Sérgio Luiz Pereira

Phylogenetics, biogeography and classification of, and character evolution in, gamebirds (Aves: Galliformes) : effects of character exclusion, data partitioning and missing data

The phylogenetic relationships, biogeography and classification of, and morpho‐behavioral (M/B) evolution in, gamebirds (Aves: Galliformes) are investigated. In‐group taxa (rooted on representatives of the Anseriformes) include 158 species representing all suprageneric galliform taxa and 65 genera. The characters include 102 M/B attributes and 4452 nucleic acid base pairs from mitochondrial cytochrome b (CYT B), NADH dehydrogenase subunit 2 (ND2), 12S ribosomal DNA (12S) and control region (CR), and nuclear ovomucoid intron G (OVO‐G). Analysis of the combined character data set yielded a single, completely resolved cladogram that had the highest levels of jackknife support, which suggests a need for a revised classification for the phasianine galliforms. Adding 102 M/B characters to the combined CYT B and ND2 partitions (2184 characters) decisively overturns the topology suggested by analysis of the two mtDNA partitions alone, refuting the view that M/B characters should be excluded from phylogenetic analyses because of their relatively small number and putative character state ambiguity. Exclusion of the OVO‐G partition (with > 70% missing data) from the combined data set had no effect on cladistic structure, but slightly lowered jackknife support at several nodes. Exclusion of third positions of codons in an analysis of a CYT B + ND2 partition resulted in a massive loss of resolution and support, and even failed to recover the monophyly of the Galliformes with jackknife support. A combined analysis of putatively less informative, “non‐coding” characters (CYT B/ND2 third position sites + CR +12S + OVO‐G sequences) yielded a highly resolved consensus cladogram congruent with the combined‐evidence cladogram. Traditionally recognized suprageneric galliform taxa emerging in the combined cladogram are: the families Megapodiidae (megapodes), Cracidae (cracids), Numididae (guineafowls), Odontophoridae (New World quails) and Phasianidae (pheasants, pavonines, partridges, quails, francolins, spurfowls and grouse) and the subfamilies Cracinae (curassows, chachalacas and the horned guan), Penelopinae (remaining guans), Pavoninae sensu lato (peafowls, peacock pheasants and argus pheasants), Tetraoninae (grouse) and Phasianinae (pheasants minus Gallus). The monophyly of some traditional groupings (e.g., the perdicinae: partridges/quails/francolins) is rejected decisively, contrasted by the emergence of other unexpected groupings. The most remarkable phylogenetic results are the placement of endemic African galliforms as sisters to geographically far‐distant taxa in Asia and the Americas. Biogeographically, the combined‐data cladogram supports the hypothesis that basal lineages of galliforms diverged prior to the Cretaceous/Tertiary (K‐T) Event and that the subsequent cladogenesis was influenced by the break‐up of Gondwana. The evolution of gamebirds in Africa, Asia and the Americas has a far more complicated historical biogeography than suggested to date. With regard to character evolution: spurs appear to have evolved at least twice within the Galliformes; a relatively large number of tail feathers (≥ 14) at least three times; polygyny at least twice; and sexual dimorphism many times.

Phylogenetic relationships and divergence times of Charadriiformes genera: multigene evidence for the Cretaceous origin of at least 14 clades of shorebirds

Comparative study of character evolution in the shorebirds is presently limited because the phylogenetic placement of some enigmatic genera remains unclear. We therefore used Bayesian methods to obtain a well-supported phylogeny of 90 recognized genera using 5 kb of mitochondrial and nuclear sequences. The tree comprised three major clades: Lari (gulls, auks and allies plus buttonquails) as sister to Scolopaci (sandpipers, jacanas and allies), and in turn sister to Charadrii (plovers, oystercatchers and allies), as in previous molecular studies. Plovers and noddies were not recovered as monophyletic assemblages, and the Egyptian plover Pluvianus is apparently not a plover. Molecular dating using multiple fossil constraints suggests that the three suborders originated in the late Cretaceous between 79 and 102 Mya, and at least 14 lineages of modern shorebirds survived the mass extinction at the K/T boundary. Previous difficulties in determining the phylogenetic relationships of enigmatic taxa reflect the fact that they are well-differentiated relicts of old, genus-poor lineages. We refrain from suggesting systematic revisions for shorebirds at this time because gene trees may fail to recover the species tree when long branches are connected to deep, shorter branches, as is the case for some of the enigmatic taxa.

