{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,2]],"date-time":"2026-06-02T10:27:26Z","timestamp":1780396046427,"version":"3.54.1"},"reference-count":62,"publisher":"Public Library of Science (PLoS)","issue":"5","license":[{"start":{"date-parts":[[2026,5,22]],"date-time":"2026-05-22T00:00:00Z","timestamp":1779408000000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS One"],"abstract":"<jats:p>\n                    Plant disease management requires sensitive and user-friendly diagnostic methods. The polymerase chain reaction (PCR) and other methods have helped diagnose diseases, but they are primarily used in high-resource settings, leaving low-resource settings without effective options. A simple probe-enhanced method for the detection of Kenyan tomato leaf curl virus (Kenyan ToLCV) isolates in host crops, including asymptomatic ones, was developed by employing loop-mediated isothermal amplification (LAMP) coupled with a rapid modified DNA extraction method. A modified alkaline polyethylene glycol (APEG) approach for sample preparation from infected tomato and chili pepper leaves was optimized for sensitivity, reproducibility, and adaptability. The modified extraction buffer contained polyethylene glycol, sodium hydroxide, polyvinylpyrrolidone, and sodium chloride. Six LAMP primers and a hybridization probe targeting the AV1 gene were designed and used for specific detection of Kenyan ToLCV isolates. Gel electrophoresis, real-time detection, and color change were used to evaluate the LAMP reaction. The probe-enhanced LAMP assay detected the virus in 2.7 minutes under optimal conditions. The analytical sensitivity reached 0.001 fg\/\u03bcL of total DNA. The optimized LAMP assay was 10\n                    <jats:sup>6<\/jats:sup>\n                    times more sensitive than conventional gold-standard PCR and did not cross-react with other tomato-infecting viruses. The modified buffer yielded a significantly shorter time-to-positive (~4.8 minutes; p\u2009&lt;\u20090.0001) compared to unmodified APEG. The probe-enhanced LAMP assay detected the virus in 50% (4\/8) of asymptomatic samples. The findings suggest the application of the developed assay for phytopathological testing in resource-poor areas and for direct field detection. This assay would provide a reliable approach for rapid plant health certification and quarantine inspection.\n                  <\/jats:p>","DOI":"10.1371\/journal.pone.0349665","type":"journal-article","created":{"date-parts":[[2026,5,22]],"date-time":"2026-05-22T17:31:46Z","timestamp":1779471106000},"page":"e0349665","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":1,"title":["Rapid detection of Kenyan tomato leaf curl virus isolates using probe-enhanced loop-mediated isothermal amplification coupled with a modified DNA extraction method"],"prefix":"10.1371","volume":"21","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-2777-4623","authenticated-orcid":true,"given":"Abigarl","family":"Ndudzo","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Florence","family":"Ng\u2019ong\u2019a","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Edith K.","family":"Avedi","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Elijah M.","family":"Ateka","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"340","published-online":{"date-parts":[[2026,5,22]]},"reference":[{"issue":"2","key":"pone.0349665.ref001","doi-asserted-by":"crossref","first-page":"233","DOI":"10.3390\/plants10020233","article-title":"Global plant virus disease pandemics and epidemics","volume":"10","author":"RA Jones","year":"2021","journal-title":"Plants"},{"issue":"2","key":"pone.0349665.ref002","doi-asserted-by":"crossref","first-page":"117","DOI":"10.1094\/PHYTO-05-22-0167-RVW","article-title":"Plant viruses of agricultural importance: current and future perspectives of virus disease management strategies","volume":"113","author":"S Tatineni","year":"2023","journal-title":"Phytopathology"},{"issue":"1","key":"pone.0349665.ref003","doi-asserted-by":"crossref","first-page":"2","DOI":"10.1186\/s12985-020-01466-z","article-title":"Metagenomic analyses and genetic diversity of Tomato leaf curl Arusha virus affecting tomato plants in Kenya","volume":"18","author":"EK Avedi","year":"2021","journal-title":"Virol J"},{"issue":"6","key":"pone.0349665.ref004","doi-asserted-by":"crossref","first-page":"513","DOI":"10.1007\/s10327-020-00950-2","article-title":"Molecular characterization of tomato leaf curl Uganda virus infecting tomato plants in Kenya","volume":"86","author":"RH Kimathi","year":"2020","journal-title":"J General Plant Pathol"},{"issue":"1","key":"pone.0349665.ref005","article-title":"Occurrence, distribution, and management of tomato yellow leaf curl virus in China","volume":"4","author":"F Li","year":"2022","journal-title":"Phytopathol Res"},{"issue":"3","key":"pone.0349665.ref006","doi-asserted-by":"crossref","first-page":"412","DOI":"10.3390\/v13030412","article-title":"Current developments and challenges in plant viral diagnostics: a systematic review","volume":"13","author":"GT Mehetre","year":"2021","journal-title":"Viruses"},{"issue":"1","key":"pone.0349665.ref007","doi-asserted-by":"crossref","first-page":"99","DOI":"10.1186\/s12985-017-0763-z","article-title":"Development and application of triple antibody sandwich enzyme-linked immunosorbent assays for begomovirus detection using monoclonal antibodies against Tomato yellow leaf curl Thailand virus","volume":"14","author":"C Seepiban","year":"2017","journal-title":"Virol J"},{"key":"pone.0349665.ref008","doi-asserted-by":"crossref","first-page":"142","DOI":"10.1186\/1743-422X-10-142","article-title":"Highly sensitive serological methods for detecting tomato yellow leaf curl virus in tomato plants and whiteflies","volume":"10","author":"Y Xie","year":"2013","journal-title":"Virol J"},{"issue":"3","key":"pone.0349665.ref009","doi-asserted-by":"crossref","first-page":"490","DOI":"10.3390\/plants12030490","article-title":"Detecting tomato leaf curl New Delhi virus causing ridge gourd yellow mosaic disease, and other begomoviruses by antibody-based methods","volume":"12","author":"P Naganur","year":"2022","journal-title":"Plants"},{"issue":"1","key":"pone.0349665.ref010","doi-asserted-by":"crossref","first-page":"1","DOI":"10.4314\/acsj.v30i1.1","article-title":"Incidence, severity and distribution of yellow leaf curl disease of tomato in Kenya","volume":"30","author":"EK Avedi","year":"1970","journal-title":"Afr Crop Sci J"},{"key":"pone.0349665.ref011","first-page":"179","article-title":"Molecular diagnostics for plant viruses towards enhanced delivery of disease-free planting materials and germplasm exchange","author":"OA Kolade","year":"2024","journal-title":"Agricultural biotechnology & biodiversity. 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