The rising moon promotes mate finding in moths
The moon increases mate finding in mothsTo investigate the impact of natural and artificial light sources on mate finding, we analyzed flight behavior in male moths, which were reliably attracted by caged virgin females (see Materials and Methods for details). Since we used these females specifically to exploit their attraction effect, we refer to them as ‘traps’ in the following. To establish a choice scenario (see below), males were released equidistantly from the traps, which were located north and south of the core release site in central Germany. Besides the stars, the moon creates the natural light environment that moths might use for visual orientation. We therefore first tested if the moon affects mate finding. We found that the percentage of males arriving within the experimental time (8 min from release, 58.6% of flights) at a trap increased significantly with the appearance of the moon (logistic regression: z = −2.06, p = 0.04, n = 58) and did not depend on the presence of clouds in front of the moon (z = −0.83, p = 0.406, n = 58). A few males reached the females later during the experimental night (13.8% of flights) and were released again on the next day. Some males never reached a trap and could therefore not be tested again in the next days (27.6% of flights). Furthermore, the time that successful males needed to reach a trap was significantly influenced by the height of the moon above or below the horizon (Fig.1; Cox PH survival model, z = 2.46, p = 0.014, n = 34): the higher the moon was above the horizon, the faster males were able to locate and reach the females. The presence of clouds in front of the moon did not play a significant role in this context either (z = −0.65, p = 0.519, n = 34), leading to the conclusion that the moon was equally well perceived if covered partly by clouds and used for effective orientation towards the females. Although the lunar phase changed during the period of the experiment from full moon to new moon, flight duration was not significantly affected by the percentage of the lit moon disk (z = 0.44, p = 0.66, n = 34). Thus, the properties of the moon that affected the flight duration of males were independent of the lunar phase.Fig. 1: Expected flight duration of a moth.Flight duration (black line) was calculated as the median flight duration predicted by the Cox PH model (p = 0.014, n = 34) for arrivals within 8 minutes after release and averaged over all individuals. Circles represent the actual measured values. Dashed lines indicate the confidence interval of the predicted duration at α = 5% level estimated by bootstrapping (5000 replicates).Full size imageIt is important to emphasize that the results were not significantly affected by traits on the individual level like body size or origin of the animal (see Supplementary Results and Discussion for details). Furthermore, a possible learning effect of animals that were released more than once was not detectable since flight duration did not decrease depending on ‘experience’ but only with the elevation of the moon (Fig. S1). Thus, the moon as an easily perceivable orientation cue increased mate finding in general but also depended on its elevation. Despite two exceptions of long flight durations at moon elevations > 20° that go back to the same animal probably for individual reasons (Fig. S1), the variance in flight duration was highest at low moon elevations (Fig. 1). This relatively high variance at low moon elevations emphasizes the question if artificial lights affected mate finding, particularly whenever the moon as a natural light cue was not yet prominent.Linking flight behavior to the light environmentWe used a calibrated digital all-sky camera to track changes in the natural and artificial components of the night sky brightness24 (Fig. 2 a–c). A similar camera system was recently used to study dung beetle behavior21. Although the impact of light pollution on the site was not strong, the night sky was also not completely pristine. Luminance (LVv) values were about 0.34 mcd/m² at zenith and 1.6 mcd/m² near the horizon under clear sky conditions when the moon was not visible. A natural (unpolluted) sky brightness is 0.25 mcd/m² at zenith and can be used as the reference value “Natural Sky Unit” (NSU) for easy comparison (see also Materials and Methods). The analysis of specific sky sectors revealed that the moon was the strongest factor determining the ambient brightness, brightening every sector of the sky as soon as it appeared above the horizon (Fig. 2d). During observation times, the course of the moon mainly progressed through the eastern part of the sky, affecting particularly the LvV values in the corresponding sectors (Fig. 2d). Furthermore, light conditions never corresponded to a non-light polluted sky, as NSU values were always greater than one. Most sectors in the south, west and north (sectors seven to 12 and one) were hardly subjected to fluctuations. Nevertheless, it is recognizable that the moon made a decisive contribution to the light environment in all directions since images with the moon above the horizon were always brighter than those with the moon below the horizon (Fig. 2d).Fig. 2: Quantification of the light environment with all-sky imagery and its impact on flight behavior of moths.a Raw RGB all-sky image with clear sky and a visible moon 26° above the horizon at 119° azimuth angle, South-east (24 July 2019, 03:23). b Same image as in a with processed luminance values. c Processed all-sky image in luminance with clear sky, a visible milky way (green patches in a ‘ribbon-shape’ across the (blue) night sky), skyglow near the horizon, and a non-visible moon 0° above the horizon at 87° azimuth angle, East (24 July 2019, 0:25). The colors of the processed image correspond to the legend in b. The black lines mark the sky segments used to quantify the light environment. The outer ring covers 5° above the horizon (85°−90° zenith angle), the inner ring 20° above the outer ring (65°−85° zenith angle). Furthermore, the sky was divided into 12 sectors of 30° width along the azimuth direction (extension by dashed line), starting with the sector marked with the small circle (counting clockwise). d Luminance in natural sky units (NSU) for each full sector of 30°. The moon icons indicate sectors in which the moon was visible, regardless of its phase. The size of each symbol encodes the rank of the frequency (n = 33). e Trap choice of arrived males depending on the position of the moon at the moment of release on the north-south axis (north = 0°). The y-axis displays choice of the southern trap at 0.0 and of the northern trap at 1.0. p = 0.022, n = 42. f Male moth affinity to northern trap in response to the direction of maximum luminance measured in the outer ring