{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,5]],"date-time":"2026-10-05T20:10:32Z","timestamp":1791231032735,"version":"4.1.0"},"reference-count":40,"publisher":"MDPI AG","issue":"3","license":[{"start":{"date-parts":[[2026,3,9]],"date-time":"2026-03-09T00:00:00Z","timestamp":1773014400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"SiciliAn MicronanOTecH Research"},{"name":"spoke 3\u2013University of Palermo S2-COMMs- Micro and Nanotechnologies for Smart and Sustainable Communities","award":["ECS_00000022"],"award-info":[{"award-number":["ECS_00000022"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Viruses"],"abstract":"<jats:p>Citrus viroid V (CVd-V), a member of the species Apscaviroid epsiloncitri (family Pospiviroidae), is a graft-transmitted pathogen that spreads through infected propagation material and has been reported in several countries. Recently, it has been detected for the first time in Italy. Although CVd-V is generally considered asymptomatic, foliar and stem symptoms have been observed in Etrog citron. In several citrus species, mixed infections of CVd-V with other viroids give synergistic effects that cause more severe symptoms. To evaluate its spread, a specific and sensitive RT-qPCR assay was developed. The assay specificity, sensitivity, and performance were compared with a conventional end-point RT-PCR, showing a higher sensitivity and no cross-reactivity with other citrus viroids. Furthermore, the assay was evaluated using two RNA extraction procedures: total RNA extraction with commercial kits and a rapid crude extract preparation using a simple extraction buffer. Moreover, RNA extracts of 111 samples, collected from commercial citrus orchards across the main Sicilian citrus-producing areas and from old varieties maintained in germplasm collection, were analyzed. The RT-qPCR revealed an overall CVd-V incidence of 8.1%. Notably, the crude extract preparation showed a sensitivity comparable with conventional total RNA extraction, with substantial savings in cost and processing time. This finding paves the way for using the developed assay with portable qPCR instruments directly in the field, as well as for routine surveillance analyses in citrus production systems.<\/jats:p>","DOI":"10.3390\/v18030335","type":"journal-article","created":{"date-parts":[[2026,3,9]],"date-time":"2026-03-09T11:06:52Z","timestamp":1773054412000},"page":"335","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["Development of a Rapid and Sensitive RT-qPCR for the Specific Detection of Citrus Viroid V and Its Field Application"],"prefix":"10.3390","volume":"18","author":[{"ORCID":"https:\/\/orcid.org\/0009-0008-9849-389X","authenticated-orcid":false,"given":"Graziella","family":"Agr\u00f2","sequence":"first","affiliation":[{"name":"Department of Agricultural, Food and Forest Sciences, University of Palermo, 90128 Palermo, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3320-2827","authenticated-orcid":false,"given":"Jes\u00fas \u00c1ngel","family":"S\u00e1nchez-Navarro","sequence":"additional","affiliation":[{"name":"Institute of Molecular and Cellular Biology of Plants, Universitat Politecnica de Valencia (UPV)\u2014Consejo Superior de Investigaciones Cient\u00edficas (CSIC), 46022 Valencia, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4954-989X","authenticated-orcid":false,"given":"Vicente","family":"Pall\u00e1s","sequence":"additional","affiliation":[{"name":"Institute of Molecular and Cellular Biology of Plants, Universitat Politecnica de Valencia (UPV)\u2014Consejo Superior de Investigaciones Cient\u00edficas (CSIC), 46022 Valencia, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2846-9009","authenticated-orcid":false,"given":"Grazia","family":"Licciardello","sequence":"additional","affiliation":[{"name":"Council for Agricultural Research and Economics (CREA), Research Centre for Olive, Fruit and Citrus Crops, 95024 Acireale, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0003-5358-7853","authenticated-orcid":false,"given":"Giuseppe","family":"Scuderi","sequence":"additional","affiliation":[{"name":"Agrobiotech Soc. Coop., Z.I Blocco Palma I, 95121 Catania, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6551-203X","authenticated-orcid":false,"given":"Antonino","family":"Catara","sequence":"additional","affiliation":[{"name":"Agrobiotech Soc. Coop., Z.I Blocco Palma I, 95121 Catania, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5215-5678","authenticated-orcid":false,"given":"Andrea Giovanni","family":"Caruso","sequence":"additional","affiliation":[{"name":"Department of Agricultural, Food and Forest Sciences, University of Palermo, 90128 Palermo, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8941-7050","authenticated-orcid":false,"given":"Stefano","family":"Panno","sequence":"additional","affiliation":[{"name":"Department of Agricultural, Food and Forest Sciences, University of Palermo, 90128 Palermo, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0885-8744","authenticated-orcid":false,"given":"Slavica","family":"Mati\u0107","sequence":"additional","affiliation":[{"name":"Department of Agricultural, Food and Forest Sciences, University of Palermo, 90128 Palermo, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5926-6300","authenticated-orcid":false,"given":"Salvatore","family":"Davino","sequence":"additional","affiliation":[{"name":"Department of Agricultural, Food and Forest Sciences, University of Palermo, 90128 Palermo, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,3,9]]},"reference":[{"key":"ref_1","unstructured":"FAO (2021). 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