Introduction of Cotton leaf curl Gezira virus into the United Arab Emirates

Abstract

Severe leaf curl and small vein thickening symptoms were observed in okra fields in Al-Ain, United Arab Emirates (UAE) during the winter season, 2013. These symptoms were reminiscent of those often associated with begomovirus infection. Based on the symptoms observed in okra plants growing in adjacent fields (20 × 20 m) on two small holding farms, the disease incidence ranged from 90 to 100%. The fields were infested with the whitefly Bemisia tabaci (Genn.), the insect vector of begomoviruses. Total DNA was extracted from four symptomatic okra leaves collected from two plants per field and used for PCR amplification of the core region of the begomovirus coat protein gene using the degenerate primers AVcore 3′-GCCHATRTAYAGRAAGCCMAGRAT-5′ and ACcore 3′-GGRTTDGARGCATGHGTACANGCC-5′. Amplicons of the expected size (~579 bp) were cloned and sequenced. BLASTn analysis of the partial coat protein sequences against the NCBI database revealed that the closet match to the four okra isolates was the Cotton leaf curl Gezira virus (CLCuGeV). CLCuGeV is a widespread Old World monopartite begomovirus described from the Nile Basin, sub-Saharan Africa, and southwestern Arabia (1). Recently, this virus has been reported in Jordan (GU945265) and Pakistan (3). To obtain the full-length viral genomic DNA for cloning and sequencing, total DNA extracts were enriched for circular dsDNA by rolling circle amplification (RCA) using Illustra TempliPhi (GE Healthcare, Life Sciences, Piscataway, NJ). The RCA products from each sample were digested with PstI, resulting in ~2.7 and 1.3 kbp molecules, respectively, and cloned into the pGEM plasmid vector (Promega, Madison, WI), linearized with PstI. Two inserts were selected from each cloning event and subjected to DNA sequencing using primer walking. The four resultant sequences of the 2.7 kbp (identified as virus genome) and the 1.3 kbp (betasatellite) inserts shared 99 to 100% nucleotide (nt) identity with each other, respectively. Therefore, only two representative genome and betasatellite sequences of 2,772 bp (KJ939446) and 1,356 bp (KM279620) were deposited in GenBank. Analysis using the Species Demarcation Tool (SDT) (v.1.0) (2) showed that the CLCuGeV UAE isolate sequence shared its highest nt identity (96 to 97%) with isolates from Egypt (AF155064), Pakistan (FR751142), and Jordan (GU945265). In contrast, it was only 93% identical to an isolate of CLCuGeV (HG530540) from the west coast of Saudi Arabia, nonetheless indicating they are all isolates of the same species. Analysis of the CLCuGeV sequence indicated that it was like other monopartite begomoviral genomes, containing a predicted hairpin, REP-binding iterons (GGTACTCA), and a TATA-box in the intergenic region. The genome contained six open reading frames encoding proteins with high homology to other CLCuGeV isolates. The 1,356-bp betasatellite shared its highest nt identity, at 97%, with the Okra leaf curl Oman betasatellite (KF267444) reported from infected okra plants in a neighboring country, Oman. The recent practice of transporting plants between the Gulf countries represents an important means and route for introducing begomoviruses among neighboring countries, compared to the long-distance aerial dispersal of viruliferous whiteflies, which is less likely because the Arabian desert poses a major barrier to long-distance whitefly flights. References: (1) A. Idris et al. Viruses 6:1219, 2014. (2) B. M. Muhire et al. Arch. Virol. 158:1411, 2013. (3) M. N. Tahir et al. PLoS ONE 6:E20366, 2011.

Molecular diagnostic development for begomovirus-betasatellite complexes undergoing diversification: A case study

“…Further, CLCuGeV from the Sahel region of Africa has recently been identified as an introduction into Pakistan (Amin et al, 2006) and certain locales in the Arabian Peninsula (Idris et al, 2014). The extensive SNPs apparent within these genomes appears to be indicative of diversification and perhaps genetic expansion, albeit, this group is poorly represented based on number of sequence in the public databases, and so requires additional attention.…”

“…An isolate of CLCuGeV has been identified in okra plants from Saudi Arabia (Al-Saleh et al, 2015;Idris et al, 2013), suggesting either viral endemism in the Arabian Peninsula, or the recent introduction on plants and/or by migrant whiteflies. The latter scenario is considered most likely, given that CLCuGeV also has been reported only recently in the United Arab Emirates (Idris et al, 2014).…”

Identification of a distinct strain of Cotton leaf curl Gezira virus infecting tomato in Oman

“…Pairwise multiple sequence alignments, using SDT (Muhire et al., ), were used calculate sequence identity values for all available CLCuGeV isolates (36 isolates available at this time). These findings showed the four sequences from tomato to have less than 94% nucleotide sequence identity to all other CLCuGeV isolates, with the highest levels (92.9%–93%) to an isolate recently identified in the United Arab Emirates (; Idris, Al‐Saleh, Amer, Abdalla, & Brown, ). This confirms the four sequences to be isolates of CLCuGeV, based on the 91% cut‐off for species demarcation of begomoviruses, as well as being isolates of a previously unidentified strain of this species, based on the 94% cut‐off for strain demarcation (Brown et al., ).…”

“…Pairwise multiple sequence alignments, using SDT (Muhire et al, 2014), were used calculate sequence identity values for all available CLCuGeV isolates (36 isolates available at this time). These findings showed the four sequences from tomato to have less than 94% nucleotide sequence identity to all other CLCuGeV isolates, with the highest levels (92.9%-93%) to an isolate recently identified in the United Arab Emirates (KJ939446; Idris, Al-Saleh, Amer, Abdalla, & Brown, 2014).…”

Minimal genomic variability in Merremia mosaic virus isolates endemic in Merremia spp and cultivated tomato in Puerto Rico

“…An equal volume of A. tumefaciens carrying pG-MerMVA11 and pG-MerMVB5 cloned components were mixed, and used to infiltrate tomato and N. benthamiana seedlings, as previously described [37]. Inoculated and mock-inoculated test plants were assayed for MerMV presence in newly developing leaves by PCR amplification using the ACcore/ AVcore primers [44].…”