Evol Ecol (2010) 24:124 DOI 10.

1007/s10682-008-9286-9 ORIGINAL PAPER

Lack of phylogenetic signal in the variation in anuran microhabitat use in southeastern Brazil
Paula C. Eterovick Camila R. Rievers Katia Kopp Milena Wachlevski Bruna P. Franco Camila J. Dias Izabela M. Barata Arquimedes D. M. Ferreira Lilian G. Afonso

Received: 3 November 2007 / Accepted: 6 November 2008 / Published online: 23 November 2008 Springer Science+Business Media B.V. 2008

Abstract Microhabitat use is an important component of anuran behavior in both the tadpole and the adult stages. It is potentially inuenced by phylogeny and extant ecological factors acting as selective pressures, such as predation, competition, or physical habitat properties. We aimed to test whether patterns of microhabitat use vary among species, habitats and sites, and how much of this variation can be explained by phylogenetic relatedness. We collected data on microhabitat use at ve different sites, where we obtained a total of 4,230 records of individual tadpoles of 34 species in 15 genera and 7 families, and a total of 1,163 records of adult individuals of 39 species in 16 genera and 8 families. Mantel tests conducted to relate species dissimilarities in microhabitat use and phylogenetic relatedness indicated a weak but signicant relationship for adult anurans, and no relationship for tadpoles. Our results suggest that microhabitat use is a plastic and variable trait, overcoming phylogenetic signal in tadpoles. In adult anurans, very little of the variation in microhabitat use can be explained by phylogenetic relatedness. Microhabitat use is not a good predictor of phylogeny, but it may be a very interesting subject to study natural selection and adaptation. Keywords Anura Phylogenetic relatedness Phylogenetic signal Microhabitat use Introduction Traits exhibited by species may be inuenced to various degrees by their phylogenetic history as well as contemporary selective pressures. Species in a given clade may show
P. C. Eterovick (&) s Graduac cia Universidade Cato lica de Minas o em Zoologia de Vertebrados, Pontif Programa de Po a Gerais, 30535-610 Belo Horizonte, Minas Gerais, Brazil e-mail: eterovick@yahoo.com P. C. Eterovick C. R. Rievers K. Kopp M. Wachlevski B. P. Franco C. J. Dias I. M. Barata A. D. M. Ferreira L. G. Afonso s Graduac cia Universidade Cato lica de Minas o em Zoologia de Vertebrados, Pontif Programa de Po a Gerais, 30535-610 Belo Horizonte, Minas Gerais, Brazil

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high similarity for traits with strong phylogenetic signal, whereas labile traits may differ even in closely related species that have diversied into different ecological niches (Richardson 2001). Certain types of traits are believed to be more conservative and show a stronger phylogenetic signal, such as morphological and physiological traits (Blomberg et al. 2003). On the other hand, behavioral traits are expected to be evolutionarily malleable and more subject to environmental effects (Blomberg et al. 2003). Understanding the roles of phylogenetic signal and contemporary selective pressures in shaping traits through evolutionary lineages has many applications, such as assessment of the usefulness of certain trait types for phylogeny reconstruction (Queiroz and Wimberger 1993), detection of radiation events and rates of evolution (Ollier et al. 2005; Vitt and Pianka 2005), and detection of labile traits which can be good subjects for studies on adaptation (Ord and Martins 2006). Phylogenetic signal can also be useful in predicting traits of a species not yet studied once its phylogenetic position is known, as well as the traits of its close relatives (Blomberg et al. 2003). Attempts have been made to relate species niche and/or morphology with evolutionary history within taxonomic groups as varied as trees (Silvertown et al. 2001), lizards (Poe 2005; Vitt and Pianka 2005), and frogs (Richardson 2001). Anuran larvae can be suitable models to study the relative contribution of ancestral trait values and contemporary selective regimes, but corresponding phenotypes will vary according to the trait considered (Richardson 2001). For instance, both family and habitat type were noticed to inuence phenotypic traits related to digestion, swim speed, and activity of tadpoles to different degrees, resulting in varied outcomes that may represent different adaptive peaks (Richardson 2001; 2002). Family was also used to explain the distribution of anuran species across habitat types in the central Amazon (Zimmerman and Simberloff 1996). However, no attempts have yet been made to understand the role of phylogenetic relatedness in explaining microhabitat use by anurans. Many aspects of anuran ecology and natural history are reected in microhabitat use. Microhabitat use can be inuenced by several factors that include evolutionary history, species interactions, and environmental pressures. Adult males select calling sites that may improve the propagation of their calls (Wells and Schwartz 1982; Lardner and Latkim 2002), confer protection against predators and water loss (Stewart and Rand 1991) or increase their attractiveness to females by containing important resources (Donnelly 1989). Females select microhabitats based on increasing their chances of reproductive success and providing available resources for their offspring (Donnelly 1989). Tadpoles select microhabitats in order to avoid predation (Kopp et al. 2006), and fulll dietary and growth-related requirements (Schiesari 2004). Tadpole microhabitats are so adaptive that they have been associated with diet and morphology to dene eco-morphological types (Altig and Johnston 1989; Altig and McDiarmid 1999). It is worth noting that anuran microhabitat use may change both ontogenetically and in response to individuals of the same or other species, making it potentially very context-dependent (e.g., Alford and Crump 1982; Alford 1986). The factors affecting microhabitat choice can act over varied time scales and in complex combinations, making the resulting patterns hard to predict. Since microhabitats are related to many aspects of anuran lives in both tadpole and adult stages, their use is expected to reect both traits that are molded by long-term evolutionary history and traits that are responsive to recent local selective pressures. This scenario makes patterns of microhabitat use an interesting subject to study phylogenetic signal and to what extent it is affected by contemporary adaptation. Nevertheless, a species pattern of microhabitat use is a complex trait which is hard to describe quantitatively with a single

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measurement, and yet the most elaborate techniques to study phylogenetic signal are designed for quantitative data (e.g. Blomberg et al. 2003; Ollier et al. 2005). The question is: with so many selective pressures potentially affecting microhabitat use, is it still possible to recover phylogenetic history from patterns of microhabitat use by tadpoles and adult frogs distributed in many different habitats? In order to evaluate the role of phylogeny in affecting microhabitat use by anurans, we rst looked for differences in microhabitat use among (a) species, (b) habitats, (lotic [running water: rivers] and lenthic [standing water: puddles, swamps, ponds] bodies of water), and (c) sites (larger spatial scale; see Fig. 1). Differences among habitats could result from differences among resident species populations and the particular species assemblage. Sites could also differ according to habitat availability, further inuencing the composition of species assemblages and overall patterns of microhabitat use. At every spatial level, species populations could also be under environmental selective pressures. Our aim was to show how much of the existing variability in microhabitat use could be explained by the species phylogeny.

