Diet of Aplastodiscus perviridis Lutz 1950 ( Anura , Hylidae ) in subtemperate forests of southern Brazil

This is an open access article distributed under the terms of the Creative Commons Attribution 4.0 International (CC BY 4.0), which permits reproduction, adaptation, and distribution provided the original author and source are credited. 1 Universidade do Vale do Rio dos Sinos, Laboratório de Ecologia de Vertebrados Terrestres. Av. Unisinos, 950, Cristo Rei, 93022-750, São Leopoldo, Rio Grande do Sul, Brazil. Diet of Aplastodiscus perviridis Lutz 1950 (Anura, Hylidae) in subtemperate forests of southern Brazil


Introduction
Studies on feeding ecology provide basic information about the biology of a species, as well as its position in the food web (Begon et al., 2006).Despite a recent significant increase in studies on amphibian ecology in the Neotropics (Ximenez and Tozetti, 2015), the number of such studies is relatively low regarding the family Hylidae when considering the high diversity of the group.
Diet composition of hylids range from generalist (Van Sluys and Rocha, 1998;Barbosa et al., 2014) to specialist trends (Parmelee, 1999).It may also vary according to morphological features (Toft, 1981), such as individual relative size and the size of prey that can be ingested (Muñoz-Guerrero et al., 2007).The species behaviour itself can also influence the diet, such as changes in prey consumption during the breeding season (Solé and Pelz, 2007).This may be related to either the duration of the reproductive period or the different distances to be traveled to the reproductive site (Duellman and Trueb, 1994;Solé and Pelz, 2007;Barbosa et al., 2014).Species with short breeding season, for example, tend to decrease prey consumption in the reproductive period (Solé and Pelz, 2007).
The arboreal anuran Aplastodiscus perviridis Lutz 1950 (Hylidae) occurs in southern and southeastern Brazil and far west of Argentina.It lives in transition areas between forests and open areas (edge) and has a short breeding season going from November to February (Kwet et al., 2010).This species has a specific reproductive behaviour, in which males display arrangement moves and calls to attract the female to an underground burrow constructed by the male itself (Haddad et al., 2005).In Rio Grande do Sul, this species occurs on the edge of native grassland areas with Araucaria Forest (with the presence of Araucaria angustifolia), using wetlands for breeding (Kwet et al., 2010).These grassland and forested areas, which are important for animal diversity in general, are under threat by human farming activities that convert natural areas into crops and apply agrochemicals (Bond-Buckup and Dreier, 2008).
Except for reproduction studies (Haddad and Prado, 2005;Haddad et al., 2005;Zina et al., 2007), distribution notes (Both et al., 2006), taxonomy (Garcia et al., 2001) and cell biology (Carvalho et al., 2009), studies on the genus Aplastodiscus are scarce, and there are no records on feeding behavior.Considering the importance of natural history studies and the limited available knowledge about this genus, the aim of this study was to contribute to filling these gaps, describing and analysing the diet composition of A. perviridis during the breeding season in areas associated to Araucaria moist forests in Rio Grande do Sul, Brazil.

Data Collection
We collected the samples in February, September and November 2015, using the nocturnal visual search method (Crump and Scott Jr., 1994).We kept the captured animals in identified plastic bags and maintained them in a refrigerated container to reduce the physiological activities that accelerate digestion.We collected the samples under the collecting permit from the Federal Authority, SISBIO (permit # -45861-1).After identification, we euthanized the specimens with topical anesthetic (lidocaine), fixed them in 10% formalin and preserved them in 70% ethanol.Afterwards, we dissected the animals to remove the gastrointestinal tract (stomach and intestine), which we maintained in 70% ethanol and screened by use of a stereomicroscope.We considered stomach and intestine contents as a single sample.We identified the extracted items (prey) mostly to the taxonomic level of order, except for hymenopterans, in which we differentiated Formicidae from non-Formicidae.When we found unidentifiable prey fragments, we grouped these under "other items", of which we quantified only volume.For each identified item (prey category), wherever possible, we calculated the number, volume and frequency of occurrence in absolute and percentage values.We calculated volume by using the area (mm²) occupied by each item with graph paper support fixed to the bottom of the Petri dish, where we evenly spread each item, maintaining a regular height of 1 mm (Hellawell and Abel, 1971).To calculate the volume (V), we multiplied the value of the area occupied by each item by its height (1 mm) (Barbosa et al., 2014).

