A test of the riverine barrier hypothesis in the largest subtropical river basin in the Neotropics.

The riverine barrier hypothesis proposes that large rivers represent geographical barriers to gene flow for terrestrial organisms, leading to population differentiation and ultimately allopatric speciation. Here we assess for the first time if the subtropical Paraná-Paraguay River system in the Del Plata basin, second in size among South American drainages, acts as a barrier to gene flow for birds. We analysed the degree of mitochondrial and nuclear genomic differentiation in seven species with known subspecies divided by the Paraná-Paraguay River axis. Only one species showed genetic differentiation concordant with the current river channel, but another five species have an east/west genetic split broadly coincident with the Paraná River's dynamic palaeochannel, suggesting this fluvial axis has had a past role in shaping present-day genetic structure. Moreover, dating analyses show that these splits have been asynchronous, with species responding differently to the riverine barrier. Comparisons informed by the geological history of the Paraná River and its influence on the ecological and climatic differences among ecoregions in the study area further bolster the finding that responses to this geographical barrier have been species-specific.

Convergent Evolution of Hemoglobin Function in High-Altitude Andean Waterfowl Involves Limited Parallelism at the Molecular Sequence Level.

A fundamental question in evolutionary genetics concerns the extent to which adaptive phenotypic convergence is attributable to convergent or parallel changes at the molecular sequence level. Here we report a comparative analysis of hemoglobin (Hb) function in eight phylogenetically replicated pairs of high- and low-altitude waterfowl taxa to test for convergence in the oxygenation properties of Hb, and to assess the extent to which convergence in biochemical phenotype is attributable to repeated amino acid replacements. Functional experiments on native Hb variants and protein engineering experiments based on site-directed mutagenesis revealed the phenotypic effects of specific amino acid replacements that were responsible for convergent increases in Hb-O2 affinity in multiple high-altitude taxa. In six of the eight taxon pairs, high-altitude taxa evolved derived increases in Hb-O2 affinity that were caused by a combination of unique replacements, parallel replacements (involving identical-by-state variants with independent mutational origins in different lineages), and collateral replacements (involving shared, identical-by-descent variants derived via introgressive hybridization). In genome scans of nucleotide differentiation involving high- and low-altitude populations of three separate species, function-altering amino acid polymorphisms in the globin genes emerged as highly significant outliers, providing independent evidence for adaptive divergence in Hb function. The experimental results demonstrate that convergent changes in protein function can occur through multiple historical paths, and can involve multiple possible mutations. Most cases of convergence in Hb function did not involve parallel substitutions and most parallel substitutions did not affect Hb-O2 affinity, indicating that the repeatability of phenotypic evolution does not require parallelism at the molecular level.

Difference between the vocalizations of two sister species of pigeons explained in dynamical terms.

Vocal communication is an unique example, where the nonlinear nature of the periphery can give rise to complex sounds even when driven by simple neural instructions. In this work we studied the case of two close-related bird species, Patagioenas maculosa and Patagioenas picazuro, whose vocalizations differ only in the timbre. The temporal modulation of the fundamental frequency is similar in both cases, differing only in the existence of sidebands around the fundamental frequency in the P. maculosa. We tested the hypothesis that the qualitative difference between these vocalizations lies in the nonlinear nature of the syrinx. In particular, we propose that the roughness of maculosa's vocalizations is due to an asymmetry between the right and left vibratory membranes, whose nonlinear dynamics generate the sound. To test the hypothesis, we generated a biomechanical model for vocal production with an asymmetric parameter Q with which we can control the level of asymmetry between these membranes. Using this model we generated synthetic vocalizations with the principal acoustic features of both species. In addition, we confirmed the anatomical predictions by making post mortem inspection of the syrinxes, showing that the species with tonal song (picazuro) has a more symmetrical pair of membranes compared to maculosa.

Multilocus coalescent analysis of haemoglobin differentiation between low- and high-altitude populations of crested ducks (Lophonetta specularioides).