Fig. 1 Map showing the location of the ve sites sampled for tadpoles and/or adult anurans in southeastern Brazil (see Table 1 for details)

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Materials and methods Study sites and sampling procedures We collected data on microhabitat use by anurans at ve sites in the state of Minas Gerais, nio Natural (RPPN) Santua rio do southeastern Brazil: the Reserva Particular do Patrimo Carac a, the Parque Estadual (PE) Serra do Brigadeiro, the Parque Nacional (PN) Serra da , and the highlands of Serra de Ouro Branco (Table 1; Canastra, the PN Serra do Cipo Fig. 1). We collected data over 14 years depending on the site (see Table 1 for details). The climate is seasonal at all sites, with the dry season extending from April to September and the wet season, from October to March. Data from some of these localities have already been used to study microhabitat selection, species distribution and niche based community structure (Eterovick 2003; Eterovick and Barros 2003; Eterovick and Barata 2006; Kopp and Eterovick 2006; Afonso and Eterovick 2007a; 2007b), but no attempts have yet been made to explain differences among species based on phylogenetic relatedness, which is addressed in this study using the whole data set for a more comprehensive approach. Differences in microhabitat use by anuran tadpoles and adults might occur among seasons, though sampling effort was always long and equivalent between dry and wet seasons at all sites (see Table 1), thereby minimizing seasonal interference in our comparisons among species and sites. Although some variation in microhabitat use by species may occur from year to year at a given site, we believe such variation would be mostly stochastic, except, maybe, in a year with very unusual weather, which was not the case during our sampling periods. So, we believe our sampling was appropriate to study specic patterns of microhabitat use, overcoming the inuences of year and season. Sampling methods were standardized among all sites: previously marked 150 m stream sections, swamps, and ponds were thoroughly searched and all individuals located were recorded during each sampling period. The types of bodies of water sampled varied among sites and this reects the existing availability of breeding habitats for anurans with a tadpole stage more than a sampling bias. All the bodies of water that could be assessed regularly to conduct the study were included. Nevertheless, sampled habitats do not cover the whole area of the parks or mountain ranges, which would represent an effort beyond the scope of a single study. Collection of data on microhabitat use Data were collected on anuran microhabitat use both in the larval and adult stages, encompassing a broad set of species and phylogenetic groups. Anuran species were sampled at ve different study sites in southeastern Brazil in order to also encompass a broad set of environmental conditions (and potentially different selective pressures). We hypothesized that, if microhabitat use is a conservative trait, more closely related species should have more similar microhabitat use. Alternatively, if closely related species, or populations of the same species at different sites, differed substantially in microhabitat use, then microhabitat use should be considered a plastic trait easily inuenced by local environmental pressures. Considering anuran spatial distribution, we described three spatial levels that we dene as microhabitats (specic places, occupied by an individual, with particular features within/by a body of water), habitats (bodies of water and their margins) and sites (different localities, each one including several bodies of water of varied types). Since habitats are closely related to reproductive modes (sensu Duellman and Trueb 1994) and have already

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Table 1 Description of the ve study sites sampled for tadpole and/or adult frog microhabitats PE Serra do Brigadeiro PN Serra da Canastra PN Serra do Cipo Serra de Ouro Branco

Site

rio do RPPN Santua Carac a 20310 21000 S, 42400 40200 W 1,0001,600 m State park 13,210 Cerrado Cerrado (montane meadows) 21.2 1,622 November 1998 February 2002 70,000 34,000 Cerrado (montane meadows), Atlantic forest 21.3 1,170 March 2004 August 2005 Adults: monthly Tadpoles and adults: monthly during the rainy season and every two months during the dry season 6 temporary ponds, 16 streams (9 permanent, 7 temporary) 4 5 6 17 National park National park None 9001,496 m 1,0001,485 m 20100 S, 46300 W 19120 19200 S, 43300 43400 W 20390 S, 43400 W 1,0001,568 m

Coordinates

20050 S, 43290 W

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Elevation

8502,070 m

Conservation unit

Private reserve

Area (ha)

10,188

Biomes

Cerrado, Atlantic forest Atlantic forest

Mean annual temperature (C) 1,300 April 2003September 2004 April 2004February 2006 1,250

21.4

20.5

23

Annual rainfall (mm)

1,150a

Sampling period

April 2003 October 2004

Sampling frequency

Tadpoles and adults: two Tadpoles and adults: monthly Tadpoles: monthly samplings during the during the rainy season and during the rainy rainy season and two every two months during the dry season and every two during the dry season season months during the dry season Adults: monthly 10 permanent streams 8 streams (7 permanent, 1 temporary)

Habitats sampled

3 ponds (2 temporary, 1 permanent), 8 permanent streams 9 1

4 permanent streams, 2 temporary swamps

Species (adults)

21

12

Species (tadpoles)

14

Elevations include the approximate range of the biomes studied in each site. Except for PE Serra do Brigadeiro, the highest limit corresponds to the highest elevation in the site. The numbers of species recorded as adults and tadpoles correspond to the species included in this study (with 4 or more individuals recorded) a rio do Carac Rainfall at the RPPN Santua a corresponds to data from the rst year of the study at this site 5

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been shown to be related to anuran phylogeny at the family level (Zimmerman and Simberloff 1996), our main focus was on the nest (microhabitat) level. We inspected each habitat (streams, ponds, swamps) once during the day and once at night each eld trip (see Table 1 for sampling schedules at each locality). We recorded features of microhabitats used by each individual tadpole and adult frog located during each eld trip to each study site. Microhabitat was considered as the exact point where the individual was located. We searched for tadpoles visually during the day and, occasionally, at night (when some species, not seen during the day, could be detected), and we characterized their microhabitats by recording six features that we observed to contribute to microhabitat heterogeneity within the bodies of water. These features were (1) water depth, (2) substrate type (silty/sandy or rocky), (3) presence or absence of current and (4) aquatic vegetation at the specic point where the tadpole was located, (5) the position of each tadpole in the water column (bottom or midwater/surface), as well as (6) the month when the record was obtained. Most of these features (14 and 6) have previously been used to characterize tadpole microhabitats and were useful in detecting microhabitat preferences by tadpole species (Eterovick and Barata 2006). Water clarity and caution allowed us to locate tadpoles before they attempted to escape. We inspected the parts of the bodies of water where visibility was poor (e.g., aquatic vegetation patches, shaded crevices) with dipnets. During sampling, we moved in an upstream direction to avoid sampling the same individual repeatedly in case tadpoles might be carried downstream by the water ow. For the most abundant species, not all tadpoles were collected, thus some individuals may have been sampled in two or more consecutive months. Nevertheless, the amount of pseudo-replication in our data would be very small in relation to our sample sizes. Only a few tadpoles were recorded at night, when not seen during the day. However, conspecics were also recorded during the day in other eld trips. Individuals of species recorded during the day were seen at night during the same eld trip in several instances (but not recorded then, to avoid pseudoreplication). Consequently, we did not attempt to classify species as diurnal or nocturnal. We identied tadpoles based on original descriptions, comparison with museum specimens, and we raised them until metamorphosis whenever necessary to conrm identication. Microhabitats used by adult anurans were recorded at night for nocturnal species and during the day for diurnal species. Most individuals recorded were calling males, ovigerous females (not amplexed, probably searching for males), or they were at the same bodies of water and fell within the adult size range for their species. Microhabitats recorded for adult frogs can be likely interpreted as microhabitats used during breeding activities. In our analyses, we looked for patterns of microhabitat use that might differ among species or populations, regardless of sex. For the microhabitat used by each individual, we recorded ve features: (1) the height above water or above ground, (2) the shortest horizontal distance to water, (3) the substrate type used (e.g., leaves, branches, soil, rocks), (4) the type of body of water (swamps/ponds or streams), and (5) the month when observation was made. We considered habitat (type of body of water) for adult anurans (but not for tadpoles) because they can choose it and such choice may be inuenced by the availability of appropriate microhabitats, whereas tadpoles can only choose microhabitats within the bodies of water where they are placed. We conducted all the analyses described in the next sections separately for tadpoles and adults. When the same species occurred at two or more of our study sites, we treated individuals of each site as from different populations, which were compared to assess the effects of site in microhabitat use. Individuals (tadpoles and adults) collected for identication purposes are deposited in the herpetological collection of the Museu de ncias Naturais of the Pontif cia Universidade Cato lica de Minas Gerais (MCNAM). Cie