Multiple gene evidence for expansion of extant penguins out of Antarctica due to global cooling

Classic problems in historical biogeography are where did penguins originate, and why are such mobile birds restricted to the Southern Hemisphere? Competing hypotheses posit they arose in tropical–warm temperate waters, species-diverse cool temperate regions, or in Gondwanaland ∼100 mya when it was further north. To test these hypotheses we constructed a strongly supported phylogeny of extant penguins from 5851 bp of mitochondrial and nuclear DNA. Using Bayesian inference of ancestral areas we show that an Antarctic origin of extant taxa is highly likely, and that more derived taxa occur in lower latitudes. Molecular dating estimated penguins originated about 71 million years ago in Gondwanaland when it was further south and cooler. Moreover, extant taxa are inferred to have originated in the Eocene, coincident with the extinction of the larger-bodied fossil taxa as global climate cooled. We hypothesize that, as Antarctica became ice-encrusted, modern penguins expanded via the circumpolar current to oceanic islands within the Antarctic Convergence, and later to the southern continents. Thus, global cooling has had a major impact on penguin evolution, as it has on vertebrates generally. Penguins only reached cooler tropical waters in the Galapagos about 4 mya, and have not crossed the equatorial thermal barrier.

Phylogenetic Relationships and Historical Biogeography of Neotropical Parrots (Psittaciformes: Psittacidae: Arini) Inferred from Mitochondrial and Nuclear DNA Sequences

Previous hypotheses of phylogenetic relationships among Neotropical parrots were based on limited taxon sampling and lacked support for most internal nodes. In this study we increased the number of taxa (29 species belonging to 25 of the 30 genera) and gene sequences (6388 base pairs of RAG-1, cyt b, NADH2, ATPase 6, ATPase 8, COIII, 12S rDNA, and 16S rDNA) to obtain a stronger molecular phylogenetic hypothesis for this group of birds. Analyses of the combined gene sequences using maximum likelihood and Bayesian methods resulted in a well-supported phylogeny and indicated that amazons and allies are a sister clade to macaws, conures, and relatives, and these two clades are in turn a sister group to parrotlets. Key morphological and behavioral characters used in previous classifications were mapped on the molecular tree and were phylogenetically uninformative. We estimated divergence times of taxa using the molecular tree and Bayesian and penalized likelihood methods that allow for rate variation in DNA substitutions among sites and taxa. Our estimates suggest that the Neotropical parrots shared a common ancestor with Australian parrots 59 Mya (million of years ago; 95% credibility interval (CrI) 66, 51 Mya), well before Australia separated from Antarctica and South America, implying that ancestral parrots were widespread in Gondwanaland. Thus, the divergence of Australian and Neotropical parrots could be attributed to vicariance. The three major clades of Neotropical parrots originated about 50 Mya (95% CrI 57, 41 Mya), coinciding with periods of higher sea level when both Antarctica and South America were fragmented with transcontinental seaways, and likely isolated the ancestors of modern Neotropical parrots in different regions in these continents. The correspondence between major paleoenvironmental changes in South America and the diversification of genera in the clade of amazons and allies between 46 and 16 Mya suggests they diversified exclusively in South America. Conversely, ancestors of parrotlets and of macaws, conures, and allies may have been isolated in Antarctica and/or the southern cone of South America, and only dispersed out of these southern regions when climate cooled and Antarctica became ice-encrusted about 35 Mya. The subsequent radiation of macaws and their allies in South America beginning about 28 Mya (95% CrI 22, 35 Mya) coincides with the uplift of the Andes and the subsequent formation of dry, open grassland habitats that would have facilitated ecological speciation via niche expansion from forested habitats.

