Ecomorphology and functional diversity of generalist and specialist bird assemblages in Amazonian white-sand ecosystem habitat patches

ABSTRACT Birds that inhabit white-sand ecosystems (WSE) in the Amazon are adapted to habitats distributed as isolated patches. These environments occur in sandy soils that are extremely poor in nutrients, have low floristic diversity and support bird assemblages restricted to WSE. We investigated whether bird species specialized in WSE have morphological or ecological traits that distinguish them from generalist birds that share the same habitat but are not restricted to WSE. We collected morphological and ecological data from 22 specialist and 102 generalist bird species from WSE and described their ecomorphological diversity using multivariate analyses and measures of functional diversity. Understory insectivorous species that move alone or in pairs, were the group with the most species among specialist birds from WSE. In contrast, canopy frugivorous species that form mono- or heterospecific groups were more frequent among generalist species. Specialist and generalist birds overlapped extensively in ecomorphological space, with specialists occupying a narrower space compared to generalists. Functional diversity, however, was not different between specialist and generalist species when controlling for the number of species within the communities. Further studies comparing the ecomorphological diversity of bird assemblages of other environments in the Amazon may highlight the ecological pressures leading to the functional diversity of specialist species in WSE observed in this study.


INTRODUCTION
Evolutionary ecologists have highlighted the study of species functions within ecosystems to connect nichebased mechanisms to assemblage patterns (Cadotte 2017;Cadotte and Tucker 2017).Species functional attributes can link population processes to environmental gradients or interactions dictating species coexistence in nature (Pigot et al. 2016;Cadotte 2017).Natural environments limit the distribution of animals and plants according to their ecological and morphological adaptations, which can be represented by their functional space (Tilman et al. 1997;Cianciaruso et al. 2009).Thus, variations in ecomorphological attributes selected through environmental filters can influence the species that coexist in biological assemblages (Petchey and Gaston 2002;Devictor et al. 2010;Clavel et al. 2011;Braga et al. 2022).
Environments with stable conditions of resource availability tend to support more diverse assemblages and require fewer specialized adaptations (Grant 1968;Cianciaruso et al. 2017).In contrast, environments with severe ecological conditions may require specific adaptations for survival, restricting the number of species that can persist (Pigot et al. 2016).Habitat insularity can also limit the number of species in an assemblage, as small, isolated habitat patches tend to harbor less species than large and connected patches (MacArthur and Wilson 1967;Rosenberg 1990;Gomes et al. 2008;Borges et al. 2016a).
Oceanic islands are examples of insular environments that impose strong selection on the number of species capable to colonize these rather severe environments (Grant 1968;Grant and Grant 2006), thus favoring phenotypes different from those found in more diverse continental environments, with greater availability of resources (Lomolino 2000).For instance, there are a number of examples of morphological trends such as dwarfism, gigantism and loss of flight capacity that have been associated to extreme environmental conditions found in oceanic islands (Boback 2003;Losos and Ricklefs 2009).
Island-like systems, habitats in an ecological context similar to islands such as mountaintops or rock outcrops, are found in several regions of the planet (Vitt et al. 1997;Brown et al. 2013;Piñeiro et al. 2021).Extreme environmental conditions of most of these systems can also impose strong ecological restrictions on species assemblages (Itescu 2019).However, the study of the ecological, morphological, and evolutionary distinction of species that inhabit these islandlike systems has received less attention when compared to studies in assemblages that occupy true islands (but see Borges et al. 2016a,b).These island-like systems provide excellent opportunities to investigate how insularity and resource limitations can act as synergetic environmental filters by selecting functional attributes of the species that occupy or specialize in these environments.
White-sand ecosystems (WSE) in the Amazon are a remarkable example of an island-like habitat with severe ecological conditions.This type of low-stature vegetation occurs on sandy soils that are extremely poor in nutrients and distributed in isolated patches surrounded by tall lowland forests (Anderson 1981;Adeney et al. 2016;Capurucho et al. 2020).The flora of white-sand ecosystems is composed of plant lineages that have adapted to extreme conditions imposed by poor soils and water stress resulting in assemblages with reduced biological diversity (Adeney et al. 2016;Capurucho et al. 2020).However, WSE host a fauna and flora quite distinct from the surrounding vegetation including species that are highly specialized in the use of these environments (Anderson 1981;Oren 1981;Borges et al. 2016a;Fine et al. 2016).
The insular distribution and harsh ecological conditions make WSE excellent models to investigate the effects of environmental filters on the adaptation and specialization of animals and plants (Fine et al. 2016;Capurucho et al. 2020).In addition, WSE patches have experienced events of expansion and shrinkage of their extension throughout the geological time, evidencing a complex and dynamic evolutionary history of taxa specialized in using these systems (Capurucho et al. 2020;Ritter et al. 2021).However, we still do not know whether specialization in the use of WSE can lead to the development of distinct ecomorphological features or a peculiar arrangement of species.
Birds stand out among the taxonomic groups that specialize in the use of WSE (Oren 1981;Alonso et al. 2013).Some of these specialist birds are abundant locally and their ecological distribution is almost entirely restricted to WSE (Alonso et al. 2013;Borges et al. 2016a).It is important to emphasize that WSE are also consistently colonized and used by generalist species that also use other close by habitats such as upland forests, flooded forests, and open environments (Borges et al. 2016a).
In this study we investigated whether the specialization of birds in the use of Amazonian WSE associates with ecomorphological and evolutionary distinctions in bird species.If the limiting factors of WSE, such as scarcity of resources and insularity exerting strong selective pressure (Anderson 1981), it is expected that the specialized species would show a different and more restricted ecomorphological diversity in the functional space than generalist species that use the same environments (Julliard et al. 2006;Hamer et al. 2015;Capurucho et al. 2020).Alternatively, the environmental filter imposed by the ecological conditions of the WSE may not be intense enough to lead to distinct morphological or functional characteristics between specialist and generalist assemblages, but still restrict the morpho-functional space of specialist birds when compared to generalist species.We investigated these alternative hypotheses through comparisons of morphological ACTA AMAZONICA and ecological traits of specialist and generalist birds found in WSE environments.