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Variability in microhabitat use among species, habitats, and sites We rst explored our dataset to conrm the existence of variability in microhabitat use among species, habitats, and sites. We conducted Principal Components Analyses (PCAs) to condense the information on microhabitat use into fewer variables to be compared among species, habitats, and sites. We used individual data for each tadpole and adult frog recorded to conduct the PCAs. Except for water depth, all tadpole microhabitat variables were treated as categorical, and among the variables used for adult frogs, substrate and month were treated as categorical variables (for variable categorization, see the section Characterization of microhabitat use by larval and adult anurans). To conduct the PCA with variables describing adult frog microhabitats, we excluded two frogs that were outliers because we recorded them very far from water (one individual of Dendropsophus rio do Carac minutus recorded 70 m from water at the RPPN Santua a and one individual of Hypsiboas polytaenius recorded 30 m from water at Ouro Branco). There is no consensus on the validity of using both categorical and continuous variables in the same PCA, but large departures from the test original assumptions may compromise the use of PCA results in statistical inference (McGarigal et al. 2000). Linearity of variables is an important assumption of PCA, and we examined ordination graphs to be sure that our dataset met this assumption (points showing an arch effect are indicative of non-linearity; McGarigal et al. 2000). We just used the scores of the two-rst axes as dependent variables to quantitatively represent the most important features explaining variability in microhabitat use. The scores did not have normal distribution, indicating lack of multivariate normality, which can reduce information provided by later principal components with lower eigenvalues (McGarigal et al. 2000). We used Kruskal-Wallis and Mann Whitney tests to test for differences in the PCA scores describing microhabitats used by tadpoles and adult frogs between species, habitats, and sites. The independent variables tested for tadpoles were site and species, and for adults, site, habitat, and species. We did not include habitat for tadpoles, because we had no records of any species in both lenthic and lotic habitats, so that the effects of habitat and species could not be evaluated separately. Since our data did not have an ideal structure for PCA (McGarigal et al. 2000; McCune et al. 2002), we also conducted the same Kruskal-Wallis and Mann Whitney tests described above using multidimensional scaling (MDS), with Kruskal loss function, to reduce the original variables (instead of the PCAs), and compared the results to be sure that the PCAs were robust to non-normality of our variables. We conducted the analyses in Systat 12 (Systat Software Inc 2007). We are aware that species are not completely independent sampling units, as they share a common ancestry somewhere in evolutionary time, but it is necessary to rst detect the existence of extant differences in microhabitat use, and then subsequently analyze the inuence of such common ancestry (next sections). Assessment of phylogenetic relatedness among studied species In order to build a matrix of phylogenetic relatedness between all species pairs, we organized the species we sampled in all study sites according to their phylogenetic relationships (see Fig. 2), extracting their topology from the cladograms provided by Faivovich et al. (2005), Frost et al. (2006), Grant et al. (2006), and Nascimento et al. (2006). Three species, recorded in the tadpole stage (Bokermannohyla sp., Bokermannohyla sp. 2 and Leptodactylus sp.; see Appendix 1), were not included in the phylogeny because they could only be identied to the generic level. We could not use actual phylogenetic distances because they are not available

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8 Fig. 2 Phylogenetic relationships among the anuran species included in the study, based on data from Faivovich et al. (2005), Frost et al. (2006), Grant et al. (2006), and Nascimento et al. (2006). Tadpoles (T) and adults (A) were analyzed separately

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for all the species pairs considered and because we used information from several phylogenetic trees (Faivovich et al. 2005; Frost et al. 2006; Grant et al. 2006; Nascimento et al. 2006) which were estimated using different data. The combined topology we obtained was entered into the software Compare 4.6b (Martins 2004) which generated random branch lengths with 1,000 repetitions. We estimated branch lengths under Brownian motion

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evolution, with variable means and variances equal to zero (Martins 2004). This method was originally described by Felsenstein (1985) to describe the evolution of phenotypes under random genetic drift. We used the randomly generated branch lengths to build a relatedness matrix among extant species pairs using Compare 4.6b (Martins 2004). Characterization of microhabitat use by larval and adult anurans We aimed to investigate whether the most closely related species have the most similar patterns of microhabitat use, regardless of site. In order to describe and compare habitat/ microhabitat use by each species (and populations of the same species at different sites), we used two approaches to build dissimilarity matrices to be compared to the phylogenetic distance matrices for tadpoles and adults. We also used the second approach to build clusters of tadpoles and adults based on microhabitat use. We used the raw data for the subsequent analyses (and not PCA scores) because the number of variables was not limiting, and the PCA axes did not explain a high proportion of the existing variation (see results for Microhabitat use variability among species, habitats and sites). Additionally, the PCAs indicated that our data did not have multicollinearity problems. Our rst approach was to characterize species/populations based on the mean values of the variables we measured to describe the microhabitats they used and months when they were recorded. The variables used for tadpoles and adults are the ones described in the earlier section Collection of data on microhabitat use, which were recorded for each individual tadpole and adult frog. We calculated the mean value for each numeric and categorical microhabitat variable for each species/population (Appendices 1, 2). For categorical variables, categories were dened as follows: binary data received values of 0 or 1, respectively, for absence or presence of current and aquatic vegetation at a tadpole microhabitat, as well as position of a tadpole on the bottom or in the water column, the habitat type used by an adult frog (lenthic or lotic), and the type of substrate at a tadpole microhabitat (silty/sandy or rocky). The mean of the binary variables was intended to express to what extent each species/ population tended to use more of the state 0 or 1, as dened. Months received values from 0 to 11 starting at the onset of the rainy season, so that October corresponded to 0, November to 1, and so on. These numbers represent number of months after the onset of the rainy season, when the rst rains triggered the reproductive activities of most of the anuran species. We had a concern that extreme values given to months at the beginning of the rainy season (the lowest values) and at the end of the dry season (the highest values), if combined, would provide means similar to those of species occurring at months with intermediate values (end of rainy season and onset of dry season). Nevertheless, this problem did not happen, because no species was active exclusively at the end of the dry season and the onset of the rainy season, so that the highest mean values obtained for month of occurrence reected higher association with the dry season (adults that were active/tadpoles that were present yearround or mostly during the dry season), whereas the smallest values were found for species that were highly dependent on the rainy season to breed/develop (see Appendices 1, 2). Adult anuran microhabitat substrates were tentatively classied according to a humidity gradient and numbered from 0 to 5, in an attempt to represent this qualitative variable as an ecologically meaningful gradient. From the moistest substrate to the driest one, based on our eld observations, water corresponded to 0, moist soil to 1, leaf litter to 2, rocks to 3, leaves hanging on vegetation to 4, and branches to 5. The mean value of substrates used by adult frogs was supposed to indicate tendencies to occupy moist sites close to the ground (lower values) or drier sites on vegetation (higher values). We interpreted these values as rough measurements of substrate use, which is a difcult variable to quantify.

