# Screening Various Bacterial-Produced Double-Stranded RNAs for Managing Asian Soybean Rust Disease Caused by Phakopsora pachyrhizi

## TL;DR

This study explores using bacterial-produced double-stranded RNA to manage soybean rust disease by targeting fungal genes, offering a sustainable alternative to fungicides.

## Contribution

The novel contribution is the large-scale production and screening of twelve dsRNAs targeting P. pachyrhizi genes, demonstrating significant disease suppression in controlled and greenhouse settings.

## Key findings

- Nine dsRNAs reduced ASR severity by 24.0% to 81.1% and fungal biomass by 50.5% to 83.1% in growth chamber studies.
- Three dsRNAs targeting ACE, CYTB1, and S12 reduced disease severity by 78.2–82.3% and fungal growth by 79.8–85.4% in greenhouse trials.
- dsRNA targeting S12 reduced spore germination to 0.0–26.6% and hyphal growth to 2.7–40.5 µm in vitro.

## Abstract

Asian soybean rust (ASR), caused by Phakopsora pachyrhizi (Syd.), poses a serious threat to global soybean production. The main approach to managing this disease has been through repeated fungicide applications which have reduced efficacy due to fungicide resistance. Recently, spray-induced gene silencing (SIGS) through exogenous application of double-stranded RNA (dsRNA) has emerged as a promising approach for plant disease management. In the present study, twelve different dsRNAs targeting genes important for P. pachyrhizi urediniospore germination, infection of the host plant or resistant to commonly used fungicides were produced in Escherichia coli on a large scale. Nine of these dsRNAs significantly reduced ASR severity (by 24.0% to 81.1%) and fungal biomass (50.5% to 83.1%) compared to the control when applied as a foliar spray in our growth chamber studies. Three of the most effective dsRNAs targeting an acyltransferase (ACE), cytochrome B (CYTB1) and a reductase (S12) also significantly reduced disease severity (78.2 to 82.3%) and fungal growth (79.8 to 85.4%) compared to the control in the greenhouse studies. Further investigation of the P. pachryrhizi urediniospore germination and hyphal growth in the presence of these dsRNAs in vitro revealed these dsRNAs reduced the spore germination rate from 72.1% to 0.0–26.6% at 4.5 h and hyphal growth from 254.0 µm to 2.7–40.5 µm at 9 h, with dsRNA targeting the S12 gene being the most effective. These results highlight the potential of SIGS using selected dsRNAs as a sustainable strategy for managing ASR through suppressing urediniospore germination and hyphal growth.