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Research Project: CHARACTERIZING, DETECTING, AND ELIMINATING PATHOGENS TO ENABLE THE SAFE INTRODUCTION OF PLANT GENETIC RESOURCES

Location: National Germplasm Resources Laboratory

Title: Next Generation Sequencing: A useful tool for detection of sugarcane viruses in quarantine programs

Author
item Mollov, Dimitre
item Malapi-wight, Martha - Animal And Plant Health Inspection Service (APHIS)

Submitted to: International Society of Sugar Cane Technologists Proceedings
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/13/2016
Publication Date: 12/5/2016
Citation: Mollov, D.S., Malapi-Wight, M. 2016. Next Generation Sequencing: A useful tool for detection of sugarcane viruses in quarantine programs. International Society of Sugar Cane Technologists Proceedings. 29:135.

Interpretive Summary: Sugarcane is an important agronomic crop in the U.S. Several virus diseases can infect sugarcane and reduce yield and cane quality. Quarantine pathogens also impact the international movement of germplasm and hinder crop improvement efforts. To prevent the introduction of sugarcane viruses in imported germplasm, rapid and reliable diagnostic methods are needed. The concern exists that newly emerging or poorly characterized viruses may not be detected by currently used methods. Recent advances have developed a technique called next generation sequencing (NGS), technology that does not require prior knowledge of the pathogen for its detection. In this report we describe the use of NGS for sugarcane virus diagnostics and its comparison to the currently used lab-based assays. The results suggest NGS may offer advantages of speed and sensitivity, as well as the ability to detect new viruses. This research will be useful to sugarcane quarantine, certification, and breeding programs.

Technical Abstract: The international exchange of sugarcane germplasm includes the risk of introducing potentially devastating pathogens that may threaten production. The USDA-APHIS Plant Germplasm Quarantine Program (PGQP) imports and tests sugarcane accessions that are used in research, variety development, and commercial production across the United States. In this study, we have compared established serological and nucleic acid-based methods to next generation sequencing (NGS) technologies for the detection of viral pathogens. Twelve samples of sugarcane (Saccharum spp.), one sample of St. Augustine grass (Stenotaphrum secundatum), and one sample of Columbus grass (Sorghum almum) of foreign and domestic origin maintained at the PGQP were sequenced using Illumina technologies. BLASTn searches against a custom sugarcane virus database identified 11 of 14 samples as positive for one or more viruses. NGS analysis detected all viruses that were identified by laboratory diagnostic tests used routinely by PGQP. Additionally, in two samples, analysis of NGS data identified Sugarcane bacilliform virus, a pathogen that was not targeted by the lab-based tests. This study shows the power of NGS as a diagnostic tool to identify rapidly multiple viruses in sugarcane samples and the possibility to apply the technology for virus identification in sugarcane quarantine, certification, and breeding programs.