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Nevertheless, we examined the values obtained (see Appendix 2) and noticed that, under this approach, they were representative of our eld observations, that is, species/populations observed using substrates in a similar way got similar values and those that were very different got the most divergent values. We used this data set to build a dissimilarity matrix between species pairs using Euclidean distance measure (Quinn and Keough 2002). Our second approach was to classify microhabitats in types, based on possible combinations of categories of their measured variables, and compare species/populations based on the number of individuals using each microhabitat type. We used this approach because many of the variables we used to characterize microhabitats were categorical, and we wanted to be sure that using the mean values of those in the rst approach would not give misleading results. Our second approach was an easier way to deal with qualitative variables, though information on temporal distribution could not be included. Nevertheless, if some microhabitat features occurred in non-random combinations in the habitats studied, this method would be more appropriate to compare specic patterns of microhabitat use. Tadpole microhabitats were classied in 24 (3 9 2 9 2 9 2) types, resulting from the combination of three depth classes (015, [1530, and [30 cm), current presence or absence, vegetation presence or absence, and two types of substrate (silty/sandy or rocky). Adult anuran microhabitats were classied in 23 types, which were either water (1), or one of two ground substrates (bare soil or leaf litter) close (01 m) or far ([1 m) from water (2 9 2), or one of three types of perching substrates (rocks, leaves or branches/trunks) close or far from water and at three different height classes (070, [70140, [140 cm) (3 9 2 9 3). Adult microhabitat types could be either in lotic or lenthic habitats, so the number of categories in the matrix was 46 (23 9 2). We pooled data from all months. We used this data set to build a dissimilarity matrix between species pairs using BrayCurtis distance measure (Quinn and Keough 2002), and also to conduct a cluster analysis (Bray Curtis distance measure, group average linkage method; McCune and Mefford 1999). We compared the matrix of phylogenetic distances (previous section) with each one of the two dissimilarity matrices generated with data on microhabitat variable means and microhabitat type use frequency, respectively, using Mantel tests. We conducted these analyses for both tadpoles and adult frogs. Three species, recorded in the tadpole stage (Bokermannohyla sp., Bokermannohyla sp. 2 and Leptodactylus sp.; see Appendix 1), were not included. For these analyses, pairs of populations of the same species (at different localities) were scored 0 in the phylogenetic distance matrix. We conducted Monte Carlo randomization tests (5,000 runs) to test for signicance. We conducted all the analyses described in this section using the software PcOrd (McCune and Mefford 1999).

Results Collection of data on microhabitat use We obtained a total of 4,230 records of individual tadpoles of 34 species in 15 genera and 7 families (Bufonidae, Cycloramphidae, Hylidae, Hylodidae, Leiuperidae, Leptodactylidae, and Microhylidae) at the study sites (Appendix 1). These include only species with four or more individuals observed. We obtained a total of 1,163 records of adult individuals of 39 species distributed in 16 genera and 8 families (Bufonidae, Centrolenidae, Cycloramphidae, Hylidae, Hylodidae, Leiuperidae, Leptodactylidae, and Microhylidae) (Appendix 2). These include only species with more than four individuals observed.

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Variability in microhabitat use among species, habitats, and sites The rst axis of the PCA conducted with variables describing tadpole microhabitats was positively associated with substrate and negatively associated with vegetation and position in the water column (we call it substrate ? vegetation ? position from here on), and the second axis was positively associated with water depth and month of occurrence (we call it depth ? month from here on). Current did not contribute much to explaining tadpole variability in microhabitat use (Table 2; Fig. 3a). Site had a signicant inuence on both substrate ? vegetation ? position (H = 1510.90, P \ 0.001) and depth ? month (H = 92.72, P \ 0.001), as did species (substrate ? vegetation ? position: H = 3105.04, P \ 0.001; depth ? month: H = 578.22, P \ 0.001). The results we obtained using the MDS were all equivalent (results not shown). Since the rst two axes of the PCA explained only 56.69% of the variability for tadpole microhabitats (Table 2), we also conducted Mann Whitney and Kruskal-Wallis tests for each microhabitat variable separately, and all of them were signicantly different among species and sites (results not shown). The rst axis of the PCA conducted with variables describing adult frog microhabitats was positively associated with substrate and height above ground/water (we call it substrate ? height from here on), and the second axis was positively associated with distance from water and negatively associated with month of occurrence (we call it distance ? month from here on) (Table 2; Fig. 3b). Site had a signicant inuence in both substrate ? height (H = 91.59, P \ 0.001) and distance ? month (H = 28.09, P \ 0.001), as did habitat (substrate ? height: U = 67,046.00, P \ 0.001; distance ? month: U = 218,140.00, P \ 0.001), and species (substrate ? height: H = 609.75, P \ 0.001; distance ? month: H = 440.29, P \ 0.001). The results we obtained using the MDS were all equivalent (results not shown). Since the rst two axes of the PCA explained only 63.62% of the variability for adult microhabitats (Table 2), we also conducted Mann Whitney and Kruskal-Wallis tests for each microhabitat variable separately, and all of them were signicantly different among species, habitats, and sites (results not shown).
Table 2 Component loadings, eigenvalues and percent of total variance explained by the rst two dimensions of the PCAs based on variables describing microhabitats used by tadpoles and adult anurans

Axis 1 Variablestadpoles Vegetation Substrate Position Depth Month Current Eigenvalue Percent of total variance explained Variablesadults Substrate Height Distance to water Month Eigenvalue 0.833 0.755 0.230 0.418 1.492 37.31 20.835 0.736 20.717 -0.379 0.218 0.409 2.112 35.20

Axis 2

-0.213 0.232 0.344 0.709 0.755 0.005 1.289 21.49 -0.007 0.128 0.784 20.649 1.052 26.31

The variables that contributed the most to each axis are in boldface

Percent of total variance explained

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Fig. 3 PCA ordination graphs showing data distribution (left) and variables (right) in relation to the rst two axes of multidimensional space describing variables related to microhabitats used by (a) tadpoles and (b) adult anurans at sites in southeastern Brazil. For variable component loadings, see Table 2

We believe there were no multicollinearity problems in tadpole and adult anuran microhabitat variables. The two variables with the highest correlation (r = 0.604 for height and substrate for adult frogs; see Fig. 3b) were still bellow the level considered to suggest potential multicollinearity problems (r [ 0.7) that might require elimination of offending variables (McGarigal et al. 2000). Inuence of phylogenetic relatedness on habitat/microhabitat use The Mantel tests conducted for tadpoles were not signicant in any of the approaches employed (r = 0.018, P = 0.326 based on microhabitat variable means, and r = 0.003, P = 0.486 based on use frequency of microhabitat types), indicating that patterns of microhabitat use do not reect phylogenetic relatedness. In the case of adult anurans, the Mantel tests were signicant in both approaches used (r = 0.074, P = 0.008 based on microhabitat variable means, and r = 0.055, P = 0.0002 based on use frequency of microhabitat types). Nevertheless, the relationship was very weak, as reected by the low value of the standardized Mantel statistic (r). The equivalent results obtained in the two approaches used to characterize microhabitat use (microhabitat variable means and microhabitat type use frequency) indicate that both methods are valid for detecting differences among species and habitats/sites.

