Volatile unsaturated hydrocarbons emitted by seedlings of Brassica species provide host location cues to Bagrada hilaris

Abstract

Bagrada hilaris Burmeister, is a stink bug native to Asia and Africa and invasive in the United States, Mexico, and more recently, South America. This species can cause serious damage to various vegetable crops in the genus Brassica, with seedlings being particularly susceptible to B. hilaris feeding activity. In this study, the role of volatile organic compounds (VOCs) emitted by seedlings of three Brassica species on the host preference of B. hilaris was evaluated. In dual choice arena and olfactometer bioassays, adult painted bugs preferred B. oleracea var. botrytis and B. napus over B. carinata. Volatiles from B. oleracea seedlings were collected and bioassayed with B. hilaris adults and late stage nymphs, using electroantennographic (EAG) and behavioral (olfactometer) techniques. When crude extracts of the VOCs from B. oleracea var. botrytis seedlings and liquid chromatography fractions thereof were bioassayed, B. hilaris adults and nymphs were attracted to the crude extract, and to a non-polar fraction containing hydrocarbons, whereas there were no responses to the more polar fractions. GC-MS analysis indicated that the main constituents of the non-polar fraction was an as yet unidentified diterpene hydrocarbon, with trace amounts of several other diterpene hydrocarbons. The major diterpene occurred in VOCs from both of the preferred host plants B. oleracea and B. napus, but not in VOCs of B. carinata. Our results suggest that this diterpene, alone or in combination with one or more of the minor compounds, is a key mediator in this insect-plant interaction, and could be a good candidate for use in lures for monitoring B. hilaris in the field.

Fig 1. Dual choice arena bioassays.

Host preference responses (% final choice) of B. hilaris adults to seedlings of B. oleracea var. botrytis, B. carinata, and B. napus. N = number of replicates; ** = P < 0.01; χ².

Fig 2. Y-shaped olfactometer bioassays.

Host preference responses (% first choice) of B. hilaris adults to seedlings of B. oleracea var. botrytis, B. carinata, and B. napus. N = number of replicates; ** = P < 0.01; χ².

Fig 3. EAG bioassays to crude extracts of VOCs from B. oleracea var. botrytis seedlings.

Electroantennogram responses (mV) (mean + SE) of antennae of B. hilaris 4^th and 5^th instar nymphs (N = 20) (A) and adults (N = 15) (B); ** = P < 0.01; one-way ANOVA followed by Tukey’s test.

Fig 4. EAG bioassays to fractions of crude extracts of VOCs from B. oleracea var. botrytis seedlings.

Electroantennogram responses (mV) (mean + SE) of B. hilaris 4^th and 5^th instar nymphs (N = 20) (A) and adults (N = 18) (B); Different letters indicate that values differ statistically at P < 0.05; one-way ANOVA followed by Tukey’s test.

Fig 5. Behavioral bioassays with B. hilaris 4^th and 5^th instar nymphs.

Responses (% first choice) to B. oleracea var. botrytis seedlings, to crude extracts of seedling VOCs, and to fractions therein, versus controls, in open vertical Y-shaped olfactometer bioassays. N = number of replicates; * = P < 0.05; χ².

Fig 6. Behavioral bioassays with B. hilaris adults.

Responses (% first choice) to B. oleracea var. botrytis seedlings, to crude extracts of seedling VOCs, and to fractions of the VOC extract, versus controls, in vertical Y-shaped olfactometer bioassays. N = number of replicates; * = P < 0.05, ** = P < 0.01; χ².

Fig 7. Representative gas chromatograms of VOCs from B. oleracea var. botrytis.

(A), B. napus (B), B. carinata (C), and B. oleracea var. botrytis non-polar fraction (D). The peak numbers indicate the diterpene hydrocarbons detected in the analysis.

Fig 8. EI mass spectrum of the major compound in VOC extracts from B. oleracea var. botrytis.

Fig 9. EI mass spectrum of the product from catalytic reduction of the major compound in VOC extracts from B. oleracea var. botrytis.