Molecular Evolution of Malacostracan Short Wavelength Sensitive Opsins.

Investigations of the molecular mechanisms behind detection of short, and particularly ultraviolet, wavelengths in arthropods have relied heavily on studies from insects due to the relative ease of heterologous expression of modified opsin proteins in model organisms like Drosophila. However, species outside of the Insecta can provide information on mechanisms for spectral tuning as well as the evolutionary history of pancrustacean visual pigments. Here we investigate the basis of spectral tuning in malacostracan short wavelength sensitive (SWS) opsins using phylogenetic comparative methods. Tuning sites that may be responsible for the difference between ultraviolet (UV) and violet visual pigment absorbance in the Malacostraca are identified, and the idea that an amino acid polymorphism at a single site is responsible for this shift is shown to be unlikely. Instead, we suggest that this change in absorbance is accomplished through multiple amino acid substitutions. On the basis of our findings, we conducted further surveys to identify spectral tuning mechanisms in the order Stomatopoda where duplication of UV opsins has occurred. Ancestral state reconstructions of stomatopod opsins from two main clades provide insight into the amino acid changes that lead to differing absorption by the visual pigments they form, and likely contribute the basis for the wide array of UV spectral sensitivities found in this order.

Physiological and behavioral evidence for multiple spectral channels in the larval stomatopod visual system.

Larval stomatopods have generally been described as having a typical larval crustacean compound eye, which lacks the visual pigment diversity and morphological specializations of the well-studied stomatopod adult eye. However, recent work has suggested that larval stomatopod eyes are more complex than previously described. In this study, we provide physiological and behavioral evidence of at least three distinct photoreceptor classes in three species of larval stomatopods: Gonodactylellus n. sp., Gonodactylaceus falcatus and Pullosquilla n. sp. First, electroretinogram recordings were used to measure the spectral sensitivity of each species. Evidence for at least three spectral classes were identified in each: an ultraviolet, peaking at 340-376 nm; a short-wavelength blue, peaking at 455-464 nm; and a long-wavelength orange, peaking at 576-602 nm. Next, the behavioral response to light was investigated. We found that each species demonstrated positive phototactic responses to monochromatic stimuli across the UV-visible spectrum. In wavelength preference trials, distinct preferences among species were identified when different colored light stimuli were presented simultaneously. All species displayed a strong response to the UV stimulus, as well as responses to blue and orange stimuli, although at different response strengths, but no response to green. The results of this study demonstrate that larval stomatopods not only have multiple physiologically active spectral classes but they also display clear and distinct responses to wavelengths across the spectrum. We propose that the spectral classes demonstrated in each are related to visually guided ecological tasks of the larvae, which may differ between species.

In situ determination of the extreme damage resistance behavior in stomatopod dactyl club.

The structure and mechanical properties of the stomatopod dactyl club have been studied extensively for its extreme impact tolerance, but a systematic in situ investigation on the multiscale mechanical responses under high-speed impact has not been reported. Here the full dynamic deformation and crack evolution process within projectile-impacted dactyl using combined fast 2D X-ray imaging and high-resolution ex situ tomography are revealed. The results show that hydration states can lead to significantly different toughening mechanisms inside dactyl under dynamic loading. A previously unreported 3D interlocking structural design in the impact surface and impact region is reported using nano X-ray tomography. Experimental results and dynamic finite-element modeling suggest this unique structure plays an important role in resisting catastrophic structural damage and hindering crack propagation. This work is a contribution to understanding the key toughening strategies of biological materials and provides valuable information for biomimetic manufacturing of impact-resistant materials in general.

Ultraviolet vision in larval Neogonodactylus oerstedii.

Stomatopod crustaceans have among the most complex eyes in the animal kingdom, with up to 12 different color detection channels. The capabilities of these unique eyes include photoreception of ultraviolet (UV) wavelengths (<400 nm). UV vision has been well characterized in adult stomatopods but has not been previously demonstrated in the comparatively simpler larval eye. Larval stomatopod eyes are developmentally distinct from their adult counterpart and have been described as lacking the visual pigment diversity and morphological specializations found in adult eyes. However, recent studies have provided evidence that larval stomatopod eyes are more complex than previously thought and warrant closer investigation. Using electroretinogram recordings in live animals we found physiological evidence of blue- and UV-sensitive photoreceptors in larvae of the Caribbean stomatopod species Neogonodactylus oerstedii. Transcriptomes of individual larvae were used to identify the expression of three distinct UV opsin mRNA transcripts, which may indicate the presence of multiple UV spectral channels. This is the first paper to document UV vision in any larval stomatopod, expanding our understanding of the importance of UV sensitivity in plankton. Larval stomatopod eyes are more complex and more similar to adult eyes than expected, showing previously uncharacterized molecular diversity and physiological functions.

