Phyllomedusa 18(1):13–26, 2019 © 2019 Universidade de São Paulo - ESALQ ISSN 1519-1397 (print) / ISSN 2316-9079 (online) doi: http://dx.doi.org/10.11606/issn.2316-9079.v18i1p13-26 Stable individual variation in ventral spotting patterns in Phyllomedusa trinitatis (Anura: Phyllomedusidae) and other Phyllomedusa species: a minimally invasive method for recognizing individuals Joanna M. Smith,1 Adam Bland,2 Eleanor Gourevitch,3 Paul A. Hoskisson,4 and J. Roger Downie3 1 School of Biological Sciences, University of Bangor. Bangor, Gwynedd, LL57 2DG, UK. E-mail: [email protected]. 2 Chester Zoo. Moston Road, Upton-by Chester, Upton, Chester CH2 1EU, UK. E-mail: [email protected]. 3 School of Life Sciences, Graham Kerr Building, University of Glasgow. Glasgow G12 8QQ, UK. E-mail: roger.downie@ glasgow.ac.uk. 4 Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde. 161 Cathedral Street, Glasgow G4 0RE, UK. Abstract Stable individual variation in ventral spotting patterns in Phyllomedusa trinitatis (Anura: Phyllomedusidae) and other Phyllomedusa species: a minimally invasive method for recognizing individuals. We describe a distinctive pattern of individually variable white ‘islands’ and dots on the ventral surfaces of the throat and forearm regions of male and female Phyllomedusa trinitatis. Crucially, we report from captive-reared individuals that these patterns are ontogenetically stable. We have used these patterns to recognize individual frogs in populations of 60 and more. Examination of the ventral surfaces of other Phyllomedusa species in museum specimens and from published accounts suggests that use of these patterns provides a generally useful, minimally invasive recognition method in this genus. We fnd that species previously classed as Phyllomedusa but now regarded as belonging to different genera lack these patterns. Keywords: individual recognition, Trinidad Leaf Frogs, ventral patterns. Resumo Variação individual estável no padrão de manchas ventrais em Phyllomedusa trinitatis (Anura: Phyllomedusidae) e em outras espécies de Phyllomedusa: um método minimamente invasivo para o reconhecimento de indivíduos. Descrevemos aqui um padrão distintivo de “ilhas” e pontos brancos individualmente variáveis nas superfícies ventrais das regiões da garganta e do antebraço de machos e fêmeas de Phyllomedusa trinitatis. Crucialmente, a partir de indivíduos criados em cativeiro relatamos que esses padrões são ontogeneticamente estáveis. Usamos esses Received 26 November 2018 Accepted 14 May 2019 Distributed June 2019 Phyllomedusa - 18(1), June 2019 13 Smith et al. padrões para reconhecer pererecas individuais em populações de 60 ou mais indivíduos. O exame das superfícies ventrais de outras espécies de Phyllomedusa em espécimes de museus e de relatos publicados sugere que o uso desses padrões fornece um método de reconhecimento minimamente invasivo e geralmente útil nesse gênero. Descobrimos que espécies anteriormente classifcadas como Phyllomedusa, mas agora consideradas pertencentes a diferentes gêneros, não possuem esses padrões. Palavras-chave: padrões ventrais, pererecas-folhas-de-trinidad, reconhecimento individual. Introduction mark-recapture studies fall drastically with each toe clipped (McCarthy and Parris 2004). Toe- Many feld studies in animal ecology, such as clipping seems especially problematic for frogs population estimations via mark-recapture with adhesive toe-pads since these cannot be methods, life history analysis and the unders- regenerated and are critical to the animals’ way tanding of interactions in social behavior require of life. Following discussions of the ethical the reliable long-term recognition of individuals. justifability of toe-clipping in the light of such In many amphibian species, this is made diffcult fndings (May 2004, Parris et al. 2010), by uniform markings that generally vary little herpetologists are keen to fnd viable alternatives between individuals and because most have that provide similar rates of recognition and moist, sheddable and permeable skin that takes return as toe-clipping (BHS 2017). dye marks poorly. Ferner (2010), when reviewing Ferner (2010) also recommended the available methods for marking amphibians, consideration of non-invasive techniques such as concluded that for adult anurans ‘toe-clipping the use of individual pattern differences where remains the method of choice’, though the use of these exist. Individual pattern recognition is passive integrated transponders (PIT) and visible made much more feasible by the advent of implant elastomer (VIE) marking has signifcant digital photography where large numbers of potential, where fnances allow (Heard et al. photographs can be easily, cheaply and quickly 2008, Sapsford et al. 2015). However, it