Metaviromic analysis of cucumber reveals diverse viral infections and development of LAMP assays for their rapid detection

Cucumber production has been significantly threatened by numerous viruses, resulting in a decrease in both the quality and quantity of the produce. However, conventional methods for identifying viruses may not provide a complete picture of co-infections of different viruses. The advent of high-throughput sequencing has made identifying known and novel viruses easy. Hence, we collected 25 cucumber samples exhibiting diverse kinds of virus-disease-like symptoms from the farmer's field, where 25.50–40 % viral disease incidence was also recorded. RNA extracted from the collected samples was used for mRNA and sRNA library preparation using pooled RNA and subjected to Illumina NOVASEQ 6000 platform to obtain the raw sequence data. The obtained raw data was analyzed using bioinformatics, revealing several virus-associated sequences in the assembled reads. Complete or near-complete genomes of five different viruses were identified, which include tomato leaf curl New Delhi virus (ToLCNDV), cucumber mosaic virus (CMV), tobacco streak virus (TSV), melon yellow spot virus (MYSV), and cucurbit aphid-borne yellows virus (CAbYV). A partial genome of cucurbit yellow stunting disorder virus (CYSDV) was also assembled. The estimation of viral abundance using Fragments Per Kilobase of transcript per Million (FPKM) mapped reads revealed that CMV (∼96 % of total FPKM) and MYSV (∼2.85 %) were the most prevalent among the identified viruses. Further phylogenies were constructed using reference sequences from the NCBI database, providing additional evidence of conformity for the identified viruses. The L genomic segment of MYSV exhibited the highest variation, with 345 single-nucleotide polymorphisms (SNPs). Further recombination analysis of the detected viruses revealed both intra and interspecific recombinants in CMV, TSV, and MYSV. The identified viruses were validated through PCR and RT-PCR assays. Additionally, loop-mediated isothermal amplification (LAMP) diagnostic assays were successfully developed for future use in screening viruses in field-collected samples. This study reports the first complete/near-complete genomes of MYSV, TSV, and CAbYV from cucumber, representing significant genomic advances from India. Overall, this study advances the virome profiling of cucumber in India and delivers complete genome data that underpin the development of rapid, field-deployable diagnostics, with potential applications in seed certification, quarantine inspection, and farm-level virus management.