A diecast mineralization process forms the tough mantis shrimp dactyl club.

Biomineralization, the process by which mineralized tissues grow and harden via biogenic mineral deposition, is a relatively lengthy process in many mineral-producing organisms, resulting in challenges to study the growth and biomineralization of complex hard mineralized tissues. Arthropods are ideal model organisms to study biomineralization because they regularly molt their exoskeletons and grow new ones in a relatively fast timescale, providing opportunities to track mineralization of entire tissues. Here, we monitored the biomineralization of the mantis shrimp dactyl club-a model bioapatite-based mineralized structure with exceptional mechanical properties-immediately after ecdysis until the formation of the fully functional club and unveil an unusual development mechanism. A flexible membrane initially folded within the club cavity expands to form the new club's envelope. Mineralization proceeds inwards by mineral deposition from this membrane, which contains proteins regulating mineralization. Building a transcriptome of the club tissue and probing it with proteomic data, we identified and sequenced Club Mineralization Protein 1 (CMP-1), an abundant mildly phosphorylated protein from the flexible membrane suggested to be involved in calcium phosphate mineralization of the club, as indicated by in vitro studies using recombinant CMP-1. This work provides a comprehensive picture of the development of a complex hard tissue, from the secretion of its organic macromolecular template to the formation of the fully functional club.

Mantis shrimp identify an object by its shape rather than its color during visual recognition.

Mantis shrimp commonly inhabit seafloor environments with an abundance of visual features including conspecifics, predators, prey and landmarks used for navigation. Although these animals are capable of discriminating color and polarization, it is unknown what specific attributes of a visual object are important during recognition. Here, we show that mantis shrimp of the species Neogonodactylus oerstedii are able to learn the shape of a trained target. Further, when the shape and color of a target that they had been trained to identify were placed in conflict, N. oerstedii tended to choose the target of the trained shape over the target of the trained color. Thus, we conclude that the shape of the target was more salient than its color during recognition by N. oerstedii, suggesting that the shapes of objects, such as landmarks or other animals, are important for their identification by the species.

Landmark navigation in a mantis shrimp.

Mantis shrimp commonly occupy burrows in shallow, tropical waters. These habitats are often structurally complex where many potential landmarks are available. Mantis shrimp of the species Neogonodactylus oerstedii return to their burrows between foraging excursions using path integration, a vector-based navigational strategy that is prone to accumulated error. Here, we show that N. oerstedii can navigate using landmarks in parallel with their path integration system, correcting for positional uncertainty generated when navigating using solely path integration. We also report that when the path integration and landmark navigation systems are placed in conflict, N. oerstedii will orientate using either system or even switch systems enroute. How they make the decision to trust one navigational system over another is unclear. These findings add to our understanding of the refined navigational toolkit N. oerstedii relies upon to efficiently navigate back to its burrow, complementing its robust, yet error prone, path integration system with landmark guidance.

Neural processing of linearly and circularly polarized light signal in a mantis shrimp Haptosquilla pulchella.

Stomatopods, or mantis shrimp, are the only animal group known to possess circular polarization vision along with linear polarization vision. By using the rhabdomere of a distally located photoreceptor as a wave retarder, the eyes of mantis shrimp are able to convert circularly polarized light into linearly polarized light. As a result, their circular polarization vision is based on the linearly polarized light-sensitive photoreceptors commonly found in many arthropods. To investigate how linearly and circularly polarized light signals might be processed, we presented a dynamic polarized light stimulus while recording from photoreceptors or lamina neurons in intact mantis shrimp Haptosquilla pulchella The results indicate that all the circularly polarized light-sensitive photoreceptors also showed differential responses to the changing e-vector angle of linearly polarized light. When stimulated with linearly polarized light of varying e-vector angle, most photoreceptors produced a concordant sinusoidal response. In contrast, some lamina neurons doubled the response frequency in reacting to linearly polarized light. These responses resembled a rectified sum of two-channel linear polarization-sensitive photoreceptors, indicating that polarization visual signals are processed at or before the first optic lobe. Noticeably, within the lamina, there was one type of neuron that showed a steady depolarization response to all stimuli except right-handed circularly polarized light. Together, our findings suggest that, between the photoreceptors and lamina neurons, linearly and circularly polarized light may be processed in parallel and differently from one another.

Medium compensation in a spring-actuated system.