seems taken and the bank of photographs can be quickly that PIT tags and VIE can have limited retention scanned on a computer screen. Ideally, what is and detection rates in adult frogs (Brannelly et needed is an individually recognizable pattern al. 2013, 2014, Bainbridge et al. 2015). difference that can quickly be distinguished and Furthermore, several studies suggest that the is stable over time. In great crested newts, effects of invasive marking techniques can Triturus cristatus (Laurenti, 1768), belly patterns infuence the expression of natural behaviors in are complex and variable and can be used for amphibians, such as courtship (Winandy and individual recognition (Hagström 1973) but Denoёl 2011), feeding ability (Davis and Ovaska ontogenetic pattern variation does occur (Arntzen 2001) and locomotory performance (Schmidt and Teunis 1993) and the complexity of the and Schwarzkopf 2010). patterns makes individual recognition tedious Evidence about the effects on survival and and time-consuming when populations are large stress following toe-clipping in frogs is variable (McNeill, pers. comm.). In their study of PIT- and contested (Narayan et al. 2011, Fisher et al. tagging compared to pattern mapping Arntzen et 2013). Even studies that suggest toe-clipping has al. (2004) suggested that non-invasive pattern minimal effects on survival advise due mapping ‘by eye’ is only more cost-effective consideration of ‘functionally important’ digits where populations have fewer than 176 indivi- and restraint in numbers of toes clipped (Grafe et duals. However, with increasing availability of al. 2011), particularly given that return rates in image analysis software, and the development of Phyllomedusa - 18(1), June 2019 14 Stable individual variation in ventral spotting patterns in Phyllomedusa trinitatis specifc pattern-matching programs for use in In 2014, 2015 and 2016, a population of P. wildlife studies, the potential for using photo- trinitatis was observed around a set of artifcial graphic evidence to study large populations of ponds at the William Beebe Research Station animals is now far greater than it was, even a (‘Simla’) in Trinidad’s Arima Valley (Downie et decade ago (Mettouris et al. 2016). al. 2013) during the early months of the rainy Here we present evidence on a reliable, season (June–August). Frogs were located at stable, individually variable pattern occurring night by call and movement in the trees and in at least some members of the neotropical bushes overhanging the ponds. Frogs were genus Phyllomedusa (Anura: Phyllomedusidae). captured, measured and photographed as above. Phyllomedusa currently comprises 16 species It was sometimes possible to photograph the (Frost 2018). Formerly, most of the species ventral markings without capturing the frog, could be assigned to one of four species thereby reducing any stress related to handling. groups: P. tarsius, P. burmeisteri, P. Sex was determined by size and behavior hypochondrialis, and P. perinesos. Following [females are signifcantly larger (90 mm SVL) the taxonomic revision by Duellman et al. than males (80 mm): Murphy 1997]. Each frog (2016), members of the P. hypochondrialis was assigned a unique number and location Group were assigned to the genus Pithecopus (there are three different pond sites) and then and P. perinesos Group to Callimedusa, leaving released where captured. Sampling was repeated P. tarsius Group and P. burmeisteri Group multiple times in each of the years (nightly in species within Phyllomedusa. In a non refereed 2016; less frequently in 2014 and 2015, usually report, one of us (JMS in Smith et al. 2007) on nights following wet days). Full measurements presented evidence that the variable ventral and photographs were taken each time and later markings found on Phyllomedusa trinitatis checked visually against our accumulating Mertens, 1926 skin could be used to recognize photograph database to determine which of the individuals. Here, we describe the ventral pattern frogs had been previously recorded and which as found in this species, provide evidence on its were new. It took about 10 min to check fve ontogenetic stability and its usefulness in a feld new photographs. Size measurements were used study in Trinidad, West Indies, and describe as a double check on identifcation. One similar patterns found in some other members of experienced observer (E.G.) both photographed the genus. the frogs and checked the database. Using one experienced observer for this task made it less Materials and Methods time consuming as the database accumulated. Two freshly-laid egg clutches (300–400 eggs Field Observations and Egg Collections in each), wrapped in leaves, were collected in July 2014 from branches overhanging