ISSN 1827-9635 (print) © Firenze University Press ISSN 1827-9643 (online) www.fupress.com/ah Acta Herpetologica 9(1): 115-117, 2014 DOI: 10.13128/Acta_Herpetol-13105 Clutch size in wild populations of Alytes muletensis Samuel Pinya1,*, Valentín Pérez-Mellado2 1 Herpetological Research and Conservation Centre, Associació per a l’Estudi de la Natura. Balearic Islands, Spain. * Corresponding author. E-mail: sampinya22@gmail.com 2 Department of Animal Biology, Universidad de Salamanca, Spain, E-mail: valentin@usal.es Submitted on 2013, 17th July; revised on 2013, 21st November; accepted on 2014, 16th April Abstract. Clutch size of genus Alytes presents strong inter- and intra-specific variability. The Mallorcan midwife toad (A. muletensis) is a threatened endemism of the island of Mallorca (Spain), and has the smallest clutch size of the genus. Most of the previously published clutch size information was obtained from animals breeding in captivity. In this study we analysed a large number of clutches from wild populations, and we compared their size (number of eggs) with previously published data from both captive and wild populations. Our results showed that clutch sizes of wild males carrying a single clutch were similar to those observed in captivity. However, multiple clutches were more common in the wild than in captivity. Keywords. Alytes muletensis, clutch size, wild populations, endemic species, Balearic Islands In Alytes species it is males that care for eggs. The female expels a strand of eggs which are fertilised by the male before he wraps them around his legs to protect them from predators. When the eggs are ready to hatch, the male wades into shallow ponds or streams to allow tadpoles to go into the water (Stebbins and Cohen, 1997). Five separate species of midwife toads are found across Western Europe, Northern Africa, and the island of Mallorca: Alytes obstetricans, Alytes cisternasii, Alytes dickhilleni, Alytes maurus and Alytes muletensis (Mar- tinez-Solano et al., 2004). The clutch size of the genus Alytes is highly variable both, inter- and intra-specifically (Table 1). The Mallorcan midwife toad, A. muletensis, is a threatened endemism of the island of Mallorca (Balearic Islands, Spain; Serra et al., 2009), with the smallest clutch size of the genus (Alcover et al., 1984, Fig. 1). As in other Alytes species, the clutch size of A. mule- tensis is fairly variable (Table 2). During studies in cap- tivity Bush (1993, 1996) observed the presence of double clutches for the first time in A. muletensis, that is, one male carrying the eggs released by two females. He also found that single clutches (less than 15 eggs) had an aver- age of 10.00 eggs (range from 4 to 14 eggs, Bush, 1993, 1996). Taking into account double clutches, the aver- age increased to 11.36 eggs (range from 4 to 27; Bush, 1996). Additionally, and also in captivity, the maximum clutch size observed by Román and Mayol (1997) had 34 eggs, while the average clutch size was 11 eggs. In order to establish demographic parameters of wild A. muleten- sis individuals, between 2007-2010, we counted the num- ber of eggs carried by 92 males from 11 natural, plus 7 artificial, breeding sites in the wild. An artificial breeding site for the purpose of this study is a man-made struc- ture such as drinking ponds for cattle or old water cis- terns/tanks being used by wild A. muletensis for breeding. These sites were established as a conservation measure for the release of captive-bred animals during the 1990s. The current sample represents the descendants of these introduced toads. On the other hand, streams and iso- lated natural pools are considered natural breeding sites. The clutch size was determined by counting the number of eggs of empty strings and egg capsules after the tad- poles had emerged, and within a short time after the males had come out of the water. 