Advances in diagnosing tomato yellow leaf curl geminivirus infection

As a result of the spread of TYLCV on tomato crops, reliable and rapid diagnostic tools to identify and isolate new sources of infection are necessary. We tested several methods, based both on antibodies and on nonradioactive DNA probes. Indirect plate-trapping ELISA was only effective in detecting the virus in purified preparations, but not in crude extracts. Dot-ELISA with chemiluminescence detection gave satisfactory results when young stems were directly squashed on membranes. A digoxigenin-labeled probe, detected with chemiluminescence, was used in leaf squashes and dot blots. Best results were obtained with dot blots of total nucleic acids prepared with a fast and safe procedure. TYLCV DNA was readily and reliably detected in spots corresponding to 15 micrograms fresh weight. When weak signals were observed, total extracts were analyzed by Southern blotting, to confirm the presence of viral DNA forms.