# Combinatorial Olfactory Signaling in Short-Distance Determines Host Plant Recognition in Locust
## 3 Results
### Volatiles from Host and Nonhost Plants Contribute to Food Selection in Locally _Food Selection_  
Volatiles derived from plants are probably the critical cues that guide host recognition by insect herbivores. However, the olfactory strategy in the insect-plant interaction remains unresolved. First, we observed that locusts preferred two host-specific volatiles and two common volatiles over the control solvent (Figure 2A,B, Video S2), whereas they refused two non-host-specific volatiles (Figure 2C, Video S3). Butyric acid has been reported to be a sensitive volatile stimulus to trigger the palp opening response in a previous study [29]. Therefore, we used butyric acid as a positive control. Similarly, butyric acid elicited an obvious DER response (Figure 2D). These results supported the contention that plant-specific volatiles confer reverse effects on locust palpation. Based on the release ratio of several volatiles in corn and cotton plants, we reproduced the mixed volatiles (Figure S1D,E). As expected, mix B, which mimicked the chemical composition in cotton, repelled locusts significantly; however, mix A, which mimicked the chemical composition in corn, did not attract locusts (Figure S1F). These results suggested that either inhibitory heptanol outperforms the feeding attraction of hexanal or more hedonic chemicals in corn are needed to mimic the feeding preference. Figure 1: Locusts discriminate host plant volatiles through palp sensation. (**A**): Schematic diagram of the duration of excitatory response (DER) behavior assay. (**B**): Examples of locust behavior in the testing arena. Top, palpation-like behavior in close proximity to wheat grinds. T0–T2, temporal order; bottom, trajectories of two locusts (track 1 and 2) between wheat grinds (odor zone) and solvent (CK zone). (**C**): PI_DER of Ant\(-\)/palp+ locusts to wheat juice and wheat grinds. \(N=\) 14–23. (**D**): PI_DER of antenna-intact locusts (Ant+, Palp\(-\)), palp-intact locusts (Ant\(-\), Palp\(+\)) and both antenna- and palp-amputated locusts (Ant\(-\), Palp\(-\)) to wheat grinds. \(N=\) 10–19. (**E**): PI_DER of locusts to plant grinds between wheat and corn or between soybean and cotton. \(N=\) 10–16. (**G**): PL_DER of locusts to plant grinds between wheat and cotton. \(N=\) 16–18. One-way ANOVA with [PERSON]’s LSD multiple comparisons tests in (**C**–**F**) and unpaired Student’s \(t\)-test in (**G**). Error bars represent the SEM. * \(p<\) 0.05, **\(p<\) 0.01, ****\(p<\) 0.0001. Figure 2: Olfactory discrimination among single odorants at close range. (**A**–**D**): PI_DER of Ant\(-\)/Palp+ locusts to host-plant-specific volatiles (**A**), common volatiles (**B**), non-host-specific volatiles (**C**), and butyric acid (**D**) as a positive control. \(N=12\)–\(25\). (**E**): PI_DER of Ant\(-\)/Palp+ locusts to wheat grinds mixed with single nonhost volatiles. \(N=14\)–\(21\). (**F**): PI_DER of Ant\(-\)/Palp+ locusts to cotton grinds mixed with single host volatiles. \(N=9\)–\(24\). (**G**): PI_DER of Ant\(-\)/Palp+ locusts to cotton grinds mixed with the common volatile hexanal. \(N=11\)–\(21\). (**H**): Working model of locust palps action in distinguishing host plants from nonhosts by detecting volatiles. Red undulating lines represent host-specific volatiles. One-way ANOVA with [PERSON]’s LSD multiple comparisons tests in (**A**–**C**,**E**–**G**), and unpaired Student’s \(t\)-test in (**D**). Error bars represent the SEM. * \(p<0.05\), **\(\textbf{{}^{*}}p<0.01\), *** \(p<0.001\). We further demonstrated the sufficiency of plant odors in manipulating locust foraging behavior. The addition of nonhost odors into the wheat grinds reduced the total preference of wheat extraction (Figure 2E), whereas supplementation of two host odors into the cotton grinds significantly increased the palpation-like behavior (Figure 2F). In addition, the attractive common volatile hexanal was sufficient to confer the hedonic valence of cotton extraction (Figure 2G).