First report of tomato spotted wilt virus on lisianthus ( Eustoma grandiflorum ) in Bulgaria
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First report of tomato spotted wilt virus on lisianthus (Eustoma grandiflorum) in Bulgaria Gancho Pasev 1 Marta Vallino 3
&
Vesela Radeva-Ivanova 1 & Rafe Lyall 2 & Amol Nankar 2 & Dimitrina Kostova 2 & Massimo Turina 3 &
Received: 24 February 2020 / Accepted: 12 November 2020 # Società Italiana di Patologia Vegetale (S.I.Pa.V.) 2020
Keywords TSWV . Lisianthus . ELISA . RT-PCR . Bulgaria
Tomato spotted wilt virus (TSWV) is one of the most destructive pathogens worldwide. In Bulgaria the virus was found to severely damage a number of solanaceous vegetables and ca. 25 ornamentals (Hristova et al. 2001). In spring 2014, about 30% of lisianthus plants (ca. 1000 plants) grown in a private farm nearby Plovdiv, Bulgaria showed yellowing and/or light brown necrosis on leaves and petals followed by subsequent wilting. To identify the causal agent of this infection, Chenopodium quinoa, Nicotiana glutinosa, and Petunia hybrida plants were sap inoculated with material from 10 symptomatic individuals. C. quinoa displayed large necrotic local lesions. Primary and systemic symptoms in N. glutinosa appeared as translucent necrotic lesions accompanied with yellow areas, while P. hybrida showed dark necrotic ring spots. Observations of lisianthus leaf sap by electron microscopy revealed the presence of isometric particles ca. 85 nm in size resembling those of tospoviruses. A TASELISA test was conducted with antisera specific for impatiens necrotic spot virus (INSV, RT-0115-0117/1) and TSWV (RT0105-0106/3) purchased from DSMZ, Germany. The samples from lisianthus and those from the inoculated indicators were positive only for TSWV. RNA was isolated from symptomatic leaves of lisianthus by TRI reagent® (Sigma-Aldrich) and
* Gancho Pasev [email protected]; [email protected] 1
Maritsa Vegetable Crops Research Institute, 4003 Plovdiv, Bulgaria
2
Center of Plant Systems Biology and Biotechnology, 4000 Plovdiv, Bulgaria
3
Institute for Sustainable Plant Protection, National Research Council of Italy, I-1013 Torino, Italy
reverse transcription (RT) was performed using random hexamer primers. The PCR amplification conducted with specific primers targeting N gene, described by Roberts et al. (2000), revealed an amplicon of about 600 bp corresponding to the expected size. Further, the amplicon was sent for sequencing. A high-confidence consensus nucleotide sequence of 567 bp (accession No. MK388405) was selected for analysis, which encoded a protein of 188 amino acids. This protein was queried against a database of all NCBI viral protein sequences by blastp and showed 99% sequence identity to TSWV nucleocapsid protein, including several accessions from Bulgaria (e.g. CAC01282.1, CAC01285.1) (Heinze et al. 2001). The results from TAS-ELISA and RT-PCR tests clearly show that the virus-specific antibodies do not have interspecies cross-reactivity. Based on our knowledge and viruses reported in literature, this appears to be a first report of TSWV on lisianthus in Bulgaria. Acknowledgements The authors GP, VR-I,
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