A reference genome for the Andean cavefish Trichomycterus rosablanca (Siluriformes, Trichomycteridae): Building genomic resources to study evolution in cave environments.

Animals living in caves are of broad relevance to evolutionary biologists interested in understanding the mechanisms underpinning convergent evolution. In the Eastern Andes of Colombia, populations from at least two distinct clades of Trichomycterus catfishes (Siluriformes) independently colonized cave environments and converged in phenotype by losing their eyes and pigmentation. We are pursuing several research questions using genomics to understand the evolutionary forces and molecular mechanisms responsible for repeated morphological changes in this system. As a foundation for such studies, here we describe a diploid, chromosome-scale, long-read reference genome for Trichomycterus rosablanca, a blind, depigmented species endemic to the karstic system of the department of Santander. The nuclear genome comprises 1 Gb in 27 chromosomes, with a 40.0× HiFi long-read genome coverage having an N50 scaffold of 40.4 Mb and N50 contig of 13.1 Mb, with 96.9% (Eukaryota) and 95.4% (Actinopterygii) universal single-copy orthologs (BUSCO). This assembly provides the first reference genome for the speciose genus Trichomycterus, serving as a key resource for research on the genomics of phenotypic evolution.

First genetic insights of Gonatodescaudiscutatus (Reptilia, Gekkota) in the Galapagos Islands and mainland Ecuador.

Studies on genetic variability amongst native and introduced species contribute to a better understanding of the genetic diversity of species along their autochthonous distribution and identify possible routes of introduction. Gonatodescaudiscutatus is a gecko native to western Ecuador and introduced to the Galapagos Islands. Despite being a successful species in human-modified habitats along its native and non-native ranges, neither the colonisation process nor the genetic diversity of this gecko is known. In this study, we analysed 55 individuals from 14 localities in western Ecuador and six localities in San Cristobal Island, Galapagos - the only island with a large, self-sustaining population. We amplified and analysed the genetic variability of two nuclear genes (Cmos and Rag2) and one mitochondrial gene (16S). Cmos and Rag2 sequences presented little to none genetic variability, while 16S allowed us to build a haplotype network. We identified nine haplotypes across mainland Ecuador, two of which are also present in Galapagos. Low genetic diversity between insular and continental populations suggests that the introduction of G.caudiscutatus on the Islands is relatively recent. Due to the widespread geographical distribution of mainland haplotypes, it was not possible to determine the source population of the introduction. This study represents the first exploration of the genetic diversity of Gonatodescaudiscutatus, utilising genetic tools to gain insights into its invasion history in the Galapagos.

Environmental niche and flight intensity are associated with molecular evolutionary rates in a large avian radiation.

BACKGROUND: Metabolic activity and environmental energy are two of the most studied putative drivers of molecular evolutionary rates. Their extensive study, however, has resulted in mixed results and has rarely included the exploration of interactions among various factors impacting molecular evolutionary rates across large clades. Taking the diverse avian family Furnariidae as a case study, we examined the association between several estimates of molecular evolutionary rates with proxies of metabolic demands imposed by flight (wing loading and wing shape) and proxies of environmental energy across the geographic ranges of species (temperature and UV radiation). RESULTS: We found weak evidence of a positive effect of environmental and morphological variables on mitochondrial substitution rates. Additionally, we found that temperature and UV radiation interact to explain molecular rates at nucleotide sites affected by selection and population size (non-synonymous substitutions), contrary to the expectation of their impact on sites associated with mutation rates (synonymous substitutions). We also found a negative interaction between wing shape (as described by the hand-wing index) and body mass explaining mitochondrial molecular rates, suggesting molecular signatures of positive selection or reduced population sizes in small-bodied species with greater flight activity. CONCLUSIONS: Our results suggest that the demands of flight and environmental energy pose multiple evolutionary pressures on the genome either by driving mutation rates or via their association with natural selection or population size. Data from whole genomes and detailed physiology across taxa will bring a more complete picture of the impact of metabolism, population size, and the environment on avian genome evolution.

Migratory connectivity then and now: a northward shift in breeding origins of a long-distance migratory bird wintering in the tropics.

