Chili pepper (Capsicum annuum L.) is an economically important vegetable crop in Indonesia (Sativa et al. 2017). During a field survey conducted in August 2024 in Cianjur, Indonesia, virus-like symptoms, including leaf yellowing, vein clearing, and mosaic, were observed in commercial chili pepper fields, with disease incidence estimated at 50-70%. Because mixed infections are common in chili pepper (Kim et al. 2014), these symptoms might not be attributed to a single viral agent. Thus, to analyze the virome of these fields, 20 symptomatic leaf samples (10 from each of two fields) were randomly collected from the fields and pooled for RNA-seq. Total RNA was extracted using the Hybrid-RTM RNA extraction kit (GeneAll, Korea). A cDNA library was prepared using the TruSeq Stranded Total RNA with Ribo-Zero Plant Kit (Illumina, USA) and analyzed on an Illumina NovaSeq 6000 platform (Macrogen, Korea). De novo assembly of 78,486,422 quality-filtered reads using the Trinity pipeline generated 84,362 contigs. BLASTn/x analyses identified contigs corresponding to cucumber mosaic virus, chilli veinal mottle virus, pepper yellow leaf curl Indonesia virus, pepper cryptic virus 2, and pepper vein yellows virus 5 (PeVYV-5). In addition, a 2,911-nt contig supported by 491,491 paired-end 150-bp reads showed the highest nucleotide identity (96.5%) to PeVYV-associated RNA (PeVYVaRNA) isolate PRO54353 from South Africa (GenBank accession no. MT321510), with nearly complete sequence coverage. To determine the complete sequence of the Cianjur isolate (PeVYVaRNA-CJ), overlapping RT-PCR fragments covering the assembled contig were amplified and sequenced, and the terminal sequences were determined by 5′ and 3′ rapid amplification of cDNA ends assays. The complete PeVYVaRNA-CJ sequence was 3,001 nt and was deposited in GenBank under accession number PZ459831. The same 20 individual field samples were tested by RT-PCR using PeVYVaRNA-specific primers (5′-TCGCTCGCCACGTTTAGAGTA-3′ and 5′-GAGGTGGGAGTACGAAGATTT-3′). Amplicons of the expected size were obtained from 16 samples, and Sanger sequencing confirmed their identity as PeVYVaRNA-derived sequences. PeVYVaRNA detection was further verified by dot blot hybridization using a specific probe (5'-ACGTTAGGCTGCGCGTTCCTTGGCCTTTATCCGGGGCAA-3'). The same 20 samples were also assayed by RT-PCR using PeVYV-5-specific primers (5′-GTCGAGGTTTCAGGACAACTGGA-3′ and 5′-GTGCGACCACAACGACAGGG-3′). PeVYV-5 was detected only in the 16 PeVYVaRNA-positive samples, indicating consistent co-detection of PeVYV-5 and PeVYVaRNA in the samples tested. PeVYVaRNA was first identified in South Africa in 2020 (Schravesande et al. 2021). Its detection in Indonesia, a geographically distant country, suggests that PeVYVaRNA may be more widely distributed than currently recognized. Because the samples contained mixed viral infections and symptom severity was not quantitatively assessed, no causal relationship between PeVYVaRNA and the observed field symptoms can be inferred from the present study. However, other polerovirus-associated RNAs have been implicated in enhanced symptom development and viral fitness during mixed infections involving helper viruses (Sanger et al. 1994). Therefore, further studies are needed to evaluate the potential epidemiological relevance of PeVYVaRNA in viral disease complexes affecting pepper. To our knowledge, this is the first report of PeVYVaRNA detected in chili pepper in Indonesia.
