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Development of indirect enzyme-linked immunosorbent assay for detection of porcine epidemic diarrhea virus specific antibodies (IgG) in serum of naturally infected pigs

BACKGROUND: Porcine epidemic diarrhea virus (PEDV) infection is a highly contagious infectious disease causing watery diarrhea, vomiting, dehydration and high mortality rate in newborn piglets. PEDV infection can cause high economic losses in pig industry. In Japan, a PEDV outbreak occurred with hig...

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Autores principales: Myint, Ohnmar, Yoshida, Ayako, Sekiguchi, Satoshi, Van Diep, Nguyen, Fuke, Naoyuki, Izzati, Uda Zahli, Hirai, Takuya, Yamaguchi, Ryoji
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6852973/
https://www.ncbi.nlm.nih.gov/pubmed/31718620
http://dx.doi.org/10.1186/s12917-019-2123-2
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author Myint, Ohnmar
Yoshida, Ayako
Sekiguchi, Satoshi
Van Diep, Nguyen
Fuke, Naoyuki
Izzati, Uda Zahli
Hirai, Takuya
Yamaguchi, Ryoji
author_facet Myint, Ohnmar
Yoshida, Ayako
Sekiguchi, Satoshi
Van Diep, Nguyen
Fuke, Naoyuki
Izzati, Uda Zahli
Hirai, Takuya
Yamaguchi, Ryoji
author_sort Myint, Ohnmar
collection PubMed
description BACKGROUND: Porcine epidemic diarrhea virus (PEDV) infection is a highly contagious infectious disease causing watery diarrhea, vomiting, dehydration and high mortality rate in newborn piglets. PEDV infection can cause high economic losses in pig industry. In Japan, a PEDV outbreak occurred with high mortality from 2013 to 2015. Even though until now, PEDV infection occurs sporadically. For the control and monitoring of PEDV infection, not only symptomatic pigs, but also asymptomatic pigs should be identified. The objective of this study is to develop and optimize novel indirect ELISA as a simple, rapid, sensitive and specific method for the detection of anti-PEDV antibodies and evaluate the efficacy of the assay as a diagnostic method for PED. RESULTS: One hundred sixty-two serum samples, consisting of 81 neutralization test (NT) positive and 81 NT negative sera, were applied to the assay. Indirect ELISA test based on whole virus antigen (NK94P6 strain) derived from Vero cell culture was evaluated by receiver operating characteristic (ROC) analysis with neutralization test (NT) as a reference method, and cut-off value was determined as 0.320 with sensitivity and specificity of 92.6 and 90.1%, respectively. The area under curve (AUC) was 0.949, indicating excellent accuracy of indirect ELISA test. There was significant positive correlation between indirect ELISA and neutralization test (R = 0.815, P < 0.05). Furthermore, the kappa statics showed the excellent agreement between these two tests (kappa value = 0.815). In addition, the sensitivity and specificity of preserved plates with different periods (1 day, 2 weeks, 1, 2, 3, 4, 5 and 6 months) after drying antigen coated plates were 100% and 80–100%, respectively. CONCLUSIONS: The developed indirect ELISA test in our study would be useful as a reliable test for serological survey and disease control of PEDV infection, and our pre-antigen coated ELISA plates can be preserved at 4 °C until at least 6 months.
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spelling pubmed-68529732019-11-21 Development of indirect enzyme-linked immunosorbent assay for detection of porcine epidemic diarrhea virus specific antibodies (IgG) in serum of naturally infected pigs Myint, Ohnmar Yoshida, Ayako Sekiguchi, Satoshi Van Diep, Nguyen Fuke, Naoyuki Izzati, Uda Zahli Hirai, Takuya Yamaguchi, Ryoji BMC Vet Res Research Article BACKGROUND: Porcine epidemic diarrhea virus (PEDV) infection is a highly contagious infectious disease causing watery diarrhea, vomiting, dehydration and high mortality rate in newborn piglets. PEDV infection can cause high economic losses in pig industry. In Japan, a PEDV outbreak occurred with high mortality from 2013 to 2015. Even though until now, PEDV infection occurs sporadically. For the control and monitoring of PEDV infection, not only symptomatic pigs, but also asymptomatic pigs should be identified. The objective of this study is to develop and optimize novel indirect ELISA as a simple, rapid, sensitive and specific method for the detection of anti-PEDV antibodies and evaluate the efficacy of the assay as a diagnostic method for PED. RESULTS: One hundred sixty-two serum samples, consisting of 81 neutralization test (NT) positive and 81 NT negative sera, were applied to the assay. Indirect ELISA test based on whole virus antigen (NK94P6 strain) derived from Vero cell culture was evaluated by receiver operating characteristic (ROC) analysis with neutralization test (NT) as a reference method, and cut-off value was determined as 0.320 with sensitivity and specificity of 92.6 and 90.1%, respectively. The area under curve (AUC) was 0.949, indicating excellent accuracy of indirect ELISA test. There was significant positive correlation between indirect ELISA and neutralization test (R = 0.815, P < 0.05). Furthermore, the kappa statics showed the excellent agreement between these two tests (kappa value = 0.815). In addition, the sensitivity and specificity of preserved plates with different periods (1 day, 2 weeks, 1, 2, 3, 4, 5 and 6 months) after drying antigen coated plates were 100% and 80–100%, respectively. CONCLUSIONS: The developed indirect ELISA test in our study would be useful as a reliable test for serological survey and disease control of PEDV infection, and our pre-antigen coated ELISA plates can be preserved at 4 °C until at least 6 months. BioMed Central 2019-11-12 /pmc/articles/PMC6852973/ /pubmed/31718620 http://dx.doi.org/10.1186/s12917-019-2123-2 Text en © The Author(s). 2019 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
spellingShingle Research Article
Myint, Ohnmar
Yoshida, Ayako
Sekiguchi, Satoshi
Van Diep, Nguyen
Fuke, Naoyuki
Izzati, Uda Zahli
Hirai, Takuya
Yamaguchi, Ryoji
Development of indirect enzyme-linked immunosorbent assay for detection of porcine epidemic diarrhea virus specific antibodies (IgG) in serum of naturally infected pigs
title Development of indirect enzyme-linked immunosorbent assay for detection of porcine epidemic diarrhea virus specific antibodies (IgG) in serum of naturally infected pigs
title_full Development of indirect enzyme-linked immunosorbent assay for detection of porcine epidemic diarrhea virus specific antibodies (IgG) in serum of naturally infected pigs
title_fullStr Development of indirect enzyme-linked immunosorbent assay for detection of porcine epidemic diarrhea virus specific antibodies (IgG) in serum of naturally infected pigs
title_full_unstemmed Development of indirect enzyme-linked immunosorbent assay for detection of porcine epidemic diarrhea virus specific antibodies (IgG) in serum of naturally infected pigs
title_short Development of indirect enzyme-linked immunosorbent assay for detection of porcine epidemic diarrhea virus specific antibodies (IgG) in serum of naturally infected pigs
title_sort development of indirect enzyme-linked immunosorbent assay for detection of porcine epidemic diarrhea virus specific antibodies (igg) in serum of naturally infected pigs
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6852973/
https://www.ncbi.nlm.nih.gov/pubmed/31718620
http://dx.doi.org/10.1186/s12917-019-2123-2
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