Diurnal moths have larger hearing organs: evidence from comparative 3D morphometric study on geometrid moths

Introduction

The animal world is either diurnal, nocturnal, or crepuscular; as the adaptations to function in one temporal activity pattern reduce the efficiency of functioning and survival in another (Fraser, Metcalfe & Thorpe, 1997). Nocturnality is presumed to be associated with adaptations to function in low-light and low-temperature conditions. However, in predominantly nocturnal taxa, like geckoes and moths, some clades have independently shifted to diurnal activity and evolved distinct adaptations suited to diurnal conditions like camouflage (Fulgione et al., 2019), photopic vision (Sondhi et al., 2021; Kojima et al., In press) aposematic (warning) wing colouration (Fiedler & Brehm, 2021) and hearing sensitivity to lower ‘non-bat’ frequency (Fullard et al., 1997). The shift to diurnality in these clades is likely driven by a combination of ecological factors such as predation, climate, and competition (Gamble et al., 2015). These factors can influence how animals send and receive information in their environment, which can lead to new selective pressures on sensory systems (Sih, Ferrari & Harris, 2011). For example, reduced bat predation during the day, prevalence of different visual predators, or changes in background sounds can all influence which sensory traits are most advantageous.

Predators are an important selective force in all terrestrial food webs. Prey species may invest proportionally more in defence, avoid interactions with their predators or follow alternative strategies for survival. For moths, the predation pressure from their most formidable predator, bats, vary in a predictable manner among nocturnal and diurnal species, influencing their daily activity pattern and hearing sensitivity (Rydell, Jones & Waters, 1995). This is reflected in the positive correlation between nocturnal flight activity and hearing sensitivity to ultrasound frequencies (ter Hofstede, Ratcliffe & Fullard, 2008). Shifting to diurnality is likely one of several strategies moths use to avoid bat predation.

Diurnal moths show marked degeneration of the ultrasonic hearing, and some of them are reported to be ‘bat-deaf’ (Fullard et al., 1997). There are reports of predator released moths with no ears, e.g., the Polynesian Pyralid Lathroteles obscura (Clarke, 1971); and extremely poor sensitivity in female wingless geometrids (Rydell et al., 1997). While hearing sensitivity is often closely linked to body size, with larger animals generally having larger hearing organs and lower frequency hearing, as observed in lizards, frogs, mammals, and birds (Hetherington, 1992; Werner & Igic, 2002; Gleich & Langemann, 2011). Larger noctuid moths often have larger tympanal organs, but may be less sensitive to higher frequency bat calls compared to smaller moth species, like the crambid moths (Fullard, 1988; Nakano & Mason, 2017). Moreover, hearing sensitivity in tympanal insects depends on more than organ size-it is also influenced by the number of sensory neurons (Loh et al., 2023; Göpfert & Wasserthal, 1999), the mechanical properties of the tympanic membrane (Lomas et al., 2011; Austin et al., 2024), and external temperature (Eberhard et al., 2014; Korsunovskaya & Zhantiev, 2007). Since diurnal species are typically active during warmer periods of the day, thermal conditions may also affect their hearing performance. Therefore, the development and function of moth hearing organs, is not solely dictated by size, but likely reflect a complex interplay of physiological, ecological and environmental factors.

Hearing organs in insects have diverse functions, such as intraspecific communication, parasitic host localization, and predator avoidance (Strauß & Lakes-Harlan, 2014). One specialized form of these organs, the tympanal organs, typically consists of a thin membrane or tympanum backed by an air-filled sac, and has evolved independently in at least seven different insect orders (Hoy & Robert, 1996). In most tympanate insects, the highest sensitivity is between 15–80 kHz, i.e., within the bat calling frequencies (Roeder & Treat, 1970; Kawahara & Barber, 2015; Hummel, Kössl & Nowotny, 2011; Miller & Surlykke, 2001). This evolutionary pressure is particularly evident in nocturnal moths, where the multiple origins of hearing organs are thought to be a defense mechanism against insectivorous bats (Roeder & Treat, 1970; Kawahara & Barber, 2015; Scoble, 1992; Ratcliffe & Fullard, 2005; ter Hofstede et al., 2013). Most moths including Noctuoidea, Geometroidea and Pyraloidea possess ultrasound sensitive ears (Fullard, 1998). As such, we would expect moths that are isolated from bats geographically or temporally to exhibit lower sensitivity to ultrasound (Fullard et al., 1997; Surlykke, 1986).

We chose geometrid moths as our model group due to well-documented diel activity patterns (Lee et al., 2024) and their use in testing hearing sensitivity in diurnal moths (Surlykke et al., 1998). While adult geometroid moths are primarily nocturnal, several unrelated lineages across families Uraniidae, Sematuridae, and Geometridae, exhibit diurnal activity (Hausmann, 2001; Kawahara et al., 2018; Õunap, Viidalepp & Truuverk, 2016). This transition has occurred independently in Lepidoptera multiple times (Sondhi et al., 2021; Kawahara et al., 2018; Kawahara & Breinholt, 2014). The strictly diurnal geometrid moth Archiearis parthenias, which is temporally isolated from bats, is sensitive to around 12 kHz and practically deaf beyond 25 kHz (Surlykke et al., 1998), comparable to sympatric noctuid moths (Rydell et al., 1997; Surlykke & Filskov, 1997). The tympanal organ is an important taxonomic character for geometrids, with ansa being a prominent unique character of the family (Cook & Scoble, 1992; Rajaei et al., 2022) (Fig. 1C, and Video S1). The ansa, thought to protect the tympanum from mechanical damage caused by pressure from surrounding organs, particularly the oesophagus and flight muscles, has a wide base that strengthens the inner wall of the cavus, helping maintain tension across the tympanum (Cook & Scoble, 1992). The variation in ansa shape and size at the species and higher taxonomic level suggests it may serve functions beyond just a mechanical structure. However, research on diel activity in moths, function of hearing organ anatomy and predator pressures, remains limited.