Study area and bird assemblage
White-sand ecosystems (WSE) have an insular distribution in the Amazon Rainforest, forming either large and extensively connected habitat patches or small isolated patches, depending on edaphic conditions and regional paleohistory (Adeney et al. 2016).WSE vary along an ecological gradient from areas with open habitats dominated by grasses and shrubs (campinas) to forest vegetation with open understory and medium canopy height (7 to 15 m) (campinaranas) (Anderson 1981).In the northwest Amazon, campinas and campinaranas occupy continuous and connected areas while smaller and isolated patches of these vegetation types are more frequent in the central and western parts of the Amazon basin (Adeney et al. 2016).
The bird assemblage analyzed in this study was defined through a species list compilation of birds that use WSE, specifically those that occupy open vegetation dominated by herbs and shrubs (campinas) located in the northwest and central portion of the Amazon basin (Borges et al. 2016b).This is the most complete database available for birds associated with WSE sampled through captures in mist nets and audiovisual censuses (for details on the sampling design see Borges et al. (2016a).
We used the categorization proposed by Borges et al. (2016b) that classified bird species according to their habitat affinity with WSE in four categories: i) sporadic; ii) regular, iii) almost restricted and iv) restricted species.These categories are based on an extensive review of the literature and a qualitative assessment of the frequency at which each species is recorded in WSE and in other environments (Borges et al. 2016b).We did not consider the sporadic species category to avoid bias in subsequent analyses and considered the almost restricted and the restricted category as a single group.Thus, we worked with two groups: 1) generalist species, which regularly use WSE patches but also use other environments, classified as regular species according to Borges et al. (2016b), and 2) specialist species, which are mostly exclusively found in WSE, equivalent to the restricted and almost restricted categories in Borges et al. (2016b), totaling 124 species within the WSE assemblage analyzed (102 generalists and 22 specialists, see Table 1).

Morphological and ecological data
We measured the morphological traits of a total of 316 individuals from specimen skins deposited in the Bird Collection of Instituto Nacional de Pesquisas da Amazônia (INPA).We sampled at least three individuals of each species and measured each attribute with a digital caliper three times to estimate their averages.We followed the recommendations of (Baldwin et al. 1931) and obtained: body mass (g) of the individual recorded at the time of preparation; tarsus and beak length (mm); beak height and width (mm) and, length of primary and secondary remiges (mm) excluding specimens in molt.We selected these morphological traits because they are associated to organism diet (beak dimensions), foraging or locomotion (wing and tarsus) and, as such, represent species functional traits in morphological space (Devictor et al. 2010).Only one of the authors (GRL) conducted measurements.
Ecological traits of analyzed species were obtained from the literature (Cohn-Haft et al. 1997;Wilman et al. 2014) and field experience from authors (GRL and SHB).In this study we adopted three qualitative functional categories, classifying the species in terms of diet, foraging stratum and sociability (Table 2).

