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Improved Diagnostic Techniques for Tomato Yellow Leaf Curl Virus in Tomato Breeding Programs

Breeding for tomatoes resistant to tomato yellow leaf curl geminivirus (TYLCV) is difficult, mainly due to the lack of an accurate system for selecting resistant plants. The variable nature of the genotypes used (wild or wild-derived genotypes and advanced breeding lines) makes the development of a reproducible selection procedure necessary during the breeding program. Serological and nucleic acid-based diagnostic techniques for detecting TYLCV were evaluated for their sensitivity, reliability, and possibility of quantification in order to select the combination of techniques that provides the most rapid and accurate characterization of the resistance level of each item. Squash blot followed by the more sensitive polymerase chain reaction (PCR) detection of inconclusive samples is recommended for the screening of new resistant sources among asymptomatic Lycopersicon spp., as they accumulate extremely low viral concentrations that cannot be detected by serological methods. A triple antibody sandwich-enzyme-linked immunosorbent assay (TAS-ELISA), modified to reduce the background, can be used in large-scale field screening of advanced breeding lines if combined with the more sensitive squash blot, which is less affected by the age and state of the tissue. Hybridization methods were also more appropriate to assess viral distribution throughout the plant than TAS-ELISA or even PCR, which fails to reliably detect TYLCV in tissues like roots.