Figure 1.
In southern AZ, M. sexta moths use the nectar resources of two plants: A. palmeri and D. wrightii.
(A) The D. wrightii flower exhibits classic characteristics of moth-adaptation with reflective corolla, sweet-smelling perfume, and sucrose-dominant nectar. These characteristics elicit an innate feeding behavior in M. sexta. (B) M. sexta also nectars from A. palmeri flowers. Moths learn to utilize A. palmeri through association of the floral scent and nectar.
Figure 2.
Characterization of the effects of bagging flowers on floral scent emissions, reflectance and gross morphology.
(A) D. wrightii corolla reflectances of unbagged (dark blue) and bagged (light blue) flowers. (B) A. palmeri petal reflectance of unbagged (orange) and bagged (yellow) flowers. For both flower species, bagging had little effect on floral reflectance. (C) The power spectrum of wind velocity fluctuations produced from a bagged (light blue) and unbagged (dark blue) D. wrightii flowers in a wind tunnel. (D) The power spectrum of wind velocity fluctuations produced from bagged (yellow) and unbagged (orange) A. palmeri umbels. For both D. wrightii and A. palmeri flowers, bagging had no effect on the turbulent wind fluctuations or energy cascade. (E) GCMS ion chromatograms of the captured headspace volatiles emitted from unbagged (dark blue) and bagged (light blue) D. wrightii flowers. Major constituents of D. wrightii floral headspace scent shown in the total ion chromatogram are monoterpenoids β-myrcene (1), E-β-ocimene (3), and geraniol (5), aromatics including benzyl alcohol (2) and methyl salicylate (4). Letters denote contamination from the polyacetate bag. (F) Ion chromatograms of the headspace volatiles from bagged (yellow) and unbagged (orange) A. palmeri umbels. Major constituents of A. palmeri floral odor shown in the total ion chromatogram are monoterpenoids α-pinene (7), camphene (8), and esters such as ethyl isovalerate and analogs (6, 9) and ethyl sorbate isomers (10 and 11). Letters denote contamination from the polyacetate bag. (G) Flower volatile concentrations measured at specific locations in a flight arena from the emitting flower using a mini photoionization detector. (Left) Schematic of the flight arena and sampling locations from the flower. (Right) Mean volatile concentrations at the locations from the flower. Volatile concentrations drop rapidly with increasing distance from the flower until at 1 m from the flower the concentration are <0.05% of the intensity near the source. Insets are photoionization traces through time, at sample points 0.05, 0.2, and 1.0 m from the source. Dashed line indicates the volatile background concentration.
Table 1.
Two-choice experimental treatments.
Figure 3.
Two-choice experiments examining the visual and olfactory floral preferences of naïve male M. sexta moths in response to live flowers or scented bagged flowers.
(Top) The percentage of moths that chose the live D. wrightii flower (Flower A) or a bagged D. wrightii flower with D. wrightii scent (Flower B). (Bottom) The percentage of moths that chose the live A. palmeri umbel (Flower A) or a bagged umbel with scent (Flower B). 20 moths were used in each two-choice experiment. In both experimental series, there were no significant differences in the first flower chosen (G-test: P>0.71). D. wrightii flower cues (odor and visual) are represented by white bars, A. palmeri flower cues (odor and visual) are represented by grey bars, and hashed bars represent the real flowers.
Figure 4.
Total time M. sexta moths attempted to feed from flowers in two-choice experiments.
There were no significant differences between mean flower feeding times between tests (one-way ANOVA: F13,121 = 0.88, P = 0.57) or between two-choice treatment groups (post-hoc Scheffé test all comparisons: P>0.97). Note: only manipulative two-choice test values are shown for clarity. Experiments are numbered according to two-choice treatments shown in Table 1.
Figure 5.
Two-choice experiments examining the visual and olfactory floral preferences of naïve male M. sexta moths.
