Identification and Characterization of Selected Taro Viruses in Kenya Using Next-Generation Sequencing

Abstract

Plant viruses are one among the biggest limiting factors of crop production globally with a potential of causing up to 100% yield loss. Identification and characterization of viruses is a critical step in formulating effective disease management strategies and programs. Several diagnostic tools, including physical, biological, serological, and molecular approaches are available for detection and analysis of viral pathogens. These methods require sequence information for the target viruses and thus making it difficult to target and study emerging viruses or even identify rare pathogens. Taro is an important food security crop affected by several viral pathogens which have not been characterized in detail because of lack of adequate sequence information of these pathogens. Insufficient sequence information on a pathogen impedes the ability to understand and develop a control strategy for such pathogens. Next generation sequencing tools, especially small RNA sequencing, is a powerful technology for robust detection and identification of both known and unknown viruses due to its ability to multiplex detections with no prior knowledge of the virus sequence. NGS offers advantages over all existing methods of identifying a viral pathogen, including removing the need for targeting a specific pathogen or requiring sequence information for that pathogen, identifying multiple pathogens in a single sample, and eliminating the need for costly and often ineffective culturing or antibody laboratory tests. Herein, PCR and RT-PCR assays using degenerate primers were employed to screen for previously known viral pathogens infecting taro germplasm in Kenya. In-depth sequencing of small RNAs isolated from both virus symptomatic and asymptomatic Taro germplasm using next-generation sequencing technology was subsequently used to detect the unknown viral pathogens. Subsequent bioinformatics analyses revealed the presence of both DNA and RNA viruses. Detected DNA viruses included the Taro Bacilliform Virus (TaBV) and Taro Bacilliform CH Virus (TaBCHV), which are badnaviruses specific to Taro, the sweet potato Badnavirus B, sugarcane bacilliform virus, and sweet potato leaf curl virus. The RNA viruses included the Colocasia Bobone Disease Virus, a rhabdovirus specific to Taro, and the East African Cassava mosaic virus, sweet potato feathery mottle, and Phaseolus vulgaris alphaendornavirus. A Citrus exocortis viroid was also detected. Interestingly, the wild Taro relatives, including tannia, had no viral sequence hits affirming that wild species possess some level of tolerance to viral infections, possibly because of having a rich reservoir of resistance genes useful in breeding cultivars with a genetically controlled resistance against numerous diseases. Moreover, the viral severity in the fields was also positively correlated with the number of viruses identified, as evidenced by the positive correlation between the viral prevalence based on the visual identification of symptoms and the TaBV PCR results. These results collectively demonstrate the reliability of the sRNA deep sequencing data in determining virus and viroid diversity. This study reports the Taro viruses and viroids circulating in Kenya and comprehensively describes the prevalence, distribution, and sequence variability of TaBV in Kenya. The study forms a basis for developing effective management strategies to support the prevention and control of Taro viruses.