Identification of miRNAs associated with Aspergillus flavus infection and their targets in groundnut (Arachis hypogaea L.)

Abstract Background: Groundnut is the major oilseed crop that suffers substantial post-harvest losses due to aflatoxin contamination by the fungus Aspergillus flavus. The interaction between A. flavus and groundnut microRNAs in combating aflatoxin contamination remains unclear. This study was carried out to identify microRNAs (miRNAs) to enhance the understanding of in vitro Seed Colonisation (IVSC) resistance mechanism in groundnut. Result: In this study, resistant (J 11) and susceptible (JL 24) genotypes of groundnut were treated with toxigenic A. flavus (strain AF-11-4), and total RNA was extracted at 1 day after inoculation (1 DAI), 2 DAI, 3 DAI and 7 DAI. Seeds of JL 24 showed higher mycelial growth than J 11 at successive days after inoculation. A total of 208 known miRNAs belonging to 36 miRNA families, with length varying from 20-24 nucleotides, were identified, along with 27 novel miRNAs, with length varying from 20-22 nucleotides. Using psRNATarget server, 952 targets were identified for all the miRNAs. The targeted genes function as disease resistant proteins encoding, auxin responsive proteins, squamosa promoter binding like proteins, transcription factors, pentatricopeptide repeat-containing proteins and growth regulating factors, etc. Through differential expression analysis, seven miRNAs (aly-miR156d-3p, csi-miR1515a, gma-miR396e, mtr-miR2118, novo-miR-n27, ptc-miR482d-3p and ppe-miR396a) were found common among 1 DAI, 2 DAI, 3 DAI and 7 DAI in J 11, whereas ten miRNAs (csi-miR159a-5p, csi-miR164a-3p, novo-miR-n17, novo-miR-n2, osa-miR162b, mtr-miR2118, ptc-miR482d-3p, ptc-miR167f-3p, stu-miR319-3p and zma-miR396b-3p) were found common among 1 DAI, 2 DAI, 3 DAI and 7 DAI in JL 24. Two miRNAs, ptc-miR482d-3p and mtr-miR2118, showed contrasting expression at different time intervals between J 11 and JL 24. These two miRNAs were found to target those genes with NBS-LRR function, making them potential candidates for marker development in groundnut breeding programs aimed at enhancing resistance against A. flavusinfection. Conclusion: This study enhances our understanding of the involvement of two miRNAs namely, ptc-miR482d-3p and mtr-miR2118, along with their NBS-LRR targets, in conferring resistance against A. flavus-induced aflatoxin contamination in groundnut under in vitro conditions.