116 S. Pinya, V. Pérez-Mellado To show clutch size variability in wild populations of A. muletensis, descriptive statistics are provided (Table 2). We used one way ANOVA to compare (1) clutch size data from Alcover et al. (1984) with that from the present study, (2) clutch size data from Bush (1996; captive popu- lations) with that from the present study, and (3) clutch size data for artificial and natural breeding sites from the present study. A chi-square test was used to compare frequencies of multiple clutches in captive (from Bush, 1996) and wild (from the present study) populations of A. muletensis. All statistical analyses were carried out with Statistica (ver. 6.0). Our data revealed an average clutch size of 12.04 eggs in the wild (Table 2). We found no significant dif- ference (F1,99 = 1.40, p = 0.24) when our data was com- pared to previously published averages for wild indi- viduals (Alcover et al., 1984), maybe because of the small sample size in the 1984 study. There was also no sig- nificant difference between clutch size data in our study (F1,319 = 0.83, p = 0.36) and that published by Bush (1996) for animals in captivity. Additionally, there was no significant difference between natural (n=41) and arti- ficial (n=51) breeding sites in the present study (F1,92 = 1.31, p = 0.26). The observed range of clutch size found in this study is the largest known for both wild and cap- tive populations, and coincides with the minimum and maximum values published previously (Alcover et al., 1984; Bush, 1996; Román and Mayol, 1997). Multiple clutches are common in natural popula- tions of continental midwife toads (Bush, 1996). Between 56-61 % of A.obstetricans and 80 % of A. cisternasii egg- carrying males carry two or more clutches simultane- ously (Reading and Clarke, 1988; Márquez 1990). In A. muletensis, double clutches are rare (12-14 % according to Bush, 1996). On the other hand, triple clutches, those with more than 28 eggs, were first reported by Román and Mayol, (1997) with a record of 34 eggs, but their fre- quency was not provided. Our data shows that, in wild populations 23.91 % of clutches should be considered multiple clutches, with 3.26 % of them consisting of triple clutches. When comparing the occurrence of multiple clutch- es, between wild individuals in this study and data for captive individuals (Bush, 1996), we found a significantly higher frequency of multiple clutches in the wild popula- tion (χ2 = 5.30, p < 0.02). The low incidence of multiple clutches in captivity may be due to the short time avail- Table 1. Clutch sizes of continental Alytes species. Average eggs per clutch, minimum and maximum eggs per clutch, and source of information. Species Average Range Reference A. obstetricans 63 35-95 Crespo, 1979 104 46-143 López-Jurado et al., 1979 51 24-142 Buchholz, 1989 38 5-120 Galán et al., 1990 27 21-33 Galán et al., 1990 55 24-77 Galán et al., 1990 34 6-53 Galán et al., 1990 77 32-171 Reading, Clarke, 1998 A. cisternasii 73 42-119 Crespo, 1979 104 46-143 López-Jurado et al., 1979 73 48-116 Malkmus, 1983 87 20-180 Marquez, 1989 A. dickhilleni 74 28-149 González-Miras, García-Cardenete & Tejedo, 2012 A. maurus - 60-70 Donaire-Barroso, Bogaerts, 2003 Table 2. Clutch size in A. muletensis: summary of literature and original (present study) data. Population type, average clutch size, sample size (when available), minimum and maximum eggs per clutch, and source of information. Population type Average N Range Variance Reference Natural 12.04 92 4-34 32.53 Present study Natural 9.78 9 7-12 - Alcover et. Al, 1984 Captive - - 10-24 - Martínez-Rica et al., 1984. Captive 11.43 - 9-15 - Tonge & Bloxam, 1989 Captive 11.36 220 4-27 14.05 Bush, 1996 Captive and Natural 11 - -34 - Román & Mayol, 1997 Fig. 1. Male A. muletensis carrying eggs (island of Mallorca, Spain). 