A cytochrome P450 from juvenile mustard leaf beetles hydroxylates geraniol, a key step in iridoid biosynthesis

Juveniles of the leaf beetle Phaedon cochleariae synthesize iridoid via the mevalonate pathway to repel predators. The normal terpenoid biosynthesis is integrated into the dedicated defensive pathway by the ω-hydroxylation of geraniol to 8-hydroxygeraniol. Here we identify and characterize the geraniol 8-hydroxylase as a P450 monooxygenase using integrated transcriptomic and proteomic analyses. In the fat body, 73 individual cytochrome P450s were identified. The double stranded RNA (dsRNA)-mediated knock down of CYP6BH5 led to a significant reduction of 8-hydroxygeraniol-glucoside in the hemolymph and, later, of the chrysomelidial in the defensive secretion. Heterologously expressed CYP6BH5 converted geraniol to 8-hydroxygeraniol. In addition to geraniol, CYP6BH5 also catalyzes other monoterpenols, such as nerol and citronellol, into the corresponding α, ω-dihydroxy compounds.HighlightsThe geraniol 8-hydroxylase in Phaedon cochleariae was identified as a cytochrome P450 CYP6BH5.RNA interference emphasized the importance of CYP6BH5 in iridoid biosynthesis.In vitro enzyme assays showed that recombinant CYP6BH5 is a substrate promiscuous enzyme, converting the ω-hydroxylation of geraniol, nerol, citronellol but not linalool.Homology modeling suggested the -OH group of the substrate plays an important role in coordinating the substrates with the enzyme’s catalytic cavity.

Insecticidal Effects of Different Application Techniques for Silica Dusts in Plant Protection on Phaedon cochleariae Fab. and Pieris brassicae L.

Since the 1900s, diatomaceous earth (DE) has been used as an alternative to chemical insecticides in stored product protection. New silica and DE formulations offer expanded possibilities for use in horticultural crops. However, many crop pests are found on the leaf underside and this is especially challenging when using silica because the substance must have direct contact with the insect to be effective. We tested three application techniques with three formulations of silica to evaluate their efficacy against different developmental stages of mustard leaf beetle (Phaedon cochleariae Fab.) and the cabbage worm of the large white butterfly (Pieris brassicae L.) on the host plant species pak choi (Brassica rapa ssp. chinensis L.). Formulations were applied manually with a powder blower, with an electrostatic spray gun, and in a closed chamber also working with electrostatic forces. The silica formulations used in the biotests were Fossil Shield 90.0s®, AE R974®, and a formulation developed at Humboldt University Berlin called AL-06-109. All formulations contained at least 60% silicon dioxide. Significant differences in efficacy were detected with different application methods and/or silica formulations. AL-06-109 electrostatic cabin-applied was the most effective combination. All formulations, if applied electrostatically, resulted in good coverage and in high plant protection against insect pests. Dusts applied manually were unevenly distributed and easily removed by wind from leaf surfaces. Electrostatic application with a spray gun resulted in even particle distribution on plants, but overspray was high. To accomplish even coverage without wasting so much active material, an enclosed mobile chamber with an electrostatic spray system and an attached exhaust system was developed.