First Report of Potato virus Y in Potato in West Java, Indonesia.

Potato (Solanum tuberosum) is an important vegetable crop in Indonesia. A small survey was conducted for virus diseases in November 2011 in Lembang, West Java, as part of assessing the sanitary status of potatoes produced in farmers' fields. Among the six potato fields surveyed, one field had nearly 20% of plants displaying stunted growth with leaves showing mild chlorotic spots and reduced size of lamina. Tubers harvested from symptomatic plants showed no necrosis symptoms. Symptomatic leaves from three representative potato plants were positive for Potato virus Y (PVY) when tested with PVY-specific immunostrips (Agdia Inc., Elkhart, IN). Leaf samples from virus-positive plants were imprinted on FTA Classic Cards (Whatman International Ltd., Maidstone, UK), air dried, and shipped to Washington State University for confirmatory diagnostic tests. Total nucleic acids were eluted from FTA cards (1) and subjected to reverse transcription (RT)-PCR using primers (PVY/Y4A and PVY/Y3S) specific to the coat protein (CP) of PVY (3). Nucleic acid extracts from samples infected with PVY ordinary strain (PVYO), tuber necrosis strain (PVYNTN), tobacco veinal necrosis strains (PVYEU-N and PVYNA-N), and a recombinant strain (PVYN:O) were included as standards to validate RT-PCR assays. The approximately 480-bp DNA fragment, representing a portion of the CP, amplified in RT-PCR was cloned into pCR2.1 (Invitrogen Corp., Carlsbad, CA). DNA isolated from four independent recombinant clones was sequenced from both orientations. Pairwise comparison of these sequences (GenBank Accession Nos. KF261310 to 13) showed 100% identity among themselves and 93 to 100% identity with corresponding sequences of reference strains of PVY available in GenBank (JQ743609 to 21). To our knowledge, this study represents the first confirmed report of PVY in potato in West Java, Indonesia. Studies are in progress to assess the prevalence of PVY in other potato-growing regions of Indonesia and document the presence of different strains of the virus (2). Since the majority of farmers in Indonesia plant seed selected from their previous potato crop, there is an increased risk of primary and secondary spread of PVY through the informal seed supply system, leading to its increased significance to potato production in Indonesia. Therefore, strengthening foundation seed potato and supply chain programs will promote the production of virus-free potatoes in Indonesia. References: (1) O. J. Alabi et al. Plant Dis. 96:107, 2012. (2) A. Karasev and S. M. Gray. Am. J. Potato Res. 90:7, 2013. (3) R. P. Singh et al. J. Virol. Methods 59:189, 1996.

A begomovirus associated with Ageratum yellow vein disease in Indonesia: evidence for natural recombination between tomato leaf curl Java virus and Ageratum yellow vein virus-[Java].

A begomovirus (2747 nucleotides) and a satellite DNA beta component (1360 nucleotides) have been isolated from Ageratum conyzoides L. plants with yellow vein symptoms growing in Java, Indonesia. The begomovirus is most closely related to Tomato leaf curl Java virus (ToLCJV) (91 and 98% in the total nucleotide and coat protein amino acid sequences, respectively), although the products of ORFs C1 and C4 are more closely related to those of Ageratum yellow vein virus-[Java] (91 and 95% identity, respectively). For this reason, the begomovirus it is considered to be a strain of ToLCJV and is referred to as ToLCJV-Ageratum. The virus probably derives from a recombination event in which nucleotides 2389-2692 of ToLCJV have been replaced with the corresponding region of the AYVV-[Java] genome, which includes the 5' part of the intergenic region and the C1 and C4 ORFs. Infection of A. conyzoides with ToLCJV-Ageratum alone produced no symptoms, but co-infection with DNAbeta induced yellow vein symptoms. Symptoms induced in Nicotiana benthamiana by ToLCJV-Ageratum, ToLCJV and AYVV-[Java] are consistent with the exchange of pathogenicity determinant ORF C4 during recombination.

Molecular Characterization of Pepper yellow leaf curl Indonesia virus in Leaf Curl and Yellowing Diseased Tomato and Pepper in Indonesia.