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The clustering of tadpoles according to similarity in microhabitat use showed that neither phylogeny nor sites were good predictors of microhabitat use by species. The species pairs that were the most similar considering microhabitat use were never the ones most closely related phylogenetically (Figs. 2, 4). Besides, even populations of the same species were very different in microhabitat use from site to site (see Scinax curicica and RPPN Santua rio do Pugliese, Pombal and Sazima, 2004 from PN Serra do Cipo Carac a, for instance; Fig. 4). In the adult stage, some related species could be noticed to be similar in microhabitat use, such as Scinax curicica, S. aff. perereca, and S. squalirostris rio do Carac (Lutz 1925) from RPPN Santua a, Bokermannohyla nanuzae (Bokermann and rio do Carac Sazima 1973) from RPPN Santua a and Bokermannohyla sp. from PN Serra da Canastra, Scinax gr. catharinae from PE Serra do Brigadeiro and S. luizotavioi (Caram rio do Carac aschi & Kisteumacher 1989) from RPPN Santua a, for instance (Figs. 2, 5). But, still, even populations of the same species might differ in adult microhabitat use from rio do Carac site to site, such as Dendropsophus minutus (Peters 1872) from RPPN Santua a, Serra de Ouro Branco, and PN Serra da Canastra (Fig. 5). Discussion As expected, microhabitats used by tadpoles and adult frogs were found to differ among species, habitats, and sites. Differences among habitats and sites may reect different

Fig. 4 Cluster analysis (BrayCurtis distances, group average linkage method) based on use frequency of 24 microhabitat types by tadpoles of 31 different species at four sites in southeastern Brazil. SC = PN Serra , CR = RPPN Santua rio do Carac do Cipo a, CN = PN Serra da Canastra, BG = PE Serra do Brigadeiro

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Fig. 5 Cluster analysis (BrayCurtis distances, group average linkage method) based on use frequency of 23 microhabitat types by adult anurans of 39 different species at ve sites in southeastern Brazil. SC = PN , CR = RPPN Santua rio do Carac Serra do Cipo a, CN = PN Serra da Canastra, BG = PE Serra do Brigadeiro, OB = Serra de Ouro Branco

anuran assemblages. Even populations of the same species, observed at different sites, showed noticeable differences in their pattern of microhabitat use, indicating that habitat and site also play a role in inuencing microhabitat use by tadpoles and adult anurans. Otherwise, the co-existence with a different set of species in each local assemblage may also inuence microhabitat use. Availability of the preferred microhabitat types may be an important factor, but species interactions and physical properties of each habitat may also inuence microhabitat choice during anurans lives. Phylogenetic relatedness explained very little of the variation in microhabitat use among adult frogs, and had no relationship with microhabitat use by tadpoles. The combined effects of specic preferences, convergence promoted by environmental conditions, and species interactions might produce patterns of microhabitat use by tadpoles and adult frogs that cannot be explained by a single factor in southeastern Brazil. Silvertown et al. (2001),

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studying meadow plant communities in the United Kingdom, found similar results: taxonomic afliation was not useful in predicting ecological features of plant species, and niche segregation could occur between species closely related or not, with no apparent patterns. Traits that lack either convergence or strong phylogenetic signals may respond to multiple adaptive peaks (Richardson 2001), and in the neotropics, the greater species diversity and habitat heterogeneity may create a higher diversity of potential adaptive peaks that make phylogenetic signal and contemporary inuences even more difcult to uncover for a particular labile trait. A weak phylogenetic signal may also result from niche segregation among related species. Losos et al. (2003) found an Anolis lizard community in Cuba to be nonrandomly structured, with species differing in ecological traits, including traits related to microhabitat use, such as perch height and diameter, as well as activity time. Phylogenetic relatedness explained less than 4% of the variation in the traits considered (Losos et al. 2003). Previous studies based on parts of the dataset analyzed in the present work, though, showed that anuran communities are not strongly structured in southeastern Brazil either in the larval (Eterovick and Barros 2003) or the adult stage (Afonso and Eterovick 2007a) considering microhabitat use. In anurans, particular components of reproductive behavior can show similarities (convergence) among different groups (Rabb 1973), like we observed in our study considering microhabitat use by calling males and females at breeding sites. On the other hand, several phenomena may prevent homologies from being expressed in the behavior of related taxa, rendering the association of phylogeny and ecological traits difcult. Species groups that show evolutionary plasticity may radiate into new adaptive zones. Frogs can occupy new habitats once they are able to exploit novel reproductive modes and patterns of microhabitat use (Andreone et al. 2005). For instance, recurrent evolution of similar ecomorphs may have occurred several times in microhylid frogs from Madagascar, including many transitions from arboreality to terrestriality as an adaptation to highaltitude habitats (Andreone et al. 2005). Ecological factors seem to play an important role in determining tadpole distribution within and among habitats. Trade-offs experienced by tadpoles in their natural habitats can inuence distribution, for instance, preventing the establishment of species with low ability to maximize growth in habitats with limited resources (Schiesari et al. 2006). In certain circumstances, a high predation pressure may confer selective advantage to the use of microhabitats that reduce predation risk (Kopp et al. 2006). Even if patterns of microhabitat use by tadpoles were not useful in recovering phylogenetic signal, morphological traits associated with ecomorphological types (Altig and Johnston 1989) remain to be tested for phylogeny recovery. Behavioral traits may evolve faster than underlying morphological traits, so that morphology may reveal phylogenetic patterns that are not behaviorally expressed. Poe (2005) successfully used morphometric characters of Anolis lizard ecomorphs to track phylogenetic relatedness even when morphologically similar species occupied different niches. Whereas some tadpole ecomorphs are highly differentiated morphologically and behaviorally, such as the semi-terrestrial Thoropa tadpoles, others can be morphologically similar (or even belong to the same species) and occupy microhabitats with varied features. Some features related to microhabitat use may be more closely associated to tadpole morphology than others, such as position in the water column and current presence or absence (see Altig and Johnston 1989; Altig and McDiarmid 1999). Other features may not be constrained by tadpole morphology. For instance, water depth may relate to temperature, which also inuences microhabitat selection by tadpoles (Ultsch et al. 1999), whereas month of occurrence depends on the reproductive patterns of adult frogs (Alford 1999).

