FREE SHIPPING over $40
OVER 9000 FIVE-STAR REVIEWS
5% OFF ALL SUBSCRIPTIONS
100% HAPPINESS GUARANTEE
Analyze Diet
0
Home / Equine Research Bank / Front Microbiol / Development and application of an indirect ELISA for detecti...
Frontiers in microbiology2022; 13; 1029444; doi: 10.3389/fmicb.2022.1029444

Development and application of an indirect ELISA for detecting equine IgG antibodies against Getah virus with recombinant E2 domain protein.

Abstract: Getah virus (GETV) disease is a mosquito-borne infectious disease that causes fever, aseptic meningitis, and abortion in a variety of animals. Currently, the epidemic trend of GETV disease increases seriously worldwide, especially in China, posing a potential threat to animal safety and public health. However, there are few reports about the epidemiological investigation of GETV disease in China as well as a lack of commercial diagnostic kit for GETV antibody. Therefore, the establishment of a rapid, sensitive and suitable GETV antibody detection method for large-scale samples is an urgent request to fully understand the prevalence of GETV disease. Here, a recombinant plasmid pET22b-GETV-E2d that contained the domain of GETV-E2 (E2d) fused to His-tag was constructed to express recombinant protein E2d (rE2d) in . The rE2d was mainly expressed in inclusion bodies. And it was purified successfully by nickel affinity column so that it could be used to develop an indirect ELISA (rE2d-ELISA). After optimizing reaction conditions of rE2d-ELISA, the cut-off value was determined as 0.396 with 100 equine sera tested by virus neutralization test (VNT). Furthermore, rE2d-ELISA method showed the positive rate of IgG antibodies against GETV was 54.3% based on testing 646 clinical serum samples obtained in Xinjiang whereas the overall coincidence rate between rE2d-ELISA and VNT was 94.0%, with 98.2% sensitivity and 92.6% specificity. The findings suggest that the developed IgG ELISA employing recombinant E2d promises was an efficient and low-cost type of antibody detection method for horse, which will benefit for prevention of GETV outbreaks in horses.
Copyright © 2022 Qiu, Cao, Shi, Zhang, Zhu, Gao, Mai, Jin and Lu.
Get Full Text
Publication Date: 2022-11-10 PubMed ID: 36439788PubMed Central: PMC9685671DOI: 10.3389/fmicb.2022.1029444Google Scholar: Lookup
The Equine Research Bank provides access to a large database of publicly available scientific literature. Inclusion in the Research Bank does not imply endorsement of study methods or findings by Mad Barn.
LinkedInCopy
  • Journal Article

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

The research paper addresses the development and application of an indirect Enzyme-Linked Immunosorbent Assay (ELISA) for detecting equine IgG antibodies against Getah virus, using a recombinant E2 domain protein. The study aids in understanding the worldwide prevalence of GETV disease and in devising efficient diagnostic methods.

Introduction

  • GETV or Getah Virus is a mosquito-borne infectious disease causing fever, aseptic meningitis, even abortion in several animals.
  • This disease is now rapidly spreading globally, making a considerable impact on animal safety and public health, particularly in China.
  • However, the limited epidemiological information and lack of commercial diagnostic kits for GETV antibody detection in China make it difficult to fully understand the spread of this disease.

Methods

  • An expression plasmid called pET22b-GETV-E2d was created in this study. It contained the E2 domain (E2d) of the GETV and was fused to a His-tag.
  • The recombinant E2d protein (rE2d) was primarily formed in inclusion bodies and was purified successfully using a nickel affinity column to be employed in an indirect ELISA (rE2d-ELISA).
  • The test’s reaction conditions were optimized, and the cut-off value was set to 0.396, ascertained through 100 horse serum samples tested by the Virus Neutralization Test (VNT).