(A) Using a green shade-cloth to mask the visual display of the flowers, the effects of scent and a visual stimulus (paper flower) were tested in isolation and simultaneously. (B) With artificial flowers, the percentages of moths that chose paper flowers emitting D. wrightii (white bars), A. palmeri scent (grey bars), or no scent (control) flowers (black bars). (C) With bagged flowers to stop the scent emissions but allowing display of the floral visual signals, the percentages of moths that chose the D. wrightii, A. palmeri, or the (control) paper flower visual display. Asterisks (*) denote a significant deviation from a random distribution (G-test: P<0.05). 20–40 moths were used in each two-choice experiment. Moths were tested individually, with each two-choice treatment using different groups of moths. D. wrightii flower cues (odor, and/or visual) are represented by white bars, A. palmeri flower cues (odor, and/or visual) are represented by grey bars, and black bars represent the paper flower (no odor) control.
Figure 6.
Two-choice experiments examining the single modality and multimodality floral display preferences in naïve male M. sexta moths.
(A) With visual (bagged flowers) versus olfactory (scented paper flowers) displays, the preferences in naïve moths using D. wrightii and A. palmeri olfactory and visual displays. (B) With olfactory-dominant (scented paper flowers) and multimodal (olfactory and visual) flowers, the percentages of moths that chose the floral displays. (C) With visual-only (bagged flowers) and multimodal (olfactory and visual) flowers, the percentages of moths that chose the bagged (visual only) or multimodal (visual and olfactory) floral displays. D. wrightii flower cues (odor, and/or visual) are represented by white bars, and A. palmeri flower cues (odor, and/or visual) are represented by grey bars. Asterisks (*) denote a significant deviation from a random distribution (G-test: P<0.05). 16–30 moths were used in each two-choice experiment. Moths were tested individually, with each two-choice treatment using different groups of moths.
Figure 7.
Two-choice experiments examining the visual floral preferences of naïve male M. sexta moths when the floral scents are similar.
(Top) The percentages of moths that chose the the A. palmeri visual display (Flower A; grey bar, hashed white) or D. wrightii visual display (Flower B; white bar) when both flower species emit the D. wrightii scent. (Bottom) The percentages of moths that chose the A. palmeri visual display (Flower A; grey bar) or D. wrightii visual display (Flower B; white bar, hashed grey) when both flower species emit the A. palmeri scent. 20 moths were used in each two-choice experiment. In both experimental series, there were no significant differences in the first flower chosen (G-test: P>0.73).
Figure 8.
Response indices calculated from the time moths spent attempting to feed from the flowers in the single modality versus control, or single modality versus multimodal two-choice experiments.
(A) A. palmeri floral signals. (B) D. wrightii floral signals. (C) A. palmeri versus D. wrightii visual (top) and olfactory (bottom) signals. Letters (A,B) or asterisks (C) denote a significant difference between two-choice treatments (unpaired t-test: P<0.05).
Figure 9.
The effects of coupled (solid bars) versus uncoupled (hashed bars) floral displays for experienced male M. sexta moths.
24 hours prior to testing, moths were assigned to one of three treatment groups: moths exposed to D. wrightii flowers (white bars), A. palmeri flowers (grey bars), or flower-naïve moths (black bars), and were re-tested the following evening. (A) Using either real flowers with their visual and olfactory signals coupled, or uncoupled, the percentages of moths (from the total number of moths tested) that chose D. wrightii or A. palmeri flowers. An asterisk (*) denotes a significant deviation from a random distribution (G-test: P<0.05). (B) The time moths spent flying before attempting to feed from the previously experienced olfactory floral cue. Letters denote a significant difference between two-choice treatments (unpaired t-test: P<0.05). (C) Two-dimensional flight tracks for experienced moths to coupled floral displays (top, blue circles) and uncoupled floral displays (bottom, green circles) in a flight arena. Circles correspond to video images captured at 0.033 s intervals. Moths were tested individually, with each two-choice treatment using different groups of moths.