RESEARCH COMMUNICATIONS CURRENT SCIENCE, VOL. 86, NO. 6, 25 MARCH 2004 845 *For correspondence. (e-mail: vgmalathi@rediffmail.com) Yellow mosaic virus infecting soybean in northern India is distinct from the species infecting soybean in southern and western India K. S. Usharani, B. Surendranath, Q. M. R. Haq † and V. G. Malathi* Plant Virology Unit, Division of Plant Pathology, Indian Agricultural Research Institute, New Delhi 110 012, India † Department of Biosciences, Jamia Millia Islamia University, New Delhi 110 025, India Genomic components of the begomovirus causing yellow mosaic disease (YMD) in soybean in Delhi were clo- ned, sequenced and infectivity proved. Nucleotide se- quence analysis of the virus isolate revealed more than 89% identity with Mungbean yellow mosaic India virus (MYMIV) and therefore it is designated as soy- bean isolate of MYMIV (MYMIV-[Sb]). Total nucleo- tide and predicted amino acid sequence analysis of MYMIV-[Sb] with other yellow mosaic virus isolates infecting legumes established dichotomy of the isolates into two species namely MYMIV and mungbean yellow mosaic virus (MYMV ). Involvement of at least two distinct viruses in the etiology of soybean YMD in India is indicated here. Y ELLOW mosaic diseases (YMD) are major constraints in improving the productivity of grain legumes in India. Yield loss per annum due to YMD was estimated to be $ 300 million taking blackgram, mungbean and soybean together 1 . YMD of soybean was first observed in North India 2,3 as early as 1970s and since then it had spread at alarming proportions. Whitefly transmission, enzyme linked immunosorbent assay (ELISA) and immuno-spe- cific electron microscopic (ISEM) studies suggested that the aetiological virus causing YMD in soybean is a bego- movirus of the family Geminiviridae. Begomoviruses have characteristic icosahedral geminate particles that encap- sidate genome of circular single-stranded DNA. They in- fect dicots and are transmitted by the whitefly Bemisia tabaci , Gennadius. They have monopartite or bipartite genome. In bipartite begomoviruses, DNA A encodes proteins required for replication, transcription and encap- sidation whereas DNA B encodes proteins required for movement functions 4 . For efficient management of YMD, the relationship bet- ween the yellow mosaic viruses (YMV) infecting different legumes needs to be understood. The present communica- tion attempts to resolve the identity of the virus causing YMD in soybean in North India and examine its relation- ship to yellow mosaic virus (YMV) isolates infecting soybean and other legumes in Indian subcontinent and Thailand. The viral inoculum from soybean plants affected by severe YMD in the fields of Indian Agricultural Research Institute (IARI), New Delhi was established in the glass- house on soybean cv. ‘Bragg’ through whitefly inocula- tion giving 24 h each of acquisition and inoculation access periods. It was readily transmissible by whitefly vector to blackgram, cowpea, mungbean and soybean wherein it produced typical yellow and golden mosaic symptoms. Total nucleic acid was isolated from infected soybean plants 17 days after inoculation by cetyl trimethyl ammonium bromide (CTAB) method 5 . Double-stranded viral replicative forms were separated from the host DNA by cesium chloride density gradient centrifugation 6 and used for cloning at HindIII, BamHI, EcoRI and PstI sites. Cloning at BamHI site in pUC18 vector yielded two dif- ferent types of recombinant clones with unit genome length (∼ 2.7 kb) insert which were identified to represent DNA A and B components of begomovirus. The putative DNA A and B clones were confirmed by Southern blot analysis using total DNA extracted from infected soybean plants. Agroinoculation of soybean plants cv. ‘Bragg’ was done by sprouted-seed method 7 . Sequencing was done for DNA A and B component clones in an automated DNA sequencer (ABI Prism, Perkin Elmer at University of Delhi, South Campus, New Delhi). The sequence data were assembled and analysed using the software programme Bioedit version 5.0.9. Multiple nucleotide and predicted amino acid sequence alignments were done in CLUSTALW (1.7) programme (http://www.ebi.ac.uk/clustalw). Comparisons were made with begomovirus DNA sequences obtained from the GenBank sequence database (Table 1) and their nomen- clature is as given in Fauquet and Stanley 8 . Dendrograms were constructed from the aligned sequences using the neighbour-joining method and bootstrap option of Tree- con version 1.3 b (500 × bootstrap replicates) 9 . The nucleotide sequences of DNA A and B components were determined for both the strands and have been depo- sited in DDJB, EMBL and GenBank database under ac- cession numbers AY049772 and AY049771 respectively. Viral sequences were aligned, ORFs were deciphered and the genome organization was found to have typical fea- tures of begomoviruses 4 . There are two virion sense genes, ORF AV2 and ORF AV1 (coat protein, CP) in DNA A and one ORF BV1 (nuclear shuttle protein, NSP) in DNA B. The complementary sense genes are ORF AC1 (repli- cation initiation protein, Rep), AC2 (transcription activa- tor protein, TrAP), AC3 (replication enhancer protein, REn), AC4 and AC5 in DNA A and ORF BC1 (move- ment protein, MP) in DNA B. Complete nucleotide se- quence analysis of DNA A showed > 89% identity with MYMIV and hence this isolate is designated as MYMIV- [Sb] as per the guidelines suggested by ICTV study group 8 . The clones were non sap-transmissible and infec- tious by agroinoculation on soybean. Typical symptoms like yellow mosaic and crinkling on the leaf and stunted