Results

  • The rE2d-ELISA method demonstrated a 54.3% positive rate for IgG antibodies against GETV based on the testing of 646 clinical serum samples from Xinjiang.
  • The overall alignment rate between rE2d-ELISA and VNT was 94.0%, with a sensitivity of 98.2% and a specificity of 92.6%.

Conclusion

  • The findings indicate that the developed IgG ELISA, which uses recombinant E2d, is a viable and cost-effective antibody detection method for horses.
  • Such a method will significantly aid in preventing GETV outbreaks among the equine population.

Cite This Article

APA
Qiu X, Cao X, Shi N, Zhang H, Zhu X, Gao Y, Mai Z, Jin N, Lu H. (2022). Development and application of an indirect ELISA for detecting equine IgG antibodies against Getah virus with recombinant E2 domain protein. Front Microbiol, 13, 1029444. https://doi.org/10.3389/fmicb.2022.1029444

Publication

Frontiers in microbiology
ISSN: 1664-302X
NlmUniqueID: 101548977
Country: Switzerland
Language: English
Volume: 13
Pages: 1029444
PII: 1029444

Researcher Affiliations

Qiu, Xiangshu
  • College of Animal Sciences, Institute of Preventive Veterinary Medicine, Zhejiang University, Hangzhou Zhejiang, China.
  • Changchun Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Changchun, Jilin, China.
Cao, Xinyu
  • Changchun Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Changchun, Jilin, China.
  • College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China.
Shi, Ning
  • Changchun Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Changchun, Jilin, China.
  • Key Laboratory of Zoonoses Research, College of Veterinary Medicine, Ministry of Education, Institute of Zoonosis, Jilin University, Changchun, China.
Zhang, He
  • Changchun Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Changchun, Jilin, China.
Zhu, Xiangyu
  • Changchun Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Changchun, Jilin, China.
Gao, Yan
  • Changchun Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Changchun, Jilin, China.
Mai, Zhanhai
  • College of Veterinary Medicine, Xinjiang Agricultural University, Ürümqi, China.
Jin, Ningyi
  • College of Animal Sciences, Institute of Preventive Veterinary Medicine, Zhejiang University, Hangzhou Zhejiang, China.
  • Changchun Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Changchun, Jilin, China.
Lu, Huijun
  • Changchun Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Changchun, Jilin, China.