Analyses
In order to understand ecomorphological differentiation, through habitat filtering of bird species in WSE, we proceeded with comparisons of morphological and ecological traits in functional spaces occupied by specialist and generalist species using WSE.For that, we controlled for the phylogenetic signal among species, as they might differ in functional position due to their phylogenetic distances alone (inertia) (Belmaker et al. 2012;Jetz et al. 2012).
We investigated the phylogenetic relationships between WSE generalist and specialist birds using the data available at www.birdtree.org,generating 10,000 phylogenetic trees through the PhylogenySubset tool, a platform with bird phylogenies from around the world (Jetz et al.2012;Belmaker and Jetz 2013).The phylogenies obtained were integrated into a single consensus tree (Maximum Clade Credibility Tree), maintaining branch lengths, using the phytools package and the consensus.edgesfunction in the R Program.This approach has been widely used in comparative studies that takes into account the phylogenetic structure of biological assemblages (Kraft et al. 2007;Podani et al. 2018).
We chose to use a principal component analysis modality that controls for phylogenetic effects since we are considering species with different phylogenetic relationships in the same analysis.Phylogenetic principal component analysis (pPCA) serves to order multivariate data considering the phylogenetic non-independence between species.Unlike common PCA scores, scores on the pPCA axes are correlated with each other and their variances do not correspond to the eigenvalues of the phylogenetically corrected axes (Revell 2009;Callaghan et al. 2019).By using a pPCA analysis as a morphometric tool, we seek to interpret results explained by the adjusted eigenvectors (Polly et al. 2013).Ordinary PCA analyses were also performed for generalist and specialist species separately to obtain the vectors (characteristics) that are most related to the two bird groups.The SYNCSA package and pca function were used in program R (Debastiani and Pillar 2012;Gianuca et al. 2014).
ACTA AMAZONICA Table 1.List of bird species that inhabit Amazonian white-sand habitat patches within categories and attributes analyzed in this study.The scientific nomenclature in this list is based on the list of birds from the Brazilian Committee of Ornithological Records (Pacheco et al. 2021).Columns with categorical ecological data were obtained from Borges et al. (2016b), Wilman et al. (2014), Cohn-Haft et al. (1997) and personal observations (see Table 2).The nine morphological measurements were obtained by only one of the authors (GRL) and values represent the mean of three individuals measured per species.WSE habitat category: GEN = generalist, SPE = specialist; Diet category: Frug = frugivore, Inse = insectivore, Carn = carnivore, Omni = omnivore, Nect = nectarivore, Gran = granivore; Foraging strata: CAN = canopy, UND = understory, GRO = ground, AER = aerial, WAT= water; Sociability: Sol/pair = solitary/pairs.We also performed a Principal Coordinate Analysis (PCoA) to explore and visualize similarities or differences including morphological and ecological categorical data.We used a dissimilarity matrix (= distance matrix) and assigned each item into a location in a low-dimensional space.For this, we used the ape package, the gowdis function (Gower 1971) and then the pcoa function in the R program (Laliberte and Legendre 2010).Thus, when including ecological data, we can better explore which attributes influenced the results the most, providing a better predictive power of morphological trends within the WSE bird assemblage.
Finally we estimated functional diversity using the Functional Richness Index (Fric) (Villeger et al. 2008) to quantify the representation of the reduced space of the specialist group visualized in the pPCA and PCA analyses.Functional richness (Fric) uses the space from the common convex covering all species in the community.We used the FD package, dbFD function by Laliberté et al. (2014), and obtained a Fric value for the generalist species assemblage and for the specialist species assemblage.However, because Fric values are strongly correlated to the number of species in the community, we used a rarefaction method (Gotelli and Colwell 2001), which allowed us to compare the Fric index while controlling by the effect of different number of species between the specialist and generalist assemblages.We extracted the Fric value 1000 times by randomly drawing 22 species from the 102 generalists, and compared it with the observed Fric value for the specialist bird community (with 22 species).

