First waikavirus infectious clones and vascular expression of green fluorescent protein from maize chlorotic dwarf virus (MCDV)

Authors: Stewart, Lucy; Willie, Kristen; XIE, WENSHUANG - The Ohio State University; Todd, Jane; TRAN, HONG HANH - The Ohio State University

Submitted to: Frontiers in Virology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/8/2023
Publication Date: 6/29/2023
Citation: Stewart, L.R., Willie, K.J., Xie, W., Todd, J.C., Tran, H. 2023. First waikavirus infectious clones and vascular expression of green fluorescent protein from maize chlorotic dwarf virus (MCDV). Frontiers in Virology. https://doi.org/10.3389/fviro.2023.1216285.
DOI: https://doi.org/10.3389/fviro.2023.1216285

Interpretive Summary: Maize chlorotic dwarf virus (MCDV) is a historically important disease-causing virus in US corn belonging to a group of viruses called waikaviruses. Other waikaviruses also cause important disease in rice (rice tungro spherical virus) or have been found in other plants, but have unknown or only partially known disease impact on host plants. A major bottleneck in understanding the basic biology and pathogenicity determinants of waikaviruses is the lack of a crucial virology research tool to dissect gene and sequence function of viruses, cDNA plasmid constructs, or "infectious clones". We report the first development of infectious clones for two strains of MCDV, the severe and mild (MCDV-S and MCDV-M1), and a green fluorescent protein (GFP)-tagged MCDV-S construct. These constructs enable virus labeling and examination of localization within plants and also allowed us to examine the role of a virus protein of unknown function, p27. By deletion of the p27-encoding gene and replacement with GFP, we discovered that p27 was dispensable for plant infection and disease symptom development, but is necessary for leafhopper transmission of the virus. The development of these tools is a milestone in waikavirus research, enabling examination of the molecular determinants of plant-virus-vector interactions and virus pathology.

Technical Abstract: Plant viruses classified in the genus Waikavirus, family Secoviridae, are positive sense single-stranded RNA viruses and include important pathogens of maize (maize chlorotic dwarf virus; MCDV) and rice (rice tungro spherical virus; RTSV). Many aspects of the molecular biology of waikaviruses remain unexplored because of experimental challenges and lack of infectious clones for low titer, phloem-limited, and obligately vector-transmitted waikaviruses. Here, we report the first development of waikavirus infectious clones for two MCDV strains, MCDV-S and MCDV-M1, and insect-free launching of infections from these clones in maize by vascular puncture inoculation. We further developed a green fluorescent protein (GFP)-tagged MCDV clone by replacing the viral p27-encoding sequence with GFP-encoding sequence. GFP-tagged virus moved systemically in plants and caused symptomatic infection, similar to wild type virus, with vascular expression of GFP. Development of waikavirus infectious clones is a major advance for this group of agriculturally significant viruses.