Conflict of Interest Statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

References

This article includes 32 references
  1. Bannai H, Nemoto M, Ochi A, Kikuchi T, Kobayashi M, Tsujimura K, Yamanaka T, Kondo T. Epizootiological Investigation of Getah Virus Infection among Racehorses in Japan in 2014.. J Clin Microbiol 2015 Jul;53(7):2286-91.
    doi: 10.1128/jcm.00550-15pmc: PMC4473224pubmed: 25972425google scholar: lookup
  2. Bannai H, Nemoto M, Tsujimura K, Ohta M. Establishment of an enzyme-linked immunosorbent assay for Getah virus infection in horses using a 20-mer synthetic peptide for the E2 glycoprotein as an antigen.. Arch Virol 2020 Feb;165(2):377-385.
    doi: 10.1007/s00705-019-04508-2pubmed: 31853643google scholar: lookup
  3. Bannai H, Nemoto M, Tsujimura K, Yamanaka T, Kokado H. Development of an enzyme-linked immunosorbent assay for Getah virus infection in horses using recombinant E2 protein as an antigen.. J Virol Methods 2019 Sep;271:113681.
    doi: 10.1016/j.jviromet.2019.113681pubmed: 31207276google scholar: lookup
  4. Brown CM, Timoney PJ. Getah virus infection of Indian horses.. Trop Anim Health Prod 1998 Aug;30(4):241-52.
    doi: 10.1023/a:1005079229232pubmed: 9760716google scholar: lookup
  5. Bryant JE, Crabtree MB, Nam VS, Yen NT, Duc HM, Miller BR. Isolation of arboviruses from mosquitoes collected in northern Vietnam.. Am J Trop Med Hyg 2005 Aug;73(2):470-3.
    doi: 10.4269/ajtmh.2005.73.470pubmed: 16103625google scholar: lookup
  6. Chen R, Mukhopadhyay S, Merits A, Bolling B, Nasar F, Coffey LL, Powers A, Weaver SC, Ictv Report Consortium. ICTV Virus Taxonomy Profile: Togaviridae.. J Gen Virol 2018 Jun;99(6):761-762.
    doi: 10.1099/jgv.0.001072pubmed: 29745869google scholar: lookup
  7. Cho B, Jeon BY, Kim J, Noh J, Kim J, Park M, Park S. Expression and evaluation of Chikungunya virus E1 and E2 envelope proteins for serodiagnosis of Chikungunya virus infection.. Yonsei Med J 2008 Oct 31;49(5):828-35.
    doi: 10.3349/ymj.2008.49.5.828pmc: PMC2615359pubmed: 18972604google scholar: lookup
  8. Doherr MG, Carpenter TE, Wilson WD, Gardner IA. Effect of different sampling techniques on odds ratio estimates using hospital-based cases and controls.. Prev Vet Med 1997 Sep;32(1-2):77-93.
    doi: 10.1016/s0167-5877(96)01146-4pubmed: 9361322google scholar: lookup
  9. Fukunaga Y, Kumanomido T, Kamada M. Getah virus as an equine pathogen.. Vet Clin North Am Equine Pract 2000 Dec;16(3):605-17.
    doi: 10.1016/s0749-0739(17)30099-8pubmed: 11219353google scholar: lookup
  10. Heo Y, Park BK, An SH, Jeon Y, Yoon YD, Kim YH. The 1986 survey on horse diseases of veterinary importance in the Republic of Korea. Res. Report Rural Dev. Administration 29, 116–120.
  11. Hu T, Zheng Y, Zhang Y, Li G, Qiu W, Yu J, Cui Q, Wang Y, Zhang C, Zhou X, Feng Z, Zhou W, Fan Q, Zhang F. Identification of a novel Getah virus by Virus-Discovery-cDNA random amplified polymorphic DNA (RAPD).. BMC Microbiol 2012 Dec 27;12:305.
    doi: 10.1186/1471-2180-12-305pmc: PMC3547691pubmed: 23268691google scholar: lookup
  12. Imagawa H, Ando Y, Kamada M, Sugiura T, Kumanomido T, Fukunaga Y, Wada R, Hirasawa K, Akiyama Y. Sero-epizootiological survey on Getah virus infection in light horses in Japan.. Nihon Juigaku Zasshi 1981 Dec;43(6):797-802.
    doi: 10.1292/jvms1939.43.797pubmed: 6283217google scholar: lookup
  13. Kuwata R, Shimoda H, Phichitraslip T, Prasertsincharoen N, Noguchi K, Yonemitsu K, Minami S, Supriyono, Tran NTB, Takano A, Suzuki K, Nemoto M, Bannai H, Yokoyama M, Takeda T, Jittapalapong S, Rerkamnuaychoke W, Maeda K. Getah virus epizootic among wild boars in Japan around 2012.. Arch Virol 2018 Oct;163(10):2817-2821.
    doi: 10.1007/s00705-018-3897-4pubmed: 29876783google scholar: lookup
  14. Lee SH, Yang D-K, Kim H-H, Jo H-Y, Choi S-S, Cho I-S. Development and evaluation of indirect ELISA for detection of antibodies to Getah virus in horse serum. J. Bacteriol. Virol. 46, 63–77.
    doi: 10.4167/jbv.2016.46.2.63google scholar: lookup
  15. Li X, Feng R, Liu J, Wang D, Yang Y, Fan J. Development of an indirect ELISA with recombinant E2 protein for detection of the antibody against Getah virus. Vet. Sci. China 42, 1037–1042.
  16. Li Y, Fu S, Guo X, Li X, Li M, Wang L, Gao X, Lei W, Cao L, Lu Z, He Y, Wang H, Zhou H, Liang G. Serological Survey of Getah Virus in Domestic Animals in Yunnan Province, China.. Vector Borne Zoonotic Dis 2019 Jan;19(1):59-61.
