Tomato Yellow Leaf Curl Kanchanaburi Research Articles

Molecular Detection and Identification of Begomovirus Infecting Cucumber (Cucumis sativus) in Terengganu, Malaysia

The genus Begomovirus from the family Geminiviridae is responsible for causing significant economic losses to many important horticultural crops, including cucumber (Cucumis sativus L.). Begomovirus infection during the early stages of plant growth can lead to complete yield loss. Hence, the identification of begomovirus species is important to design a precise resistant breeding strategy. This study aims to detect the presence of begomovirus in typical symptomatic cucumber leaves, identify the species of begomovirus present, and investigate the evolutionary relationships with other reported begomoviruses using phylogenetic analysis. Leaf samples from symptomatic cucumber plants were collected from the Green World Genetics (GWG) research station and a farm in Lembah Bidong, Rhu Tapai, Setiu, Terengganu. To detect the presence of begomovirus, PCR was carried out using universal primers targeting DNA-A, DNA-B, and betasatellite regions. DNA-A and betasatellite fragments were amplified, but not DNA-B. The amplified partial sequences of DNA-A were then analysed and compared with other begomovirus sequences in the GenBank database managed by the National Centre for Biotechnology Information (NCBI). The newly isolated DNA-A sequence from cucumber was 100% identical to the tomato leaf curl New Delhi virus (ToLCNDV). The phylogenetic tree was divided into two groups: group A, consisting of the newly isolated DNA-A sequence from cucumber, ToLCNDV, followed by squash leaf curl China virus (SLCCNV) and a monopartite begomovirus Ageratum yellow vein virus (AYVV); and group B, consisting of tomato yellow leaf curl Kanchanaburi virus (TYLCKaV), pepper yellow leaf curl Indonesia virus (PepYLCIV), and pepper yellow leaf curl Aceh virus (PepYLCAV). For the first time in cucumber, betasatellite is reported in association with ToLCNDV, a bipartite old-world begomovirus. This study provides a basis for the selection and breeding of begomovirus-resistant cucumber varieties in the future.

Current Status and Complexity of Three Begomovirus Species in Pepper Plants in Lowlands and Highlands in Java Island, Indonesia.

Three primary species from the Begomovirus genus, Pepper yellow leaf curl Indonesia virus (PepYLCIV), Tomato yellow leaf curl Kanchanaburi virus (TYLCKaV), and Tomato leaf curl New Delhi virus (ToLCNDV), are suspected of spreading throughout pepper production centers, and plants are infected by a single species or a combination of two or three species. This study was conducted to provide complete information about the symptoms, incidence and severity, whitefly biotypes, as well as the dominance status of the three Begomovirus species in pepper-producing areas in Java. A DNA analysis was carried out on leaf samples to identify Begomovirus species and biotypes of B. tabaci collected from 18 areas (16 districts) in lowlands (<400 m asl) and highlands (>700 m asl). The DNA analysis showed that B. tabaci biotype B was the most commonly detected in all locations compared to the A, AN, and Q biotypes. The incidence of begomovirus infection was at a high level, 93% and 88.78% in the lowlands and highlands, respectively. However, the severity of begomovirus infection was significantly higher in the lowlands (54.50%) than in the highlands (38.11%). A single infection of PepYLCIV was most dominant in all locations sampled and caused severe infection, followed by a mixed infection with TYLCKaV. Therefore, the current status of begomovirus infection, especially PepYLCIV, can provide advice to farmers using more tolerant and resistant varieties as well as a breeding strategy for resistant pepper varieties.

Identification of begomoviruses associated with the insect vector Bemisia tabaci and various host plants on Java Island, Indonesia.