RESULTS
The WSE bird assemblage analyzed is composed by 124 bird species (Table 1), 102 generalists (82 genera, 37 families and 18 orders) and 22 specialists (21 genera, 13 families and 4 orders).The proportions of species distributed among the major lineages (orders) were similar between generalist (17.5%) and specialist (19%) birds (Figure 1).Specialist species are grouped in different lineages when compared to generalist species and are represented by lineages throughout the bird clade (e.g., from Tinamiformes and Caprimulgiformes to Passeriformes, Figure 1).
Most specialist birds are insectivorous or omnivorous (Figure 2).In contrast, frugivorous and nectarivorous species were proportionately more diverse among generalist species (Figure 2).There is a clear predominance of understory species among specialist birds, while canopy species are more diverse among generalist species (Figure 2).Indeed, only five species that use the vegetation canopy are WSE specialists.Only one tinamou (Crypturellus duidae) forages on the ground within specialist species, and other 10 species within the group of generalists (Figure 2).Solitary species appear to be more common among specialists, while habitat generalist species ACTA AMAZONICA Figure 1.Consensus phylogenetic tree of the 102 generalist bird species (terminals with black line) and 22 species specialized in Amazonian white-sand habitat patches (terminals with red line).Data from "A Global Phylogeny of Birds" (www.birdtree).This figure is in color in the electronic version.
Figure 2. Percentage of Amazonian white-sand ecosystem bird species distributed in the major categories of diet, foraging strata and sociability.Numbers above columns are number of species within each category and total number of species for the specialist and generalist groups.Percentages within each category were calculated separately for specialist and generalist species.This figure is in color in the electronic version.
include a slightly higher proportion of species that congregate in mono-or heterospecific flocks (Figure 2).
The first two components of pPCA accounted for 74.43% of the variations in morphological data with PC1 (61.22%) and PC2 (13.21%).Generalist and specialist species showed a wide overlap in the multidimensional morphological space, although specialist species showed reduced morphological variability (Figure 3a).On the axis 1 of the pPCA, weight (59%) was the variable that most grouped the species in the multidimensional space followed by the length of the primary wing feathers (0.77) and the tarsus length (0.68).On the axis 2, tail length (0.93) was the only morphological variable to obtain a positive value on the axis, all other morphological traits have negative values on the axis (Figure 3a).The PCA analysis without phylogenetic control showed a similar pattern to the pPCA analysis but with an even higher morphological variability between species groups (Figure 3b).
PCA analyses performed separately for each species group indicated that for the generalist species PC1 represented 61% of the morphological variations, with longer primary wing feathers for generalist species.The same value of primary wing feathers was found for specialist birds in PC1.The morphological attribute that most influenced the multidimensional space for both generalist and specialist species was the length of the primary feathers.The generalist species have longer wings, wider and heavier beaks than the specialist species.Finally, although there are some marked differences in the categorical functional traits used in this study (Figure 2), the multivariate analysis (PCoA) considering morphological and categorical ecological traits did not discriminate between specialist and generalist species based on morphology, diet, foraging strata, and sociability (Figure 4).Functional richness measured by Fric was similar for the WSE generalist assemblage (102 species; Fric = 0.93) and for the specialist bird assemblage (22 species; Fric = 0.97), although slightly higher for specialists.Using rarefaction to take the effect of the difference in number of species between generalists and specialists, there was no significant difference in the Fric index (Figure 5) among assemblages.Functional richness as measured by the Fric index was similar between groups, with specialist birds having the same functional diversity as generalists (Figure 5).