    doi: 10.1089/vbz.2018.2273pubmed: 29957135google scholar: lookup
  17. Li YY, Liu H, Fu SH, Li XL, Guo XF, Li MH, Feng Y, Chen WX, Wang LH, Lei WW, Gao XY, Lv Z, He Y, Wang HY, Zhou HN, Wang GQ, Liang GD. From discovery to spread: The evolution and phylogeny of Getah virus.. Infect Genet Evol 2017 Nov;55:48-55.
    doi: 10.1016/j.meegid.2017.08.016pubmed: 28827175google scholar: lookup
  18. Li XD, Qiu FX, Yang H, Rao YN, Calisher CH. Isolation of Getah virus from mosquitos collected on Hainan Island, China, and results of a serosurvey.. Southeast Asian J Trop Med Public Health 1992 Dec;23(4):730-4.
    pubmed: 1338481
  19. Lin H, Zhou H, Gao L, Li B, He K, Fan H. Development and application of an indirect ELISA for the detection of antibodies to porcine epidemic diarrhea virus based on a recombinant spike protein.. BMC Vet Res 2018 Aug 20;14(1):243.
    doi: 10.1186/s12917-018-1570-5pmc: PMC6102851pubmed: 30126390google scholar: lookup
  20. Liu H, Li LX, Bu YP, Liu Y, Sun XT, Shi N, Deng XY, Lu RG, Hu B, Jin NY, Yan XJ. Rapid Visual Detection of Getah Virus Using a Loop-Mediated Isothermal Amplification Method.. Vector Borne Zoonotic Dis 2019 Oct;19(10):741-746.
    doi: 10.1089/vbz.2018.2434pubmed: 30964395google scholar: lookup
  21. Lu G, Chen R, Shao R, Dong N, Liu W, Li S. Getah virus: An increasing threat in China.. J Infect 2020 Mar;80(3):350-371.
    doi: 10.1016/j.jinf.2019.11.016pubmed: 31790706google scholar: lookup
  22. Lu G, Ou J, Ji J, Ren Z, Hu X, Wang C, Li S. Emergence of Getah Virus Infection in Horse With Fever in China, 2018.. Front Microbiol 2019;10:1416.
    doi: 10.3389/fmicb.2019.01416pmc: PMC6596439pubmed: 31281304google scholar: lookup
  23. Morey SH, Kashyap RS, Purohit HJ, Taori GM, Daginawala HF. An approach towards peptide-based antibody detection for diagnosis of Chikungunya infection.. Biomarkers 2010 Sep;15(6):546-52.
    doi: 10.3109/1354750X.2010.494200pubmed: 20550431google scholar: lookup
  24. Morita K, Igarashi A. Oligonucleotide fingerprint analysis of strains of Getah virus isolated in Japan and Malaysia.. J Gen Virol 1984 Nov;65 ( Pt 11):1899-908.
    doi: 10.1099/0022-1317-65-11-1899pubmed: 6094708google scholar: lookup
  25. Shi N, Li LX, Lu RG, Yan XJ, Liu H. Highly Pathogenic Swine Getah Virus in Blue Foxes, Eastern China, 2017.. Emerg Infect Dis 2019 Jun;25(6):1252-1254.
    doi: 10.3201/eid2506.181983pmc: PMC6537705pubmed: 31107236google scholar: lookup
  26. Shi N, Qiu X, Cao X, Mai Z, Zhu X, Li N, Zhang H, Zhang J, Li Z, Shaya N, Lu H, Jin N. Molecular and serological surveillance of Getah virus in the Xinjiang Uygur Autonomous Region, China, 2017-2020.. Virol Sin 2022 Apr;37(2):229-237.
    doi: 10.1016/j.virs.2022.02.004pmc: PMC9170979pubmed: 35527224google scholar: lookup
  27. Shi N, Zhu X, Qiu X, Cao X, Jiang Z, Lu H, Jin N. Origin, genetic diversity, adaptive evolution and transmission dynamics of Getah virus.. Transbound Emerg Dis 2022 Jul;69(4):e1037-e1050.
    doi: 10.1111/tbed.14395pubmed: 34812572google scholar: lookup
  28. Shortridge KF, Mason DK, Watkins KL, Aaskov JG. Serological evidence for the transmission of Getah virus in Hong Kong.. Vet Rec 1994 May 14;134(20):527-8.
    doi: 10.1136/vr.134.20.527pubmed: 8085315google scholar: lookup
  29. Verma P, Bhatnagar S, Kumar P, Chattree V, Parida MM, Hoti SL, Ali S, Rao DN. Analysis of antibody response (IgM, IgG, IgG3) to Chikungunya virus using panel of peptides derived from envelope protein for serodiagnosis.. Clin Chem Lab Med 2014 Feb;52(2):297-307.
    doi: 10.1515/cclm-2013-0363pubmed: 24088615google scholar: lookup
  30. Voss JE, Vaney MC, Duquerroy S, Vonrhein C, Girard-Blanc C, Crublet E, Thompson A, Bricogne G, Rey FA. Glycoprotein organization of Chikungunya virus particles revealed by X-ray crystallography.. Nature 2010 Dec 2;468(7324):709-12.
    doi: 10.1038/nature09555pubmed: 21124458google scholar: lookup
  31. Wekesa SN, Inoshima Y, Murakami K, Sentsui H. Genomic analysis of some Japanese isolates of Getah virus.. Vet Microbiol 2001 Nov 8;83(2):137-46.
    doi: 10.1016/s0378-1135(01)00417-5pubmed: 11557154google scholar: lookup
  32. Zhai YG, Wang HY, Sun XH, Fu SH, Wang HQ, Attoui H, Tang Q, Liang GD. Complete sequence characterization of isolates of Getah virus (genus Alphavirus, family Togaviridae) from China.. J Gen Virol 2008 Jun;89(Pt 6):1446-1456.
    doi: 10.1099/vir.0.83607-0pubmed: 18474561google scholar: lookup

