A validated SYBR-based RT-qPCR assay reveals seasonal dynamics and within-stool heterogeneity of sugarcane yellow leaf virus in sugarcane

Authors: MILLER, ANNABEL - Louisiana State University; MORALES, JOAO - Louisiana State University; ROLLINS, MARY-BETH - Louisiana State University; ESCOBAR, WILLIAM - Louisiana State University; CHOI, JIYEONG - Cornell University; SINGH, RAGHUWINDER - Louisiana State University; NAVIA URRUTIA, MONICA - University Of Florida; Abrahamian, Peter; GAMA, ANDRE - Louisiana State University; FLASCO, MADISON - Louisiana State University

Submitted to: Tropical Plant Pathology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/5/2026
Publication Date: 6/8/2026
Citation: Miller, A., Morales, J., Rollins, M., Escobar, W., Choi, J., Singh, R., Navia Urrutia, M., Abrahamian, P., Gama, A., Flasco, M. 2026. A validated SYBR-based RT-qPCR assay reveals seasonal dynamics and within-stool heterogeneity of sugarcane yellow leaf virus in sugarcane. Tropical Plant Pathology. https://doi.org/10.1007/s40858-026-00814-8.
DOI: https://doi.org/10.1007/s40858-026-00814-8

Interpretive Summary: Sugarcane yellow leaf virus (SCYLV) is ubiquitous in sugarcane and can reduce yields by as much as half. In Louisiana, the virus does not show obvious symptoms, which makes it easy to spread undetected. Therefore, a reliable method to detect SCYLV in breeding, certification, and production programs is essential. RT-PCR assays are available but often may not be sensitive enough to avoid false negative results. In this study, a more accurate and sensitive one-step test RT-qPCR (quantitative) test was developed. Leaves from two infected sugarcane varieties were tested for SCYLV monthly during both the 2024 and 2025 growing seasons. The virus was consistently found in the most emerging leaf, called the top visible dewlap leaf, indicating this is reliable tissue to test. The quantity of virus in infected plants varied by month and variety, indicating this is a factor in developing optimum virus detection protocols. Furthermore, plants that became infected after planting had lower virus levels compared to those grown from already infected material. This research provides a highly sensitive test for SCYLV and reports the best tissue to collect and optimal timing of testing. This is critical for protecting sugarcane yields and the industry’s sustainability.

Technical Abstract: Sugarcane yellow leaf virus (SCYLV) occurs in all sugarcane producing regions and can reduce yield by up to 50%. In Louisiana infections are asymptomatic and scalable, reliable detection is paramount to avoid unintentional disease spread. While reverse-transcription polymerase chain reaction (RT-PCR) assays are available, they may lack sensitivity. This study aimed to develop a reliable and sensitive assay appropriate for state certification and routine virus detection. Using an RNA transcript encoding the coat protein of SCYLV, a one-step, SYBR green-based quantitative RT-PCR (RT-qPCR) assay was developed that can detect as few as seven viral copies from three genotypes. Monthly sampling conducted from April to August in 2024 (cultivar LCP 85-384) and April to September in 2025 (cultivar L 01-299) showed that SCYLV is consistently detectable in the top visible dewlap leaf (TVD) in two cultivars. In LCP 85-384, virus titer varied in April (p = 0.002), but in L 01-299, variation occurred in June (p < 0.001), contributing to significant differences in titer across months. Current Louisiana certification protocols call for 50 leaves collected per 5-10 acres, likely underestimating incidence as our research revealed that SCYLV was not evenly distributed in across canes in newly infected stools. Those stools exhibited non-ubiquitous distribution (56.1 ± 7.0%; F1,30 = 20.2, p < 0.001) between canes compared to plants resulting from already infected planting material (least squares mean = 95.4 ± 5.3%). Stools that were originally SCYLV-negative and infected post planting displayed a significantly lower viral load than stools planted with SCYLV-positive material (p = 0.002). This study provides a sensitive RT-qPCR assay for SCYLV detection further validated using field collected samples, confirms the TVD is adequate tissue for detection throughout the growing season, and emphasizes the criticality of thorough collection methods for reliable detection of this economically important virus.