To better understand the genetic diversity of cucurbit-infecting poleroviruses in Taiwan, a survey was conducted in 2008 and 2009 as a prelude to screening germplasm for resistance. Out of 102 cucurbit samples showing yellowing symptoms collected across eight counties, 29 were identified as infected by polerovirus(es) by reverse transcription polymerase chain reaction (RT-PCR) using a set of universal polerovirus primers. Sequence analysis of a 1AE4-kb PCR product spanning part of the RNA-dependent RNA polymerase (RdRp) gene, the intergenic region (IR) and the complete coat protein (CP) gene of 13 of the 29 samples revealed the presence of three distinct Polerovirus species, namely Cucurbit aphid-borne yellows virus (CABYV), Melon aphid-borne yellows virus (MABYV) and Suakwa aphid-borne yellows virus (SABYV). In addition to the common strain of CABYV (CABYV-C) a recombinant strain of CABYV (CABYV-R) was identified. The recombinant strain probably arose by recombination in the IR, a recombination hot-spot of poleroviruses, between ancestors of MABYV and CABYV-C. RT-PCR protocols based on sets of specific primers were developed to distinguish between the different cucurbit-infecting polerovirus species and the two CABYV strains. These findings are discussed in relation to strategies for breeding for durable resistance against poleroviruses.

Pronounced yellowing symptoms on the lower leaves of tomato plants, similar to those caused by nitrogen deficiency, were observed in the spring of 1998 in The Asian Vegetable Research and Development Center and in farmers' fields in southern Taiwan. However, the brittleness of the discolored leaves, occasional upward leaf rolling, and abundance of whiteflies on these plants suggested the involvement of Tomato chlorosis virus (ToCV) and Tomato infectious chlorosis virus (TICV) that belong to the group of whitefly-transmitted, phloem-limited criniviruses (family Closteroviridae). Leaves of symptomatic and healthy plants were collected, and total nucleic acids were extracted from 0.2 g of leaf tissue (1). The total nucleic acids were precipitated by ethanol and dissolved in 160 μl of sterile water. Eight microliters of total nucleic acids were observed on positively charged nylon membranes (Roche Diagnostic GmbH, Roch Applied Science, Germany). Two digoxigenin-labeled riboprobes, transcribed from pTIC8-44 (complementary to the 3′-end region of TICV RNA 1) and pToC 78 (corresponding to the coat protein region of ToCV RNA 2), were used in hybridization tests to detect TICV and ToCV, respectively (2). Six of seventeen symptomatic tomato plant samples were positive with the ToCV probe, whereas none of the 13 samples reacted with the TICV probe. Similar symptoms as described above for tomato were observed on zinnia plants in the same locations. Five of eight zinnia samples gave a positive reaction with the ToCV probe. One of the ToCV positive samples also gave a positive reaction with the TICV probe. Electron microscopic examination from leaf-dip preparations of ToCV-positive leaf tissues, stained in 1% uranyl acetate, showed the presence of flexuous filamentous particles approximately 800 to 850 nm long. To our knowledge, this is the first evidence of the presence of ToCV and TICV in zinnia and ToCV in tomato in Taiwan. References: (1) A. Hadidi et al. J. Virol. Methods 30:261, 1990. (2) G. C. Wisler et al. Phytopathology 88:402, 1998.

Tospovirus isolates from tomato and watermelon in Taiwan and from peanut in India reacted neither with antibodies to tomato spotted wilt virus (TSWV), nor with antibodies to impatiens necrotic spot virus (INSV), in ELISA and in electroblot immunoassays. Additionally, all three tospovirus isolates had an N protein with a molecular weight of 32 kDa as compared to 29 kDa for the N protein to TSWV and INSV. The observed physicochemical and serological differences suggest the generation of a new, third species in the genus tospovirus for which we propose the name groundnut bud necrosis virus.

Production of tomato (Lycopersicon esculentum) in Bangladesh, Malaysia, Myanmar, Vietnam, and Laos has been severely affected by yellow leaf curl disease. Tomato leaf samples were collected from symptomatic tomato plants from farmers' fields in the five countries from 1997 to 1999. DNA was extracted from all samples, four from Vietnam, two each from Malaysia, Laos, and Myanmar, and seven from Bangladesh. Virus DNA was amplified by polymerase chain reaction (PCR) using the begomovirus-specific degenerate primer pair PAL1v 1978/PAR1c 715(1), which amplifies the top part of DNA A. All samples gave the expected 1.4-kb PCR product. The PCR product of one sample per country was cloned and sequenced. Based on the sequences of the 1.4-kb DNA products amplified by the first primer pair, specific primers were designed to complete each of the DNA A sequences. Computer-assisted sequence comparisons were performed with begomovirus sequences available in the laboratory at the Asian Vegetable Research and Development Center, Shanhua, Tainan, and in the GenBank sequence database. The five DNA species resembled DNA A of begomoviruses. For the detection of DNA B two degenerate primer pairs were used, DNABLC1/DNABLV2 and DNABLC2/DNABLV2 (DNABLC1: 5′-GTVAATGGRGTDCACTTCTG-3′, DNABLC2: 5′-RGTDCACTT CTGYARGATGC-3′, DNABLV2: 5′-GAGTAGTAGTGBAKGTTGCA-3′), which were specifically designed to amplify DNA B of Asian tomato geminiviruses. Only the virus associated with yellow leaf curl of tomato in Bangladesh was found to contain a DNA B component, which was detected with the DNABLC1/DNABLV2 primer pair. The DNA A sequence derived from the virus associated with tomato yellow leaf curl from Myanmar (GenBank Accession No. AF206674) showed highest sequence identity (94%) with tomato yellow leaf curl virus from Thailand (GenBank Accession No. X63015), suggesting that it is a closely related strain of this virus. The other four viruses were distinct begomoviruses, because their sequences shared less than 90% identity with known begomoviruses of tomato or other crops. The sequence derived from the virus associated with tomato yellow leaf curl from Vietnam (GenBank Accession No. AF264063) showed highest sequence identity (82%) with the virus associated with chili leaf curl from Malaysia (GenBank Accession No. AF414287), whereas the virus associated with yellow leaf curl symptoms in tomato in Bangladesh (GenBank Accession No. AF188481) had the highest sequence identity (88%) with a tobacco geminivirus from Yunnan, China (GenBank Accession No. AF240675). The sequence derived from the virus associated with tomato yellow leaf curl from Laos (GenBank Accession No. AF195782) had the highest sequence identity (88%) with the tomato begomovirus from Malaysia (GenBank Accession No. AF327436). This report provides further evidence of the great genetic diversity of tomato-infecting begomoviruses in Asia. Reference: M. R. Rojas et al. Plant Dis. 77:340, 1993.