Begomoviruses are economically important plant viruses and are transmitted by Bemisia tabaci which is a complex of various cryptic species. However, it is uncertain whether most begomoviruses that infect host plants are transmitted by B. tabaci at a similar rate. We compared the begomovirus profiles that were detected in a total of 37 whitefly populations and 52 host plants on Java Island, Indonesia. Seven begomovirus species were detected in B. tabaci at different rates: pepper yellow leaf curl Indonesia virus (PepYLCIV, 56.8%), tomato yellow leaf curl Kanchanaburi virus (TYLCKaV, 46.0%), tomato leaf curl New Delhi virus (ToLCNDV, 21.6%), squash leaf curl China virus (SLCCNV, 21.6%), ageratum yellow vein China virus (AYVCNV, 2.7%), mungbean yellow mosaic India virus (MYMIV, 2.7%), and okra enation leaf curl virus (OELCuV, 2.7%). The begomoviruses were detected at different rates in three cryptic species of B. tabaci. In addition, six begomovirus species were detected in the various host plants at different rates: PepYLCIV (67.3%), TYLCKaV (53.9%), ToLCNDV (13.5%), MYMIV (11.5%), AYVCNV (3.9%), and Tomato yellow leaf curl Thailand virus (TYLCTHV) (1.9%). By comparing the virus presence between whiteflies and plants, five begomoviruses (AYVCNV, MYMIV, PepYLCIV, ToLCNDV, and TYLCKaV) were detected in both samples, but their sequence similarity was highly variable depending on the begomovirus themselves; TYLCKaV was highest (99.4%-100%) than any other viruses. Our study suggests B. tabaci acquire begomoviruses at different rates from plants. This study provides important information on the potential variation in the begomovirus transmission mechanism.

Incidence and Molecular Identification of Begomoviruses Infecting Tomato and Pepper in Myanmar.

In Myanmar, yellow mosaic and leaf curl diseases caused by whitefly-transmitted begomoviruses are serious problems for vegetables such as tomatoes and peppers. To investigate the incidence of begomoviruses in Myanmar between 2017 and 2019, a field survey of tomato and pepper plants with virus-like symptoms was conducted in the Naypyitaw, Tatkon, and Mohnyin areas of Myanmar. Among the 59 samples subjected to begomovirus detection using polymerase chain reaction, 59.3% were infected with begomoviruses. Complete genome sequences using rolling circle amplification identified five begomovirus species: tomato yellow leaf curl Thailand virus (TYLCTHV), tomato yellow leaf curl Kanchanaburi virus (TYLCKaV), tobacco leaf curl Yunnan virus (TbLCYnV), chili leaf curl Pakistan virus (ChiLCV/PK), and tobacco curly shoot Myanmar virus (TbCSV-[Myanmar]). Excluding the previously reported TYLCTHV, three begomoviruses (ChiLCV/PK, TYLCKaV, and TbLCYnV) were identified in Myanmar for the first time. Based on the 91% demarcation threshold of begomovirus species, TbCSV-[Myanmar] was identified as a new species in this study. Among these, ChiLCV/PK and TbCSV-[Myanmar] were the most predominant in tomato and pepper fields in Myanmar. Identification of begomovirus species may be helpful for predicting the origin of viruses and preventing their spread.

Tomato yellow leaf curl Kanchanaburi virus

This datasheet on Tomato yellow leaf curl Kanchanaburi virus covers Identity, Distribution, Hosts/Species Affected, Vectors &amp; Intermediate Hosts, Further Information.

Simultaneous detection of virus infecting tobacco plant using multiplex-PCR in Klaten, Indonesia

Tobacco is one of the important plantation commodities in Indonesia. The multiple virus attack in the field causes crop damage to crop failure. Therefore, detection is needed quickly and accurately as a basis for control strategies. This study aimed to determine the viruses that attack tobacco plants in Klaten, Indonesia through simultaneous detection using multiplex-PCR. The study was conducted by observation in the field then detection in the Laboratory. Based on Multiplex-PCR detection results using primers of MJ 1/MJ 2, Krusty/Homer, and CMV P1/P2 showed that tobacco plants infected with several viruses from the genus Potyvirus, Begomovirus and Cucumovirus. Based on the results of the nucleotide sequencing blast, it was found that the tobacco isolate was positively infected by the Tomato yellow leaf curl Kanchanaburi virus (TYLCV), Cucumber mosaic virus (CMV) and Potato virus (PVY).

Identification of the determinant of tomato yellow leaf curl Kanchanaburi virus infectivity in tomato

Geminiviruses cause devastating diseases in solanaceous crops, with the bipartite begomoviruses tomato yellow leaf curl Kanchanaburi virus (TYLCKaV) and pepper yellow leaf curl Thailand virus (PYLCThV) major threats in Southeast Asia. To determine the molecular mechanism of geminivirus infection, we constructed infectious clones of TYLCKaV and PYLCThV. Both constructs infected Nicotiana benthamiana, but only TYLCKaV could infect Solanum lycopersicum ‘A39’. A genome-swapping of TYLCKaV with PYLCThV revealed the TYLCKaV-B genome segment as the determinant of TYLCKaV infectivity in tomato. We constructed five geminivirus clones with chimeric TYLCKaV-B and PYLCThV-B genome segments to narrow down the region determining TYLCKaV infectivity in tomato. Only chimeric clones carrying the TYLCKaV intergenic region (IR) showed infectivity in S. lycopersicum ‘A39’, indicating that the IR of TYLCKaV-B is essential for TYLCKaV infectivity in tomato. Our results provide a foundation for elucidating the molecular mechanism of geminivirus infection in plants.

