Multiplex Real Time RT-PCR for detection of Tomato Chlorosis Virus (ToCV) and Tomato Infectious Chlorosis Virus (TICV) and their Whitefly Vectors

The whiteflies Bemisia tabaci and Trialeurodes vaporariorum (Hemiptera, Aleyrodidae) are harmful pests of vegetable and ornamental crops in many countries. In particular, B. tabaci is vector of different begomiviruses, pathogenic agents of Tomato yellow leaf curl disease (TYLCD), responsible of severe losses in tomato crops. During recent years, two new emerging whitefly transmitted viruses are spreading on tomato. They are: Tomato chlorosis virus (ToCV) and Tomato infectious chlorosis virus (TICV), belonging to the genus Crinivirus, family Closteroviridae, inducing symptoms often confused with nutritional or physiological disorders: interveinal yellowing, red and/or brown necrotic flecking and brittleness on older leaves. Firstly identified in USA, TICV and ToCV rapidly spread worldwide now being reported in Europe, North America, Africa and Asia. Both viruses are transmitted in semi-persistent manner; especially, TICV is transmitted by T. vaporariorum and ToCV by B. tabaci, T. vaporariorum and T. abutilonea. Since similar symptoms are induced on plants by the two viruses and different vectors are involved in their transmission, a sensitive diagnosis and virus discrimination are essential for disease control. For these reasons, a multiplex TaqMan RT-PCR assay is being developed for a simultaneous detection of the two viruses in their insect vectors, while trying to characterize the latter ones at specific and eventually also at biotype level. The method is proving to be successful in detecting the viruses in extracts from individual insect, sensibly helping also in identification of vector populations, and demonstrated to be useful for wider future monitoring and epidemiology studies as well as for applied phytosanitary purposes.