117Clutch size in Alytes muletensis able for a male to obtain the second clutch from a female (Bush, 1996). Males in wild populations may have longer time intervals during which they can accept additional clutches. In addition, the operational sex-ratio during the breeding season also determines the potential for multi- ple clutches. If more females are available males will be able to mate with more than one female in a short period of time and hence brood more than one clutch (Bush, 1996). It is also possible that wild individuals breed at higher densities than captive individuals, and wild recep- tive males may be able to obtain a second clutch more frequently simply because they are more likely to encoun- ter more than one gravid female. AKNOWLEDGEMENTS We would like to thank Joan Mayol, Joan A. Oliver, Mª Antònia Vanrell and Joan C. Salom of the Balearic Environmen- tal Agency, who gave us administrative support and provided the capture permits (19/2007, 44/2008, 07/2009, 13/2010) for our study with the Mallorcan midwife toad. We are also grateful to all the people who provided support during the field-work, specially Xavier Manzano and Jaume Bonnin. Montserrat Car- bonell made an excellent revision of English language and gave us very useful suggestions. REFERENCES Alcover, J.A., Mayol, J., Jaume, D., Alomar, G., Jurado J. (1984): Biologia i Ecologia de les poblacions de Baleaphryne muletensis a la muntanya mallorqui- na. In: Història Biològica del Ferreret. Monografies científiques 3, pp. 129-152. Hemmer, H., Alcover, J.A., Eds, Editorial Moll, Palma de Mallorca. Buchholz, S. (1989): Untersuchungen zur Fortpflanzungs- biologie und Populationsdynamik einer Freilandpopu- lation von Alytes obstetricans (Amphibia, Anura, Disco- glossidae). Diplomarbeit, Universität Würzburg, 81 S. Bush, S.L. (1993): Courtship and male parental care in the Majorcan midwife toad, Alytes muletensis. Unpub- lished doctoral dissertation. University of East Anglia, Norwich. Bush, S. L. (1996): Why is double clutching rare in the Majorcan midwife toad?. Anim. Behav. 52: 913-922. Crespo, E.G. (1979): Contribuçao para o conhecimento da biología dos Alytes ibéricos, Alytes obstetricians boscai Lataste 1879 e A. cisternasii Boscá 1879: a pro- blamática da especiaçao de A. cisternasii. Unpublished doctoral dissertation. University of Lisboa. Donaire-Barroso, D., Bogaerts, S. (2003): Datos sobre tax- onomía, ecología y biología de Alytes maurus (Pasteur & Bons, 1962) (Anura; Discoglossidae). Bull. Soc. Cat. Herp. 16: 25-41. Galán, P., Vences, M., Glaw, F., Fernádenz-Arias, B., García-París, M. (1990): Beobachtungen zur Biologie von Alytes obstetricans in Nordwestiberien. Herpeto- fauna 12: 17-24. González-Miras, E., García-Cardenete, L., Tejedo M. (2012): Historia Natural. In: Seguimiento de Alytes dickhilleni: Informe final. Monografías SARE 02, pp. 17-22. Bosch, J., González-Miras, E., Eds. Asociación Herpetológica Española, Ministerio de Agricultura, Alimentación y Medio Ambiente, Madrid. Malkmus, R. (1983): Zur Fortpflanzungsbiologie von A. cisternasii. Weinstadt 5: 30-34. Márquez, R. (1990): Male parental care, sexual selection, and the mating systems of the midwife toads Alytes obstetricans and Alytes cisternasii. Unpublished doc- toral dissertation. University of Chicago. Martínez-Rica J.P., Pardo, M.P., Cervantes, J. (1984): La reproducción y la conducta en cautividad del sapillo balear, Baleaphryne muletensis. In: Història Biològica del Ferreret. Monografies científiques 3, pp. 175-191. Hemmer, H., Alcover, J.A., Eds, Editorial Moll, Palma de Mallorca. Martínez-Solano, I., Gonçalves, H.A., Arntzen, W., García-París, M. (2004): Phylogenetic relationships and biogeography of midwife toads (Discoglossidae: Alytes). J. Biogeogr. 31: 603-618 Reading, C.J., Clarke, T. (1988): Multiple clutches, egg mortality and mate choice in the midwife toad, Alytes obstetricans. Amphibia-Reptilia 9: 357-364 Román, A., Mayol, J. (1997): La recuperación del ferreret, Alytes muletensis. Documents tècnics de Conservació 1. Conselleria de Medi Ambient, Ordenació del Ter- ritori i Litoral, Palma de Mallorca. Serra, J.M., Griffiths, R., Bosch, J., Beebee, T., Schmidt, B., Tejedo, M, Lizana, M., Martínez-Solano, I., Salvador, A., García-París, M., Gil, E.L., Arntzen, J.W. (2009). Alytes muletensis. In: IUCN 2013. IUCN Red List of Threatened Species. Version 2013.2. . Downloaded on 24 March 2014. Stebbins, R.C., Cohen N.W. (1995): A Natural History of Amphibians. Princeton University Press. Chichester, West Sussex. Tonge, S., Bloxam, Q. (1989): Breeding the Mallorcan midwife toad Alytes muletensis in captivity. Int. Zoo Yb 28: 45-53. Acta Herpetologica Vol. 8, n. 2 - December 2013 Firenze University Press Journal of the Societas Herpetologica Italica ACTA HERPETOLOGICA