First Report of Southern Tomato Virus in Tomato ( <i>Solanum lycopersicum</i> ) in Malaysia

The southern tomato virus (STV), a double-stranded RNA virus with an approximate genome size of 3.5 kb, is a resilient seed-transmitted pathogen classified within the genus Amalgavirus (Sabanadzovic et al. 2009). The virus was originally identified in the United States and Mexico and has since been documented in numerous tomato-producing regions globally, including several countries in Asia, such as China and Korea (Wei et al. 2024; Oh et al. 2018). STV has been detected in both asymptomatic and symptomatic tomato plants, with the latter showing chlorotic spots, interveinal necrosis, mottling, and yellowing in recent studies (Harju et al. 2021; Gaafar et al. 2019). Given the high rate of STV transmission vertically and the variable symptoms observed in specific cultivars, it is therefore imperative to elucidate the symptoms associated with local tomato cultivars. In September 2023, 17 tomato plants (Solanum lycopersicum hybrid Tymoti F1) exhibiting virus-like symptoms such as leaf curling, distortion, mottling and yellowing were collected from a tomato cultivation area in Selangor, Malaysia (GPS coordinate 3.0041467, 101.7006063). Total RNA was extracted from 100 mg of each sample using TRIzol (Thermo Fisher Scientific, USA), and equal molar amounts (250 ng/µl) of RNA from each plant were pooled to prepare a 150 bp paired-end RNA library with the Fast RNA-seq Lib Prep Kit V2 (ABclonal, Inc., USA) that was sequenced on the Illumina NovaSeq platform (NovogeneAIT Genomics Singapore Pte Ltd, Singapore). De novo assembly with SPAdes version 4.0.0 (Bankevich et al., 2012) produced 39,457 scaffolds, of which 3,629 scaffolds between 1–15 kb were selected for BLASTx analysis. Four scaffolds ranging from 280 to 882 bp produced the highest sequence match to the southern tomato virus (STV) isolates from the United States (Accession no. ABC96787) and Slovenia (Accession no. USN17665) with 98 to 100% amino acid identity. Mapping of reads to the STV reference genome (Accession no. NC011591) using Bowtie2 implemented in Geneious Prime® 2025.2.1 yielded 1,177 mapped reads with 50x mean coverage, covering 98.5% of the viral genome. The presence of STV was confirmed in individual tomato plants using RNA extracted with the same method previously used for HTS. Complementary DNA was synthesized using the RevertAid First Strand cDNA Synthesis Kit (Thermo Fisher Scientific, USA), followed by PCR employing Taq DNA polymerase (Promega, USA) with the published STV-specific primers STV-F and STV-R (Sabanadzovic et al., 2009). Four out of the 17 tomato leaf samples yielded the expected PCR product size of 440 bp, indicating approximately 24% STV incidence in the tested samples. These PCR products were subsequently subjected to direct sequencing (Apical Scientific Sdn. Bhd., Malaysia). BLASTn analysis of the 357 bp trimmed sequences showed that three had over 99% nucleotide identity with STV isolates from Korea (Accession no. OR596867), while one shared 99.8% identity with an isolate from China (Accession no. PV582988). All four sequences were deposited into GenBank with accession numbers PV661099 to PV661102. To our knowledge, this is the first report of STV in Malaysia. STV may spread through seed movement across Southeast Asia, highlighting the need to screen seeds from major local tomato cultivars for surveillance. More research is essential to assess the distribution, incidence, and impact of STV in Malaysian and regional tomato cultivation areas.