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.
Influenza A viruses evolve at a high rate requiring continuous monitoring to maintain the efficacy of vaccines and antiviral drugs. We performed next generation sequencing analysis of 100 influenza A/H3N2 isolates collected in four Asian countries (Japan, Lebanon, Myanmar, and Vietnam) during 2012-2015. Phylogenetic analysis revealed several reassortment events leading to the circulation of multiple clades within the same season. This was particularly evident during the 2013 and 2013/2014 seasons. Importantly, our data showed that certain lineages appeared to be fitter and were able to persist into the following season. The majority of A/H3N2 viruses continued to harbor the M2-S31N mutation conferring amantadine-resistance. In addition, an S31D mutation in the M2-protein, conferring a similar level of resistance as the S31N mutation, was detected in three isolates obtained in Japan during the 2014/2015 season. None of the isolates possessed the NA-H274Y mutation conferring oseltamivir-resistance, though a few isolates were found to contain mutations at the catalytic residue 151 (D151A/G/N or V) of the NA protein. These variations did not alter the susceptibility to neuraminidase inhibitors and were not detected in the original clinical specimens, suggesting that they had been acquired during their passage in MDCK cells. Novel polymorphisms were detected in the PB1-F2 open-reading frame resulting in truncations in the protein of 24-34 aminoacids in length. Thus, this study has demonstrated the utility of monitoring the full genome of influenza viruses to allow the detection of the potentially fittest lineages. This enhances our ability to predict the strain(s) most likely to persist into the following seasons and predict the potential degree of vaccine match or mismatch with the seasonal influenza season for that year. This will enable the public health and clinical teams to prepare for any related healthcare burden, depending on whether the vaccine match is predicted to be good or poor for that season.