Ty-2 and Ty-3a Conferred Resistance are Insufficient Against Tomato Yellow Leaf Curl Kanchanaburi Virus from Southeast Asia in Single or Mixed Infections of Tomato.

Tomato yellow leaf curl virus (TYLCV), a monopartite begomovirus that originated in the eastern Mediterranean, has spread worldwide, becoming a serious threat to tomato (Solanum lycopersicum L.) production. Southeast Asia is considered one of the hotspots for begomovirus diversity, and a wide variety of local begomovirus species distinct from TYLCV have been identified. In this study, the protection effect of introgressions of single TYLCV Ty resistance genes, Ty-2 and Ty-3a, in tomato was examined against inoculations of the bipartite begomoviruses Tomato yellow leaf curl Kanchanaburi virus (TYLCKaV) and Pepper yellow leaf curl Indonesia virus (PepYLCIV) isolated from Indonesia. Our findings suggest that Ty-2 in the heterozygous state was found to be ineffective against PepYLCIV and TYLCKaV, whereas Ty-3a in the heterozygous state was effective against PepYLCIV and partially effective against TYLCKaV. Quantification of viral DNAs showed correlation between symptom expression and viral DNA accumulation. Moreover, mixed infections of TYLCKaV and PepYLCIV caused notably severe symptoms in tomato plants harboring Ty-3a. In cases of mixed infection, quantifying viral DNAs showed a relatively high accumulation of PepYLCIV, indicating that Ty-3a loses its effectiveness against PepYLCIV when TYLCKaV is also present. This study demonstrates the lack of effectiveness of Ty resistance genes against single and mixed infections of distinct local begomoviruses from Southeast Asia.

Identification and characterization of Ageratum yellow vein Malaysia virus (AYVMV) and an associated betasatellite among begomoviruses infecting Solanum lycopersicum in Malaysia.

The study describes results of a survey of tomato fields for the presence of begomoviruses from different regions of Peninsular Malaysia. An ORF-based (C2 and C3) study was performed to determine the distribution of begomoviruses associated with a severe leaf curl disease in tomato-growing areas of Peninsular Malaysia. Viral DNA was isolated from symptomatic tomato plants, and begomovirus association was confirmed by PCR using DNA-A degenerate primers. The C2 and C3 sequences of the putative begomoviruses were similar to two corresponded ORFs of different geographically separated strains of begomoviruses: Pepper yellow leaf curl Indonesia virus and Tomato yellow leaf curl Kanchanaburi virus. The present study also identified a unique isolate, Ageratum yellow vein Malaysia virus (AYVMV) among above mentioned survey. It has a single-stranded DNA component and its associated betasatellite. The single-stranded DNA component is consisting of 2750 nt with six open reading frames and an organization resembling that of monopartite geminiviruses. The full length of viral single-stranded DNA component genome obtained using next generation sequencing (NGS) showed the highest sequence identity (99%) with Ageratum yellow vein virus (AYVV-BA). The betasatellite component genome obtained by NGS has 1342 nt and showed the highest sequence identity (91%) with the Pepper yellow leaf curl betasatellite. Following ICTV guidelines, Ageratum yellow vein Malaysia virus was assigned the abbreviation AYVMV with sequence and phylogenetic analysis indicating that it might have evolved by recombination of two or more viral ancestors.

