Identification and mapping of the occurrence of begomoviruses infecting squash (Cucurbita moschata Duch.) in the Philippines

Squash leaf curl disease in the Philippines caused by the bipartite begomovirus species, Squash leaf curl Philippines virus (SLCuPV or SLCPHV), formerly named as Squash leaf curl China virus-Philippines (SLCCNV-PH), has been identified in virus isolate from Muñoz, Nueva Ecija in central Luzon. Later, SLCuPV isolates from Laguna and Batangas in southern Luzon and Benguet in Cordillera Autonomous Region (CAR) have been identified. This study aimed to identify and map the occurrence of squash-infecting begomovirus species in the country. Begomovirus was detected by indirect ELISA in 59% (145/245) samples from plants with symptons of leaf curling, puckering, enation, mosaic and/or yellowing. Single or mixed infections of begomovirus with ZYMV, PRSV-W or CMV were detected in very few samples. Presence of begomovirus was confirmed by PCR and the virus was efficiently detected using primer pairs, TYRepF/TYRepR and TYBF/TYBR that amplify the viral DNA-A or DNA-B component, respectively. BLAST analysis of partial REP gene sequence (744-887 nucleotides) showed that most isolates detected in this study have 93-99% identity with SLCuPV (SLCuPV-PH, Accession no. EU487041 or EU487033.1) and SLCuPV-TW (DQ866135 or EF19974). Virus isolates with highest identity (96-99%) to SLCuPV-PH were found in Ilocos Norte (P180) (MK850838) and Occidental Mindoro (P173) (MK850839) in Luzon, and from Davao (P247) (MK850840) in Mindanao. Isolates from Batangas (P37) (MK850841) and Bulacan (P210) (MK850842) in Luzon were closets to both SLCuPV-PH ans SLCuPV-TW with 95% and 94% identity, respectively. Isolates from Cavite (P131) (MK850843) in Luzon and from Cebu (P215) (MK850844) in the Visayas have the highest identity (95-96%) with SLCuPV. On the other hand, isolate with highest identity (94%) to SLCCNV-CN (AM260206), SLCCNV-PH (EU487031) and SLCCNV�IN (AY184487) was found in Cebu (P211) (MK850845) while isolate P156 (MK850846) from Zamboanga in Mindanao was closest (97%) to SLCCNV-IN. Phylogenetic analysis of partial Rep sequences showed that five isolates from Luzon, one from the Visayas and one from Mindanao clustered with SLCuPV, while the other Visayas and Mindanao isolates clustered with SLCCNV. The identity of the squash-infecting begomoviruses will be fully identified by nucleotide sequence analysis using the full-length DNA-A component. Knowledge of the identity of the virus species and their occurrence in the country are important information in developing strategies for breeding squash with begomovirus resistance.