DISCUSSION
White-sand ecosystems are island-like environments where ecological factors may limit the colonization of species not adapted to extreme ecological conditions (Adeney et al. 2016).Here, we describe and compare morphological and ecological traits of specialist and generalist species that use WSE.We did not find evidence for differentiated ecomorphological adaptations in the specialist species group associated with WSE when compared with the generalist species that also use WSE habitats besides other environments.Generalist and specialist species showed a wide overlap in the multidimensional morphological space, although specialist species showed reduced morphological variability.However, functional diversity among specialist birds is not different from functional diversity of generalists, therefore even with a reduced number of species in the specialist assemblage, functional diversity is not reduced when compared to the assemblage of generalist species.
Generalist species are more flexible in habitat use and have a greater capacity to colonize new niches and occupy the same functional space as specialist species (Pigot et al. 2020).On the other hand, despite low taxonomic diversity  of specialists, they are more abundant locally than generalists and may represent 30% to 50% of individuals caught in mist nets (Borges et al. 2016b).This high degree of species dominance, with high local abundances, may be interpreted as a consequence of specialization and higher ability to occupy and explore resources in habitats with such stressful conditions as in WSE.In white-sand ecosystems plant communities are mostly structured by soil characteristics with flooding and oligotrophic soils imposing stressful conditions on plant communities (Damasco et al. 2012).As such, specialist species may have responded stronger to environmental filters than have generalist species in WSE.
Our results point to a group of specialist species from WSE mainly composed by insectivorous species that live solitarily in the understory of the shrub vegetation.We also show that the morphology of most species in the WSE community with narrow beaks is consistent with the insectivorous diet of most specialist species.Specialization should result in restrictions on habitats used rather than items fed within a given area (Morse 1971).The restrictions exerted by the patchy and harsh environment in WSE may result in a higher dependence on more reliable and constant resources such as insects, in opposition to a highly seasonal resource such as fruits, and, as a consequence, morphology related to foraging should be less variable.We show that bird species in the specialist group have mostly narrow beaks when compared to generalist birds that have wider and heavier beaks.On the other hand, generalist species will thrive primarily through behavioral plasticity switching environments when resources are scarce.The low diversity of frugivores among the specialist species may be additional evidence of an environmental filter, since the WSE are dominated by xeromorphic vegetation (Vicentini 2004) that likely provides scarce and intermittent fruit availability.
Because our comparison includes two groups of species that use WSE, it is difficult to disentangle the effects and intensity of selective pressures imposed by the stressful environmental conditions of the WSE itself.Future studies should further investigate specialization of WSE specialist birds by, for example, comparing ecomorphological traits with other groups of species that do not use WSE habitats.Furthermore, our data, do not allow comparisons of the ecomorphology of species specialized in WSE with that of their closest relatives.For instance, available phylogenies suggest that the closest relatives of birds specialized in WSE occupy extra-Amazonian biomes such as tepuis, Cerrado, Caatinga and Atlantic Forest (Capurucho et al. 2013;Matos et al. 2016;Crouch et al. 2018;Ribas and Aleixo 2019).The ancestors of these species may have occupied white sand ecosystems in periods of expansion of this environment and, subsequently, may have experienced a period of isolation long enough to have become independent species (Gubili et al. 2016;Capurucho 2020).Ecomorphological comparisons between specialist birds and their relatives from other environments may shed light on ecomorphological adaptations on a broader temporal scale than that adopted in the present study.
Here we show that the functional space of WSE specialist species is more restricted than that of generalists in the use of the ecomorphological space, a pattern consistent with the ecological restrictions imposed by this environment (e.g.Anderson 1981;Adeney et al. 2016;Capurucho et al. 2020).However, we also show that despite reduced taxonomic diversity of specialist species and occupation of a reduced functional space, this group does not have a reduced ACTA AMAZONICA functional diversity when compared to the more diverse group of generalists.Specializing in such a harsh and fragile habitat can have a high evolutionary cost, and the behavior that increases movement and dispersal between patches to access resources in other habitat patches can be more advantageous than developing specific morphological structures.In addition, the dynamics of historical landscape changes resulting in expansion and increase of connectivity between patches of white-sand ecosystems may have contributed to increasing the dispersive capacity of these birds.In this sense, the behavioral functional trait of habitat use may be more relevant to specialization than the morphological or ecological traits considered in this study.However, morpho-functional comparisons between specialist bird species and their closest relatives that inhabit other environments are essential to better understand the evolution of specialization among birds in the Amazon white-sand ecosystems.

CONCLUSIONS
A number of studies in the last decade have increased our knowledge about Amazonian white-sand ecosystems, but the ecomorphology of white-sand ecosystem specialist birds was not previously investigated.Here, we show that specialist and generalist species that use white-sand habitat patches show a wide overlap in the multidimensional morphological space, but specialist species have reduced morphological variability.Moreover, functional diversity is not reduced in the specialist species assemblage, as may be expected when compared to the assemblage of generalist species.Specializing in a patchy, harsh and fragile habitat can have a high evolutionary cost, and the behavior that increases movement and dispersal between patches to access resources in other habitat patches can be more advantageous than evolving specific habitat specializations.This study brings new insights into habitat specialization of white-sand ecosystem birds, and opens new questions on the role of environmental filters on Amazonian bird communities.

Figure 3 .
Figure 3. Analysis of morphological data of Amazonian white-sand ecosystem birds using: A -phylogenetic principal component analysis (pPCA); and B -principal component analysis without phylogenetic control.Blue dots represent generalist species (102 species) and red dots represent specialist species (22 species).This figure is in color in the electronic version.

Figure 4 .Figure 5 .
Figure 4. Principal coordinate analysis (PCoA) with morphological and ecological traits of specialist and generalist Amazonian white-sand ecosystem birds.Blue dots represent generalist species (102 species) and red dots represent specialist species (22 species).This figure is in color in the electronic version.

Table 2 .
Ecological functional traits for Amazonian white-sand ecosystem birds used in this study.