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Through their unique use of sophisticated laryngeal echolocation bats are considered sensory specialists amongst mammals and represent an excellent model in which to explore sensory perception. Although several studies have shown that the evolution of vision is linked to ecological niche adaptation in other mammalian lineages, this has not yet been fully explored in bats. Recent molecular analysis of the opsin genes, which encode the photosensitive pigments underpinning color vision, have implicated high-duty cycle (HDC) echolocation and the adoption of cave roosting habits in the degeneration of color vision in bats. However, insufficient sampling of relevant taxa has hindered definitive testing of these hypotheses. To address this, novel sequence data was generated for the SWS1 and MWS/LWS opsin genes and combined with existing data to comprehensively sample species representing diverse echolocation types and niches (SWS1 n = 115; MWS/LWS n = 45). A combination of phylogenetic analysis, ancestral state reconstruction, and selective pressure analyses were used to reconstruct the evolution of these visual pigments in bats and revealed that although both genes are evolving under purifying selection in bats, MWS/LWS is highly conserved but SWS1 is highly variable. Spectral tuning analyses revealed that MWS/LWS opsin is tuned to a long wavelength, 555-560 nm in the bat ancestor and the majority of extant taxa. The presence of UV vision in bats is supported by our spectral tuning analysis, but phylogenetic analyses demonstrated that the SWS1 opsin gene has undergone pseudogenization in several lineages. We do not find support for a link between the evolution of HDC echolocation and the pseudogenization of the SWS1 gene in bats, instead we show the SWS1 opsin is functional in the HDC echolocator, Pteronotus parnellii. Pseudogenization of the SWS1 is correlated with cave roosting habits in the majority of pteropodid species. Together these results demonstrate that the loss of UV vision in bats is more widespread than was previously considered and further elucidate the role of ecological niche specialization in the evolution of vision in bats.

The long history and great ecological and morphological diversity of reptiles (all amniotes except mammals and birds) is matched by their visual system diversity. Although less known than in other amniotes, visual pigments have been studied in all extant reptile orders except Sphenodontia. There have been no additions to the five visual pigments present in the ancestral vertebrate, although there have been multiple independent losses. Crocodylians retain three visual pigments, many lizards as well as Testudines four or five and snakes one to three. Adaptive pigment evolution includes tuning site amino acid substitutions and switches between chromophore types that together generate ultraviolet to infrared spectral sensitivity. Reptiles present some of the best evidences of evolutionary rod–cone and cone–rod transmutation with, for example typically cone visual pigments expressed in rod-like photoreceptors. Reptile visual pigments show evidence of substantial adaptive evolution, at least some of which is associated with major ecological shifts. Key Concepts Reptiles have no more than five visual pigments, as few as one and typically at least three. Up to four of the ancestral vertebrate visual pigments have been lost independently in different reptile lineages, and no visual opsin gene duplications have been identified so far. Testudines have between four and five visual pigments and a wide range of photoreceptor oil droplets, representing one of the most complex visual pigment systems in tetrapod vertebrates. Despite many of them being nocturnal, no rhodopsin 1 has been detected in geckos. No sws1 or rh2 opsin genes can be detected in genomic sequence data for crocodylians. Among squamates, some lizards have a mixture of A1 and A2 chromophores incorporated in their visual pigments, allowing a wide colour sensitivity, including infrared vision. In the green anole (Anolis carolinensis), only cone opsins have been reported (SWS1, SWS2, RH2 and LWS) in studies of the eye. However, a 515-nm tuned rh1 rhodopsin gene occurs in the genome of this species. Extant snakes have lost two visual pigments (SWS2 and RH2), likely as an adaptation to a low light environment inhabited by a snake ancestor. Extreme burrowing in scolecophidians (blind, worm and thread snakes) is correlated with the loss of the SWS1 and LWS visual pigments and cones, rendering these snakes rod (RH1) monochromats. The expression of RH1, SWS1 and LWS pigments in snake lineages with seemingly all-cone and all-rod retinae suggests multiple transmutations from cone to rod and vice versa in snakes. Similar transmutations likely occurred in some lizards and perhaps crocodylians. Visual pigment spectral sensitivity in reptiles has been found to correlate with the light transmission properties of the ocular media (e.g. snakes) and photoreceptor oil droplets (e.g. testudines), that is the pigments are not sensitive to wavelengths of light filtered out by these structures. Keywords: cones; photopic vision; rods; scotopic vision; oil droplets; opsins; crocodylians; Testudines; lizards; snakes

Much of what is known about the molecular evolution of vertebrate vision comes from studies of mammals, birds and fish. Reptiles (especially snakes) have barely been sampled in previous studies despite their exceptional diversity of retinal photoreceptor complements. Here, we analyze opsin gene sequences and ocular media transmission for up to 69 species to investigate snake visual evolution. Most snakes express three visual opsin genes (rh1, sws1, and lws). These opsin genes (especially rh1 and sws1) have undergone much evolutionary change, including modifications of amino acid residues at sites of known importance for spectral tuning, with several tuning site combinations unknown elsewhere among verte- brates. These changes are particularly common among dipsadine and colubrine “higher” snakes. All three opsin genes are inferred to be under purifying selection, though dN/dS varies with respect to some lineages, ecologies, and retinal anatomy. Positive selection was inferred at multiple sites in all three opsins, these being concentrated in transmembrane domains and thus likely to have a substantial effect on spectral tuning and other aspects of opsin function. Snake lenses vary substantially in their spectral transmission. Snakes active at night and some of those active by day have very transmissive lenses, whereas some primarily diurnal species cut out shorter wavelengths (including UVA). In terms of retinal anatomy, lens transmission, visual pigment spectral tuning and opsin gene evolution the visual system of snakes is exceptionally diverse compared with all other extant tetrapod orders.

