzl72@126.com
更新日期/Last Update: 2018-11-05