Herpetology Notes, volume 13: 443-447 (2020) (published online on 30 May 2020)
New accounts of biofluorescence in several anuran genera (Hylidae, Microhylidae, Ranidae, Leptodactylidae) with comments on intraspecific variation
Courtney Whitcher1,*
Several species of tropical frogs separated by a Materials and Methods geographic distance of over 18,000 km were recently Frogs were collected between 19 May 2019 and 30 found to biofluoresce (Taboada et al., 2017a,b; May 2019 on night surveys at the Belize Foundation Deschepper et al., 2018; Gray, 2019; Saporito, 2019), for Research and Environmental Education (BFREE) including the hylids Boana punctata (Schneider, in the Toledo region of Belize (ca. 16.5558°N, 1799), B. atlantica (Caramaschi and Velosa, 88.7077°W). Fluorescence was observed by shining 1996), and B. rufitela (Fouquette, 1961), and the UV (360–380 nm) and Violet (400–415 nm) excitation rhacophorid Philautus macroscelis (Boulenger, 1896). wavelengths on living specimens using a NIGHTSEA® Biofluorescence, the ability to absorb light and reemit it Dual Fluorescent Protein Flashlight (NDT, Lexington, at a greater wavelength, has been examined in a range Massachusetts, USA). Filter glasses that corresponded of organisms, including insects, plants, fish, reptiles, to each excitation (emission 415 nm longpass2 and 460 birds, and mammals (Sloggett, 2018; Cummings et al., nm longpass, respectively) were used to distinguish 2018; Kohler et al., 2019; Wilkinson et al., 2019). Until between reflectance and biofluorescence. All photos recently this phenomenon had never been reported in were taken through the 460 nm longpass filter, focusing amphibians. The three anurans originally found to be on the frog with the Violet light positioned above it. biofluorescent presented fluorescence across all of Photos were taken indoors during daylight hours with their skin. It was not until Gray (2019) found lateral regular ceiling lights on, and all frogs were released the patterning that the prospect of intraspecific pattern subsequent evening. variation of fluorescent skin could be considered. Here I document biofluorescence in six additional frog Results and Discussion species and from three additional families, including the hylids Bromeliohyla bromeliacia (Schmidt, 1933), Green fluorescent flanks and/or inguinal spots were Dendropsophus microcephalus (Cope, 1886), and found in Smilisca baudinii (Fig. 1D), Lithobates juliani Smilisca baudinii (Duméril and Bibron, 1841), the (Fig. 2D), and Bromeliohyla bromeliacia (Fig. 1F). The microhylid Gastrophryne elegans (Boulenger, 1882), extent and variation of the fluorescence in these species the ranid Lithobates juliani (Hillis and de Sá, 1988), was variable. Fluorescent arm bands were observed and the leptodactylid Leptodactylus fragilis (Brocchi, in Gastrophryne elegans (Fig. 1B). Dendropsophus 1877). I also examine intraspecific variation in three of microcephalus presented weak fluorescence all over these species. in some specimens (Fig. 2B), and no fluorescence in others. Finally, faint fluorescent patches were found
2 A longpass filter is a glass filter that interferes with some 1 Department of Biological Science, Florida State University, wavelengths and transmits others across the target spectrum 319 Stadium Drive, Tallahassee, Florida 32306, USA. (ultraviolet, visible, or infrared). Thus, a 415 nm longpass * Corresponding author. E-mail: [email protected] filter selectively passes wavelengths between 410–420 nm. 444 Courtney Whitcher
Figure 1. Natural light (left) and fluorescent (right) photos of Gastrophryne elegans (top), Smilisca baudinii (middle), and Bromeliohyla bromeliacia (bottom). Photos by the author.