Temporal variation in the connectivity of populations of migratory animals has not been widely documented, despite having important repercussions for population ecology and conservation. Because the long-distance movements of migratory animals link ecologically distinct and geographically distant areas of the world, changes in the abundance and migratory patterns of species may reflect differential drivers of demographic trends acting over various spatial scales. Using stable hydrogen isotope analyses (δ2H) of feathers from historical museum specimens and contemporary samples obtained in the field, we provide evidence for an approximately 600 km northward shift over 45 years in the breeding origin of a species of songbird of major conservation concern (blackpoll warbler, Setophaga striata) wintering in the foothills of the eastern Andes of Colombia. Our finding mirrors predictions of range shifts for boreal-breeding species under warming climate scenarios and habitat loss in the temperate zone, and underscores likely drivers of widespread declines in populations of migratory birds. Our work also highlights the value of natural history collections to document the effects of global change on biodiversity.

An extinct hummingbird species that never was: a cautionary tale about sampling issues in molecular phylogenetics.

The selection of species and individuals for molecular analyses critically affects inferences in various fields of systematic biology including phylogenetics, phylogeography, and species delimitation. Especially in areas like the Neotropical region where molecular analyses have recovered substantial within-species divergence and unexpected affinities of populations (Turchetto-Zolet et al. 2013), biases resulting from incomplete taxonomic or geographic sampling may compromise the understanding of phylogenetic relationships (Avendaño et al. 2017). Here we describe a case in which assessments of the validity of a potentially extinct species of Neotropical bird were likely compromised because within-species variation was not accounted for in phylogenetic analyses evaluating the alternative hypothesis that the only known specimen may represent a hybrid.

Assessing seasonal changes in animal diets with stable-isotope analysis of amino acids: a migratory boreal songbird switches diet over its annual cycle.

Tools to study seasonal changes in animal diets are needed to address a wide range of ecological questions. This is especially true of migratory animals that experience distinct environments where diets may be substantially different. However, tracking diets of individuals that move vast distances has proven difficult. Compound-specific isotope analysis has emerged as a valuable tool to study diets but has been little used to study dietary changes of migratory animals. Using this technique, we quantify seasonal variation in the annual diet of a migratory songbird (gray-cheeked thrush, Catharus minimus) and test the hypothesis that migrants change their diet in response to the energetic requirements of different periods of the annual cycle. By measuring δ13C and δ15N values of amino acids from feathers grown on the breeding grounds, blood formed during migration and claw grown on the wintering grounds, we found that migration is associated with greater consumption of fruit, compared to the breeding or wintering periods. This was confirmed by the lower trophic position of blood compared to feather and claw, by a decrease in the δ15N value of the source amino acid phenylalanine in blood as a function of days of stopover, and by the positive correlation between δ15N and δ13C values of phenylalanine in blood, and not in feather or claw. This study illustrates how isotopic analysis of amino acids can contribute to understand food webs, seasonal dietary changes and metabolic routing of nutrients in migratory animals.

Issues and Perspectives in Species Delimitation using Phenotypic Data: Atlantean Evolution in Darwin's Finches.

Progress in the development and use of methods for species delimitation employing phenotypic data lags behind conceptual and practical advances in molecular genetic approaches. The basic evolutionary model underlying the use of phenotypic data to delimit species assumes random mating and quantitative polygenic traits, so that phenotypic distributions within a species should be approximately normal for individuals of the same sex and age. Accordingly, two or more distinct normal distributions of phenotypic traits suggest the existence of multiple species. In light of this model, we show that analytical approaches employed in taxonomic studies using phenotypic data are often compromised by three issues: 1) reliance on graphical analyses that convey little information on phenotype frequencies; 2) exclusion of characters potentially important for species delimitation following reduction of data dimensionality; and 3) use of measures of central tendency to evaluate phenotypic distinctiveness. We outline approaches to overcome these issues based on statistical developments related to normal mixture models (NMMs) and illustrate them empirically with a reanalysis of morphological data recently used to claim that there are no morphologically distinct species of Darwin's ground-finches (Geospiza). We found negligible support for this claim relative to taxonomic hypotheses recognizing multiple species. Although species limits among ground-finches merit further assessments using additional sources of information, our results bear implications for other areas of inquiry including speciation research: because ground-finches have likely speciated and are not trapped in a process of "Sisyphean" evolution as recently argued, they remain useful models to understand the evolutionary forces involved in speciation. Our work underscores the importance of statistical approaches grounded on appropriate evolutionary models for species delimitation. We discuss how NMMs offer new perspectives in the kind of inferences available to systematists, with significant repercussions on ideas about the phenotypic structure of biodiversity.

