Herpetology Notes, volume 8: 173-177 (2015) (published online on 10 April 2015)

Diet composition and niche overlap in two sympatric species of (Anura, , Leiuperinae) in coastal subtemperate wetlands

Mateus de Oliveira1,*, Marco Silva Gottschalk2, Daniel Loebmann3, Mauricio Beux dos Santos4, Stéfanie Miranda3, Camila Rosa3 and Alexandro Marques Tozetti1

Abstract. Physalaemus Fitzinger, 1826 is the most speciose genus of Leiuperinae , with more than 40 recognized species. They are widely distributed in the Neotropical region, especially in open areas, and are among the most abundant species in several anuran communities. Nevertheless, our comprehension of the diet composition and trophic relationships of these species remains poor. In this study, we describe and compare the diet of sympatric and syntopic populations of Physalaemus gracilis and P. biligonigerus that inhabit the southernmost coastal zone of Brazil. The stomach content analyses revealed 13 distinct prey categories in the diet of P. gracilis and 12 in the diet of P. biligonigerus. On one hand the diet compositions of the species were significantly different according to both numeric and volumetric evaluations. On the other hand, Coleoptera and Formicidae were the most important prey for both species. The niche breadth was higher for P. gracilis than P. biligonigerus, and niche overlap was relatively low for both species (28%). Our results indicate that P. biligonigerus has a less generalist diet than P. gracilis, with a remarkable consumption of ants. Although both species are morphologically similar and sympatric, they appear to present different feeding strategies or preferences based on the low values of niche overlap.

Keywords: Coleoptera, diet, Formicidae, Physalaemus biligonigerus, Physalaemus gracilis, restinga

Introduction The Neotropical frogs of the genus Physalaemus Fitzinger, 1826 currently represent 45 species distributed The assessment of information on the use of food throughout Central and South America (Frost, 2013). resources is important for understanding the trophic Among those species, Physalaemus gracilis and relationships in communities (Pianka, 1974; Physalaemus biligonigerus present high overlap in Bianchi, 2009). With this information available, their distribution and are sympatric in southern Brazil, generalizations about the ecological aspects of organisms Uruguay and Argentina (Langone, 1994; Lavilla et can be made, such as foraging strategies and adaptations al., 2010; IUCN, 2013). Additionally, studies have to different habitats (Pianka, 1975). demonstrated that both species also share microhabitats for foraging (Oliveira et al., 2013) and reproduction (Kwet, 2002). However, studies on the diets of these species are scarce (Rosa et al., 2002). Previous studies 1 Universidade do Vale do Rio dos Sinos – UNISINOS, have considered these species as predators of arthropods Laboratório de Ecologia de Vertebrados Terrestres, São (Achaval and Olmos, 2003). The present study aimed to Leopoldo, RS, Brazil. describe and compare the diet of sympatric populations 2 Universidade Federal de Pelotas – UFPel, Instituto de of P. gracilis and P. biligonigerus in habitats from Biologia, Departamento de Ecologia, Zoologia e Genética, coastal restingas of southern Brazil. Pelotas, RS, Brazil. 3 Universidade Federal do Rio Grande, Instituto de Ciências Biológicas, Laboratório de Vertebrados Ectotérmicos, Rio Materials and Methods Grande, RS, Brazil. Study Site: were collected in coastal 4 Programa de Pós-graduação em Biodiversidade Animal, Universidade Federal de Santa Maria (UFSM), Santa Maria, environments of wetland and restinga habitats (between RS, Brazil. the coordinates 32°00’ and 32°14’S, 52°09’ and * Corresponding author e-mail: *[email protected] 52°12’W), in the city of Rio Grande, southern Brazil. 174 Mateus de Oliveira et al.