In the above context, we wanted to ask, using a non-destructive 3D geometric morphometry: (1) Does the size of the tympanal organ show an allometric relationship with body size? (2) Is there a difference in the size of the tympanal organs between diurnal and nocturnal moths? (3) How do the shape and size of the ansa vary between diurnal and nocturnal species? The overarching question guiding this study is whether these differences are linked to specialized hearing adaptations for detecting predators. We hypothesize that the structure of the tympanal organs is driven more by diel activity patterns than by body size, with tympanal organs being smaller in diurnal moths due to reduced anti-bat function. Also, we hypothesize that the ansa plays a crucial role in auditory function—not just for mechanical strength—meaning its size and shape should vary depending on the moth’s activity pattern.

Materials and Methods

We selected dried museum specimens of 19 geometrid species (seven diurnal, one cathemeral and 11 nocturnal). The study specimens with diurnal vs nocturnal species pairs, were chosen based on their relatedness classified in the same genus, same tribe, or same subfamily (Murillo-Ramos et al., 2019; Brehm et al., 2022; Murillo-Ramos et al., 2023). All study specimens are males, dry specimens deposited in museums, and they have been collected during the years 1993–2021, from different geographical locations, listed in Table S1.

The authors received permission from the National Biodiversity Authority, India to export and study specimens collected from India at the Finnish Museum of Natural History (Permit no: NBA/Tech Appl/9/INBA1202203315/22/22-23/132S).

Results

The phylogenetic linear regression revealed no significant allometric relationship of tympanal organ volume with body size in geometrid moths (adjusted R-square = 0.06, p > 0.05) (Fig. 2A). This suggests that the size of the tympanal organ does not scale proportionally with the overall body size in this group. Notably, the diurnal moths were found to be significantly smaller in body size compared to their nocturnal counterparts (Fig. 2B). Despite the absence of a significant allometric relationship, we observed that diurnal moths possess significantly larger tympanal organs than nocturnal species (p < 0.05) (Fig. 3).

In terms of the ansa, no significant differences were found in the overall shape or size across taxa (Fig. 4). However, we did observe some evidence of convergence in the size of the ansa among the nocturnal species. The inter-quartile range for the ansa size in diurnal moths was 0.197, while for nocturnal moths it was 0.09 (Fig. 5). The micro-CT scans also reveal that the ansa is hollow with a narrow tube-like structure in the middle (Fig. 6 and Video S2). This indicates that while the ansa size does not vary significantly across all species, there may be a trend towards similar shape characteristics within nocturnal geometrid moths, possibly reflecting a shared evolutionary adaptation related to hearing sensitivity.

Discussion

In this study, we investigated how auditory structures in moths change as a function of body size, and the extent to which certain morphological traits predict hearing sensitivity. Our findings revealed several interesting patterns in the tympanal organ and ansa characteristics of geometrid moths. Despite no significant allometric relationship between tympanal organ volume and body size, we found that diurnal moths tend to have larger tympanal organs—despite being smaller overall—pointing to potential evolutionary adaptations that help them navigate different abiotic and biotic factors, such as avoiding diurnal predators. These include the flight sounds and calls of insectivorous birds, or the rustling vegetation sounds created by birds or other terrestrial predators like lizards. Such diverse acoustic cues may have shaped the tympanal organ morphology in diurnal moths (Yack et al., 2020).

In essence, our result means that the tympanal organ, originally assumed to function as an anti-bat hearing organ in nocturnal species, has adapted a new function in diurnal environment. The structure could still have a hearing function, potentially being adapted to different predator pressure on lower “non-bat” frequency and the larger size could serve more effectively this purpose.

Interestingly, the shape and size of the ansa do not differ significantly across taxa, though there is some evidence of convergence in the size of the ansa among nocturnal moth species. This suggests that the ansa may be playing a previously unexplored role in auditory function.

These findings highlight the complex evolutionary dynamics between predator-prey interactions and morphological adaptations in moths. Moths seem to be fine-tuning their auditory systems in response to their specific ecological niches, whether to optimizing hearing for bat evasion or adapting to the demands of diurnal activity. This research offers new perspectives on how sensory organs might contribute to survival and how predators could play a role in shaping highly specialized traits over time.

Conclusion

In conclusion, this study uses 3D geometric morphometry to hearing related insect cuticular structures in geometrid moths, offering new insights into the complex interplay of ecological pressures, evolutionary constraints, and morphological adaptations in this group. The lack of a significant allometric relationship between tympanal organ size and body size, alongside the presence of larger tympanal organs in diurnal moths, suggests that environmental factors, particularly predator-prey interactions, likely contribute to adaptations in auditory structures. The potential for convergent evolution, especially in the size and shape of the ansa across nocturnal species, further underscores the role of selective pressures in shaping auditory morphology. We gain valuable insights into how body size influences both hearing capabilities and evolutionary trends. Ultimately, our study emphasizes the importance of considering environmental and phylogenetic constraints when interpreting the evolution of auditory structures and opens avenues for further investigation into the convergent mechanisms of hearing across taxa.

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