A novel bipartite begomovirus, Blechum interveinal chlorosis virus (BleICV), was characterized at the genome level. Comparative analyses revealed that BleICV coat protein (CP) gene promoter is highly divergent from the equivalent region of other begomoviruses (BGVs), with the single exception of Tomato chino La Paz virus (ToChLPV) with which it shares a 23-bp phylogenetic footprint exhibiting dyad symmetry. Systematic examination of the homologous CP promoter segment of 132 New World BGVs revealed the existence of a quasi-palindromic DNA segment displaying a strongly conserved ACTT-(N7)-AAGT core. The spacer sequence between the palindromic motifs is constant in length, but its sequence is highly variable among viral species, presenting a relaxed consensus (TT)GGKCCCY, which is similar to the Conserved Late Element or CLE (GTGGTCCC), a putative TrAP-responsive element. The homologous CP promoter region of Old World BGVs exhibited a distinct organization, with the putative TATA-box overlapping the left half of the ACTT-N7 composite element. Similar CP promoter sequences, dubbed “TATA-associated composite element” or TACE, were found in viruses belonging to different Geminiviridae genera, hence hinting unsuspected evolutionary relationships among those lineages. To get cues about the TACE function, the regulatory function of the CLE was explored in distinct experimental systems. Transgenic tobacco plants harboring a GUS reporter gene driven by a promoter composed by CLE multimers expressed high beta-glucuronidase activity in absence of viral factors, and that expression was increased by begomovirus infection. On the other hand, the TrAP-responsiveness of a truncated CP promoter of Tomato golden mosaic virus (TGMV) was abolished by site-directed mutation of the only CLE present in it, whereas the artificial addition of one CLE to the -125 truncated promoter strongly enhanced the transactivation level in tobacco protoplasts. These results indicate that the CLE is a TrAP-responsive element, hence providing valuable clues to interpret the recurrent association of the CLE with the TACE. On the basis of the aforesaid direct evidences and the insights afforded by the extensive comparative analysis of BleICV CP promoter, we propose that the TACE might be involved in the TrAP-mediated derepression of CP gene in vascular tissues.

Soybean (Glycine max Merr.) is an alternative crop during the summer in Sinaloa, a northern state of Mexico. During the last 4 years, symptoms of yellowing, curled leaves, and stunting have been observed on soybean plantings, and a scrutiny of field samples collected in 2003 identified a begomovirus related to Pepper golden mosaic virus in symptomatic plants (4). A new survey was conducted during the summer of 2004 when the soybean disease was prevalent in the region. Affected plants appeared as patches displaying symptoms ranging from mild to severe yellow mosaic with leaf deformation and stunted growth in several parcels of commercial fields of northern Sinaloa. More than 100 samples from symptomatic soybean plants and weeds growing within the same fields were collected and analyzed for the presence of begomoviruses using DNA hybridization with the coat protein gene of Pepper huasteco yellow vein virus as a probe. Thirty-eight soybean, 12 Rhynchosia sp., and 14 sunflower hybridization-positive samples were subsequently used for polymerase chain reaction (PCR) amplification with the degenerate primers pRep-DGR and pCP70-Mot (1). PCR products were cloned into pGEM-T Easy vector (Promega, Madison, WI) and sequenced. The amplified viral DNA (915 nt) from two soybean plants, Sb1 and Sb2 (GenBank Accession Nos. AY955101 and AY957561, respectively), one isolate from Rhynchosia minima (GenBank Accession No. AY955102), and one from Heliantus annum (GenBank Accession No. AY957560) were sequenced and compared with DNA sequences available at NCBI database using BLAST. The highest sequence similarity was obtained with the two known isolates of Rhynchosia golden mosaic virus, RhGMV [Honduras] (GenBank Accession No. AF239671), and RhGMV [Chiapas] (GenBank Accession No. AF408199), displaying a nucleotide identity of approximately 89% with the Sinaloa isolates. Sequence comparisons of the latter isolates showed that viruses in the weeds were 97% identical to one of the soybean isolates, RhGMV-Sb1, but differed significantly (88% of nucleotide identity) from the second soybean isolate, RhGMV-Sb2. The complete genome A sequence of RhGMV-Sb1 was determined using PCR amplification of viral DNA with four degenerate primers recently described (2), cloning of overlapping PCR products into pGEM-T Easy vector (Promega) and sequencing. The 2,604-bp DNA-A of RhGMV-Sb1 (GenBank Accession No. DQ347950) was compared with the homologous genome of RhGMV [Chiapas] and RhGMV [Honduras] using the CLUSTAL alignment method (MegAlign, DNASTAR software, London) and an overall nucleotide identity of 89.2 and 88.6%, respectively, was determined. Current taxonomic criteria for begomoviruses establish that a DNA-A sequence identity lower than 93% with other isolates of a virus is indicative of a separate strain (3). Therefore, the virus identified in this study is a new strain of RhGMV that is provisionally named Rhynchosia golden mosaic virus-Soybean [Mexico:Sinaloa:2004]. This is the first soybean-infecting begomovirus from the American continent whose genome A has been completely characterized as of today. References: (1) J. T. Ascencio-Ibañez et al. Plant Dis. 86:692, 2002. (2) R. De La Torre-Almaraz et al. Plant Dis. 90:378, 2006. (3) C. Fauquet et al. Arch. Virol. 150:2151, 2005. (4) J. Mendez-Lozano et al. Plant Dis. 90:109, 2006.