on the venter of Leptodactylus fragilis (Fig. 2F). All Gastrophryne elegans, Smilisca baudinii, and fluorescence observed in these species was greatest Bromeliohyla bromeliacia had the most pronounced when excited with the Violet 400–415 nm light. fluorescence (Fig. 1). It is likely that peak excitation New accounts of biofluorescence in several anuran genera 445
Figure 2. Natural light (left) and fluorescent (right) photos of Dendropsophus microcephalus (top), Lithobates juliani (middle), and Leptodactylus fragilis (bottom). Photos by author.
for these species’ fluorescence is in the range of 400– fluoresced when excited at 365 nm, and Philautus 415 nm. Boana punctata and B. atlantica both have macroscelis fluoresced with excitation of 395 nm an excitation maximum of 390–430 nm, B. rufitela (Taboada et al., 2017a,b; Deschepper et al., 2018; Gray, 446 Courtney Whitcher
2019). The excitation maximum for these three species patterns and UV sensitive optical ranges (Yovanovich is most similar to that of B. punctata and B. atlantica. et al., 2019), a key question is what role biofluorescence A comparison of the fluorescence mechanisms across may be playing in anuran communication. Specifically, species could provide great insight into the evolution fluorescence may be functionally significant in mate of biofluorescence in anurans. Very weak fluorescence choice or predation on anurans. To ascertain a function was found in Dendropsophus microcephalus, Lithobates of fluorescence, certain criteria must be met: (1) juliani, and Leptodactylus fragilis (Fig. 2). It is likely organisms must fluoresce; (2) potential recipients of a that the peak excitation for the fluorescence of these fluorescent signal must be able to perceive fluorescence; species is greater than 415 nm as it was only seen when (3) fluorescence must make a meaningful contribution excited by 400–415 nm light and not when excited by in the visual signalling process; and (4) a behavioural 360–380 nm light. These species should be tested for response must be related to the fluorescent signal. The fluorescence with a longer wavelength light. biofluorescent signal of Boana punctata was found to I also documented intraspecific variation in five make a meaningful contribution to the visual signalling specimens of Smilisca baudinii (Fig. 3). The number in that species, contributing nearly 30% of the light and size of inguinal spots that fluoresced, and the extent emanating from the frog (i.e., reflected light 70%, to which this fluorescence covered the flanks, was fluorescence 30%) under twilight conditions (Taboada variable. et al., 2017a). This contribution, coupled with the UV With an increased number of species, across a range sensitivity of many anuran optical ranges (Yovanovich of localities and families presenting both fluorescent et al., 2019) gives reason to test the broader significance
Figure 3. Intraspecific variation in fluorescence of Smilisca baudinii. Each photo is a different individual. All photos were taken with the same camera, filter, and excitation light. Photos by the author. New accounts of biofluorescence in several anuran genera 447 of biofluorescence in frogs. Frog eyes have both blue- References sensitive rods with a maximum absorbance sensitivity Amézquita, A., Hödl, W. (2004): How, when, and where to at approximately 430 nm and green-sensitive rods with perform visual displays: the case of the Amazonian frog Hyla a maximum absorbance sensitivity at approximately parviceps. Herpetologica 60(4): 420–429. 500 nm (Yovanovich et al., 2017). As fluorescence Cummings, M.E., Endler, J.A. (2018): 25 Years of sensory drive: reemits light at a longer wavelength than absorbed, the evidence and its watery bias. Current Zoology 64(4): 471– the fluorescence present in many of the anurans tested 484. shifts the wavelength of the light emitted closer to Deschepper, P., Jonckheere, B., Matthys, J. (2018): A light in the dark: the discovery of another fluorescent frog in the Costa the maximum sensitivity of the green-sensitive rods, Rican rainforests. Wilderness & Environmental Medicine 29(3): of which there are more in the retina, or to the range 421–422. of wavelengths in which