Mechanical Inoculation and Host Range Determination of Tomato Virus

Purified viral particles served for mechanical inoculation of laboratory test plants and tomato plants harboring the Tm-2² resistance gene. Symptomatic tomato plants from the initial experiments served for sap-mechanical inoculation assays. Tomato fruit and leaves were ground in 0.01 M phosphate buffer (pH 7.0) and inoculated to a variety of potential host plants (pre-dusted with carborundum) for extended host range determination. Host range testing was performed in three sets of experiments (> four plants per species). Inoculation of a range of commercial tomato varieties harboring the Tm-2² resistance gene, as certified by the 'Tomato Genetic Resource Center' (TGRC) http://tgrc.ucdavis.edu/Data/Acc/dataframe.aspx?start=AccSearch.aspx&navstart=nav.html, was performed with 8 replicates.
The virus was maintained on systemically infected tomato plants (harboring the Tm-2² resistance gene) cv’s: Ikram, Odelia, Bruno, Olympiacos, Magi, Tory which served as propagation hosts. In addition to the registered resistance to ToMV by the seed-companies, the authenticity of the presence of Tm-2² was further confirmed by DNA sequencing of the resistance gene in Odelia and Ikram cultivars. All virus-infected plants were kept in an insect-proof growth chamber inside a glasshouse and sprayed regularly with insecticides to prevent any infestation.