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When Eterovick and Fernandes (2001) reported some consistency in microhabitat use by tadpoles within representatives of the family Leptodactylidae, studies on amphibian phylogeny were scarce and this family was already suspected to be paraphyletic (Zug et al. 2001). The new amphibian phylogeny proposed by Frost et al. (2006) and Grant et al. (2006) shed some light on the subject and rendered a new interpretation of the data presented: the species reported by Eterovick and Fernandes (2001) to use microhabitats in a similar way belong to two different monophyletic families: Cycloramphidae and Leptodactylidae, and so represent cases of convergence. On the other hand, we found Hylidae to show relatively high variation and diversication in microhabitat use, corroborating the results of Eterovick and Fernandes (2001). In the case of adult anurans, we were able to nd a signicant relationship between phylogenetic relatedness and patterns of microhabitat use, but still the amount of variation explained by phylogenetic relatedness was very small. Adult anurans may have more control over microhabitat use than tadpoles, given that the latter are restricted to the body of water where they are placed and the microhabitats it contains, whereas adults can migrate from one body of water to another. Besides, if a tadpole does not manage to survive by being unable to use the most suitable microhabitats at a given site, it will leave no descendants at all. On the other hand, an adult that is unable to use the appropriate microhabitats and is preyed upon or prevented from remaining at a given habitat by competitors, may still have already left some offspring or can migrate to another habitat, respectively. Our results suggest that local selective pressures may be harsher on molding microhabitat use by tadpoles, to the point of completely overcoming phylogenetic signal. It is possible to consider microhabitat use as a very plastic behavioral trait, as corroborated by our records of the same species showing different patterns of microhabitat use at different sites. Our results show that microhabitat use, both by adult anurans and tadpoles, cannot be predicted by site or species relatedness alone. Microhabitat use in anurans is highly variable and thus is not useful for phylogenetic inferences at most levels. Nonetheless, it may respond relatively rapidly to local selective pressures imposed by other species or the physical environment, constituting an interesting trait to be explored in studies focusing on natural selection and adaptation. In this context, it would be possible to correlate microhabitat traits with possible inuencing variables without taking phylogeny into account. Characterizing these variables for particular lineages is an appealing subject for future studies.
Acknowledgments We are grateful to A. S. Castro, C. A. N. Ventura, F. S. F. Leite, F. F. R. de Oliveira, and P. H. C. de Souza for help during eld work, to G. Andrade, D. C. Rossa-Feres, M. D. Boone, D. C. Adams, L. K. Erdtmann, E. R. Wild and two anonymous reviewers for helpful suggestions in the manuscript, to E. R. Wild for English revision, to D. L. Viana for making the map, to C. Paganini for permit and logistics rio do Carac to work at the RPPN Santua a, to the Instituto Nacional de Florestas (IEF) for permit and logistics to work at the PE Serra do Brigadeiro, to the Instituto Brasileiro do Meio Ambiente e dos Recursos veis (Ibama) for collecting permits (111/98, 128/2004, 046/05, 288/2005), and to the Fundo Naturais Renova ` Pesquisa (FIP) of the Pontif cia Universidade Cato lica de Minas Gerais (PUC-Minas) and de Incentivo a ` Pesquisa do Estado de Minas Gerais (Fapemig CRA 405/03) for nancial support. o de Amparo a Fundac a We dedicate this work to Dr. Wolney Lobato for his support and incentive to research at PUC-Minas.

Appendix 1 See Table 3

123

Table 3 Description of microhabitats used by larvae of anuran species at four sites in southeastern Brazil Depth (cm) Vegetation Current Substrate Position Month n

Species

Site

Family Bufonidae 12.0 8.8 (730) 25.4 20.6 (275) 14.2 9.7 (530) 11.5 4.1 (619) 15.0 7.3 (625) 0.2 0.9 (04) 0.00 0.00 1.00 0.00 0.94 0.24 0.11 0.32 0.16 0.38 0.95 0.23 0.00 0.00 0.00 0.00 0.25 0.46 0.00 0.00 0.25 0.46 0.00 0.00 0.07 0.27 0.00 0.00 0.92 0.27 0.00 0.00 0.00 0.00 0.00 0.00 0.51 0.50 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.17 0.41 0.8 0.4 (01) 6.6 3.2 (19) 1.0 0.0 (11) 4.3 1.4 (25) 3.1 1.0 (26) 2.0 0.0 (02) 6 111 195 8 19 18

Rhinella pombali (Baldissera, Caramaschi and Haddad 2004)

CR

Evol Ecol (2010) 24:124

R. rubescens (Lutz 1925)

SC

CN

Family Cycloramphidae

ril Odontophrynus americanus (Dume and Bibron 1841)

SC

Proceratophrys cururu Eterovick and Sazima 1998

SC

Thoropa megatympanum Caramaschi and Sazima 1984 20.3 9.0 (842) 17.3 8.8 (240) 24.1 13.1 (0.5100) 23.1 18.7 (693) 29.3 16.3 (580) 34.3 20.6 (5120) 30.9 14.1 (580) 27.6 3.6 (2032) 13.3 4.7 (823) 19.3 15.3 (440) 11.2 2.2 (412) 41.4 30.2 (473) 0.96 0.19 1.00 0.00 0.00 0.00 0.75 0.5 0.00 0.00 1.00 0.00 0.25 0.44 0.17 0.37 0.07 0.26 0.17 0.38 0.45 0.50 0.11 0.31 0.00 0.00 0.15 0.36 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.06 0.23 0.47 0.50 0.67 0.49 0.00 0.00 0.89 0.32 0.51 0.51 0.52 0.50 0.89 0.31 0.80 0.40 0.72 0.45 0.02 0.15 0.00 0.00 0.00 0.00 0.25 0.5 0.00 0.00 0.00 0.00

SC

Family Hylidae 0.00 0.00 0.00 0.00 0.03 0.17 0.00 0.00 0.01 0.09 0.03 0.18 0.93 0.26 0.72 0.46 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 3.6 0.9 (25) 3.7 3.1 (110) 3.6 4.1 (010) 5.5 2.2 (19) 6.3 2.6 (18) 5.2 2.8 (18) 2.0 2.4 (08) 2.0 0.0 (22) 7.9 1.1 (79) 4.5 3.1 (29) 9.0 0.0 (99) 7.1 1.1 (68) 18 55 355 56 138 117 388 25 9 4 23 7 17

Bokermannohyla alvarengai (Bokermann 1956)

SC

B. gr. circumdata

CR

B. martinsi (Bokermann 1964)

CR

B. saxicola (Bokermann 1964)

SC

Bokermannohyla sp.