Citations

This article has been cited 4 times.
  1. Irekeola AA, Shueb RH. Global seroprevalence and distribution of Getah virus in domestic and wild animals: A systematic review and meta-analysis. Vet World 2025 Nov;18(11):3464-3475.
    doi: 10.14202/vetworld.2025.3464-3475pubmed: 41472765google scholar: lookup
  2. Liu Z, Yang F, Fang M, Wu Q, Fan K, Huang D, Ye Y, Wan G, Song D. Rapid and Sensitive One-Tube Detection of Getah Virus Using RT-LAMP Combined with Pyrococcus furiosus Argonaute. Vet Sci 2025 Jan 24;12(2).
    doi: 10.3390/vetsci12020093pubmed: 40005853google scholar: lookup
  3. Zhong D, Zheng J, Ma Z, Wang Y, Wei J. Rapid Detection of Getah Virus Antibodies in Horses Using a Recombinant E2 Protein-Based Immunochromatographic Strip. Animals (Basel) 2024 Aug 8;14(16).
    doi: 10.3390/ani14162309pubmed: 39199843google scholar: lookup
  4. Jiang Z, Qin Y, Zhang L, Xing G, Shi Z, Song W, Dobrikov GM, Chen J, Su S. Development and application of a colloidal-gold immunochromatographic strip for detecting Getah virus antibodies. Appl Microbiol Biotechnol 2024 Jun 1;108(1):355.
    doi: 10.1007/s00253-024-13168-5pubmed: 38822832google scholar: lookup
Subscribe to our newsletter
Join 99,344 horse owners like you who receive our email updates on horse health and nutrition.