First report of naturally occurring recombinant non-coding DNA satellite associated with Tomato yellow leaf curl Kanchanaburi virus on eggplant in Indonesia

Abstract. Kandito A, Hartono S, Sulandari S, Somowiyarjo S, Widyasari YA. 2019. First report of naturally occurring recombinant non-coding DNA satellite associated with Tomato yellow leaf curl Kanchanaburi virus on eggplant in Indonesia. Biodiversitas 20: 129-136. Begomovirus is a viral genus which is a major impediment in the cultivation of plants, especially Solanaceae. Tomato yellow leaf curl Kanchanaburi virus (TYLCKaV) is a member of the Begomovirus genus that is widely reported to infect eggplant and results in severe symptoms of chlorosis. In addition to the type of virus that infects, the severity of symptoms can also be caused by the presence of betasatellite associated with several begomovirus species. The presence of betasatellite that is related to several begomovirus species can lead to more severe symptoms. The aim of this study was to identify satellites associated with begomovirus in Indonesia. Eggplant samples from Bantul, Special Region of Yogyakarta show with mosaic and severe chlorotic symptoms due to Begomovirus infection. Total DNA samples were subject to PCR amplification using universal primer for begomovirus PALIV1978/PAR1C715 and specific primer for betasatellite β01/β02. The PCR amplification produced a DNA band measuring ± 1600bp, identical size of begomovirus. The amplification of the betasatellite specific primer produced a DNA band featuring ± 1300bp, identical size of betasatellites that were associated with begomovirus in the sample. The results of DNA sequencing, suggested that begomovirus in this study had a close relationship with TYLCKaV from Thailand and Indonesia. Characterization of the satellite-based on nucleotide sequence revealed the presence of stem-loop structures, satellite conservative regions, and adenine-rich regions that resembled structures present in the betasatellite, which were associated with Tomato leaf curl virus and Ageratum yellow vein virus, with no ORF found. These results indicated that the TYLCKaV could be associated with a non-coding satellite. This result is the first report regarding the TYLCKaV association with a non-coding satellite in Indonesia.

A survey of mixed Begomovirus infection in solanaceae and fabaceae at different altitudes in East Java, Indonesia

A multiplex primer set was developed to detect four Begomoviruses in East Java, Indonesia, i.e. Tomato leaf curl New Delhi virus (ToLCNDV), Tomato yellow leaf curl Kanchanaburi virus (TYLCKaV), Pepper yellow leaf curl Indonesia virus (PepYLCIV) and Mungbean yellow mosaic India virus (MYMIV). Survey at different altitudes found that begomoviruses infecting pepper, tomato and long bean were more variable, while in eggplant and string bean were more uniform. As a single virus, TYLCKaV infected eggplant, and sometimes tomato and pepper; PepYLCIV infected pepper, tomato and long bean; ToLCNDV only infected long bean and tomato at low frequency; and MYMIV infected beans. Mixed infection occurred more frequently in the low altitude areas. Subsequent examination indicated that Cucumber mosaic virus (CMV) and potyviruses were also responsible for diseased fabaceous. Our data suggest a relationship between altitudes and virus species occurrence. However, which viral species infects a crop is mainly influenced by the crop rather than by altitude.

Detection and identification of Begomovirus infecting Cucurbitaceae and Solanaceae in Yogyakarta, Indonesia

Abstract. Subiastuti AS, Hartono S, Daryono BS. 2019. Detection and Identification of Begomovirus infecting Cucurbitaceae and Solanaceae in Yogyakarta, Indonesia. Biodiversitas 20: 738-744. Begomovirus genome has high plasticity that led to evolve rapidly. Begomovirus is one of a remarkably successful group of emerging viruses as the results from combination of many factors. Planting systems in Indonesia which often overlapping two or more plant species in one land has high possibility for occurring mixed infection. It is also suggested has high contribution to increase Begomovirus diversity. The aim of this research is to do preliminary identification of Begomovirus infected-Solanaceae and Cucurbitaceae in Yogyakarta based core coat protein (CP) gene sequence. A total of 50 melon, 50 chili, 30 eggplants, and 30 watermelon samples which showed Begomovirus symptoms were observed from several fields in Yogyakarta and Purworejo, Indonesia during 2016. Almost 90% of infected samples for each plant were tested by PCR and showed positive for Begomovirus. Based on coat protein (CP) gene nucleotide sequence identity, Begomovirus infected Solanaceae in Indonesia has close relationship with Pepper yellow leaf curl Indonesia virus (PepYLCIV) and Tomato yellow leaf curl Kanchanaburi virus (TYLCVKaV), while in Cucurbitaceae has close relationship with Squash leaf curl China virus (SLCCV) Tomato leaf curl New Delhi virus (ToLCNDV). All collected isolates showed highest sequence identity with isolates from South-East Asia and China. However, further analysis that including full genome characterization is still needed to explain Begomovirus evolution in Indonesia.

