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Establishment of reverse transcription recombinase–aided amplification-lateral-flow dipstick and real-time fluorescence–based reverse transcription recombinase–aided amplification methods for detection of the Newcastle disease virus in chickens
Newcastle disease is an acute and highly contagious disease of poultry caused by Newcastle disease virus infection, which does great harm to the poultry industry all over the world. To diagnose the disease simply and quickly, 2 detection methods were established based on reverse transcription recomb...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Elsevier
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7597694/ https://www.ncbi.nlm.nih.gov/pubmed/32616233 http://dx.doi.org/10.1016/j.psj.2020.03.018 |
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author | Wang, Wenjing Wang, Chunguang Bai, Yun Zhang, Peng Yao, Shanshan Liu, Jingru Zhang, Tie |
author_facet | Wang, Wenjing Wang, Chunguang Bai, Yun Zhang, Peng Yao, Shanshan Liu, Jingru Zhang, Tie |
author_sort | Wang, Wenjing |
collection | PubMed |
description | Newcastle disease is an acute and highly contagious disease of poultry caused by Newcastle disease virus infection, which does great harm to the poultry industry all over the world. To diagnose the disease simply and quickly, 2 detection methods were established based on reverse transcription recombinase–aided amplification (RT-RAA) technology. One is reverse transcription recombinase–aided amplification-lateral flow dipstick (RT-RAA-LFD) that is to combine RT-RAA with lateral flow dipstick; the other is real-time fluorescence-based reverse transcription recombinase–aided amplification (RF-RT-RAA) that is the combination of RT-RAA and exo probe. In this study, the reaction conditions such as reaction temperature and reaction time of the 2 methods were optimized, and their specificity and sensitivity were tested. The results showed that the RT-RAA-LFD method could be used to complete reaction within 23 min, and its lowest detectable limit was 10(2) copies/μL, 10 times higher than that of the conventional PCR method (10(3) copies/μL); the RF-RT-RAA method could be used to complete reaction within 26 min, and its lowest detectable limit was 10 copies/μL, 100 times higher than that of conventional PCR method (10(3) copies/μL), and it was as sensitive as real-time fluorescence–based quantitative PCR (10 copies/μL). The 2 methods had no cross reaction to the nucleic acid of other avian pathogens and showed good specificity. A total of 86 clinical samples suspected of the Newcastle disease virus were tested by conventional PCR, real-time fluorescence–based quantitative PCR, RT-RAA-LFD, and RF-RT-RAA. Based on the commonly used conventional PCR method, the other 3 detection methods had a coincidence rate of higher than 93%. In summary, RT-RAA-LFD and RF-RT-RAA had high specificity, sensitivity, and efficiency, which were suitable for clinical and laboratory diagnosis, respectively, and provided technical support for the prevention and control of Newcastle disease. |
format | Online Article Text |
id | pubmed-7597694 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | Elsevier |
record_format | MEDLINE/PubMed |
spelling | pubmed-75976942020-11-03 Establishment of reverse transcription recombinase–aided amplification-lateral-flow dipstick and real-time fluorescence–based reverse transcription recombinase–aided amplification methods for detection of the Newcastle disease virus in chickens Wang, Wenjing Wang, Chunguang Bai, Yun Zhang, Peng Yao, Shanshan Liu, Jingru Zhang, Tie Poult Sci Immunology, Health and Disease Newcastle disease is an acute and highly contagious disease of poultry caused by Newcastle disease virus infection, which does great harm to the poultry industry all over the world. To diagnose the disease simply and quickly, 2 detection methods were established based on reverse transcription recombinase–aided amplification (RT-RAA) technology. One is reverse transcription recombinase–aided amplification-lateral flow dipstick (RT-RAA-LFD) that is to combine RT-RAA with lateral flow dipstick; the other is real-time fluorescence-based reverse transcription recombinase–aided amplification (RF-RT-RAA) that is the combination of RT-RAA and exo probe. In this study, the reaction conditions such as reaction temperature and reaction time of the 2 methods were optimized, and their specificity and sensitivity were tested. The results showed that the RT-RAA-LFD method could be used to complete reaction within 23 min, and its lowest detectable limit was 10(2) copies/μL, 10 times higher than that of the conventional PCR method (10(3) copies/μL); the RF-RT-RAA method could be used to complete reaction within 26 min, and its lowest detectable limit was 10 copies/μL, 100 times higher than that of conventional PCR method (10(3) copies/μL), and it was as sensitive as real-time fluorescence–based quantitative PCR (10 copies/μL). The 2 methods had no cross reaction to the nucleic acid of other avian pathogens and showed good specificity. A total of 86 clinical samples suspected of the Newcastle disease virus were tested by conventional PCR, real-time fluorescence–based quantitative PCR, RT-RAA-LFD, and RF-RT-RAA. Based on the commonly used conventional PCR method, the other 3 detection methods had a coincidence rate of higher than 93%. In summary, RT-RAA-LFD and RF-RT-RAA had high specificity, sensitivity, and efficiency, which were suitable for clinical and laboratory diagnosis, respectively, and provided technical support for the prevention and control of Newcastle disease. Elsevier 2020-04-15 /pmc/articles/PMC7597694/ /pubmed/32616233 http://dx.doi.org/10.1016/j.psj.2020.03.018 Text en © 2020 Published by Elsevier Inc. on behalf of Poultry Science Association Inc. http://creativecommons.org/licenses/by-nc-nd/4.0/ This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). |
spellingShingle | Immunology, Health and Disease Wang, Wenjing Wang, Chunguang Bai, Yun Zhang, Peng Yao, Shanshan Liu, Jingru Zhang, Tie Establishment of reverse transcription recombinase–aided amplification-lateral-flow dipstick and real-time fluorescence–based reverse transcription recombinase–aided amplification methods for detection of the Newcastle disease virus in chickens |
title | Establishment of reverse transcription recombinase–aided amplification-lateral-flow dipstick and real-time fluorescence–based reverse transcription recombinase–aided amplification methods for detection of the Newcastle disease virus in chickens |
title_full | Establishment of reverse transcription recombinase–aided amplification-lateral-flow dipstick and real-time fluorescence–based reverse transcription recombinase–aided amplification methods for detection of the Newcastle disease virus in chickens |
title_fullStr | Establishment of reverse transcription recombinase–aided amplification-lateral-flow dipstick and real-time fluorescence–based reverse transcription recombinase–aided amplification methods for detection of the Newcastle disease virus in chickens |
title_full_unstemmed | Establishment of reverse transcription recombinase–aided amplification-lateral-flow dipstick and real-time fluorescence–based reverse transcription recombinase–aided amplification methods for detection of the Newcastle disease virus in chickens |
title_short | Establishment of reverse transcription recombinase–aided amplification-lateral-flow dipstick and real-time fluorescence–based reverse transcription recombinase–aided amplification methods for detection of the Newcastle disease virus in chickens |
title_sort | establishment of reverse transcription recombinase–aided amplification-lateral-flow dipstick and real-time fluorescence–based reverse transcription recombinase–aided amplification methods for detection of the newcastle disease virus in chickens |
topic | Immunology, Health and Disease |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7597694/ https://www.ncbi.nlm.nih.gov/pubmed/32616233 http://dx.doi.org/10.1016/j.psj.2020.03.018 |
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