Reduction of aflatoxin contamination in maize kernels mediated by host-induced gene silencing

Aflatoxins produced by some species of Aspergillus pose a significant threat to global food safety, because they contaminate a variety of crops including maize. In this study, we used host-induced gene silencing (HIGS) to reduce aflatoxin accumulation in maize upon infection with Aspergillus flavus and Aspergillus parasiticus. Two RNAi constructs were designed and introduced into maize, one targeting two essential fungal genes involved in ergosterol biosynthesis (CYP51A and CYP51B) and the other targeting three aflatoxin biosynthesis genes (AflC, AflD, and AflR). The presence of RNAi transgenes was confirmed in 17 and 14 independent lines possessing the CYP51A/CYP51B RNAi construct and the AflC/AflD/AflR RNAi construct, respectively. Most transgenic lines had single-locus integrations and one or two copies of the transgene. Transgene expression varied considerably among the lines. Small RNA sequencing confirmed the production of siRNAs from the HIGS trigger sequences. Levels of CYP51A, AflC, AflD, and AflR transcripts in Aspergillus spp. were successfully decreased upon infection of transgenic maize. However, significant reduction in the level of CYP51B transcripts was not achieved and the CYP51A/CYP51B RNAi transgene had no discernible effect on fungal growth in infected maize. In contrast, we observed significant reductions in aflatoxin accumulation in four AflC/AflD/AflR RNAi transgenic lines infected with A. flavus and in two transgenic lines infected with A. parasiticus. Notably, one AflC/AflD/AflR transgenic line exhibited minimal aflatoxin contamination upon kernel infection with A. parasiticus. These results demonstrate the potential of HIGS to reduce aflatoxin production in mature corn kernels.