Development of a LAMP assay with a portable device for real-time detection of begomoviruses under field conditions

The emergence of begomovirus infection is one of the most important problems affecting production of a variety of vegetable crops worldwide. Infection by begomoviruses has been detected and spread rapidly on Cucurbitaceae and Solanaceae plants in Indonesia. A rapid and simple detection assay for begomoviruses under field conditions for routine sampling of plants is needed. Primers for a loop-mediated isothermal amplification (LAMP) assay were designed based on the sequences of three Indonesian begomoviruses, namely Tomato leaf curl New Delhi virus (ToLCNDV), Pepper yellow leaf curl Indonesia virus (PepYLCIV), and Tomato yellow leaf curl Kanchanaburi virus (TYLCKaV), infecting Cucurbitaceae and Solanaceae plants. LAMP assays using a Genelyzer™ III portable fluorometer with a toothpick method successfully detected these begomoviruses in infected melon, pepper, and eggplant samples. LAMP assays conducted during a field survey for detection of the three begomoviruses on 104 fresh leaves indicated that most of the samples were positive; the findings were confirmed by PCR using universal primers of begomovirus as a common detection method. These results demonstrate that this simple and rapid LAMP assay using a fluorometer portable device may be used to achieve real-time detection of begomoviruses under field conditions.

First Report of Tomato yellow leaf curl Kanchanaburi virus Infecting Eggplant in China

First Report of Tomato yellow leaf curl Kanchanaburi virus Infecting Eggplant in China

Highly Efficient Agroinoculation Method for Tomato Plants with &lt;i&gt;Tomato Yellow Leaf Curl Kanchanaburi Virus&lt;/i&gt;

Tomato yellow leaf curl disease caused by begomoviruses is a serious threat to tomato (Solanum lycopersicum L.) production. If begomoviruses, transmitted by whitefly (Bemisia tabaci), infect tomato plants during early growth, production can be almost entirely lost. Tomato yellow leaf curl Kanchanaburi virus (TYLCKaV), a bipartite Begomovirus, is emerging as an important threat to solanaceous crop production in Southeast Asia. The lack of mechanical transmission of some begomoviruses is a major experimental constraint. In this study, an agroinoculation method using TYLCKaV in tomato plants was established. Partial tandem repeats of TYLCKaV DNA A and DNA B were constructed and cloned to a binary pGreenII vector, and their infectivity was tested. Co-inoculation of TYLCKaV DNA A and DNA B to Nicotiana benthamiana L. produced typical begomoviral symptoms, and both of the viral DNA components accumulated in the upper uninoculated leaves, suggesting systemic infection of TYLCKaV. Two agroinoculation methods were conducted on tomatoes. First, excised sections of tomato shoots were agroinoculated with a soaking procedure. Although two Agrobacterium tumefaciens strains were tested, approximately 40% of inoculated plants only showed viral symptoms for EHA105. Second, agrobacterium from a cultured petri dish was directly inoculated with a colony inoculation procedure. When EHA105 was used, approximately 92% of inoculated plants showed viral symptoms. Sequencing the recovered viral DNA from the upper uninoculated leaf clarified that TYLCKaV had successfully infected the tomato plants. The colony inoculation procedure is labor-saving, and viral symptoms develop in susceptible tomato plants within approximately a month from sowing the seeds. This method could contribute to simple and speedy evaluation of TYLCKaV resistance of tomato plants.

First report of Tomato yellow leaf curl Kanchanaburi virus in eggplant and tomato in Cambodia

First report of <i>Tomato yellow leaf curl Kanchanaburi virus</i> in eggplant and tomato in Cambodia

First Report of Tomato yellow leaf curl Kanchanaburi virus Infecting Eggplant in Laos.

