Presence of Tomato yellow leaf curl virus Infecting Squash (Curcubita pepo) in Cuba

In a survey conducted in Havana Province during January and February 2003, symptoms of curling and light yellowing of leaves were found in squash plants (Curcubita pepo). DNA from leaves of six symptomatic squash plants was extracted (1) and hybridized at high stringency with specific probes of the intergenic region of Tomato yellow leaf curl virus (TYLCV, genus begomovirus) isolated in Cuba by using a nonradioactive hybridization kit (AlkPhos Direct Labeling and Detection Systems; Amersham Pharmacia Biotech Inc., Piscataway, NJ). Three samples were positive in the nonradioactive analyses. The same samples were positive using polymerase chain reaction (PCR) when the DNA was analyzed with degenerate primers PAL1v1978/PAR1c715 for DNA-A (5), specific primers, ORITY1/ORITY2, for the intergenic region (2), and overlapping specific primers for TYLCV (3). Fragments of 1.4, 0.750, and 2.8 kb were cloned using pGem-T Easy (Promega, Madison, WI), and the six clones obtained were sequenced using the Terminator Cycl...