Mantis shrimp strikes are one of the fastest animal movements, despite their occurrence in a water medium with viscous drag. Since the strike is produced by a latch-mediated spring-actuated system and not directly driven by muscle action, we predicted that strikes performed in air would be faster than underwater as a result of reduction in the medium's drag. Using high-speed video analysis of stereotyped strikes elicited from Squilla mantis, we found the exact opposite: strikes are much slower and less powerful in air than in water. S. mantis strikes in air have a similar mass and performance to latch-mediated spring-actuated jumps in locusts, suggesting a potential threshold for the energetics of a 1-2 g limb rotating in air. Drag forces induced by the media may be a key feature in the evolution of mantis shrimp strikes and provide a potential target for probing the braking system of these extremely fast movements.

Path integration error and adaptable search behaviors in a mantis shrimp.

Mantis shrimp of the species Neogonodactylus oerstedii occupy small burrows in shallow waters throughout the Caribbean. These animals use path integration, a vector-based navigation strategy, to return to their homes while foraging. Here, we report that path integration in N. oerstedii is prone to error accumulated during outward foraging paths and we describe the search behavior that N. oerstedii employs after it fails to locate its home following the route provided by its path integrator. This search behavior forms continuously expanding, non-oriented loops that are centered near the point of search initiation. The radius of this search is scaled to the animal's positional uncertainty during path integration, improving the effectiveness of the search. The search behaviors exhibited by N. oerstedii bear a striking resemblance to search behaviors in other animals, offering potential avenues for the comparative examination of search behaviors and how they are optimized in disparate taxa.

Gaze stabilization in mantis shrimp in response to angled stimuli.

Gaze stabilization is a fundamental aspect of vision and almost all animals shift their eyes to compensate for any self-movement relative to the external environment. When it comes to mantis shrimp, however, the situation becomes complicated due to the complexity of their visual system and their range of eye movements. The stalked eyes of mantis shrimp can independently move left and right, and up and down, whilst simultaneously rotating about the axis of the eye stalks. Despite the large range of rotational freedom, mantis shrimp nevertheless show a stereotypical gaze stabilization response to horizontal motion of a wide-field, high-contrast stimulus. This response is often accompanied by pitch (up-down) and torsion (about the eye stalk) rotations which, surprisingly, have no effect on the performance of yaw (side-to-side) gaze stabilization. This unusual feature of mantis shrimp vision suggests that their neural circuitry for detecting motion is radially symmetric and immune to the confounding effects of torsional self-motion. In this work, we reinforce this finding, demonstrating that the yaw gaze stabilization response of the mantis shrimp is robust to the ambiguous motion cues arising from the motion of striped visual gratings in which the angle of a grating is offset from its direction of travel.

Context-dependent scaling of kinematics and energetics during contests and feeding in mantis shrimp.

Measurements of energy use, and its scaling with size, are critical to understanding how organisms accomplish myriad tasks. For example, energy budgets are central to game theory models of assessment during contests and underlie patterns of feeding behavior. Clear tests connecting energy to behavioral theory require measurements of the energy use of single individuals for particular behaviors. Many species of mantis shrimp (Stomatopoda: Crustacea) use elastic energy storage to power high-speed strikes that they deliver to opponents during territorial contests and to hard-shelled prey while feeding. We compared the scaling of strike kinematics and energetics between feeding and contests in the mantis shrimp Neogonodactylus bredini We filmed strikes with high-speed video, measured strike velocity and used a mathematical model to calculate strike energy. During contests, strike velocity did not scale with body size but strike energy scaled positively with size. Conversely, while feeding, strike velocity decreased with increasing size and strike energy did not vary according to body size. Individuals most likely achieved this strike variation through differential compression of their exoskeletal spring prior to the strike. Post hoc analyses found that N. bredini used greater velocity and energy when striking larger opponents, yet variation in prey size was not accompanied by varying strike velocity or energetics. Our estimates of energetics inform prior tests of contest and feeding behavior in this species. More broadly, our findings elucidate the role behavioral context plays in measurements of animal performance.

Complex gaze stabilization in mantis shrimp.

Almost all animals, regardless of the anatomy of the eyes, require some level of gaze stabilization in order to see the world clearly and without blur. For the mantis shrimp, achieving gaze stabilization is unusually challenging as their eyes have an unprecedented scope for movement in all three rotational degrees of freedom: yaw, pitch and torsion. We demonstrate that the species Odontodactylus scyllarus performs stereotypical gaze stabilization in the yaw degree of rotational freedom, which is accompanied by simultaneous changes in the pitch and torsion rotation of the eye. Surprisingly, yaw gaze stabilization performance is unaffected by both the torsional pose and the rate of torsional rotation of the eye. Further to this, we show, for the first time, a lack of a torsional gaze stabilization response in the stomatopod visual system. In the light of these findings, we suggest that the neural wide-field motion detection network in the stomatopod visual system may follow a radially symmetric organization to compensate for the potentially disorientating effects of torsional eye movements, a system likely to be unique to stomatopods.