Origin and cross-century dynamics of an avian hybrid zone.

BACKGROUND: Characterizations of the dynamics of hybrid zones in space and time can give insights about traits and processes important in population divergence and speciation. We characterized a hybrid zone between tanagers in the genus Ramphocelus (Aves, Thraupidae) located in southwestern Colombia. We evaluated whether this hybrid zone originated as a result of secondary contact or of primary differentiation, and described its dynamics across time using spatial analyses of molecular, morphological, and coloration data in combination with paleodistribution modeling. RESULTS: Models of potential historical distributions based on climatic data and genetic signatures of demographic expansion suggested that the hybrid zone likely originated following secondary contact between populations that expanded their ranges out of isolated areas in the Quaternary. Concordant patterns of variation in phenotypic characters across the hybrid zone and its narrow extent are suggestive of a tension zone, maintained by a balance between dispersal and selection against hybrids. Estimates of phenotypic cline parameters obtained using specimens collected over nearly a century revealed that, in recent decades, the zone appears to have moved to the east and to higher elevations, and may have become narrower. Genetic variation was not clearly structured along the hybrid zone, but comparisons between historical and contemporary specimens suggested that temporal changes in its genetic makeup may also have occurred. CONCLUSIONS: Our data suggest that the hybrid zone likey resulted from secondary contact between populations. The observed changes in the hybrid zone may be a result of sexual selection, asymmetric gene flow, or environmental change.

On the importance of geographic and taxonomic sampling in phylogeography: A reevaluation of diversification and species limits in a Neotropical thrush (Aves, Turdidae).

Phylogeographic studies seeking to describe biogeographic patterns, infer evolutionary processes, and revise species-level classification should properly characterize the distribution ranges of study species, and thoroughly sample genetic variation across taxa and geography. This is particularly necessary for widely distributed organisms occurring in complex landscapes, such as the Neotropical region. Here, we clarify the geographic range and revisit the phylogeography of the Black-billed Thrush (Turdus ignobilis), a common passerine bird from lowland tropical South America, whose evolutionary relationships and species limits were recently evaluated employing phylogeographic analyses based on partial knowledge of its distribution and incomplete sampling of populations. Our work employing mitochondrial and nuclear DNA sequences sampled all named subspecies and multiple populations across northern South America, and uncovered patterns not apparent in earlier work, including a biogeographic interplay between the Amazon and Orinoco basins and the occurrence of distinct lineages with seemingly different habitat affinities in regional sympatry in the Colombian Amazon. In addition, we found that previous inferences about the affinities and taxonomic status of Andean populations assumed to be allied to populations from the Pantepui region were incorrect, implying that inferred biogeographic and taxonomic scenarios need re-evaluation. We propose a new taxonomic treatment, which recognizes two distinct biological species in the group. Our findings illustrate the importance of sufficient taxon and geographic sampling to reconstruct evolutionary history and to evaluate species limits among Neotropical organisms. Considering the scope of the questions asked, advances in Neotropical phylogeography will often require substantial cross-country scientific collaboration.

Toxic metals and associated sporulated bacteria on Andean hummingbird feathers.