CN

Bokermannohyla sp. 2

CN

Dendropsophus minutus (Peters 1872)

CR

D. seniculus (Cope 1868)

CR

Hypsiboas albopunctatus (Spix 1824)

SC

H. cipoensis (Lutz 1968)

SC

H. lundii (Burmeister 1856)

SC

123

H. polytaenius (Cope 1868)

CR

18

Table 3 continued
Depth (cm) 45.5 26.6 (1190) 23.4 10.2 (551) 27.2 15.1 (370) 20.5 11.1 (360) 21.1 9.6 (441) 28.3 17.1 (670) 75.3 39.4 (20130) 30.0 15.9 (277) 30.0 18.0 (579) 47.3 17.8 (1068) 14.4 6.4 (436) 12.9 9.8 (150) 0.17 0.37 0.25 0.44 0.07 0.26 0.43 0.50 1.00 0.00 0.00 0.00 0.99 0.08 0.00 0.00 0.02 0.14 0.03 0.17 0.02 0.15 0.17 0.38 0.10 0.30 0.32 0.47 0.25 0.43 0.00 0.00 0.00 0.00 1.00 0.00 0.97 0.16 0.00 0.00 0.01 0.08 0.67 0.47 0.00 0.00 0.44 0.50 0.38 0.49 0.41 0.49 0.00 0.00 0.04 0.21 0.20 0.40 0.51 0.50 0.00 0.00 0.02 0.13 0.21 0.41 0.22 0.41 0.60 0.49 0.01 0.08 0.07 0.25 0.25 0.43 0.97 0.18 0.00 0.00 0.00 0.00 0.13 0.35 0.33 0.48 0.87 0.34 3.5 1.2 (15) 2.4 2.3 (18) 5.8 4.5 (010) 7.7 1.9 (39) 3.1 2.8 (09) 5.5 2.3 (39) 4.8 4.1 (010) 3.1 3.2 (09) 3.5 1.2 (17) 6.5 4.0 (010) 5.1 4.1 (010) 0.33 0.52 0.00 0.00 0.33 0.52 1.00 0.00 4.3 2.7 (29) Vegetation Current Substrate Position Month n 6 31 93 344 34 1045 23 160 571 69 42 127

123
21.0 9.7 (740) 13.1 5.7 (119) 15.3 8.4 (441) 7.9 4.0 (417) 0.00 0.00 0.1 0.31 0.4 0.5 0.70 0.47 0.00 0.00 0.08 0.29 0.03 0.18 0.00 0.00 0.00 0.00 0.44 0.51 0.73 0.45 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 6.2 1.4 (49) 2.0 0.8 (13) 1.5 0.9 (14) 3.6 1.4 (15) 27 23 30 10

Species

Site

Phasmahyla jandaia (Bokermann and Sazima 1978)

SC

Phyllomedusa megacephala (Miranda-Ribeiro 1926)

SC

Scinax canastrensis Cardoso and Haddad 1982

CN

S. gr. catharinae

BG

S. curicica Pugliese, Pombal and Sazima 2004

SC

CR

S. machadoi (Bokermann and Sazima 1973)

SC

CR

S. aff. perereca

CR

S. squalirostris (Lutz 1925)

CR

Family Hylodidae

Crossodactylus bokermanni Caramaschi and Sazima 1985

CR

Hylodes uai Nascimento, Pombal and Haddad 2001

CR

Family Leiuperidae

Physalaemus aff. olfersii

CR

Family Leptodactylidae

Leptodactylus camaquara Sazima and Bokermann 1978

SC

L. cunicularius Sazima and Bokermann 1978

SC

Evol Ecol (2010) 24:124

L. furnarius Sazima and Bokermann 1978

SC

Table 3 continued
Depth (cm) 11.3 5.3 (420) 11.6 7.3 (424) 8.8 3.8 (413) 1.00 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.21 0.43 0.57 0.51 1.00 0.00 0.00 0.00 1.2 0.4 (12) 2.0 0.0 (22) 0.19 0.40 0.28 0.46 0.43 0.51 0.00 0.00 1.7 0.9 (14) Vegetation Current Substrate Position Month n 21 14 8

Species

Site

L. syphax Bokermann 1969

SC

Leptodactylus sp.

CN

Family Microhylidae

Evol Ecol (2010) 24:124

Elachistocleis ovalis (Schneider 1799)

CR

Values presented are mean SD (range). Month number of months after the onset of the rainy season (October). Qualitative variables were scored as detailed in the text. SC , CR RPPN Santua rio do Carac Serra do Cipo a, CN Parque Nacional da Serra da Canastra, BG Parque Estadual da Serra do Brigadeiro

19

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Appendix 2

123
Height (m) Habitat (lenthic/ lotic) Distance from water (m) Month n 0.14 0.36 0.36 0.52 1.00 0.00 1.00 0.00 1.00 0.00 1.00 0.00 1.00 0.00 1.00 0.00 1.00 0.00 1.00 0.00 1.00 0.00 1.00 0.00 0.90 0.32 0.05 0.09 (00.35) 0.08 0.18 (00.5) 0.60 0.95 (02) 0.50 0.00 (0.50.5) 0.35 0.47 (01) 0.23 0.38 (01.2) 2.21 1.37 (04) 1.50 1.00 (13) 1.12 3.61 (015) 0.32 0.52 (02.5) 0.18 0.35 (00.7) 1.68 1.75 (0.35) 0.72 1.26 (04) 1.08 2.71 (010) 7.89 2.66 (011) 6.13 4.70 (011) 1.50 0.58 (12) 0.00 0.00 (00) 1.5 0.58 (12) 3.19 3.39 (08) 6.04 4.59 (111) 11.00 0.00 (1111) 6.00 4.18 (011) 6.54 3.94 (011) 4.00 3.56 (19) 6.50 4.93 (211) 5.70 4.00 (211) 5.77 3.56 (111) 28 8 4 5 4 16 24 4 17 98 4 6 10 13 0.00 0.00 (00) 1.75 1.04 (0.53) 3.00 0.00 (33) 1.75 0.91 (0.93) 0.78 0.79 (03) 1.16 0.50 (02) 0.88 0.25 (0.51) 0.87 0.74 (02) 0.77 0.70 (03) 0.85 1.14 (02.4) 2.67 1.63 (15) 1.23 1.42 (0.025) 3.75 0.96 (35) 1.38 0.64 (0.52.26) 1.00 0.00

See Table 4

Table 4 Description of microhabitats used by adult individuals of anuran species at ve sites in southeastern Brazil

Species

Site Substrate

Family Bufonidae

Rhinella pombali

CR 0.61 0.57 (02) 0.002 0.01 (00.05)

R. rubescens

CR 0.75 1.39 (03)

Family Centrolenidae

Hyalinobatrachium uranoscopum ller 1924) (Mu

OB 4.00 0.00 (44)

BG 4.00 0.00 (44)

Hyalinobatrachium sp.