Smashing mantis shrimp strategically impact shells.

Many predators fracture strong mollusk shells, requiring specialized weaponry and behaviors. The current shell fracture paradigm is based on jaw- and claw-based predators that slowly apply forces (high impulse, low peak force). However, predators also strike shells with transient intense impacts (low impulse, high peak force). Toward the goal of incorporating impact fracture strategies into the prevailing paradigm, we measured how mantis shrimp (Neogonodactylus bredini) impact snail shells, tested whether they strike shells in different locations depending on prey shape (Nerita spp., Cenchritis muricatus, Cerithium spp.) and deployed a physical model (Ninjabot) to test the effectiveness of strike locations. We found that, contrary to their formidable reputation, mantis shrimp struck shells tens to hundreds of times while targeting distinct shell locations. They consistently struck the aperture of globular shells and changed from the aperture to the apex of high-spired shells. Ninjabot tests revealed that mantis shrimp avoid strike locations that cause little damage and that reaching the threshold for eating soft tissue is increasingly difficult as fracture progresses. Their ballistic strategy requires feed-forward control, relying on extensive pre-strike set-up, unlike jaw- and claw-based strategies that can use real-time neural feedback when crushing. However, alongside this pre-processing cost to impact fracture comes the ability to circumvent gape limits and thus process larger prey. In sum, mantis shrimp target specific shell regions, alter their strategy depending on shell shape, and present a model system for studying the physics and materials of impact fracture in the context of the rich evolutionary history of predator-prey interactions.

Circularly polarized light detection in stomatopod crustaceans: a comparison of photoreceptors and possible function in six species.

A combination of behavioural and electrophysiological experiments have previously shown that two species of stomatopod, Odontodactylus scyllarus and Gonodactylaceus falcatus, can differentiate between left- and right-handed circularly polarized light (CPL), and between CPL and linearly polarized light (LPL). It remains unknown if these visual abilities are common across all stomatopod species, and if so, how circular polarization sensitivity may vary between and within species. A subsection of the midband, a specialized region of stomatopod eyes, contains distally placed photoreceptor cells, termed R8 (retinular cell number 8). These cells are specifically built with unidirectional microvilli and appear to be angled precisely to convert CPL into LPL. They are mostly quarter-wave retarders for human visible light (400-700 nm), as well as being ultraviolet-sensitive linear polarization detectors. The effectiveness of the R8 cells in this role is determined by their geometric and optical properties. In particular, the length and birefringence of the R8 cells are crucial for retardation efficiency. Here, our comparative studies show that most species investigated have the theoretical ability to convert CPL into LPL, such that the handedness of an incoming circular reflection or signal could be discriminated. One species, Haptosquilla trispinosa, shows less than quarter-wave retardance. Whilst some species are known to produce circularly polarized reflections (some Odontodactylus species and G. falcatus, for example), others do not, so a variety of functions for this ability are worth considering.

Spectral absorption of visual pigments in stomatopod larval photoreceptors.

Larval stomatopod eyes appear to be much simpler versions of adult compound eyes, lacking most of the visual pigment diversity and photoreceptor specializations. Our understanding of the visual pigment diversity of larval stomatopods, however, is based on four species, which severely limits our understanding of stomatopod eye ontogeny. To investigate several poorly understood aspects of stomatopod larval eye function, we tested two hypotheses surrounding the spectral absorption of larval visual pigments. First, we examined a broad range of species to determine if stomatopod larvae generally express a single, spectral class of photoreceptor. Using microspectrophotometry (MSP) on larvae captured in the field, we found data which further support this long-standing hypothesis. MSP was also used to test whether larval species from the same geographical region express visual pigments with similar absorption spectra. Interestingly, despite occupation of the same geographical location, we did not find evidence to support our second hypothesis. Rather, there was significant variation in visual pigment absorption spectra among sympatric species. These data are important to further our understanding of larval photoreceptor spectral diversity, which is beneficial to ongoing investigations into the ontogeny, physiology, and molecular evolution of stomatopod eyes.

Hiding opaque eyes in transparent organisms: a potential role for larval eyeshine in stomatopod crustaceans.