Human activities in the Sabana de Bogotá, Colombia, release toxic metals such as lead (Pb) and chromium (Cr) into the environment polluting the air, water, and soil. Because birds are in contact with these pollutants and their sources, they may serve as bioindicator organisms. We evaluated the use of hummingbird feathers obtained from individuals captured in three sites of the Sabana de Bogotá as bioindicators of toxic metal pollution using spectrophotometric and spectroscopic methods based on single-feather samples. We also characterized the bacterial microbiota associated with hummingbird feathers by molecular identification using the 16S rRNA with a special focus on sporulated bacteria. Finally, we described the interactions which naturally occur among the feathers, their associated bacteria, and pollutants. We found differences in Pb and Cr concentrations between sampling sites, which ranged from 2.11 to 4.69 ppm and 0.38 to 3.00 ppm, respectively. This may reflect the impact of the activities held in those sites which release pollutants to the environment. Bacterial assemblages mainly consisted of sporulated bacilli in the Bacillaceae family (65.7 % of the identified morphotypes). We conclude that the feathers of wild tropical birds, including hummingbirds, can be used as lead and chromium bioindicators and that bacteria growing on feathers may in fact interact with these two toxic metals.

Niche-tracking migrants and niche-switching residents: evolution of climatic niches in New World warblers (Parulidae).

Differences in life-history traits between tropical and temperate lineages are often attributed to differences in their climatic niche dynamics. For example, the more frequent appearance of migratory behaviour in temperate-breeding species than in species originally breeding in the tropics is believed to have resulted partly from tropical climatic stability and niche conservatism constraining tropical species from shifting their ranges. However, little is known about the patterns and processes underlying climatic niche evolution in migrant and resident animals. We evaluated the evolution of overlap in climatic niches between seasons and its relationship to migratory behaviour in the Parulidae, a family of New World passerine birds. We used ordination methods to measure seasonal niche overlap and niche breadth of 54 resident and 49 migrant species and used phylogenetic comparative methods to assess patterns of climatic niche evolution. We found that despite travelling thousands of kilometres, migrants tracked climatic conditions across the year to a greater extent than tropical residents. Migrant species had wider niches than resident species, although residents as a group occupied a wider climatic space and niches of migrants and residents overlapped extensively. Neither breeding latitude nor migratory distance explained variation among species in climatic niche overlap between seasons. Our findings support the notion that tropical species have narrower niches than temperate-breeders, but does not necessarily constrain their ability to shift or expand their geographical ranges and become migratory. Overall, the tropics may have been historically less likely to experience the suite of components that generate strong selection pressures for the evolution of migratory behaviour.

Molecular systematics of the new world screech-owls (Megascops: Aves, Strigidae): biogeographic and taxonomic implications.

Megascops screech-owls are endemic to the New World and range from southern Canada to the southern cone of South America. The 22 currently recognized Megascops species occupy a wide range of habitats and elevations, from desert to humid montane forest, and from sea level to the Andean tree line. Species and subspecies diagnoses of Megascops are notoriously difficult due to subtle plumage differences among taxa with frequent plumage polymorphism. Using three mitochondrial and three nuclear genes we estimated a phylogeny for all but one Megascops species. Phylogenies were estimated with Maximum Likelihood and Bayesian Inference, and a Bayesian chronogram was reconstructed to assess the spatio-temporal context of Megascops diversification. Megascops was paraphyletic in the recovered tree topologies if the Puerto Rican endemic M. nudipes is included in the genus. However, the remaining taxa are monophyletic and form three major clades: (1) M. choliba, M. koepckeae, M. albogularis, M. clarkii, and M. trichopsis; (2) M. petersoni, M. marshalli, M. hoyi, M. ingens, and M. colombianus; and (3) M. asio, M. kennicottii, M. cooperi, M. barbarus, M. sanctaecatarinae, M. roboratus, M. watsonii, M. atricapilla, M. guatemalae, and M. vermiculatus. Megascops watsonii is paraphyletic with some individuals more closely related to M. atricapilla than to other members in that polytypic species. Also, allopatric populations of some other Megascops species were highly divergent, with levels of genetic differentiation greater than between some recognized species-pairs. Diversification within the genus is hypothesized to have taken place during the last 8 million years, with a likely origin in Central America. The genus later expanded over much of the Americas and then diversified via multiple dispersal events from the Andes into the Neotropical lowlands.

