Environmental Fate of Double-Stranded RNA in Agricultural Soils

Abstract

A laboratory soil degradation study was conducted to determine the biodegradation potential of a DvSnf7 dsRNA transcript derived from a Monsanto genetically modified (GM) maize product that confers resistance to corn rootworm (CRW; Diabrotica spp.). This study provides new information to improve the environmental assessment of dsRNAs that become pesticidal through an RNAi process. Three agricultural soils differing in their physicochemical characteristics were obtained from the U.S., Illinois (IL; silt loam), Missouri (MO; loamy sand) and North Dakota (ND; clay loam), and exposed to the target dsRNA by incorporating insect-protected maize biomass and purified (in vitro-transcribed) DvSnf7 RNA into soil. The GM and control (non-GM maize) materials were added to each soil and incubated at ca. 22°C for 48 hours (h). Samples were collected at 12 time intervals during the incubation period, extracted, and analyzed using QuantiGene molecular analysis and insect bioassay methods. The DT50 (half-life) values for DvSnf7 RNA in IL, MO, and ND soils were 19, 28, and 15 h based on QuantiGene, and 18, 29, and 14 h based on insect bioassay, respectively. Furthermore, the DT90 (time to 90% degradation) values for DvSnf7 RNA in all three soils were <35 h. These results indicate that DvSnf7 RNA was degraded and biological activity was undetectable within approximately 2 days after application to soil, regardless of texture, pH, clay content and other soil differences. Furthermore, soil-incorporated DvSnf7 RNA was non-detectable in soil after 48 h, as measured by QuantiGene, at levels ranging more than two orders of magnitude (0.3, 1.5, 7.5 and 37.5 µg RNA/g soil). Results from this study indicate that the DvSnf7 dsRNA is unlikely to persist or accumulate in the environment. Furthermore, the rapid degradation of DvSnf7 dsRNA provides a basis to define relevant exposure scenarios for future RNA-based agricultural products.

RNAi Efficiency, Systemic Properties, and Novel Delivery Methods for Pest Insect Control: What We Know So Far

“…Likewise, when maize seedlings were irrigated with dsRNA of Kunitz-type trypsin inhibitors (dsKTI), this resulted in a high mortality rate with the Asian corn borer, Ostrinia furnacalis (Li et al, 2015a). Although interesting, Dubelman et al (2014) reported that the soil persistency of dsRNA can be short, with a rapid breakdown within 2–3 days. Therefore, the dsRNA stability in the soil is still a matter of question.…”

“…Supply of dsRNA through irrigation water, root drench, or trunk injection would be a great strategy for pest insects, such as root feeders, for which no efficient control method is available at this moment. Moreover, the delivery of dsRNA through irrigation or trunk injections holds low environmental risks due to the rapid breakdown of dsRNAs within 2–3 days in the soil and plant debris (Petrick et al, 2013; Dubelman et al, 2014), as well as due to the localized application fashion.…”

RNA Interference for Mosquito and Mosquito-Borne Disease Control

“…Many different delivery approaches have been used for per os RNAi trigger delivery in larvae, including naked dsRNA in buffer or water [126,127]. Although naked dsRNA can be internalized at high concentration in sterile conditions in the lab, dsRNA in field settings could be subject to rapid environmental degradation [21,22]. To prevent RNAi trigger degradation, abiotic and biotic delivery systems including the use of Effectene ® (Qiagen) liposomes, chitosan nanoparticles, E. coli expression systems, and Pichia pastoris expression systems have been explored [27,47,49,75,122,123,128,129].…”

“…In order to achieve these same detrimental phenotypes in wild populations, RNAi triggers must be delivered to the target species and life stage with consideration for environmental and abiotic factors including: UV, ribonucleases, microbes, dissipation and dilution in aqueous environs and on solid substrates [20,21,22,23]. RNAi triggers must also be delivered to target mosquito species using field feasible applications.…”

RNA ‘Information Warfare’ in Pathogenic and Mutualistic Interactions

“…Naturally, the transgenic nature of HIGS and its mode of action have been under considerable scrutiny [98][99][100]. The environmental persistence of dsRNAs in agricultural soils has been explored and found to be low, reducing the chance of off-target effects [101]. Already, a few field crop varieties with effective antiviral HIGS have been approved and widely grown [102].…”