Department of Plant Sciences, University of Arizona, Tucson 85721
Genetics Department, Cairo University, Egypt
Virology Laboratory, Research Institute, Universidad Del Valle, Guatemala
In 2000, geminivirus-like symptoms were widespread in muskmelon (Cucumis melo L.) fields (70 to 80% incidence) in Zacapa Valley, Guatemala. Muskmelon fields were infested with the whitefly Bemisia tabaci (Genn.), and plants exhibited patchy foliar chlorosis, leaf curling, and reduced fruit set, which is reminiscent of symptoms caused by certain whitefly-transmitted geminiviruses. Quarantine restrictions prevented experimental transmission experiments from being carried out with the whitefly vector or biolistic inoculation. Leaves collected from six symptomatic plants were assessed for the presence of begomovirus DNA by polymerase chain reaction (PCR) with the use of degenerate primers that amplify the core region of the coat protein (CP) gene of most begomoviruses (1). PCR products of the expected size (approximately 576 bp) were obtained from all three melon samples. The core CP amplicons were cloned, and their nucleotide sequences were compared. Nucleotide sequences of core CP fragments shared 99.7% identity, suggesting the presence of a single begomovirus in all assayed symptomatic melon plants. Two additional pairs of degenerate primers were used to obtain contiguous viral fragments containing the CP gene, the common region of the A component (CR-A; approximately 2,100 bp), and a fragment containing the CR of the B component (CR-B; approximately 1,100 bp), respectively (2). At least three amplicons obtained with each primer pair were cloned and their nucleotide sequence was determined. Virus-specific PCR primers were then designed within the CP open reading frame and used to obtain fragments that overlapped with the 2,100-bp fragment to yield an apparent full-length A component of 2,662 nucleotides (accession no. AF325497). CR-A and CR-B (accession no. AF325498) sequences (161 nucleotides) shared 98.1% identity and contained an identical directly repeated, replication-associated protein (REP) binding site: GGTGT CCT GGTGT. Nucleotide sequence alignment, with CLUSTAL W, of the melon virus A-component with that of other well-studied begomoviruses revealed that its closest relatives were members of the Squash leaf curl virus (SLCV) group. The melon virus from Guatemala shared its greatest sequence identity, 83.1%, with SLCV extended (SLCV-E) (accession no. M38183), indicating that it is a new, previously unidentified begomovirus species, herein referred to as Melon chlorotic leaf curl virus (MCLCV). The next closest relatives of MCLCV were SLCV restricted (SLCV-R; 78.6%) (S. G. Lazarowitz, unpublished) Cucurbit leaf curl virus-Arizona (CuLCV-AZ; accession no. AF256199; 74.1%) (3), Cabbage leaf curl virus (CaLCV; 72.0%), Bean calico mosaic virus (BCMoV; 71.7%), and Texas pepper virus-Tamaulipas (71.4%). Additionally, the theoretical REP binding element, GGTGT, is 100% identical among MCLCV and BCMoV, CaLCV, CuLCV-AZ, SLCV-E, and SLCV-R. On the basis of shared nucleotide sequence identities with other begomoviruses described to date and the presence of B. tabaci in melon fields, it is likely that MCLCV also is whitefly-transmitted. Collectively, CP and CR sequences suggest that MCLCV is a new species of the SLCV lineage that contains other bipartite begomoviruses indigenous to Central America, Mexico, and the U.S. Sunbelt states.
References: (1) S. D. Wyatt and J. K. Brown. Phytopathology 86:1288, 1996. (2) A. M. Idris and J. K Brown. Phytopathology 88:648, 1998. (3) J. K. Brown et al. Plant Dis. 84:809, 2000.