Andika Septiana Suryaningsih, Hee-Seong Byun, Seok-Yeong Jang et al.· Plant Disease· 0 citations
Melon (Cucumis melo L.) is an economically important cucurbit crop in Indonesia, with a production of 117,793 tons cultivated on 7,039 ha (FAOSTAT, 2023). In 2023, virus-like symptoms including interveinal yellowing and chlorosis were observed in open-field melons in Kediri District, Indonesia. To identify associated viruses, two symptomatic leaf samples (IND-ME10, IND-ME11) were analyzed by high-throughput sequencing (HTS) and reverse transcription (RT)-PCR. Total RNA was extracted using the RNeasy Plant Mini Kit (Qiagen, Germany) and sequenced on an Illumina NovaSeq 6000 platform (Macrogen, Korea). De novo assembly of 79,400,902 clean reads using Trinity (v. r20140717) generated 254,870 contigs. BLAST analyses against the GenBank viral database identified six virus-associated large contigs corresponding to cucurbit yellow stunting disorder virus (CYSDV), Cucumis melo alphaendornavirus (CmEV), and tomato leaf curl New Delhi virus (ToLCNDV). The CYSDV contig (31,881 mapped reads; 281.7× depth) comprised nearly complete RNA1 (8,594 nt) and RNA2 (7,781 nt), sharing 99% and 97% nucleotide identity (100% coverage), respectively, with CYSDV isolate GDBL from China (PV232312, PV232313). Three CmEV contigs (717–2,914 mapped reads; 32.9–114.7× depth) had a nearly complete genome of 15,073 nt, showing 96% nucleotide identity (100% coverage) with the United States CmEV isolate CL-01 (NC_029064) (Sabanadzovic et al. 2016). Two ToLCNDV contigs (DNA-A, DNA-B) shared 96% nucleotide identity with previously reported Indonesian isolates (AB613825, AB613826) (Mizutani et al. 2011). RT-PCR and PCR assays confirmed that sample IND-ME11 was positive for CYSDV and CmEV, whereas IND-ME10 was positive for CmEV and ToLCNDV. Sanger-sequenced amplicons of CYSDV and CmEV from IND-ME11 shared 99% nucleotide identity with corresponding HTS-derived sequences. The obtained sequences were deposited in GenBank under accession numbers PX959674 (9,157 nt, CYSDV_RNA1), PX959675 (7,779 nt, CYSDV_RNA2), PX959676 (15,073 nt, CmEV), and PX959677 (2,739 nt, ToLCNDV_DNA-A from IND-ME10). Because multiple viruses were detected, we compared the observed symptoms with those previously reported for the detected viruses. Sample IND-ME11, co-infected with CYSDV and CmEV, showed interveinal yellowing and chlorosis. While CmEV (genus Alphaendornavirus, family Endornaviridae) is generally regarded as asymptomatic in melon (Sabanadzovic et al. 2016; Zeng et al. 2020), CYSDV (genus Crinivirus, family Closteroviridae) characteristically induces severe foliar yellowing and chlorotic symptoms in cucurbits (Wintermantel et al. 2017). Sample IND-ME10 was infected with CmEV and ToLCNDV. Due to the mixed infections and restricted sample size, definitive symptom causality was not assessed. CYSDV isolates divide into two phylogenetic groups with 73–76% nucleotide identities. The Indonesian isolate identified in this study grouped with Asian CYSDV isolates, which have been proposed as cucurbit chlorotic virus (CuCV) (Che et al. 2023). To the best of our knowledge, this is the first report of CYSDV and CmEV in melon in Indonesia. Given the limited sample size, broad conclusions regarding regional prevalence or distribution cannot be made; further large-scale surveys are warranted to determine their economic impact on Indonesian cucurbit production.
Hae-Ryun Kwak, Hee-Seong Byun, Jin-Ju Song et al.· Plant Disease· 0 citations
Gibson Assembly provides an efficient and reliable approach for constructing an infectious clone of ToMMV, which reproduced disease symptoms and pathogenicity, providing a valuable tool for future research on ToMMV biology.
T. N. T. Vo, Marjia Tabassum, Bupi Nattanong et al.· Molecular Biology Reports· 1 citation
We use cookies to run the site and, with your consent, for analytics and to show ads.
See our Cookie Policy.
