Recombinant defense protease inhibitors from Vigna unguiculata show bioactivity against non-host related pests

The long-term use of chemical pesticides has caused resistance, environmental pollution and other problems, in contrast, biological pesticides are safer and more environmentally friendly. Plant defense substances are important sources of biopesticides for pest control. Although pests have developed adaptations to the host plant, the defense substances derived from plants still have activities against non-host related pests. The defense pathways and substances of Vigna unguiculata were identified by transcriptome sequencing under the treatments of mechanical damage and mite feeding. MAPK pathway related genes were identified as important regulators during the activation of plant defense. Among substances with potential direct activity, a total of 15 serine protease inhibitors and 8 cysteine protease inhibitors were screened out. Since the gene expression levels of VuCPI649 and VuSPI680 were significantly up-regulated in mite feeding group, the soluble proteins of VuCPI649 and VuSPI680 were obtained through heterologous expression in Escherichia coli. Biological activity assays showed that recombinant VuCPI649 and VuSPI680 could significantly inhibit the hatching of the eggs of Tetranychus urticae, and recombinant VuCPI649 had insecticidal effect on the larvae of Tuta absoluta through feeding. Subsequently, the protease genes with specific expression patterns in T. urticae and T. absoluta were screened out as potential targets, and the interaction and binding sites between protease inhibitors and proteases were predicted. In conclusion, recombinant VuCPI649 and VuSPI680 are expected to become new types of insecticidal and acaricidal active substances.