Opaque screening pigments are a fundamental requisite for preserving resolution in image-forming eyes. Possession of any type of image-forming eye in a transparent, pelagic animal will thus undermine the ability of that animal to be invisible in the water column. Transparent, pelagic animals must therefore deal with the trade-off between the ability to see and the ability of other animals to see them. Stomatopod larvae, like many transparent crustaceans, possess specialized optics in their compound eyes that minimize the volume of the opaque retina. Though the volumes of these retinas are reduced, their opacity remains conspicuous to an observer. The light reflected from structures overlying the retinas of stomatopod crustacean larval eyes, referred to here as eyeshine, is hypothesized to further reduce the visibility of opaque retinas. Blue or green wavelengths of light are most strongly reflected in stomatopod larval eyeshine, suggesting a putative spectral matching to the light environment against which the larval eyes are viewed. We tested the efficacy of stomatopod crustacean larval eyeshine as an ocular camouflaging mechanism by photographing larvae in their natural light environment and analysing the contrast of eyes with the background light. To test for spectral matching between stomatopod larval eyeshine and the background light environment, we characterized the spectrum of eyeshine and calculated its performance using radiometric measurements collected at the time of each photographic series. These results are the first to demonstrate an operative mirror camouflage matched in both spectrum and radiance to the pelagic background light environment.

Out of the blue: the evolution of horizontally polarized signals in Haptosquilla (Crustacea, Stomatopoda, Protosquillidae).

The polarization of light provides information that is used by many animals for a number of different visually guided behaviours. Several marine species, such as stomatopod crustaceans and cephalopod molluscs, communicate using visual signals that contain polarized information, content that is often part of a more complex multi-dimensional visual signal. In this work, we investigate the evolution of polarized signals in species of Haptosquilla, a widespread genus of stomatopod, as well as related protosquillids. We present evidence for a pre-existing bias towards horizontally polarized signal content and demonstrate that the properties of the polarization vision system in these animals increase the signal-to-noise ratio of the signal. Combining these results with the increase in efficacy that polarization provides over intensity and hue in a shallow marine environment, we propose a joint framework for the evolution of the polarized form of these complex signals based on both efficacy-driven (proximate) and content-driven (ultimate) selection pressures.

Investigation of the ultrastructures and retinal arrangements of larval stomatopod eyes.

Though the transparent apposition eyes of larval stomatopod crustaceans lack most of the unique retinal specializations known from their adult counterparts, increasing evidence suggests that these tiny pelagic organisms possess their own version of retinal complexity. In this paper, we examined the structural organization of larval eyes in six species of stomatopod crustaceans across three stomatopod superfamilies using transmission electron microscopy. The primary focus was to examine retinular cell arrangement of the larval eyes and characterize the presence of an eighth retinular cell (R8), which is typically responsible for UV vision in crustaceans. For all species investigated, we identified R8 photoreceptor cells positioned distal to the main rhabdom of R1-7 cells. This is the first evidence that R8 photoreceptor cells exist in larval stomatopod retinas, and among the first identified in any larval crustacean. Considering recent studies that identified UV sensitivity in larval stomatopods, we propose that this sensitivity is driven by this putative R8 photoreceptor cell. Additionally, we identified a potentially unique crystalline cone structure in each of the species examined, the function of which is still not understood.

Colour vision in stomatopod crustaceans.

The stomatopod crustaceans, or mantis shrimps, are colourful marine invertebrate predators. Their unusual compound eyes have dorsal and ventral regions resembling typical crustacean apposition designs separated by a unique region called the midband that consists of from two to six parallel rows of ommatidia. In species with six-row midbands, the dorsal four rows are themselves uniquely specialized for colour analysis. Rhabdoms of ommatidia in these rows are longitudinally divided into three distinct regions: an apical ultraviolet (UV) receptor, a shorter-wavelength middle tier receptor and a longer-wavelength proximal tier receptor. Each of the total of 12 photoreceptors has a different spectral sensitivity, potentially contributing to a colour-vision system with 12 channels. Mantis shrimps can discriminate both human-visible and UV colours, but with limited precision compared to other colour-vision systems. Here, we review the structure and function of stomatopod colour vision, examining the types of receptors present in a species, the spectral tuning of photoreceptors both within and across species, the neural analysis of colour and the genetics underlying the multiple visual pigments used for colour vision. Even today, after many decades of research into the colour vision of stomatopods, much of its operation and its use in nature remain a mystery. This article is part of the theme issue 'Understanding colour vision: molecular, physiological, neuronal and behavioural studies in arthropods'.