Identification and Molecular Characterization of Viruses Infecting Cucurbits in Pakistan

Abstract

Cucurbits are grown throughout the North-West Frontier Province of Pakistan as summer and winter crops. Plants having mosaic, mottling, chlorosis and leaf distortion symptoms were frequently found in most of the cucurbit fields during the survey. Using dot immunobinding assay, Cucumber green mottle mosaic virus (CGMMV), Zucchini yellow mosaic virus (ZYMV), Watermelon mosaic virus (WMV) and Papaya ringspot virus (PRSV) were found infecting cucurbits. CGMMV was widespread, infecting 46.9% of the samples tested followed by ZYMV (14.8%), WMV (12.5%) and PRSV (7.8%). Multiple infections were common with 42% of the samples being infected with two viruses and 8% with three viruses. The nucleotide sequences of the coat protein (CP) genes of these four viruses were determined and deduced amino acid sequence comparisons revealed 88.3-99% similarity of the ZYMV-Pak isolate with other isolates of ZYMV reported worldwide. The amino acid sequence identity of Pakistani isolates of WMV, CGMMV and PRSV ranged from 96.8 to 98.4%, 98.1 to 99.4% and 79.3 to 84.2%, respectively, with other isolates reported elsewhere. Little variability was observed in the sequences of WMV and CGMMV. ZYMV-Pak was very close to the USA isolate, and the PRSV-Pak isolate was close to Indian isolates of PRSV possibly reflecting the geographical relationship between these isolates.

Evaluation of Low Temperature Induced Mutants of Cucumber green mottle mosaic virus for Cross-protection in Cucurbits

“…The stability of the mild strain in this case should not be a concern as tobamoviruses are known to have the most stable genome among the plant viruses, but one must always need to take the necessary precautions by continually evaluating the actual inocula used for crossprotection. Although, the previous findings Ali et al, [24] indicate very low level of diversity in the sequence of CGMMV isolates reported so far, one must also investigate the diversity of the virus in the area concerned. Some sort of variability do occur in genome of CGMMV, which is also reflected by different symptom expressions reported for watermelon isolate from Japan and CGMMV-Pk.…”

“…Five viruses were found infecting the crops, with CGMMV being the most widespread [24]. The dominance of CGMMV could be attributed to the contact transmission of the virus, coupled with the poor phytosanitary measures.…”

“…Cucumber green mottle mosaic virus has been reported as the most prevalent and dominant virus infecting cucurbit crops in KPK, Pakistan [24] inducing mosaic symptoms that later on turns into yellow mosaic (Figure 1a), mottle and severe puckering. In an attempt to obtain mild isolate for cross-protection, assay hosts including C. amaranticolor, C. quinoa, N. benthamiana, N. Glutinosa, N. Rustica, C. sativus and V. unguiculata were mechanically inoculated with CGMMV.…”

“…sativus seedlings were mechanically inoculated with the selected isolates (hereafter termed Pk-47 and Pk-81) by rub inoculation as described previously by Ali et al, [24]. Infection was confirmed by DIBA assay 10 days after inoculation.…”

“…Sap from infected leaves was coated on carbon-formvar coated grids as described previously [24] and were examined with JEOL 100S electron microscope.…”