Tomato yellow leaf curl Kanchanaburi virus (TYLCKaV) is a bipartite begomovirus (genus Begomovirus, family Geminiviridae) reported to infect tomato and eggplant in Thailand and Vietnam (1,2). In April 2013, eggplant (Solanum melongena L.) plants exhibiting yellow mosaic symptoms were found in a suburb of Vientiane, Laos. Three symptomatic samples were collected. Total DNA was extracted from leaves by the CTAB method, and used as template for PCR using the degenerate primer pair AV494/CoPR (3). The PCR results suggested that the plants were infected by a begomovirus. The begomoviral genome was amplified by rolling circle amplification (RCA) with TempliPhi kit (GE Healthcare) following the manufacturer's protocol. RCA product was digested with the endonucleases BamH I, EcoR I, Hind III, Kpn I, Pst I, and Xba I, respectively. The fragments about 2.1 kbp (with Pst I digestion) and 1.5 kbp (with Xba I digestion) in size were cloned and sequenced. The sequence of the 2.1-kbp fragment showed similarity with begomovirus DNA-A component. A pair of primers for amplification of the full-length DNA-A, AF (5'-CTTCATCGTTTCTCAGCATCAT-3') and AR (5'-CACTTGCACACGATCTCTAAGA-3') were designed from the 2.1-kbp sequence. The full-length DNA-A was 2,752 nucleotides and encoded six putative ORFs (GenBank Accession No. KF218820). The sequence of the 1.5-kbp fragment shared similarity with begomoviruses DNA-B. The begomoviral circular DNA-B was amplified using the pair of primers BF (5'-GTAACAGCCGAAGTGCACG-3') and BR (5'-AATGGAGAGACACCAGTCTGCC-3') designed from the 1.5-kbp sequence. PCR yielded a product of expected size (~1.4 kbp). The full-length DNA-B sequence was obtained by assembling the two sequences. The DNA-B was 2,734 nucleotides and encoded two putative ORFs (GenBank Accession No. KF218821). The sequences of DNA-A and DNA-B of isolate Laos shared the highest nucleotide sequences identities at 99.0% and 98.0% with those of TYLCKaV-[TH:Kan 1:01] (AF511529), and [TH:Kan 2:Egg:01] (AF511527), respectively. The results indicated that the virus associated with eggplant yellow mosaic disease was an isolate of TYLCKaV. To our knowledge, this is the first report of this begomovirus in Laos. Our results indicate that this virus may be spreading in Southeast Asia and scientists there should be aware of this virus when developing begomovirus-resistant varieties of tomato or eggplant.

Emergence and diversity of begomoviruses infecting solanaceous crops in East and Southeast Asia

Over the past three decades diseases caused by whitefly-transmitted geminiviruses (begomoviruses) have emerged to be important constraints to the production of solanaceous crops, particularly tomato (Solanum lycopersicum) and peppers (Capsicum spp.), in many tropical and subtropical regions of the world. The most studied of these is Tomato yellow leaf curl virus (TYLCV), which has spread to many other areas from its likely origin in the Mediterranean basin region. The virus is usually associated with the polyphagous and virus-vectoring-efficient B-biotype of its vector whitefly (Bemisia tabaci). However, in Southeast and East Asia, a wide variety of distinct local begomovirus species have been identified from tomato and pepper crops over this period, and TYLCV was detected in Japan only in about 1996, China in 2006 and Korea in 2008, despite B-biotype whiteflies being present in several of the countries of the region since at least the early 1990s. Continental Southeast Asia appears to be a major center of diversity for begomoviruses and some species may have spread across the region; Tomato yellow leaf curl Thailand virus (TYLCTHV) appears to have spread from the Thailand–Myanmar region into southern China and is now displacing the local tomato-infecting species in Taiwan, and Tomato yellow leaf curl Kanchanaburi virus (TYLCKaV) appears to have spread from the Thailand–Vietnam region to Java, Indonesia. Since many of the native tomato- or pepper-infecting begomoviruses and associated satellite DNAs have also been detected in local weed species, it seems likely that their ancestors originated in these weed hosts, but with the expansion and intensification of tomato and pepper production in the region, there was selection for recombinant or mutant forms with greater virulence on tomato and/or pepper. Expansion and intensification of these crops may also have resulted in increased populations of local, and if present, B- or Q-biotype whiteflies, aiding the increase and spread of local begomovirus species.

Bahasa Indonesia

Yellow and yellow mosaic symptoms was observed on eggplant ( Solanum melongena ) around Bogor, Bandung, Pati, Rembang, and Bantul during a survey in early 2013. Polymerase chain reaction based detection was carried out using total DNA isolated from symptomatic leaf samples and a pair of Begomovirus -universal primers, SPG1/SPG2. The expected size (~900 bp) amplicon was detected from all five symptomatic samples, indicating the presence of Begomovirus infection. Each amplicon was sequenced followed by basic local alignment search tool analysis. It was revealed that the sequence had the highest homology (98.8%) with Tomato yellow leaf curl Kanchanaburi virus (TYLCKaV) sequence. This is the first report on TYLCKaV infection on eggplant in Indonesia.

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.