猪捷申病毒5型VP1蛋白的原核表达及间接ELISA检测方法的建立

doi:10.11843/j.issn.0366-6964.2025.02.036

Abstract

Abstract:

The aim of this study was to establish an indirect ELISA method for the detection of porcine teschovirus (PTV) type 5 and to provide a material reserve for its prevention and control. PTV is an acute virulent pathogen of pigs, which is characterized by high morbidity and mortality. The clinical manifestations include encephalitis, pneumonia, reproductive disorders, myocarditis, and diarrhea. PTV can be divided into various subtypes. The clinical symptoms vary from each other. Therefore, lacking of efficient and specific serological detection methods blocks the control and prevention of PTV-5. This study inserted the VP1 gene of PTV-5 into pET-30a plasmid and obtained the VP1 protein. After optimizing the conditions, an indirect ELISA method against the VP1 of PTV-5 was successfully established. The method has no cross-reactivity with the standard positive sera of common porcine viruses. The lowest detection dilution was 1∶3 200, and the coefficients of variation were less than 10% both intra-and inter-batch, showing good specificity, sensitivity, and reproducibility of the method. A total of 279 clinical samples collected from the northeast were detected by this method. The results showed that the positive rate was 67.74%, which was in consistent with previous report. The indirect ELISA assay established in this study has a good overall evaluation and provides technical support for the clinical detection of PTV-5.

Key words: porcine teschovirus, VP1, prokaryotic expression, indirect ELISA

CLC Number:

S855.3

SHAO Yongheng, NI Minting, GAO Mengling, TANG Jiao, ZHANG Gengxin, LIN Shengyu, LIU Guangliang, CHEN Jianing, WANG Wenhui. Prokaryotic Expression of VP1 Protein to Porcine Teschovirus Type 5 and the Establishment of an Indirect ELISA Detection Method[J]. Acta Veterinaria et Zootechnica Sinica, 2025, 56(2): 883-889.

Figures/Tables 4

References 17

1	申识川, 王文芳, 王冠杰. 猪捷申病的流行、诊断与防治[J]. 养殖与饲料, 2019, (12): 94- 96.
	SHEN S C , WANG W F , WANG G J . Epidemiology, diagnosis and control of porcine teschovirus[J]. Animals Breeding and Feed, 2019, (12): 94- 96.
2	张晓杰, 刘业兵, 汤德元, 等. 猪捷申病毒HB株的分离与全基因序列分析[J]. 畜牧兽医学报, 2012, 43 (6): 937- 942.
	ZHANG X J , LIU Y B , TANG D Y , et al. Isolation and sequence analysis of complete genome of porcine teschovirus HB strain[J]. Acta Veterinaria et Zootechnica Sinica, 2012, 43 (6): 937- 942.
3	胡峰, 刘杉杉, 蔺文成, 等. 猪捷申病毒8型rVP1-ELISA检测方法的建立[J]. 中国兽医科学, 2012, 42 (3): 258- 263.
	HU F , LIU S S , LIN W C , et al. Establishment of an indirect ELISA based on VP1 gene of porcine teschovirus type 8[J]. Chinese Veterinary Science, 2012, 42 (3): 258- 263.
4	王丽瑄, 周群, 付能胜, 等. 藏猪源捷申病毒的检测和遗传演化分析[J]. 畜牧兽医学报, 2021, 52 (1): 185- 194.
	WANG L X , ZHOU Q , FU N S , et al. Detection and genetic evolution of teschovirus of tibetan pig origin in Sichuan[J]. Acta Veterinaria et Zootechnica Sinica, 2021, 52 (1): 185- 194.
5	VILLANOVA F , CUI S J , AI X , et al. Analysis of full-length genomes of porcine teschovirus (PTV) and the effect of purifying selection on phylogenetic trees[J]. Arch Virol, 2016, 161 (5): 1199- 1208. doi: 10.1007/s00705-015-2744-0
6	ZELL R , SEITZ S , HENKE A , et al. Linkage map of protein-protein interactions of Porcine teschovirus[J]. J Gen Virol, 2005, 86 (Pt 10): 2763- 2768.
7	ZELL R , DELWART E , GORBALENYA A E , et al. ICTV virus taxonomy profile: Picornaviridae[J]. J Gen Virol, 2017, 98 (10): 2421- 2422. doi: 10.1099/jgv.0.000911
8	KIRKEGAARD K . Mutations in VP1 of poliovirus specifically affect both encapsidation and release of viral RNA[J]. J Virol, 1990, 64 (1): 195- 206. doi: 10.1128/jvi.64.1.195-206.1990
9	SCHMIDTKE M , SELINKA H C , HEIM A , et al. Attachment of coxsackievirus B3 variants to various cell lines: mapping of phenotypic differences to capsid protein VP1[J]. Virology, 2000, 275 (1): 77- 88. doi: 10.1006/viro.2000.0485
10	刘芹防, 刘杉杉, 张超范, 等. 检测猪捷申病毒抗体的间接免疫荧光方法的建立与初步应用[J]. 中国预防兽医学报, 2011, 33 (1): 70- 72.
	LIU Q F , LIU S S , ZHANG C F , et al. Establishment of the indirect immunofluorescence assay for detecting the antibody of porcine teschovirus[J]. Chinese Journal of Preventive Veterinary Medicine, 2011, 33 (1): 70- 72.
11	庄金秋, 梅建国, 李峰, 等. 猪捷申病毒检测方法研究进展[J]. 动物医学进展, 2013, 34 (7): 80- 83.
	ZHUANG J Q , MEI J G , LI F , et al. Progress on detection methods of porcine teschovirus[J]. Progress in Veterinary Medicine, 2013, 34 (7): 80- 83.
12	BANGARI D S , POGRANICHNIY R M , GILLESPIE T , et al. Genotyping of Porcine teschovirus from nervous tissue of pigs with and without polioencephalomyelitis in Indiana[J]. J Vet Diagn Invest, 2010, 22 (4): 594- 597. doi: 10.1177/104063871002200415
13	MALIK Y S, BHAT S, VLASOVA A N, et al. Teschovirus[M]//MALIK Y S, SINGH R K, PAL YADAV M. Emerging and Transboundary Animal Viruses. Singapore: Springer, 2020: 123-136.
14	QIU Z , WANG Z T , ZHANG B K , et al. The prevalence of porcine teschovirus in the pig population in northeast of China[J]. J Virol Methods, 2013, 193 (1): 209- 214. doi: 10.1016/j.jviromet.2013.06.005
15	KNOWLES N J, REUTER G. Porcine caliciviruses[M]//Diseases of Swine. 10th ed. New York: Wiley, 2012: 493-500.
16	BOVA T O , DEREV'IANKO S V , SOROKA V I . Express method for diagnostics of enzootic encephalomyelitis (Teschen disease) in pigs[J]. Mikrobiol Z, 2005, 67 (6): 49- 56.
17	王瑶, 田志军, 王斌, 等. 一株猪捷申病毒的分离鉴定及血清学初步调查[J]. 黑龙江畜牧兽医, 2011, (2): 14- 17.
	WANG Y , TIAN Z J , WANG B , et al. Isolation and identification of porcine teschovirus and preliminary serological survey[J]. Heilongjiang Animal Science and Veterinary Medicine, 2011, (2): 14- 17.

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Prokaryotic Expression of VP1 Protein to Porcine Teschovirus Type 5 and the Establishment of an Indirect ELISA Detection Method

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