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Limited research in Saudi Arabia has devolved into the prevalence and genetic diversity of begomoviruses. Utilizing Illumina MiSeq sequencing, we obtained 21 full-length begomovirus sequences (2.7–2.8 kb) from eight cucumber plants grown in fields and greenhouses. We found that two complete begomovirus genomes were variants of the Boushehr strain of tomato yellow leaf curl virus (TYLCV) with nucleotide (nt) sequence identities of 94.7-95.9%. Another full-length genome was a variant of TYLCV-Iran with 94.6% identity. Five full-length sequences closely matched the DNA-A of watermelon chlorotic stunt virus (WmCSV) isolates with 97.9-98.7% nt sequence identities, while five sequences had their highest nt sequence identities (95.8-96.3%) with the DNA-B of WmCSV isolates. Simultaneously, four sequences were 99.1-99.6% identical to the DNA-A of tomato leaf curl Palampur virus (ToLCPalV). Four sequences matched the DNA-B of ToLCPalV reported from Iran and Saudi Arabia with identities ranging from 96.2-100%. Four plants showed a mixed infection of these begomoviruses. Most ORFs showed evidence of negative selection pressure, suggesting that purifying selection plays a crucial role in shaping the diversity of these begomoviruses. Additionally, potential intra- and interspecies recombination events were detected in the TYLCV and WmCSV DNA-B genomic regions. The ToLCPalV isolates identified in this study formed a cluster with the other ToLCPalV isolates reported from Saudi Arabia, Iran and Iraq, representing a unique lineage distinct from ToLCPalV reported from Southeast Asia. High mutation rate and robust selection facilitated the independent evolution of ToLCPalV without recombination. Overall, this study offers valuable insights into the diversity and evolutionary dynamics of begomoviruses infecting cucumber crops in Al-Ahsa, Saudi Arabia.

Citrus viroid infection is emerging as a serious threat because of its efficient systemic movement within the host plant and its quick spread due to contaminated pruning tools. A survey was conducted to investigate the primary distribution and molecular characterization of Citrus bent leaf viroid (CBLVd) and its variants in different citrus cultivars. A total of 154 symptomatic citrus samples were collected and detected by RT‒PCR with newly designed specific primers with the incidence of 36.33%. During biological indexing study on Etrog citron, expressions of reduced leaf size, yellowing with a light green pattern, and bending were observed. Amplified products were sequenced and analyzed using a nucleotide BLAST search, which showed 98% homology with other CBLVd isolates. The results of the phylogenetic tree analysis showed the presence of two main groups (A and B), with the predominant variants of CBLVd, i.e., CVd-I-LSS (Citrus viroid Low Sequence Similarity) sequences, clustering in subgroup A1 along with newly detected CVd-I-LSS from Palestinian sweet lime (Citrus limettioides), which has been identified as a new host of CVd-I-LSS in Pakistan. Further analysis of the sequences in subgroup A1 showed that the variant of CVd-I-LSS infecting citrus cultivars had a close relationship with isolates reported from China, Japan, and Iran, which may have resulted from the exchange of planting material. This study also unveiled the variability in nucleotide sequences of CBLVd, which made it unable to be detected by old primers. The results of this study indicate that the widespread presence of divergent variants of CBLVd is a major concern for the citrus industry in Pakistan and other countries where virulent isolates of CBLVd are prevalent. These findings suggest the need for future research on effective management and quarantine measures to stop the spread of CBLVd.

Spinach is a vegetable crop which is widely grown over a large area especially in Punjab province of Pakistan. Leaf curling and enations on spinach plant collected shown to be associated with the begomovirus Pedilanthus leaf curl virus (PeLCV) and Shahdadpur strain of Cotton leaf curl Multan betasatellite (CLCuMBSha). Defective molecules of half and quarter size derived from monopartite begomoviruses are usually generated by the deletion of virion-sense sequences. Characterization of defective molecules of PeLCV from spinach revealed that the molecules of half the size are derived from the deletion of complementary-sense genes while quarter size molecule appears to have evolved by further deletion. This is the first report of the begomovirus-betasatellite complex on spinach and unusual defective molecules derived from deletion of complementary-sense genes in Pakistan.

Cotton leaf curl disease (CLCuD) in Pakistan and northwestern India is caused by monopartite begomoviruses in association with an essential, disease-specific satellite, Cotton leaf curl Multan betasatellite (CLCuMB). Following a recent upsurge in CLCuD problems in Sindh province (southern Pakistan), sequences of clones of CLCuMB were obtained from Sindh and Punjab province (central Pakistan), where CLCuD has been a problem since the mid-1980s. The sequences were compared to all sequences of CLCuMB available in the databases. Analysis of the sequences shows extensive sequence variation in CLCuMB, most likely resulting from recombination. The range of sequence variants differ between Sindh, the Punjab and northwestern India. The possible significance of the findings with respect to movement of the CLCuD between the three regions is discussed. Additionally, the lack of sequence variation within the only coding sequence of CLCuMB suggests that the betasatellite is not involved in resistance breaking which became a problem after 2001 in the Punjab and subsequently also in northwestern India.

Cotton leaf curl disease (CLCuD) is a severe disease of cotton that occurs in Africa and Pakistan/northwestern India. The disease is caused by begomoviruses in association with specific betasatellites that differ between Africa and Asia. During survey of symptomatic cotton in Sindh (southern Pakistan) Cotton leaf curl Gezira virus (CLCuGV), the begomovirus associated with CLCuD in Africa, was identified. However, the cognate African betasatellite (Cotton leaf curl Gezira betasatellite) was not found. Instead, two Asian betasatellites, the CLCuD-associated Cotton leaf curl Multan betasatellite (CLCuMB) and Chilli leaf curl betasatellite (ChLCB) were identified. Inoculation of the experimental plant species Nicotiana benthamiana showed that CLCuGV was competent to maintain both CLCuMB and ChLCB. Interestingly, the enations typical of CLCuD were only induced by CLCuGV in the presence of CLCuMB. Also in infections involving both CLCuMB and ChLCB the enations typical of CLCuMB were less evident. This is the first time an African begomovirus has been identified on the Indian sub-continent, highlight the growing threat of begomoviruses and particularly the threat of CLCuD causing viruses to cotton cultivation in the rest of the world.