Biology subjects

Okechukwu, E. C.

Publications and source records attributed to Okechukwu, E. C..

2 recordsLinked to original sources

bioRxiv2025

Advancing RNAi-Based Strategies Against Downy Mildews: Insights Into dsRNA Uptake and Gene Silencing

Downy mildew (DMs) dieases are caused by destructive obligate pathogens with limited control options, posing a significant threat to global agriculture. RNA interference (RNAi) has emerged as a promising, environmentally sustainable strategy for disease management. In this study, we evaluated the efficacy of dsRNA-mediated RNAi in suppressing key biological functions in DM pathogens of Arabidopsis thaliana, pea and lettuce DM pathogens, Hyaloperonospora arabidopsidis (Hpa), Peronospora viciae f. sp. pisi (Pvp) and Bremia lactucae (Bl), respectively. We specifically targeted the cellulose synthase 3 (CesA3) and the beta tubulin (BTUB) genes. Silencing CesA3 impaired spore germination and infection across multiple species, while BTUB silencing reinforced the potential of dsRNA-mediated inhibition. Reduction in gene expression levels correlated well with the sporulation assays confirming the effectiveness of dsRNA-mediated gene silencing. We used dsRNAs that were chemically synthesized, in vitro transcribed (IVT) or produced in E. coli. We found that the length and concentration of these dsRNAs significantly affected uptake efficiency, spore germination, and sporulation, with higher concentrations enhancing inhibitory effects. Confocal microscopy using Cy-5-labelled short-synthesized dsRNA (SS-dsRNA) provided direct evidence of spore uptake, confirming the potential of SS-dsRNA for pathogen control. However, species-specific sequence variations influenced dsRNA efficacy, underscoring the importance of target sequence design. Multiplexed RNAi impacted silencing synergisticly, further reducing germination and sporulation in Hpa. Additionally, we demonstrated that SS-dsRNA-mediated gene silencing is sustained over time, with a significant reduction in gene expression level at 4, 7, 10 and 11dpi. This indicates the durability and efficacy of this approach. Taken together, these findings demonstrate the potential of dsRNA-mediated gene silencing as a precision tool for managing DM pathogens.

plant biology↗

bioRxiv2025

Pea-Saving Partners: Bacillus and Pseudomonas combat downy mildew in pea crops

Downy mildew (DM) is a destructive disease that significantly reduces the yield and quality of important pulses (legumes) and horticultural crops, particularly during humid and cool seasons. This disease is caused by obligate and host-specific oomycete pathogens. Controlling the pathogen is challenging due to its long-term survival as spores and its rapid mutation. Use of chemical pesticides has been the most effective method to control DM pathogens, but their environmental hazards are a global concern. Current research is focused on exploring the potential of microbial biological control agents (MBCA), particularly rhizobacteria strains of the genera Bacillus and Pseudomonas, which have shown suppression of plant pathogens. However, to date, no MBCA has been reported to be effective against DM pathogens in pulses. We investigated the effectiveness of Bacillus and Pseudomonas strains as potential biopesticides against the pea downy mildew pathogen Peronospora viciae f. sp. pisi (Pvp). In our study, in vitro bioassays showed 100% inhibition of Pvp spore germination compared to the control. In planta antagonism assays further demonstrated significant suppression (>80%) of Pvp sporulation in pea plants sprayed with strains of Bacillus velezensis or P. fluorescens or their filtrates. The drench application also showed significant effects where either a Pseudomonas or cold-adapted Bacillus strain was used. We observed a synergistic effect for the dual foliar application of the microbes compared to individual application (27.6 to 46.7% suppression). Furthermore, the results from the molecular biomass analysis were consistent with the results of the sporulation assays. This demonstrates the strong interactive and promotive benefits of using Bacillus and Pseudomonas as biocontrol agents Based on these results, we conclude that these MBCAs could be effective in combatting Pvp infections in the field.

plant biology↗