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Seroprevalence of Getah virus in Pigs in Eastern China Determined with a Recombinant E2 Protein-Based Indirect ELISA
Getah virus (GETV), in the genus Alphavirus and the family Togaviridae, has been detected throughout the world. GETV causes high morbidity and mortality in newborn piglets, entailing serious economic losses. Therefore, the experimental work on GETV detection is necessary. However, due to the influen...
Autores principales: | , , , , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2022
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9607375/ https://www.ncbi.nlm.nih.gov/pubmed/36298726 http://dx.doi.org/10.3390/v14102173 |
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author | Sun, Qing Xie, Yixuan Guan, Zhixin Zhang, Yan Li, Yuhao Yang, Yang Zhang, Junjie Li, Zongjie Qiu, Yafeng Li, Beibei Liu, Ke Shao, Donghua Wang, Jiaxiang Ma, Zhiyong Wei, Jianchao Li, Peng |
author_facet | Sun, Qing Xie, Yixuan Guan, Zhixin Zhang, Yan Li, Yuhao Yang, Yang Zhang, Junjie Li, Zongjie Qiu, Yafeng Li, Beibei Liu, Ke Shao, Donghua Wang, Jiaxiang Ma, Zhiyong Wei, Jianchao Li, Peng |
author_sort | Sun, Qing |
collection | PubMed |
description | Getah virus (GETV), in the genus Alphavirus and the family Togaviridae, has been detected throughout the world. GETV causes high morbidity and mortality in newborn piglets, entailing serious economic losses. Therefore, the experimental work on GETV detection is necessary. However, due to the influence of a variety of unavoidable factors, the ELISA test for the primary screening of animal diseases has low accuracy in detection results. Therefore, we optimized a recombinant E2 (rE2) protein-based enzyme-linked immunosorbent assay (ELISA) for the detection of GETV antibodies in pig serum. The E2 protein was successfully expressed and purified with SDS-PAGE. A Western blotting analysis of sera from infected pigs showed strong reaction with a viral antigen of ~46 KDa corresponding to the E2 glycoproteins. By using chessboard titration and comparing the P/N values, we found that the optimal concentration of coated antigen was found to be 24.5 μg/mL, and the optimal dilution of serum specimens was 1:100. The best working dilution of the horseradish peroxidase (HRP)-conjugated goat anti-pig immunoglobulin (IgG) was 1:5000. The optimal coating conditions were 12 h at 4 °C. The optimal incubation conditions for serum specimens, blocking, and reaction with the secondary antibody were all 1 h at 37 °C. We also investigated the seroprevalence of GETV in 133 serum specimens collected in Eastern China, and 37.59% of the samples tested positive for anti-GETV IgG antibodies, indicating that the seroprevalence of GETV is high in pig populations in China. The seroprevalence was significantly lower in spring (April; 24.24%, 16/66) than in autumn (October; 50.75%, 34/67), which suggested that the presence of anti-GETV antibodies in pigs was seasonal. In conclusion, we improved an rE2 ELISA that detected pig antibodies against GETV after experimental and natural infections. This should be useful in the diagnosis and surveillance of GETV infections. |
format | Online Article Text |
id | pubmed-9607375 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-96073752022-10-28 Seroprevalence of Getah virus in Pigs in Eastern China Determined with a Recombinant E2 Protein-Based Indirect ELISA Sun, Qing Xie, Yixuan Guan, Zhixin Zhang, Yan Li, Yuhao Yang, Yang Zhang, Junjie Li, Zongjie Qiu, Yafeng Li, Beibei Liu, Ke Shao, Donghua Wang, Jiaxiang Ma, Zhiyong Wei, Jianchao Li, Peng Viruses Article Getah virus (GETV), in the genus Alphavirus and the family Togaviridae, has been detected throughout the world. GETV causes high morbidity and mortality in newborn piglets, entailing serious economic losses. Therefore, the experimental work on GETV detection is necessary. However, due to the influence of a variety of unavoidable factors, the ELISA test for the primary screening of animal diseases has low accuracy in detection results. Therefore, we optimized a recombinant E2 (rE2) protein-based enzyme-linked immunosorbent assay (ELISA) for the detection of GETV antibodies in pig serum. The E2 protein was successfully expressed and purified with SDS-PAGE. A Western blotting analysis of sera from infected pigs showed strong reaction with a viral antigen of ~46 KDa corresponding to the E2 glycoproteins. By using chessboard titration and comparing the P/N values, we found that the optimal concentration of coated antigen was found to be 24.5 μg/mL, and the optimal dilution of serum specimens was 1:100. The best working dilution of the horseradish peroxidase (HRP)-conjugated goat anti-pig immunoglobulin (IgG) was 1:5000. The optimal coating conditions were 12 h at 4 °C. The optimal incubation conditions for serum specimens, blocking, and reaction with the secondary antibody were all 1 h at 37 °C. We also investigated the seroprevalence of GETV in 133 serum specimens collected in Eastern China, and 37.59% of the samples tested positive for anti-GETV IgG antibodies, indicating that the seroprevalence of GETV is high in pig populations in China. The seroprevalence was significantly lower in spring (April; 24.24%, 16/66) than in autumn (October; 50.75%, 34/67), which suggested that the presence of anti-GETV antibodies in pigs was seasonal. In conclusion, we improved an rE2 ELISA that detected pig antibodies against GETV after experimental and natural infections. This should be useful in the diagnosis and surveillance of GETV infections. MDPI 2022-09-30 /pmc/articles/PMC9607375/ /pubmed/36298726 http://dx.doi.org/10.3390/v14102173 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Sun, Qing Xie, Yixuan Guan, Zhixin Zhang, Yan Li, Yuhao Yang, Yang Zhang, Junjie Li, Zongjie Qiu, Yafeng Li, Beibei Liu, Ke Shao, Donghua Wang, Jiaxiang Ma, Zhiyong Wei, Jianchao Li, Peng Seroprevalence of Getah virus in Pigs in Eastern China Determined with a Recombinant E2 Protein-Based Indirect ELISA |
title | Seroprevalence of Getah virus in Pigs in Eastern China Determined with a Recombinant E2 Protein-Based Indirect ELISA |
title_full | Seroprevalence of Getah virus in Pigs in Eastern China Determined with a Recombinant E2 Protein-Based Indirect ELISA |
title_fullStr | Seroprevalence of Getah virus in Pigs in Eastern China Determined with a Recombinant E2 Protein-Based Indirect ELISA |
title_full_unstemmed | Seroprevalence of Getah virus in Pigs in Eastern China Determined with a Recombinant E2 Protein-Based Indirect ELISA |
title_short | Seroprevalence of Getah virus in Pigs in Eastern China Determined with a Recombinant E2 Protein-Based Indirect ELISA |
title_sort | seroprevalence of getah virus in pigs in eastern china determined with a recombinant e2 protein-based indirect elisa |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9607375/ https://www.ncbi.nlm.nih.gov/pubmed/36298726 http://dx.doi.org/10.3390/v14102173 |
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