Critical thermal limits of Poecilia caucana (Steindachner, 1880) (Cyprinodontiformes: Poeciliidae)

Aunque la temperatura tiene grandes repercusiones en la biología de los peces, se desconocen los rangos de tolerancia térmica de la mayoría de los peces dulceacuícolas. Esta falta de información impide pronosticar respuestas al cambio climático y limita los análisis comparativos que podrían enriquecer estudios evolutivos y biogeográficos. Utilizamos la metodología del crítico térmico para cuantificar la capacidad de aclimatación y la tolerancia térmica en la especie neotropical dulceacuícola Poecilia caucana. Para peces aclimatados a 20C, 25ºC y 28ºC, los críticos térmicos mínimos (CTmin) fueron 12,52 ± 0,62ºC, 13,41 ± 0,56ºC y 14,24 ± 0,43ºC, respectivamente, y los críticos térmicos máximos (CTmax) fueron 38,43 ± 0,64ºC, 40,28 ± 0,92ºC y 41,57 ± 0,27ºC, respectivamente. Tanto el CTmin como el CTmax cambiaron significativamente con las temperaturas de aclimatación, indicando que P. caucana es efectivamente aclimatable. Comparada con otras especies de peces dulceacuícolas, la capacidad de aclimatación de P. cuacana fue baja para CTmin y promedio para CTmax. Estos resultados, en conjunto con los datos de otras especies, pueden ser utilizados para responder preguntas ecológicas y evolutivas más generales.(AU)

Although temperature has far-reaching effects on fish biology, the thermal tolerance ranges of most freshwater fish species are unknown. This lack of information precludes forecasting responses to climatic change and does not allow for comparative analyses that may inform evolutionary and biogeographic studies. We used the critical thermal methodology to quantify acclimation capacity and thermal tolerance in the Neotropical freshwater species Poecilia caucana . For fish acclimated to 20C, 25ºC, and 28ºC, critical thermal minima (CTmin) were 12.52 ± 0.62ºC, 13.41 ± 0.56ºC and 14.24 ± 0.43ºC respectively, and critical thermal maxima (CTmax) were 38.43 ± 0.64ºC, 40.28 ± 0.92ºC and 41.57 ± 0.27ºC, respectively. Both CTmin and CTmax changed with acclimation temperatures, indicating that P. caucana was effectively acclimatable. Relative to values reported for other freshwater fish species, the acclimation capacity of P. caucana for CTmin was low, but it was average for CTmax. The data, together with similar work in other species, can be used in analyses focusing on broad ecological and evolutionary questions.(AU)

Critical thermal limits of Poecilia caucana (Steindachner, 1880) (Cyprinodontiformes: Poeciliidae)

Aunque la temperatura tiene grandes repercusiones en la biología de los peces, se desconocen los rangos de tolerancia térmica de la mayoría de los peces dulceacuícolas. Esta falta de información impide pronosticar respuestas al cambio climático y limita los análisis comparativos que podrían enriquecer estudios evolutivos y biogeográficos. Utilizamos la metodología del crítico térmico para cuantificar la capacidad de aclimatación y la tolerancia térmica en la especie neotropical dulceacuícola Poecilia caucana. Para peces aclimatados a 20C, 25ºC y 28ºC, los críticos térmicos mínimos (CTmin) fueron 12,52 ± 0,62ºC, 13,41 ± 0,56ºC y 14,24 ± 0,43ºC, respectivamente, y los críticos térmicos máximos (CTmax) fueron 38,43 ± 0,64ºC, 40,28 ± 0,92ºC y 41,57 ± 0,27ºC, respectivamente. Tanto el CTmin como el CTmax cambiaron significativamente con las temperaturas de aclimatación, indicando que P. caucana es efectivamente aclimatable. Comparada con otras especies de peces dulceacuícolas, la capacidad de aclimatación de P. cuacana fue baja para CTmin y promedio para CTmax. Estos resultados, en conjunto con los datos de otras especies, pueden ser utilizados para responder preguntas ecológicas y evolutivas más generales.