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The Rioverde Valley is an important farming area of the San Luis Potosi State in the north-central region of Mexico, where a variety of horticultural crops (i.e., tomato, pepper, cucumber, and watermelon) are annually cultivated. In the summer of 2005, a number of plants exhibiting a variety of symptoms, including leaf yellowing, curling, and stunted growth, were observed in several tomato (Lycopersicon esculentum L.) fields. The presence of whiteflies (Bemisia tabaci Genn.) and symptoms seemed to suggest a begomoviral etiology. Leaves of 12 symptomatic tomato plants and seven plants of the weed Solanum rostratum (Dunal) growing into the same area were collected in July and September from several fields throughout the Rioverde area and assessed for the presence of begomoviruses (genus Begomovirus, family Geminiviridae) by PCR using the degenerate primers prRepDGR (CCTCCTCTAGCASWTCTNCCGTC), SL2050 (2), and prC889 (3). Amplicons of 1.4 kb were derived from viral DNA-A present in all examined S. rostratum and tomato samples, which were cloned into pGEM-T Easy Vector (Promega, Madison, WI) and subsequently analyzed by restriction fragment length polymorphism (RFLP) using MspI and HinfI. Several restriction fragment patterns were observed among the cloned PCR products, hence indicating the occurrence of different begomoviruses in the sampled fields. Sequencing of amplicons derived from one S. rostratum plant revealed the concurrent presence of Tomato severe leaf curl virus (ToSLCV; GenBank Accession No. DQ347946; [2]) and a distinct virus (GenBank Accession No. EF501978) displaying a high sequence identity with Tomato golden mottle virus from Guatemala (ToGMoV-GT94-R2; GenBank Accession No. AF32852). Restriction fragment patterns identical to that of the ToGMoV-like isolate were found in PCR clones from three additional S. rostratum plants and five tomato samples. A set of partially overlapping PCR products of 1.8 and 1.4 kb encompassing the complete DNA-A component of ToGMoV were obtained from one tomato sample by using two pairs of degenerate primers, prRepQGR-rev and prCP70 (1) and prRepDGR and prC889. Amplicons were cloned, sequenced, and compared with viral sequences available in the GenBank database using BlastN and Clustal V alignments (MegAlign, DNASTAR, Madison, WI). The 2,614-bp DNA-A sequence of the Rioverde isolate (GenBank Accession No. DQ520943) displays 93% sequence identity with the Guatemalan isolate of ToGMoV. In addition, a number of B. tabaci specimens of unidentified biotype were collected in one tomato field and total DNA was isolated from them by a modified Dellaporta method. Amplification of viral DNA present in the whiteflies was carried out and the PCR products were cloned and sequenced. One of the begomoviral DNA-A genomes isolated from the whiteflies (GenBank Accession No. EF501976) displayed 99% sequence identity with the virus isolated from plants. Previously, ToGMoV had been found only in Central America ( http://gemini.biosci.arizona.edu/viruses ), but this report considerably expands its known geographical distribution. References: (1) R. De La Torre-Almaraz et al. Plant Dis. 90:378, 2006. (2) J. A. Mauricio-Castillo et al. Plant Dis. 90:1116, 2006. (3) S. D. Wyatt and J. K. Brown. Phytopathology 86:1288, 1996.

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