both rods have sensitivity, Gray, R.J. (2019): Biofluorescent lateral patterning on the likely increasing the amount of optical stimulus for mossy bushfrog (Philautus macroscelis): the first report of the receiver. Additionally, as foot flagging has been biofluorescence in a rhacophorid frog. Herpetology Notes 12: documented in other hylids (Amézquita and Hödl, 2004), 363–364. the fluorescence on the inner thigh of Smilisca baudinii Kohler, A.M., Olson, E.R., Martin, J.G., Anich, P.S. (2019): Ultraviolet fluorescence discovered in New World flying and Bromeliohyla bromeliacia could be being presented squirrels (Glaucomys). Journal of Mammalogy 100(1): 21–30. during this behaviour. As Gray (2019) suggested, these Saporito, R. (2019): A field-based survey of fluorescence in non-acoustic signals could be used to attract a mate tropical tree frogs using an LED UV-B flashlight. Herpetology without increasing detectability by a predator. Notes 12: 987–990. Thompson et al. (2019) suggested that biofluorescence Sloggett, J.J. (2018): Field observations of putative bone-based in anurans was not widespread in nature. My observations fluorescence in a gecko. Current Zoology 64(3): 319–320. and those in the recent literature indicate that the contrary Taboada, C., Brunetti, A.E., Pedron, F.N., Carnevale Neto, F., Estrin, D.A., Bari, S.E., Faivovich, J. (2017a): Naturally may be true, and they highlight the importance of testing occurring fluorescence in frogs. Proceedings of the National additional frogs with a range of excitation wavelengths. Academy of Sciences 114(14): 3672–3677. I suggest that species tested with 385 nm excitation Taboada, C., Brunetti, A.E., Alexandre, C., Lagorio, M.G., should also be tested with longer excitation wavelengths, Faivovich, J. (2017b): Fluorescent frogs: a herpetological as I found fluorescence with 400–415 nm excitation and perspective. South American Journal of Herpetology 12(1): the likelihood of fluorescence at even longer excitation 1–13. wavelengths. As acoustic factors have been the focus Thompson, M.E., Saporito, R.A., Ruiz-Valderrama, D.H., Medina- Rangel, G.F., Donnelly, M.A. (2019): A field-based survey of of anuran communication research, understanding fluorescence in tropical tree frogs using an LED UV-B flashlight. biofluorescence could create a new paradigm for how Herpetology Notes 12: 987–990. we think about anuran sexual selection. The discovery Wilkinson, B.P., Johns, M.E., Warzybok, P. (2019): Fluorescent of biofluorescence, and its apparent widespread nature, ornamentation in the Rhinoceros Auklet Cerorhinca in these amphibians provides a glimpse into an aspect monocerata. Ibis 161(3): 694–698. of frog ecology that has never been properly studied. Yovanovich, C.A.M., Koskela, S.M., Nevala, N., Kondrashev, Further exploratory work will provide great insight S.L., Kelber, A., Donner, K. (2017): The dual rod system of into amphibian optics and communication, despite the amphibians supports colour discrimination at the absolute visual threshold. Philosophical Transactions of the Royal Society B outcome of functionality. 372: 20160066. Yovanovich, C.A.M., Pierotti, M.E.R., Kelber, A., Jorgewich- Acknowledgements. I would like to thank the Rabosky Lab at the Cohen, G., Ibáñez, R., Grant, T. (2020). Lens transmittance University of Michigan for support in the field surveys that aided shapes ultraviolet sensitivity in the eyes of frogs from diverse in the collection of these frogs. In addition, these data could not ecological and phylogenetic backgrounds. Proceedings of the have been collected without the facilities provided by the Belize Royal Society 287: 20192253. Foundation for Research and Environmental Education (BFREE) and its welcoming staff. Data collection was done under Belize Forest Department Scientific Collection/Research Permit Ref. No. FD/WL/1/19 (19) and tissue samples collected from individual specimens are accessioned in the University of Michigan Museum of Zoology Research Collection (Field Numbers IAH 1000–IAH 1250). Costs of travel and lodging were defrayed in part by an Anna Olcott Smith Award from the University of Michigan Rackham Accepted by Hinrich Kaiser Graduate School.