Although temperature has far-reaching effects on fish biology, the thermal tolerance ranges of most freshwater fish species are unknown. This lack of information precludes forecasting responses to climatic change and does not allow for comparative analyses that may inform evolutionary and biogeographic studies. We used the critical thermal methodology to quantify acclimation capacity and thermal tolerance in the Neotropical freshwater species Poecilia caucana . For fish acclimated to 20C, 25ºC, and 28ºC, critical thermal minima (CTmin) were 12.52 ± 0.62ºC, 13.41 ± 0.56ºC and 14.24 ± 0.43ºC respectively, and critical thermal maxima (CTmax) were 38.43 ± 0.64ºC, 40.28 ± 0.92ºC and 41.57 ± 0.27ºC, respectively. Both CTmin and CTmax changed with acclimation temperatures, indicating that P. caucana was effectively acclimatable. Relative to values reported for other freshwater fish species, the acclimation capacity of P. caucana for CTmin was low, but it was average for CTmax. The data, together with similar work in other species, can be used in analyses focusing on broad ecological and evolutionary questions.

Genetic differentiation, niche divergence, and the origin and maintenance of the disjunct distribution in the Blossomcrown Anthocephala floriceps (Trochilidae).

Studies of the origin and maintenance of disjunct distributions are of special interest in biogeography. Disjunct distributions can arise following extinction of intermediate populations of a formerly continuous range and later maintained by climatic specialization. We tested hypotheses about how the currently disjunct distribution of the Blossomcrown (Anthocephala floriceps), a hummingbird species endemic to Colombia, arose and how is it maintained. By combining molecular data and models of potential historical distributions we evaluated: (1) the timing of separation between the two populations of the species, (2) whether the disjunct distribution could have arisen as a result of fragmentation of a formerly widespread range due to climatic changes, and (3) if the disjunct distribution might be currently maintained by specialization of each population to different climatic conditions. We found that the two populations are reciprocally monophyletic for mitochondrial and nuclear loci, and that their divergence occurred ca. 1.4 million years before present (95% credibility interval 0.7-2.1 mybp). Distribution models based on environmental data show that climate has likely not been suitable for a fully continuous range over the past 130,000 years, but the potential distribution 6,000 ybp was considerably larger than at present. Tests of climatic divergence suggest that significant niche divergence between populations is a likely explanation for the maintenance of their disjunct ranges. However, based on climate the current range of A. floriceps could potentially be much larger than it currently is, suggesting other ecological or historical factors have influenced it. Our results showing that the distribution of A. floriceps has been discontinous for a long period of time and that populations exhibit different climatic niches have taxonomic and conservation implications.

The drivers of tropical speciation.

Since the recognition that allopatric speciation can be induced by large-scale reconfigurations of the landscape that isolate formerly continuous populations, such as the separation of continents by plate tectonics, the uplift of mountains or the formation of large rivers, landscape change has been viewed as a primary driver of biological diversification. This process is referred to in biogeography as vicariance. In the most species-rich region of the world, the Neotropics, the sundering of populations associated with the Andean uplift is ascribed this principal role in speciation. An alternative model posits that rather than being directly linked to landscape change, allopatric speciation is initiated to a greater extent by dispersal events, with the principal drivers of speciation being organism-specific abilities to persist and disperse in the landscape. Landscape change is not a necessity for speciation in this model. Here we show that spatial and temporal patterns of genetic differentiation in Neotropical birds are highly discordant across lineages and are not reconcilable with a model linking speciation solely to landscape change. Instead, the strongest predictors of speciation are the amount of time a lineage has persisted in the landscape and the ability of birds to move through the landscape matrix. These results, augmented by the observation that most species-level diversity originated after episodes of major Andean uplift in the Neogene period, suggest that dispersal and differentiation on a matrix previously shaped by large-scale landscape events was a major driver of avian speciation in lowland Neotropical rainforests.

Asynchrony of seasons: genetic differentiation associated with geographic variation in climatic seasonality and reproductive phenology.