Data Analysis
We assessed the importance of each prey in the species' diet using the Index of Relative Importance (IRI): IRI = (%N +%P) %FO, where %N is the relative abundance of each prey in the diet; %P is the mass percentage of each prey in the diet; and %FO is the relative frequency of occurrence of prey (Pinkas et al., 1971;Krebs, 1999).The higher the value of IRI, the greater is the importance of each prey in the diet.
We calculated the amplitude of trophic niche through Levins' Trophic Niche Amplitude Index (B) (Krebs, 1999), defined by: B = 1 / Σp i 2 , in which p is the proportion of individuals of a given resource i (taxon) found in the diet.To facilitate comparisons among species, we calculated the standardized Levins' index (Bsta), which limits the index on a scale from 0 to 1 according to the following equation: Bsta = (B-1) / (n-1), where n is the number of resources (prey categories) recorded.Values near zero are assigned to specialized diet, while those closer to 1, to a generalist diet.

Results
We captured 42 males of Aplastodiscus perviridis, in Garibaldi, Farroupilha and São Francisco de Paula (five without gastrointestinal content, representing 12% of the total sample).Altogether, we found 98 food items classified into 12 categories (Table 1).According to IRI, the most important category in the diet was Acarina, followed by Araneae, Coleoptera-adults, Lepidoptera-larvae, Orthoptera, Hemiptera and Coleoptera-larvae (Table 1).Other groups, such as Lepidoptera-adult, non Formicidae, Formicidae, Pseudoscorpionida and Dermaptera, had little relevance in the diet of A. perviridis, presenting a value of IRI < 30.
The trophic niche breadth (Bsta) was 0.27.Plant material content had high frequency in the stomach/intestines of frogs (Table 1).However, we could not identify a relatively large proportion of items due to their high level of fragmentation ("other items").

Discussion
Some items consumed by Aplastodiscus perviridis were also recorded in the diet of other frog species of the Hylidae family.Coleoptera-adult prey also showed high importance to Boana leptolineata (Braun and Braun, 1977) in a habitat that is similar to that of this study (Bar-bosa et al., 2014).A high consumption of Araneae was also recorded for Boana pulchella (Duméril and Bibron, 1841) in coastal areas of Rio Grande do Sul and Uruguay (Da Rosa et al., 2011;Oliveira, 2014).Moreover, the presence of Acarina was also identified in the diet of Boana raniceps (Cope, 1862) in Pantanal (Sabagh et al., 2010) and of B. pulchella in Uruguay (Da Rosa et al., 2011).It is worth reminding that, in terms of biomass, it is believed that this prey category may not be of much importance, as mentioned in other studies on Neotropical hylids (Muñoz-Guerrero et al., 2007;Antoniazzi et al., 2013).
The presence of larvae in the diet (Lepidoptera and Coleoptera-larvae), which are not very movable prey, suggests that A. perviridis individuals make an active search for them.In addition, when not vocalizing, A. perviridis probably uses the same shelter as the Coleoptera-larvae (e.g.fallen trunks, decomposing material) (Haddad and Prado, 2005;Haddad et al., 2005), which would increase the probability of encountering this prey.The number of prey that could not be identified due the advanced digestion degree (Other items) does not compromise the conclusions of this work, since this category is distributed in many of the individuals, besides being common in diet works (Siqueira et al., 2006), mainly in intestinal contents (Santana and Juncá, 2007;Moser et al., 2017).
There is a tendency in hylids with short reproductive period to reduce the ingestion of prey, prioritizing reproduction over foraging (Duellman and Trueb, 1994;Solé and Pelz, 2007).However, even A. perviridis having a short reproductive period, it does not seem to reduce foraging during reproduction, because the average amount of prey in the gastrointestinal tract was similar to that observed in another hylids with long-reproductive period, such as B. pulchella (Oliveira, 2014).
The trophic niche breadth of the studied populations of A. perviridis was smaller than that of B. leptolineata, Bsta = 0.51 (Barbosa et al., 2014) and B. raniceps, Bsta = 0.64 (Sabagh et al., 2010) and similar to that of B. pulchella, Bsta = 0.31 (Oliveira, 2014) from Brazil.This suggests that A. perviridis was more specialized in the use of food resources during the study period.

Table 1 .
Prey categories consumed by Aplastodiscus perviridis in Araucaria Forest in Rio Grande do Sul, southern Brazil.FO = frequency of occurrence of each prey category; IRI = Index of relative importance; N = number of individuals; V = total volume of prey (mm 3 ); (%) = percentage related to total.