Yellowing and leaf curl symptoms were observed in tomato and pepper fields near Bogor, Java, Indonesia in 2000. Samples were collected from one diseased tomato (Lycopersicum esculentum) and three diseased chili pepper (Capsicum annuum) plants. Viral DNA was extracted (2) and tested for the presence of geminiviral DNA-A, DNA-B, and associated satellite DNA using polymerase chain reaction (PCR) with previously described primers (1,3,4). The begomovirus DNA-A general primer pair PAL1v1978/PAR1c715 amplified the predicted 1.4-kb DNA fragment from the tomato and two of the chili samples. DNA-B and satellite DNA were not detected using PCR with DNA-B general primer pairs (DNABLC1/DNABLV2 and DNABLC2/DNABLV2) and satellite detection primer pair (Beta01/Beta02). The amplicons from the tomato and from one of the chili samples were cloned and sequenced. On the basis of the 1.4-kb DNA sequences, specific primers were designed to complete the DNA-A sequences. Following sequence assembly, the full-length DNA-A nucleotide sequences were determined as 2,744 nt (GenBank Accession No. DQ083765) for the tomato- and 2,743 nt (GenBank Accession No. DQ083764) for the chili-infecting begomoviruses. Sequence comparisons and analyses were conducted using the DNAMAN sequence analysis software (Lynnon Corporation, Quebec, Canada). The DNA-A of both begomoviruses contained six open reading frames, including two in the virus sense and four in the complementary sense, and the geminivirus conserved nanosequence-TAATATTAC in the loop of the hairpin structure of the intergenic region. Because of their high nucleotide sequence identities of 99%, the tomato- and chili-infecting begomovirus are considered the same virus. When compared by using BLAST with available gem-iniviral sequences in the GenBank database, the DNA-A sequences of the tomato and the chili isolates showed highest nucleotide sequence identity (95%) with the partially sequenced Pepper yellow leaf curl Indonesia virus (GenBank Accession No. AB189849) in the 1,842 nt to 660 nt region and in the 1,841 nt to 659 nt region, respectively. Comparisons with full-length DNA-A sequences of begomoviruses available in the GenBank database indicated high sequence identities of 76 and 77% for the tomato and chili isolates, respectively, with an eggplant isolate of Tomato yellow leaf curl Kanchanaburi virus (GenBank Accession No. AF511530) from Thailand. According to our knowledge, this is the first report of full-length DNA-A sequence of the Pepper yellow leaf curl Indonesia virus and its natural occurrence in tomato and pepper in the Bogor area of Indonesia. References: (1) R. W. Briddon et al. Virology 312:106, 2003. (2) R. L. Gilbertson et al. J. Gen. Virol. 72:2843, 1991. (3) S. K. Green et al. Plant Dis. 85:1286, 2001. (4) M. R. Rojas et al. Plant Dis. 77:340, 1993.

Pseudorecombination between infectious cloned DNA components of tomato mottle and bean dwarf mosaic geminiviruses.

A newly described whitefly-transmitted geminivirus infecting tomato plants in Florida induces yellow mottling symptoms on leaves, and stunted and distorted growth. The DNA-A and DNA-B components were cloned from extracts of field-infected tomato tissue; excised monomers or uncut tandem dimers of these clones were infectious when co-inoculated on to Nicotiana benthamiana by rub-inoculation. Tomato plants inoculated directly with the DNA-A and DNA-B dimers, or indirectly by sap or graft transmission from N. benthamiana plants previously infected with the dimers, developed symptoms similar to those observed in field-infected plants. This tomato geminivirus is different from previously characterized geminiviruses, and has been named tomato mottle geminivirus (ToMoV). DNA sequence comparisons revealed that ToMoV is closely related to bean dwarf mosaic geminivirus (BDMV) and abutilon mosaic geminivirus. Infectious pseudorecombinants were made by exchanging the cloned infectious DNA components of ToMoV and BDMV and inoculating N. benthamiana plants. The presence of the inoculated DNA components in systemically infected plants was confirmed by characterization of DNA-A and DNA-B fragments amplified by the polymerase chain reaction. This is the first report of pseudorecombination between two distinct geminiviruses. The implications of this finding in geminivirus evolution are discussed.