of 5°. Each circle indicates an observed arrival, p = 0.753, n = 41. g Male moth affinity to northern trap as in f but with luminance measured in the inner ring of 20°, p = 0.065, n = 41. e–g The line represents the prediction of the logistic model, providing a probability value for arriving at the northern trap (north prone = 1; south prone = 0). Dashed lines indicate the confidence interval of the prediction at α = 5% level estimated by bootstrapping (5000 replicates).Full size imageDue to the design of the experiment with one trap located in the north and the other in the south of a central release site, we were able to investigate the choice behavior of males, especially in respect of the possible influence of the cardinal position of the moon as it was almost exclusively visible in the southern hemisphere of the sky (Fig. 2d). Although the moon continued to move south during the night, the moon’s cardinal position never overlapped with the exact direction of the southern trap. The only parameter that had a significant effect on choice behavior was indeed the cardinal position of the moon (Fig. 2e, logistic regression, z = −2.3, p = 0.022, n = 42). The more southern the moon’s position was, the more likely males flew to the southern trap. However, while some clouds in front of the moon had no significant effect on choice behavior (z = 0, p = 1, n = 42), moon above the horizon showed a tendency to affect males (z = −1.82, p = 0.069, n = 42). The results indicate that despite the general increase of ambient brightness by the moon, it is its position that mainly influenced the flight direction of males. Thus, moths preferred a flight direction with the prominent compass cue ahead to steer their flight towards the females but it is important to emphasize that moon and trap had an angular difference of at least 23° (80.8° to the moon’s mean cardinal direction). Therefore, males that chose to fly towards the southern trap did not fly directly towards the direction of the moon.As the moon represents a natural distant light source, we tested whether distant artificial light sources or skyglow might elicit a comparable effect on the behavior of male moths and if such light sources might mask the moon. To evaluate the light environment with regards to these aspects, we defined sky segments of particular interest that occurred due to the location of the experimental field (Fig. 2c). For each arrival at a trap, the brightest sector of the environment was determined and placed on a north-south axis of maximum 180 degrees (Fig. 2f, g). If we look at the brightest sector of the environment and distinguish between the area close to the horizon, i.e. “outer ring” (Fig. 2f) and the one above, i.e. “inner ring” (Fig. 2g), we can observe differences in trap choice. The line indicates the logistic regression model and provides the probability of arriving at the northern trap. For the Lv in the area close to the horizon no effect of maximum Lv on trap choice could be found (logistic regression, z = 0.31, p = 0.753, n = 41). For the segment further above the horizon the probability of flying to the southern trap increased with maximum Lv but the results are marginally not significant (z = −1.85, p = 0.065, n = 41). Our results for trap selection indicate that distant artificial lights of the surroundings did not attract males and support the hypothesis that the moon, once it appears above the horizon and stands out from the general light (pollution) near the horizon (above five degrees), is used as an effective visual cue with moths rather flying towards than away from.Digital cameras are suitable to measure the dynamics of night-time lighting conditions25,26, and allow researchers to track changes in artificial lighting conditions and brightness of the sky simultaneously27. However, it is not straightforward to distinguish between ALAN and natural light sources like the moon with luminance images when the moon is close to the horizon and thus in the section of the sky where most light pollution occurred. Yet, once the moon rose higher than 5° and thus stood out distinctly from the light-polluted horizon, it could be clearly identified on the images (Fig. 2b). In this context, it is particularly remarkable that the speed at which the females were reached increased reliably only above a similar threshold (Fig. 1), with the only exceptions of two flights with long durations at a moon elevation greater than 20° (Fig. 1); both flights originated from the same individual (Fig. S1). Thus, the high variance of flight durations at low moon elevations (Fig. 1) supports our hypothesis that the moon, as an orientation cue, can be masked by artificial light for the animals as well. Yet, this hypothesis needs to be explicitly tested in future experiments. In general, the possible consequences of light pollution are still uncertain28, especially because the amount of artificial light emitted during the night continues to increase exponentially worldwide18. But regardless of this, the moon is the decisive orientation cue as soon as it is visibly silhouetted against the horizon despite possible diffuse light pollution.Another interesting next research project would be to investigate the relevance of polarized light, as this could provide an explanation for the occasional fast flights at times of low lunar elevations (cf. Figure 1). Furthermore, it might explain why flight duration was not significantly affected by clouds in front of the moon since the polarization pattern extends over the whole sky and is therefore not shielded completely by scattered clouds29. For dung beetles it has been already shown that they are capable of using the polarization signal for navigation16,30,31 and it has been proposed that moths might be capable of utilizing the same signal32. At the same time, it has already been demonstrated that urban skyglow can diminish the lunar polarization signal33, making a detailed investigation of the interplay between these two factors and the significance for moth orientation particularly exciting to understand underlying mechanisms.Our results confirm that moths use the moon as an orientation cue, supporting the notion of Vickers & Baker34 that pheromones alone are not sufficient for successful (and fast) orientation. Since flight duration decreased as a function of lunar elevation, we conclude that the moon contributes to mating success, especially when it can be easily perceived. Since nocturnal landscapes around the world have been drastically restructured in terms of light intensity and light spectrum due to the rapid spread and increase of electrical lighting18, a deeper understanding of orientation mechanisms even in the absence of the moon as an easily perceivable cue could provide a valuable contribution to counteract insect decline. More
