Development of a Propidium Monoazide-Based Viability Quantitative PCR Assay for Red Sea Bream Iridovirus Detection
Red sea bream iridovirus (RSIV) is an important aquatic virus that causes high mortality in marine fish. RSIV infection mainly spreads through horizontal transmission via seawater, and its early detection could help prevent disease outbreaks. Although quantitative PCR (qPCR) is a sensitive and rapid...
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MDPI AG
2023-02-01
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| Series: | International Journal of Molecular Sciences |
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| Online Access: | https://www.mdpi.com/1422-0067/24/4/3426 |
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| author | Kyung-Ho Kim Gyoungsik Kang Won-Sik Woo Min-Young Sohn Ha-Jeong Son Chan-Il Park |
| author_facet | Kyung-Ho Kim Gyoungsik Kang Won-Sik Woo Min-Young Sohn Ha-Jeong Son Chan-Il Park |
| author_sort | Kyung-Ho Kim |
| collection | DOAJ |
| description | Red sea bream iridovirus (RSIV) is an important aquatic virus that causes high mortality in marine fish. RSIV infection mainly spreads through horizontal transmission via seawater, and its early detection could help prevent disease outbreaks. Although quantitative PCR (qPCR) is a sensitive and rapid method for detecting RSIV, it cannot differentiate between infectious and inactive viruses. Here, we aimed to develop a viability qPCR assay based on propidium monoazide (PMAxx), which is a photoactive dye that penetrates damaged viral particles and binds to viral DNA to prevent qPCR amplification, to distinguish between infectious and inactive viruses effectively. Our results demonstrated that PMAxx at 75 μM effectively inhibited the amplification of heat-inactivated RSIV in viability qPCR, allowing the discrimination of inactive and infectious RSIV. Furthermore, the PMAxx-based viability qPCR assay selectively detected the infectious RSIV in seawater more efficiently than the conventional qPCR and cell culture methods. The reported viability qPCR method will help prevent the overestimation of red sea bream iridoviral disease caused by RSIV. Furthermore, this non-invasive method will aid in establishing a disease prediction system and in epidemiological analysis using seawater. |
| first_indexed | 2024-03-11T08:42:55Z |
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| institution | Directory Open Access Journal |
| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-11T08:42:55Z |
| publishDate | 2023-02-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | International Journal of Molecular Sciences |
| spelling | doaj.art-62edae8d8d0848e29beac330bf7f28952023-11-16T20:59:47ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672023-02-01244342610.3390/ijms24043426Development of a Propidium Monoazide-Based Viability Quantitative PCR Assay for Red Sea Bream Iridovirus DetectionKyung-Ho Kim0Gyoungsik Kang1Won-Sik Woo2Min-Young Sohn3Ha-Jeong Son4Chan-Il Park5Department of Marine Biology and Aquaculture, College of Marine Science, Gyeongsang National University, 2, Tongyeonghaean-ro, Tongyeong 53064, Republic of KoreaDepartment of Marine Biology and Aquaculture, College of Marine Science, Gyeongsang National University, 2, Tongyeonghaean-ro, Tongyeong 53064, Republic of KoreaDepartment of Marine Biology and Aquaculture, College of Marine Science, Gyeongsang National University, 2, Tongyeonghaean-ro, Tongyeong 53064, Republic of KoreaDepartment of Marine Biology and Aquaculture, College of Marine Science, Gyeongsang National University, 2, Tongyeonghaean-ro, Tongyeong 53064, Republic of KoreaDepartment of Marine Biology and Aquaculture, College of Marine Science, Gyeongsang National University, 2, Tongyeonghaean-ro, Tongyeong 53064, Republic of KoreaDepartment of Marine Biology and Aquaculture, College of Marine Science, Gyeongsang National University, 2, Tongyeonghaean-ro, Tongyeong 53064, Republic of KoreaRed sea bream iridovirus (RSIV) is an important aquatic virus that causes high mortality in marine fish. RSIV infection mainly spreads through horizontal transmission via seawater, and its early detection could help prevent disease outbreaks. Although quantitative PCR (qPCR) is a sensitive and rapid method for detecting RSIV, it cannot differentiate between infectious and inactive viruses. Here, we aimed to develop a viability qPCR assay based on propidium monoazide (PMAxx), which is a photoactive dye that penetrates damaged viral particles and binds to viral DNA to prevent qPCR amplification, to distinguish between infectious and inactive viruses effectively. Our results demonstrated that PMAxx at 75 μM effectively inhibited the amplification of heat-inactivated RSIV in viability qPCR, allowing the discrimination of inactive and infectious RSIV. Furthermore, the PMAxx-based viability qPCR assay selectively detected the infectious RSIV in seawater more efficiently than the conventional qPCR and cell culture methods. The reported viability qPCR method will help prevent the overestimation of red sea bream iridoviral disease caused by RSIV. Furthermore, this non-invasive method will aid in establishing a disease prediction system and in epidemiological analysis using seawater.https://www.mdpi.com/1422-0067/24/4/3426red sea bream iridoviruspropidium monoazideviability qPCRseawater concentration |
| spellingShingle | Kyung-Ho Kim Gyoungsik Kang Won-Sik Woo Min-Young Sohn Ha-Jeong Son Chan-Il Park Development of a Propidium Monoazide-Based Viability Quantitative PCR Assay for Red Sea Bream Iridovirus Detection International Journal of Molecular Sciences red sea bream iridovirus propidium monoazide viability qPCR seawater concentration |
| title | Development of a Propidium Monoazide-Based Viability Quantitative PCR Assay for Red Sea Bream Iridovirus Detection |
| title_full | Development of a Propidium Monoazide-Based Viability Quantitative PCR Assay for Red Sea Bream Iridovirus Detection |
| title_fullStr | Development of a Propidium Monoazide-Based Viability Quantitative PCR Assay for Red Sea Bream Iridovirus Detection |
| title_full_unstemmed | Development of a Propidium Monoazide-Based Viability Quantitative PCR Assay for Red Sea Bream Iridovirus Detection |
| title_short | Development of a Propidium Monoazide-Based Viability Quantitative PCR Assay for Red Sea Bream Iridovirus Detection |
| title_sort | development of a propidium monoazide based viability quantitative pcr assay for red sea bream iridovirus detection |
| topic | red sea bream iridovirus propidium monoazide viability qPCR seawater concentration |
| url | https://www.mdpi.com/1422-0067/24/4/3426 |
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