Development and validation of a multiplex real-time RT-PCR for detection of citrus and hibiscus-infecting Citrus leprosis virus C2
Benjamin Adducci: USDA-APHIS-PPQ-S&T-CPHST
<div>Citrus leprosis is one of the destructive viral diseases of citrus in the Americas. <em>Citrus leprosis virus</em> C2 (CiLV-C2), the second member of the genus <em>Cilevirus </em>was discovered on sweet orange in Colombia in 2013. A <em>Cilevirus</em> infecting ornamental hibiscus was reported from Hawaii in 2013 and from Tampa, Florida in 2017. Based on amino acid sequence identity, Hibiscus-infecting <em>Cilevirus</em> is taxonomically reclassified as a hibiscus strain of <em>Citrus leprosis virus</em> C2 (CiLV-C2H). In order to detect both citrus and hibiscus-infecting CiLV-C2, a multiplex real-time RT-PCR (qRT-PCR) assay was developed for simultaneous detection and discrimination of CiLV-C2 and CiLV-C2H with a plant internal control. Reported here are the results of developing and validating the multiplex qRT-PCR assay on the ABI QuantStudio 7 real-time PCR platform and improvement of the internal control for the assay. The described multiplex assay can be used for a quick and reliable detection of CiLV-C2 and CiLV-C2H in single or dual infection.</div>
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