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First report of Tomato leaf curl New Delhi virus infecting Bitter Gourd in Pakistan M. Tahir and M.S. Haider* School of Biological Sciences, University of the Punjab, Quaid-i-Azam Campus, Lahore, Pakistan Accepted for publication 31/01/05 Bitter Gourd (Momordica charantia; family Cucurbitaceae) is a vegetable cultivated in many areas of Pakistan. Plants showing yellow blotch symptoms were observed in several fields in the vicinity of Lahore, Pakistan, with an average incidence of 60-70%. Leaf samples were collected from 4 symptomatic and from 2 apparently healthy (symptomless) plants and used for DNA extraction. The presence of a begomovirus was confirmed by PCR amplification using a degenerate primer pair, designed to conserved regions of the coat protein genes from published sequences of begomoviruses from the Old World (Haider, 1996; virion-sense primer 5'-ATG(C/A/T)(G/C) (G/C/A)AAGCG(A/T)(C/A)C(A/C)G(G/C)(A/C)GATAT-3'; complementary-sense primer 5'-TTAATT(T/G/C)(C/G/A)(A/T/C)(A/T/G)A(C/T)(A/T/C)(G/C)(C/A/T)(A/G)TCATA(G/A)AA(A/G)TA-3'). An amplification product of the expected size (approx. 750 bp) was produced from symptomatic samples but not from the symptomless plants. The PCR product was cloned and sequenced (Acc. No. AJ854186). The sequence showed the highest levels of sequence identity (95%) to Tomato leaf curl New Delhi virus (ToLCNDV; U15015), indicating that the virus of M. charantia is a strain of ToLCNDV. Recent reports indicate that many Old World begomoviruses are associated with a single-stranded DNA satellite (DNA β; Briddon et al., 2003). Attempts to identify the presence of a DNA β in infected M. charantia using universal DNA β primers (Briddon et al., 2002) produced a 0.6 kb band that may be a defective DNA β. Efforts are now underway to sequence this and to produce full-length clone(s) of the virus. To the best of our knowledge this is the first report of ToLCNDV infecting M. charantia. 
Figure 1: Phylogenetic dendrogram derived from an alignment of the partial nucleotide sequences of the coat protein genes of selected begomoviruses with the sequence obtained from M. charantia. The tree was rooted on the DNA A sequence of Tomato mottle virus (ToMoV), a distantly related begomovirus originating from the new World. The numbers at nodes indicate bootstrap confidence values (100 replicates). The position of the sequence originating from M. charantia is indicated with an arrow. Acknowledgements The authors would like to thank Dr Rob Briddon (Higher Education Commission [Pakistan] - Foreign Faculty Hiring Programme) for assistance in preparing the manuscript. References Briddon RW, Bull SE, Amin I, Idris AM, Mansoor S, Bedford ID, Dhawan P, Rishi N, Siwatch SS, Abdel-Salam AM, Brown JK, Zafar Y, Markham PG, 2003. Diversity of DNA; a satellite molecule associated with some monopartite begomoviruses. Virology 312, 106-121. Briddon RW, Bull SE, Mansoor S, Amin I, Markham PG, 2002. Universal primers for the PCR-mediated amplification of DNA b; a molecule associated with some monopartite begomoviruses. Molecular Biotechnology 20, 315-318. Haider MS, 1996. Characterization of whitefly-transmitted geminiviruses from Pakistan. Ph.D thesis, University of London. |