Location: National Germplasm Resources LaboratoryTitle: Detection by next generation sequencing of a multi-segmented viral genome from sugarcane associated with Ramu stunt disease Author
|Marron-largo, Clarissa - Animal And Plant Health Inspection Service (APHIS)|
|Kuniata, Lastus - New Britain Palm Oil|
Submitted to: Virus Genes
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/18/2015
Publication Date: 1/4/2016
Citation: Mollov, D.S., Marron-Largo, C., Kuniata, L. 2016. Detection by next generation sequencing of a multi-segmented viral genome from sugarcane associated with Ramu stunt disease. Virus Genes. 52:152-155.
Interpretive Summary: Sugarcane is an important agronomic crop for the U.S. agriculture. Several virus diseases affect sugarcane and reduce yield and cane quality. Additionally quarantine diseases impact the economic trade of germplasm and hinder cultivar improvement. Ramu stunt disease has a devastating impact on sugarcane yield and has only been reported from Papua New Guinea. It is a major quarantine concern to the rest of the world were sugarcane is grown. In this report Ramu stunt virus (RmSV), a virus associated with the Ramu stunt disease, was characterized and its genomic structure was described. The completion of the RmSV genome sequence facilitates reliable detection methods that can be utilized to screen sugarcane genetic material, develop better control measures in the trade and production industries, and implement reliable quarantine measures. This will ultimately aid in disease management efforts of Ramu stunt disease.
Technical Abstract: Ramu stunt disease of sugarcane was first reported in Papua New Guinea in the mid 1980's. The disease can reduce sugarcane yields significantly and causes severe stunting and mortality in highly susceptible cultivars. The causal agent of Ramu stunt has been investigated but its characterization has not been completed. Sugarcane cv. Ragnar from Papua New Guinea with symptoms of Ramu stunt was analyzed by next generation sequencing. Total RNA was extracted and whole transcriptome shotgun sequencing was performed using an Illumina platform. Over thirty-seven million reads with an average length of 100 nucleotides were obtained. More than eighteen thousand contigs were assembled and subjected to BLASTX analysis. Twenty-one contigs were virus related and six were associated with plant viruses. The BLAST algorithms revealed sequence similarity to Tenuivirus and Phlebovirus, genera of viruses whose members contain genomes consisting of multiple RNA segments. The six contigs derived from the RNA sequencing data correspond to six RNAs that compose the Ramu stunt virus genome. Primers were designed for each of the six RNAs and RT-PCR amplicons were obtained only from the symptomatic sugarcane. There was concordance between the sequence data of the contigs obtained from the NGS and that of the amplicons obtained by RT-PCR. The NGS approach allowed us to determine the complete genomic sequence of Ramu stunt virus. It is likely that this virus is the causal agent of Ramu stunt disease.