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Development of a protocol for the simultaneous identification of Tomato spotted wilt virus and thrips species in individual thrips
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- URLhttps://db.koreascholar.com/Article/Detail/364684
Thrips from the genus Frankliniella (Thysanoptera, Thripidae) are a serious insect pest of various crops, including vegetables, fruits and ornamental plants. Thrips cause significant economic damage plants directly by feeding, and indirectly by acting as vectors for the tospoviruses such as Tomato spotted wilt virus (TSWV) and Chrysanthemum stem necrosis virus (CSNV) in chrysanthemum. To investigate the associations of thrips with tospoviruses and their ability to transmit, we have developed a protocol for identifying tospovirus and thrips species simultaneously in an individual thrips sample was successfully conducted. Total RNA was extracted from thrips according to manufacturer’s insturctions of RNeasy mini kit (Quiagen co.), and then TSWV was identified by reverse transcription-polymerase chain reaction (RT-PCR) using TSWV specific primers for the N genes. The residual genomic DNA in thrips RNA extract was used as a template to identify thrips species by PCR using universal primers for ITS2 region and subsequently digesting the PCR product by an restriction enzyme (RsaI) In addition, a classification into the species of thrips was confirmed using the nucleotide sequence of PCR products. The developed protocol was applied to investigate the occurrence of viruliferous thrips species in thrips populations collected from chrysanthemum fields. In this study, most of thrips were identified to Frankliniella spp. and thirps that acquire TSWV was 14 % of 65 thrips.