The region has a subtemperate climate with an average is the number of resources (prey species) recorded. annual temperature of 18.1 °C; the average annual Values close to 0 are assigned to a specialized diet, rainfall being 1162 mm (Maluf, 2000). whereas values closer to 1 indicate a generalist diet. The standardization allows for better interpretation and Data Collection: The specimens were obtained from understanding of the data comparison because the main the Herpetological Collection of Universidade Federal advantage is that the pattern is not influenced by the total do Rio Grande – FURG and were collected between number of prey categories that may be related to sample December 2008 and December 2011. To ensure the size (Krebs, 1999). For the analysis of overlap in niche preservation of their stomach contents, only individuals dimensions and/or the degree of similarity between the that were euthanized shortly after capture and fixed diets of pairs of species, we used Pianka’s niche overlap with 10% formalin and preserved in 70% alcohol were index (Ojk) (Pianka, 1973), with values ranging from 0 used. For stomach content analysis, we exclusively used (no overlap) to 1 (complete overlap) (Krebs, 1999). This individuals obtained from areas with the same habitat is calculated by the following expression: type (coastal wetlands and restinga habitat). Anurans were dissected to assess stomach contents that were o o o u u 2 u ppppO 2 extracted and preserved in 70% ethanol until analysis jk 1n ikij 1n ij ¦¦¦ 1n ik using a stereomicroscope. Food items were identified to the lowest possible taxonomic level. After prey , where O represents the index of overlap of Pianka’s identification and quantification, we measured the jk niche between species j and k; p is the proportion of volume (V) based on the area (mm²) occupied by each ij resource i in total resources used by species j; p is the item and its height (V = area x height). For this, each ik proportion of resource i in total resources used by species item was crushed and spread evenly on a Petri dish over k; and n is the total number of resource categories for a grid scale forming a regular layer of 1 mm (Hellawell species j and k. and Abel, 1971). To test the similarity of the composition of the diet between species, an analysis of similarity (ANOSIM) Data Analysis: Numeric and volumetric percentages was performed using samples as individual species of each category of prey in each stomach, as well (Clarke and Gorley, 2006). To that end, we generated as the percentage frequency of occurrence, were a similarity matrix using the Bray-Curtis similarity calculated on the set of individuals. To determine the coefficient from the diet composition (abundance and relative importance of each prey category in the diet of volume). The test compares ANOSIM similarities the species, we used the index of relative importance between two or more groups of sampling units (factors) (IRI) (Krebs, 1999) from Pinkas et al. (1971) using the and generates a statistical R, which varies between -1 following formula: IRI = (% N +% P) FO%, where % and 1 (Clarke and Gorley, 2001). In the context of this N is equal to the relative abundance of each prey in the study, the R value of zero represents the null hypothesis diet; % P is the percent volume of each prey in the diet; (no difference between a set of samples), which means % FO is the relative frequency of occurrence of prey. that the similarities between and within samples are The higher the value, the greater the importance of each the same, and the value of R indicates that within the prey item in the diet. sample, there is more similarity between the first set of In order to determine the level of specialization of the factors than between the levels of each of the factors. diet, the trophic niche breadth was calculated according to the Levins Index of trophic niche breadth (B) (Krebs, Results 1999). This index is defined as follows: B = 1 / Σpi2, where p equals the proportion of individuals of a given We analyzed the stomach contents from 93 specimens, resource i (taxon) found in the diet. This index allows of which 44 belonged to Physalaemus gracilis (4 us to measure the amplitude or “diversity” of the diet empty stomachs) and 49 to P. biligonigerus (3 empty considering the quantitative distribution of each prey stomachs). Sixteen prey categories were identified item. As the Levins index (B) varies from 1 to n, we (Table 1), with 13 categories for P. gracilis and 12 for used a standardized Levins index (Bsta) to allow direct P. biligonigerus. Significant differences were observed comparisons with other studies. A standardized Levins in the diet of the study species, both numerically (R = index (Bsta) limits the index on a scale from 0 to 1 using 0.21, p = 0.0001) and volumetrically (R = 0.2282, p = the following equation: Bsta = (B-1) / (n-1), where n 0.0001). Isopoda, Amphipoda, Blattodea and larvae of Diet composition and niche overlap in two sympatric species of Physalaemus 175

Lepidoptera were found exclusively in the diet of P. Discussion gracilis, whereas Pseudoescorpionida, Homoptera and Although phylogenetically related and sympatric, Termitidae occurred only in the diet of P. biligonigerus the two species showed significant differences in the (Table 1). composition of their diets. Coleoptera larvae and The IRI revealed that for P. gracilis, Coleoptera larvae Isopoda were the most important preys for Physalaemus was the most important prey category (IRI = 1050.9) gracilis, whereas Formicidae was more important for P. followed by Isopoda (IRI = 872.4) and Formicidae biligonigerus. This is the first detailed study revealing (IRI = 272). For P. biligonigerus, greater values of IRI that the larvae of Coleoptera is a representative prey item were registered for Formicidae (IRI = 10122.5) and for Physalaemus (Santana and Juncá, 2007). Moreover, Coleoptera larvae (IRI = 815) (Table 1). even for species where there are records of consumption The trophic niche breadth was higher for P. gracilis of Coleoptera, this prey item is rarely consumed in the (Bsta = 0.23) than P. biligonigerus (Bsta = 0.04). larval form (M. Oliveira, unpublished data). Regarding the niche trophic overlap, the Pianka index Coleoptera and Formicidae have been reported as (O ) showed an overlap of 28% (O = 0.28) between jk jk important prey in the diet of many anurans, which should the species. 1 CAPTIONS

2 Table 1. Diet composition of P. gracilis and P. biligonigerus in coastal environments in southern

Table3 1. DietBrazil. composition N = Numberof P. gracilis of and individuals P. biligonigerus found; in coastal V environments = Volume in of southern the set Brazil. of N individuals = Number of inindividuals mm³; F = found; V = Volume of the set of individuals in mm³; F = Frequency of occurrence; (%) = Percentage with respect to total; IRI = index4 of relativeFrequency importance. of occurrence; (%) = Percentage with respect to total; IRI = index of relative importance.

