Fig. 3

Heptanal elicits obvious avoidance behaviors in both Plutella xylostella larvae and adults. Preference index of P. xylostella wild-type (WT) and CRISPR/Cas9 PxylOR16 knockout (PxylOR16^−/−) adults and larvae for heptanal. Preference indexes = (number of choices at treatment – number of choices at control) / number of total choices. A Preference index of WT and PxylOR16^−/− mutant female and male moths for control (filter paper vs filter paper) and for filter paper + paraffin oil vs filter paper + heptanal in a Y-tube olfactometer (n = 16). WT male moths exhibited significant avoidance to heptanal at doses of 100 ng, 1 μg, and 10 μg (left). Avoidance was also observed for WT female moths, but only at doses of 1 μg and 10 μg (Welch’s t-test; NS, no significant difference, P > 0.05; *** P < 0.001) (right). PxylOR16^−/− mutant female and male moths did not show a preference for one side of the device (Welch’s t-test; NS, no significant difference, P > 0.05). B Preference index of WT and PxylOR16^−/− mutant female and male moths for blank (one side blank vs a piece of Brassica pekinensis), for control (a piece of B. pekinensis vs a piece of B. pekinensis), and for a piece of B. pekinensis + paraffin oil vs a piece of B. pekinensis + heptanal in a Y-tube olfactometer (n = 16). WT and PxylOR16^−/− mutant female and male moths were significantly attracted to B. pekinensis, compared with blank. WT male moths were significantly less attracted to B. pekinensis + heptanal at doses of 100 ng, 1 μg, and 10 μg than to B. pekinensis alone (with paraffin oil) (left). Female moths were also less attracted to B. pekinensis + heptanal than to B. pekinensis alone but only at heptanal doses of 1 μg and 10 μg (Welch’s t-test; NS, no significant difference; P > 0.05; *** P < 0.001) (right). The PxylOR16^−/− mutant female and male moths lost their avoidance responses to heptanal significantly, compared with WT adult female and male moths (Welch’s t-test; NS, no significant difference; P > 0.05). C Preference indexes of 10 WT and mutant female and male third instar larvae for blank (one side blank vs a piece of B. pekinensis), for control (a piece of B. pekinensis vs a piece of B. pekinensis), and for a piece of B. pekinensis + paraffin oil vs a piece of B. pekinensis + heptanal in a 10-cm diameter plastic Petri dish (n = 28). Both sexes of WT and mutant larvae were attracted to B. pekinensis. Significantly more female and male third instar larvae were located in the control area (B. pekinensis + paraffin oil) than in the heptanal-supplemented area (B. pekinensis + heptanal) at all heptanal doses (100 ng, 1 μg, and 10 μg on filter paper) except for 10 ng (Welch’s t-test; NS, no significant difference, P > 0.05; *** P < 0.001). As shown for PxylOR16^−/− adults, detection of heptanal was abolished in PxylOR16^−/− larvae (Welch’s t-test; NS, no significant difference, P > 0.05)