BG 4.75 0.50 (45)

Family Cycloramphidae

Thoropa miliaris (Spix 1824)

BG 3.00 0.00 (33)

Family Hylidae

Aplastodiscus arildae (Cruz and Peixoto 1987)

BG 4.29 0.69 (25)

A. leucopygius (Cruz and Peixoto 1985)

BG 4.25 0.50 (45)

Bokermannohyla martinsi

CR 4.12 0.99 (35)

B. nanuzae (Bokermann and Sazima 1973)

CR 4.12 0.93 (15)

B. saxicola

SC

B. gr. circumdata

OB 4.50 0.55 (45)

CR 4.70 0.48 (45)

Evol Ecol (2010) 24:124

BG 4.54 0.52 (45)

Table 4 continued
Height (m) Habitat (lenthic/ lotic) 1.00 0.00 1.00 0.00 0.05 0.10 (02) 1.10 7.00 (070) 1.26 1.40 (04) 0.41 0.71 (02) 0.21 0.33 (01) 1.50 1.26 (03.5) 2.18 1.35 (06) 0.00 0.00 (00) 0.09 0.26 (01) 0.37 0.64 (01.5) 2.87 8.14 (030) 1.73 1.36 (05) 0.19 0.20 (00.5) 3.50 4.36 (110) 1.97 1.81 (06) 0.00 0.00 0.75 0.50 0.92 0.28 0.00 0.00 0.00 0.00 0.37 0.70 (02) 0.00 0.00 (00) 0.18 0.50 (03) 2.08 2.52 (0.210) 0.08 0.10 (00.2) 1.52 1.51 (16) 0.13 0.34 0.00 0.00 0.81 0.41 1.00 0.00 0.43 0.54 1.00 0.00 0.00 0.00 0.33 0.49 1.00 0.00 0.88 1.81 (06) 0.95 0.98 (03) 1.67 0.49 (12) 1.54 0.52 (12) 1.25 0.96 (02) 3.53 2.93 (011) 1.92 0.28 (12) 3.45 1.04 (36) 2.68 2.11 (18) 3.43 1.72 (16) 6.27 1.80 (18) 1.20 0.84 (02) 1.72 1.57 (05) 4.40 5.60 (011) 6.44 3.33 (111) 4.19 3.34 (011) 3.86 4.91 (011) 1.00 1.16 (02) 3.38 1.71 (07) 1.27 0.47 (12) 9.50 1.73 (811) 7.04 2.56 (011) 2.00 1.35 (06) 3.00 1.16 (24) 1.62 1.28 (06) Distance from water (m) Month n

Species

Site Substrate

Bokermannohyla sp. 0.50 0.39 (01) 0.11 0.06 (0.050.2) 0.00 0.00 0.15 0.26 (01.5) 0.33 0.59 (02) 0.40 0.27 (01) 0.31 0.47 (01.5) 1.25 0.96 (0.22.5) 0.51 0.47 (0.052) 0.05 0.07 (00.15) 0.21 0.41 (01.1) 0.61 0.89 (02) 0.27 0.07 (0.050.3) 0.00 0.00 0.85 0.75 (0.052.5) 0.62 0.50 2.16 1.02 (14) 1.93 0.83 (13) 0.46 0.41 (01.7) 0.00 0.00 (00) 0.29 0.14 (0.20.5) 0.40 0.44 (03) 0.37 0.38 (01.5) 0.29 0.34 (02) 1.00 0.00 0.25 0.50 0.00 0.00

CN 4.26 0.88 (35)

0.77 0.69 (02)

15 11 4 102 13 11 19 7 40 5 18 5 16 21 7 4 42 11 4 131 24 4 34

Dendropsophus minutus

CN 4.00 1.41 (05)

Evol Ecol (2010) 24:124

OB 4.25 0.50 (45)

CR 3.01 1.77 (05) 4.45 1.21 (15) 4.57 0.54 (45)

D. seniculus

CR 2.39 2.02 (05)

Hypsiboas albopunctatus

SC

CN 2.21 1.75 (05)

H. cipoensis

SC

H. aff. cipoensis

CN 4.27 0.45 (45)

H. faber (Wied-Neuwied 1821)

OB 2.00 2.74 (05)

CR 1.11 2.03 (05)

H. pardalis (Spix 1824)

BG 3.80 1.10 (35)

H. polytaenius

OB 4.81 0.40 (45)

CR 3.86 1.28 (15)

Phasmahyla sp.

OB 4.29 0.49 (45)

Phyllomedusa burmeisteri Boulenger 1882

CR 4.75 0.50 (45)

Scinax curicica

CR 3.62 1.67 (05)

S. eurydice (Bokermann 1968)

CR 0.73 0.47 (01)

S. luizotavioi (Caramaschi and Kisteumacher 1989) 4.00 0.00 (44)

OB 4.50 0.58 (45)

CR 4.48 0.60 (15)

S. aff. perereca

CR 4.00 1.02 (15)

S. squalirostris

SC

0.31 0.34 (0.010.6) 0.00 0.00

CR 3.94 0.55 (15)

21

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Table 4 continued
Height (m) Habitat (lenthic/ lotic) 0.05 0.13 (00.5) 0.20 0.50 (04) 1.88 2.77 (0.26) 0.09 0.38 (04) 0.00 0.00 0.00 0.00 (00) 0.39 1.05 (04.7) 1.25 0.96 (02) 1.84 2.26 (09) 0.25 0.50 (01) 0.00 0.00 (00) 0.15 0.07 (0.10.2) 0.00 0.00 (00) 0.00 0.00 0.00 0.00 (00) 8.47 2.38 (111) 5.28 3.83 (010) 2.00 0.00 (22) 7.16 3.75 (011) 6.25 0.71 (57) 0.83 0.72 (02) 1.89 1.72 (08) 1.50 0.58 (08) 2.32 0.98 (15) 7.50 1.00 (79) 1.85 0.37 (12) 1.50 0.58 (12) 0.5 0.58 (01) 1.40 0.55 (12) Distance from water (m) Month n

123
0.54 0.33 (0.021.5) 0.94 0.24 0.63 0.47 (02) 0.98 0.73 (0.42) 0.08 0.24 (01.5) 0.48 0.45 (01.5) 0.00 0.00 0.05 0.14 (00.6) 0.15 0.19 (00.4) 0.03 0.07 (00.3) 0.00 0.00 (00) 0.01 0.02 (00.06) 0.00 0.00 (00) 0.00 0.00 (00) 0.00 0.00 (00) 0.50 0.58 0.00 0.00 0.00 0.00 1.00 0.00 0.03 0.18 0.00 0.00 0.00 0.00 0.00 0.00 1.00 0.00 1.00 0.00 0.50 0.58 1.00 0.00 17 120 4 114 8 12 27 4 31 4 20 4 4 5 0.75 1.50 (03) 1.00 0.00 (11)

Species

Site Substrate

S. gr. catharinae

OB 4.53 0.51 (45)

BG 4.16 0.87 (35)

S. gr. ruber

OB 4.00 0.00 (44)

Family Hylodidae

Crossodactylus bokermanni

CR 2.93 1.03 (05)

Hylodes uai

CR 1.25 0.71 (13)

Family Leiuperidae

Physalaemus cuvieri Fitzinger 1826

OB 0.75 0.45 (01)

CR 0.93 1.54 (04)

P. evangelistai Bokermann 1967

CR 3.25 2.22 (05)

P. aff. olfersii

CR 1.45 1.12 (15)

Pseudopaludicola mineira Lobo 1994

SC

P. saltica Cope 1887

OB 1.30 0.92 (14)

Family Leptodactylidae

Leptodactylus jolyi Sazima and Bokermann 1978

SC

L. ocellatus (Linnaeus 1758)

OB 0.00 0.00 (00)

Family Microhylidae

Elachistocleis ovalis

CR 0.00 0.00 (00)

Evol Ecol (2010) 24:124

Values presented are mean SD (range). Month number of months after the onset of the rainy season (October). Qualitative variables were scored as detailed in the text. OB Serra de Ouro Branco. The other sites are abbreviated as in Appendix 1

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