P. gracilis (N = 44; 4 empty) P. biligonigerus (N = 49; 3 empty) N (%) V (%) F (%) IRI N (%) V (%) F (%) IRI Arachnida Pseudoescorpionida 0 (0) 0 (0) 0 (0) 0 1 (0.1) 2 (0.1) 1 (2.2) 0.4 Araneae 7 (3.4) 105 (5.9) 4 (10) 85.8 6 (0.6) 54 (1.8) 3 (6.5) 15.7 Acarina 2 (1) 2 (0.1) 2 (5) 4.5 8 (0.8) 15 (0.5) 5 (10.9) 13.9 Unidentified 3 (1.4) 8 (0.5) 1 (2.5) 4.1 2 (0.2) 5 (0.2) 1 (2.2) 0.8 Malacostraca Amphipoda 30 (14.4) 88 (5) 3 (7.5) 125.2 0 (0) 0 (0) 0 (0) 0 Entognatha Collembola 7 (3.4) 9 (0.5) 1 (2.5) 8.1 4 (0.4) 7 (0.2) 2 (4.3) 2.7 Insecta Blattodea 1 (0.5) 21 (1.2) 1 (2.5) 3.9 0 (0) 0 (0) 0 (0) 0 Coleoptera (larvae) 102 (49) 118 (6.6) 9 (22.5) 1050.9 79 (7.7) 390 (13.2) 18 (39.1) 815 Diptera 8 (3.8) 86 (4.8) 6 (15) 118.8 4 (0.4) 22 (0.7) 3 (6.5) 7.4 Hemiptera 8 (3.8) 63.25 (3.6) 7 (17.5) 116.5 13 (1.3) 40 (1.4) 5 (10.9) 28.4 Homoptera 0 (0) 0 (0) 0 (0) 0 5 (0.5) 11 (0.4) 2 (4.3) 3.7 Isopoda 18 (8.7) 571 (32.1) 9 (22.5) 872.4 0 (0) 0 (0) 0 (0) 0 Lepidoptera (larvae) 1 (0.5) 15 (0.8) 1 (2.5) 3.1 0 (0) 0 (0) 0 (0) 0 Isoptera Termitidae 0 (0) 0(0) 0 (0) 0 57 (5.6) 53 (1.8) 5 (10.9) 79.6 Hymenoptera Formicidae 21 (10.1) 47.1 (2.7) 10 (25) 272 848 (82.6) 1503 (50.8) 35 (76.1) 10122.5 Arthropoda (fragments) - 482 (27.1) 27 (67.5) -- -- 567 (19.2) 23 (50) -- Other - 66 (3.7) 4 (10) -- -- 0 (0) 0 (0) -- Vegetal fragments - 95 (5.4) 13 (32.5) -- -- 291 (9.8) 25 (54.3) -- 5

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 176 Mateus de Oliveira et al. be related to the high availability of these organisms in Acknowledgments. We would like to thank FAPERGS, CAPES various environments (Clarke, 1974). Despite some and CNPq for financial support. speculation, the consumed prey items may indicate the distinct foraging tactics of the study species. For References example, the presence of larvae of Coleoptera and Achaval, F., Olmos, A. (2003): Anfibios y reptiles del Uruguay. 2a Isopoda suggests that the Physalaemus gracilis are edição. Montevideo, Graphis. 136 pp. foraging in specific microenvironments, as the prey Bianchi, R.C. (2009): Ecologia de mesocarnívoros em uma área species have a conspicuous distribution in habitats (e.g., no Pantanal Central, Mato Grosso do Sul. Tese de Doutorado. under logs). It is likely that P. biligonigerus adopts a Centro de Ciências Biológicas e da Saúde, Universidade Federal foraging tactic directed to the search of ant nests. 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(1994): Ranas y sapos del Uruguay (reconocimiento Although there are ecological similarities between the y aspectos biológicos). Museo Damaso Antonio Larrañaga, Ser. Divul. 5:1-123. two species (Kwet, 2002; pers obs), we recorded a small Lavilla, E., Kwet, A., Segalla, M.V., Langone, J., Baldo, D. (2010): trophic niche overlap. In comparative terms, Oliveira Physalaemus gracilis. In: IUCN 2013. IUCN Red List of (2014) recorded a greater overlap than that of the Threatened Species. Version 2013.1. . present study in the same habitat, even when comparing Maluf, J.R.T. (2000): Nova classificação climática do Estado do ecologically and phylogenetically distinct species Rio Grande do Sul. Revista Brasileira de Agrometeorologia 8: (Leptodactylus latrans X Physalaemus biligonigerus 141-150. = 0.83 and Hypsiboas pulchellus X Odontophrynus Oliveira, M.C.L.M, Dos Santos M.B., Loebmann, D., Hartman, A., maisuma = 0.84). Even preliminary, this assessment Tozetti, A.M. 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Accepted by Mirco Solé