The dominant hypothesis for the evolutionary origin of snakes from ‘lizards’ (non-snake squamates) is that stem snakes acquired many snake features while passing through a profound burrowing (fossorial) phase. To investigate this, we examined the visual pigments and their encoding opsin genes in a range of squamate reptiles, focusing on fossorial lizards and snakes. We sequenced opsin transcripts isolated from retinal cDNA and used microspectrophotometry to measure directly the spectral absorbance of the photoreceptor visual pigments in a subset of samples. In snakes, but not lizards, dedicated fossoriality (as in Scolecophidia and the alethinophidian Anilius scytale) corresponds with loss of all visual opsins other than RH1 (kmax 490– 497 nm); all other snakes (including less dedicated burrowers) also have functional sws1 and lws opsin genes. In contrast, the retinas of all lizards sampled, even highly fossorial amphisbaenians with reduced eyes, express functional lws, sws1, sws2 and rh1 genes, and most also express rh2 (i.e. they express all five of the visual opsin genes present in the ancestral vertebrate). Our evidence of visual pigment complements suggests that the visual system of stem snakes was partly reduced, with two (RH2 and SWS2) of the ancestral vertebrate visual pigments being eliminated, but that this did not extend to the extreme additional loss of SWS1 and LWS that subsequently occurred (probably independently) in highly fossorial extant scolecophidians and A. scytale. We therefore consider it unlikely that the ancestral snake was as fossorial as extant scolecophidians, whether or not the latter are para- or monophyletic.

In 1934, Gordon Walls forwarded his radical theory of retinal photoreceptor ‘transmutation’. This proposed that rods and cones used for scotopic and photopic vision, respectively, were not fixed but could evolve into each other via a series of morphologically distinguishable intermediates. Walls' prime evidence came from series of diurnal and nocturnal geckos and snakes that appeared to have pure-cone or pure-rod retinas (in forms that Walls believed evolved from ancestors with the reverse complement) or which possessed intermediate photoreceptor cells. Walls was limited in testing his theory because the precise identity of visual pigments present in photoreceptors was then unknown. Subsequent molecular research has hitherto neglected this topic but presents new opportunities. We identify three visual opsin genes,rh1,sws1andlws, in retinal mRNA of an ecologically and taxonomically diverse sample of snakes central to Walls' theory. We conclude that photoreceptors with superficially rod- or cone-like morphology are not limited to containing scotopic or photopic opsins, respectively. Walls' theory is essentially correct, and more research is needed to identify the patterns, processes and functional implications of transmutation. Future research will help to clarify the fundamental properties and physiology of photoreceptors adapted to function in different light levels.

We examined lens and brille transmittance, photoreceptors, visual pigments, and visual opsin gene sequences of viperid snakes with and without infrared-sensing pit organs. Ocular media transmittance was high in both groups. Contrary to previous reports, both small and large single cones occur in pit vipers. Non-pit vipers differ from pit vipers in having a two-tiered retina, but few taxa have been examined for this poorly understood feature. All vipers sampled express rh1, sws1 and lws visual opsin genes. Opsin spectral tuning varies but not in accordance with the presence/absence of pit organs, and not always as predicted from gene sequences. The visual opsin genes were generally under purifying selection, with positive selection at spectral tuning amino acids in RH1 and SWS1 opsins, and at retinal pocket stabilization sites in RH1 or LWS (and without substantial differences between pit and nonpit vipers). A lack of evidence for a sensory trade-off between viperid eyes (in the aspects examined) and pit organs might be explained by the high degree of neural integration of vision and infrared detection; the latter represents an elaboration of an existing sense with addition of a novel sense organ, rather than involving the evolution of a wholly novel sensory system.

Dermal phototaxis has been reported in a few aquatic vertebrate lineages spanning fish, amphibians and reptiles. These taxa respond to light on the skin of their elongate hind-bodies and tails by withdrawing under cover to avoid detection by predators. Here, we investigated tail phototaxis in sea snakes (Hydrophiinae), the only reptiles reported to exhibit this sensory behaviour. We conducted behavioural tests in 17 wild-caught sea snakes of eight species by illuminating the dorsal surface of the tail and midbody skin using cold white, violet, blue, green and red light. Our results confirmed phototactic tail withdrawal in the previously studied Aipysurus laevis, revealed this trait for the first time in A. duboisii and A. tenuis, and suggested that tail photoreceptors have peak spectral sensitivities between blue and green light (457–514 nm). Based on these results, and an absence of photoresponses in five Aipysurus and Hydrophis species, we tentatively infer that tail phototaxis evolved in the ancestor of a clade of six Aipysurus species (comprising 10% of all sea snakes). Quantifying tail damage, we found that the probability of sustaining tail injuries was not influenced by tail phototactic ability in snakes. Gene profiling showed that transcriptomes of both tail skin and body skin lacked visual opsins but contained melanopsin (opn4x) in addition to key genes of the retinal regeneration and phototransduction cascades. This work suggests that a nonvisual photoreceptor (e.g., Gq rhabdomeric) signalling pathway underlies tail phototaxis, and provides candidate gene targets for future studies of this unusual sensory innovation in reptiles.