Many organisms exhibit distinct breeding seasons tracking food availability. If conspecific populations inhabit areas that experience different temporal cycles in food availability spurred by variation in precipitation regimes, then they should display asynchronous breeding seasons. Thus, such populations might exhibit a temporal barrier to gene flow, which may potentially promote genetic differentiation. We test a central prediction of this hypothesis, namely, that individuals living in areas with more asynchronous precipitation regimes should be more genetically differentiated than individuals living in areas with more similar precipitation regimes. Using mitochondrial DNA sequences, climatic data, and geographical/ecological distances between individuals of 57 New World bird species mostly from the tropics, we examined the effect of asynchronous precipitation (a proxy for asynchronous resource availability) on genetic differentiation. We found evidence for a positive and significant cross-species effect of precipitation asynchrony on genetic distance after accounting for geographical/ecological distances, suggesting that current climatic conditions may play a role in population differentiation. Spatial asynchrony in climate may thus drive evolutionary divergence in the absence of overt geographic barriers to gene flow; this mechanism contrasts with those invoked by most models of biotic diversification emphasizing physical or ecological changes to the landscape as drivers of divergence.

Sensitivity of metrics of phylogenetic structure to scale, source of data and species pool of hummingbird assemblages along elevational gradients.

Patterns of phylogenetic structure of assemblages are increasingly used to gain insight into the ecological and evolutionary processes involved in the assembly of co-occurring species. Metrics of phylogenetic structure can be sensitive to scaling issues and data availability. Here we empirically assess the sensitivity of four metrics of phylogenetic structure of assemblages to changes in (i) the source of data, (ii) the spatial grain at which assemblages are defined, and (iii) the definition of species pools using hummingbird (Trochilidae) assemblages along an elevational gradient in Colombia. We also discuss some of the implications in terms of the potential mechanisms driving these patterns. To explore how source of data influence phylogenetic structure we defined assemblages using three sources of data: field inventories, museum specimens, and range maps. Assemblages were defined at two spatial grains: coarse-grained (elevational bands of 800-m width) and fine-grained (1-km(2) plots). We used three different species pools: all species contained in assemblages, all species within half-degree quadrats, and all species either above or below 2000 m elevation. Metrics considering phylogenetic relationships among all species within assemblages showed phylogenetic clustering at high elevations and phylogenetic evenness in the lowlands, whereas those metrics considering only the closest co-occurring relatives showed the opposite trend. This result suggests that using multiple metrics of phylogenetic structure should provide greater insight into the mechanisms shaping assemblage structure. The source and spatial grain of data had important influences on estimates of both richness and phylogenetic structure. Metrics considering the co-occurrence of close relatives were particularly sensitive to changes in the spatial grain. Assemblages based on range maps included more species and showed less phylogenetic structure than assemblages based on museum or field inventories. Coarse-grained assemblages included more distantly related species and thus showed a more even phylogenetic structure than fine-grained assemblages. Our results emphasize the importance of carefully selecting the scale, source of data and metric used in analysis of the phylogenetic structure of assemblages.

Non-monophyly and deep genetic differentiation across low-elevation barriers in a Neotropical montane bird (Basileuterus tristriatus; Aves: Parulidae).

Most widespread birds of Neotropical cloud forests exhibit phenotypic variation that is partitioned geographically suggesting allopatric divergence, but little is known about the extent to which such phenotypic differentiation is consistent with genetic variation. We studied geographic patterns of genetic differentiation in the Three-striped Warbler (Basileuterus tristriatus), a polytypic and widespread understory bird of the foothills and mid-elevation zone of the tropical Andes and adjacent mountains of Central and South America. We sequenced mitochondrial DNA for 196 samples covering the entire range of B. tristriatus, as well as 22 samples of its putative closest relatives: the Three-banded (B. trifasciatus) and Santa Marta (B. basilicus) warblers. We found deep genetic structure across the range of B. tristriatus, which consisted of ten major clades including B. trifasciatus, a species that was nested within B. tristriatus. In contrast, B. basilicus was not closely related to B. tristriatus but part of a clade of Myiothlypis warblers. Geographic boundaries among clades were clearly related to lowland gaps separating subspecies groups. The subspecies melanotis of the mountains of Central America was sister to a large clade including B. t. tacarcunae, and the rest of South American clades, including B. trifasciatus. Five clades are found in the northern Andes, where no signs of gene flow were found across barriers such as the Táchira Depression or the Magdalena valley. Our study highlights the importance of valleys in promoting and maintaining divergence in a lower montane forest bird. The substantial genetic and phenotypic differentiation, and the paraphyly uncovered in B. tristriatus, may call for revising its species boundaries.