Hypoxia is a key factor determining survival, and haemoglobins are targets of selection in species native to high-altitude regions. We studied population genetic structure and evaluated evidence for local adaptation in the crested duck (Lophonetta specularioides). Differentiation, gene flow and time since divergence between highland and lowland populations were assessed for three haemoglobin genes (α(A) , α(D) , ß(A) ) and compared to seven reference loci (six autosomal introns and mtDNA). Four derived amino acid replacements were found in the globin genes that had elevated Φ(ST) values between the Andean highlands and Patagonian lowlands. A single ß(A) -globin polymorphism at a site known to influence O(2) affinity was fixed for different alleles in the two populations, whereas three α(A) - and α(D) -globin polymorphisms exhibited high heterozygosity in the highlands but not in the lowlands. Coalescent analyses supported restricted gene flow for haemoglobin alleles and mitochondrial DNA but nonzero gene flow for the introns. Simulating genetic data under a drift-migration model of selective neutrality, the ß(A) -globin fell outside the 95% confidence limit of simulated data, suggesting that directional selection is maintaining different variants in the contrasting elevational environments, thereby restricting migration of ß(A) -globin alleles. The α(A) - and α(D) -globins, by contrast, did not differ from the simulated values, suggesting that variants in these genes are either selectively neutral, or that the effects of selection could not be differentiated from background levels of population structure and linkage disequilibrium. This study illustrates the combined effects of selection and population history on inferring levels of population divergence for a species distributed across an altitudinal gradient in which selection for hypoxia resistance has likely played an important role.

Detecting areas of endemism with a taxonomically diverse data set: plants, mammals, reptiles, amphibians, birds, and insects from Argentina.

The idea of an area of endemism implies that different groups of plants and animals should have largely coincident distributions. This paper analyses an area of 1152 000 km2 , between parallels 21 and 32°S and meridians 70 and 53°W to examine whether a large and taxonomically diverse data set actually displays areas supported by different groups. The data set includes the distribution of 805 species of plants (45 families), mammals (25 families), reptiles (six families), amphibians (five families), birds (18 families), and insects (30 families), and is analysed with the optimality criterion (based on the notion of endemism) implemented in the program NDM/VNDM. Almost 50% of the areas obtained are supported by three or more major groups; areas supported by fewer major groups generally contain species from different genera, families, or orders. © The Willi Hennig Society 2011.

Rapid speciation via the evolution of pre-mating isolation in the Iberá Seedeater.

Behavioral isolation can catalyze speciation and permit the slow accumulation of additional reproductive barriers between co-occurring organisms. We illustrate how this process occurs by examining the genomic and behavioral bases of pre-mating isolation between two bird species (Sporophila hypoxantha and the recently discovered S. iberaensis) that belong to the southern capuchino seedeaters, a recent, rapid radiation characterized by variation in male plumage coloration and song. Although these two species co-occur without obvious ecological barriers to reproduction, we document behaviors indicating species recognition by song and plumage traits and strong assortative mating associated with genomic regions underlying male plumage patterning. Plumage differentiation likely originated through the reassembly of standing genetic variation, indicating how novel sexual signals may quickly arise and maintain species boundaries.

Behaviour of resampling methods under different weighting schemes, measures and variable resampling strengths.

We compared general behaviour trends of resampling methods (bootstrap, bootstrap with Poisson distribution, jackknife, and jackknife with symmetric resampling) and different ways to summarize the results for resampling (absolute frequency, F, and frequency difference, GC') for real data sets under variable resampling strengths in three weighting schemes. We propose an equivalence between bootstrap and jackknife in order to make bootstrap variable across different resampling strengths. Specifically, for each method we evaluated the number of spurious groups (groups not present in the strict consensus of the unaltered data set), of real groups, and of inconsistencies in ranking of groups under variable resampling strengths. We found that GC' always generated more spurious groups and recovered more groups than F. Bootstrap methods generated more spurious groups than jackknife methods; and jackknife is the method that recovered more real groups. We consistently obtained a higher proportion of spurious groups for GC' than for F; and for bootstrap than for jackknife. Finally, we evaluated the ranking of groups under variable resampling strengths qualitatively in the trajectories of "support" against resampling strength, and quantitatively with Kendall coefficient values. We found fewer ranking inconsistencies for GC' than for F, and for bootstrap than for jackknife. © The Willi Hennig Society 2009.

Old divergence and restricted gene flow between torrent duck (Merganetta armata) subspecies in the Central and Southern Andes.

AIM: To investigate the structure and rate of gene flow among populations of habitat-specialized species to understand the ecological and evolutionary processes underpinning their population dynamics and historical demography, including speciation and extinction. LOCATION: Peruvian and Argentine Andes. TAXON: Two subspecies of torrent duck (Merganetta armata). METHODS: We sampled 156 individuals in Peru (M. a. leucogenis; Chillón River, n = 57 and Pachachaca River, n = 49) and Argentina (M. a. armata; Arroyo Grande River, n = 33 and Malargüe River, n = 17), and sequenced the mitochondrial DNA (mtDNA) control region to conduct coarse and fine-scale demographic analyses of population structure. Additionally, to test for differences between subspecies, and across genetic markers with distinct inheritance patterns, a subset of individuals (Peru, n = 10 and Argentina, n = 9) was subjected to partial genome resequencing, obtaining 4,027 autosomal and 189 Z-linked double-digest restriction-associated DNA sequences. RESULTS: Haplotype and nucleotide diversities were higher in Peru than Argentina across all markers. Peruvian and Argentine subspecies showed concordant species-level differences (ΦST mtDNA = 0.82; ΦST autosomal = 0.30; ΦST Z chromosome = 0.45), including no shared mtDNA haplotypes. Demographic parameters estimated for mtDNA using IM and IMa2 analyses, and for autosomal markers using ∂a∂i (isolation-with-migration model), supported an old divergence (mtDNA = 600,000 years before present (ybp), 95% HPD range = 1.2 Mya to 200,000 ybp; and autosomal ∂a∂i = 782,490 ybp), between the two subspecies, characteristic of deeply diverged lineages. The populations were well-differentiated in Argentina but moderately differentiated in Peru, with low unidirectional gene flow in each country. MAIN CONCLUSIONS: We suggest that the South American Arid Diagonal was preexisting and remains a current phylogeographic barrier between the ranges of the two torrent duck subspecies, and the adult territoriality and breeding site fidelity to the rivers define their population structure.

Evolution of Vocal Diversity through Morphological Adaptation without Vocal Learning or Complex Neural Control.

The evolution of complex behavior is driven by the interplay of morphological specializations and neuromuscular control mechanisms [1-3], and it is often difficult to tease apart their respective contributions. Avian vocal learning and associated neural adaptations are thought to have played a major role in bird diversification [4-8], whereas functional significance of substantial morphological diversity of the vocal organ remains largely unexplored. Within the most species-rich order, Passeriformes, "tracheophones" are a suboscine group that, unlike their oscine sister taxon, does not exhibit vocal learning [9] and is thought to phonate with tracheal membranes [10, 11] instead of the two independent sources found in other passerines [12-14]. Here we show tracheophones possess three sound sources, two oscine-like labial pairs and the unique tracheal membranes, which collectively represent the largest described number of sound sources for a vocal organ. Birds with experimentally disabled tracheal membranes were still able to phonate. Instead of the main sound source, the tracheal membranes constitute a morphological specialization, which, through interaction with bronchial labia, contributes to different acoustic features such as spectral complexity, amplitude modulation, and enhanced sound amplitude. In contrast, these same features arise in oscines from neuromuscular control of two labial sources [15-17]. These findings are supported by a modeling approach and provide a clear example for how a morphological adaptation of the tracheophone vocal organ can generate specific, complex sound features. Morphological specialization therefore constitutes an alternative path in the evolution of acoustic diversity to that of oscine vocal learning and complex neural control.

Barcoding the butterflies of southern South America: Species delimitation efficacy, cryptic diversity and geographic patterns of divergence.

Because the tropical regions of America harbor the highest concentration of butterfly species, its fauna has attracted considerable attention. Much less is known about the butterflies of southern South America, particularly Argentina, where over 1,200 species occur. To advance understanding of this fauna, we assembled a DNA barcode reference library for 417 butterfly species of Argentina, focusing on the Atlantic Forest, a biodiversity hotspot. We tested the efficacy of this library for specimen identification, used it to assess the frequency of cryptic species, and examined geographic patterns of genetic variation, making this study the first large-scale genetic assessment of the butterflies of southern South America. The average sequence divergence to the nearest neighbor (i.e. minimum interspecific distance) was 6.91%, ten times larger than the mean distance to the furthest conspecific (0.69%), with a clear barcode gap present in all but four of the species represented by two or more specimens. As a consequence, the DNA barcode library was extremely effective in the discrimination of these species, allowing a correct identification in more than 95% of the cases. Singletons (i.e. species represented by a single sequence) were also distinguishable in the gene trees since they all had unique DNA barcodes, divergent from those of the closest non-conspecific. The clustering algorithms implemented recognized from 416 to 444 barcode clusters, suggesting that the actual diversity of butterflies in Argentina is 3%-9% higher than currently recognized. Furthermore, our survey added three new records of butterflies for the country (Eurema agave, Mithras hannelore, Melanis hillapana). In summary, this study not only supported the utility of DNA barcoding for the identification of the butterfly species of Argentina, but also highlighted several cases of both deep intraspecific and shallow interspecific divergence that should be studied in more detail.