Mitochondrial and Nuclear DNA Sequences Support a Cretaceous Origin of Columbiformes and a Dispersal-Driven Radiation in the Paleogene

Phylogenetic relationships among genera of pigeons and doves (Aves, Columbiformes) have not been fully resolved because of limited sampling of taxa and characters in previous studies. We therefore sequenced multiple nuclear and mitochondrial DNA genes totaling over 9000 bp from 33 of 41 genera plus 8 outgroup taxa, and, together with sequences from 5 other pigeon genera retrieved from GenBank, recovered a strong phylogenetic hypothesis for the Columbiformes. Three major clades were recovered with the combined data set, comprising the basally branching New World pigeons and allies (clade A) that are sister to Neotropical ground doves (clade B), and the Afro-Eurasian and Australasian taxa (clade C). None of these clades supports the monophyly of current families and subfamilies. The extinct, flightless dodo and solitaires (Raphidae) were embedded within pigeons and doves (Columbidae) in clade C, and monophyly of the subfamily Columbinae was refuted because the remaining subfamilies were nested within it. Divergence times estimated using a Bayesian framework suggest that Columbiformes diverged from outgroups such as Apodiformes and Caprimulgiformes in the Cretaceous before the mass extinction that marks the end of this period. Bayesian and maximum likelihood inferences of ancestral areas, accounting for phylogenetic uncertainty and divergence times, respectively, favor an ancient origin of Columbiformes in the Neotropical portion of what was then Gondwana. The radiation of modern genera of Columbiformes started in the Early Eocene to the Middle Miocene, as previously estimated for other avian groups such as ratites, tinamous, galliform birds, penguins, shorebirds, parrots, passerine birds, and toucans. Multiple dispersals of more derived Columbiformes between Australasian and Afro-Eurasian regions are required to explain current distributions.

Low number of mitochondrial pseudogenes in the chicken (Gallus gallus) nuclear genome: implications for molecular inference of population history and phylogenetics

BackgroundMitochondrial DNA has been detected in the nuclear genome of eukaryotes as pseudogenes, or Numts. Human and plant genomes harbor a large number of Numts, some of which have high similarity to mitochondrial fragments and thus may have been inadvertently included in population genetic and phylogenetic studies using mitochondrial DNA. Birds have smaller genomes relative to mammals, and the genome-wide frequency and distribution of Numts is still unknown. The release of a preliminary version of the chicken (Gallus gallus) genome by the Genome Sequencing Center at Washington University, St. Louis provided an opportunity to search this first avian genome for the frequency and characteristics of Numts relative to those in human and plants.ResultsWe detected at least 13 Numts in the chicken nuclear genome. Identities between Numts and mitochondrial sequences varied from 58.6 to 88.8%. Fragments ranged from 131 to 1,733 nucleotides, collectively representing only 0.00078% of the nuclear genome. Because fewer Numts were detected in the chicken nuclear genome, they do not represent all regions of the mitochondrial genome and are not widespread in all chromosomes. Nuclear integrations in chicken seem to occur by a DNA intermediate and in regions of low gene density, especially in macrochromosomes.ConclusionThe number of Numts in chicken is low compared to those in human and plant genomes, and is within the range found for most sequenced eukaryotic genomes. For chicken, PCR amplifications of fragments of about 1.5 kilobases are highly likely to represent true mitochondrial amplification. Sequencing of these fragments should expose the presence of unusual features typical of pseudogenes, unless the nuclear integration is very recent and has not yet been mutated. Metabolic selection for compact genomes with reduced repetitive DNA and gene-poor regions where Numts occur may explain their low incidence in birds.

Combined Nuclear and Mitochondrial DNA Sequences Resolve Generic Relationships within the Cracidae (Galliformes, Aves)

The Cracidae is one of the most endangered and distinctive bird families in the Neotropics, yet the higher relationships among taxa remain uncertain. The molecular phylogeny of its 11 genera was inferred using 10,678 analyzable sites (5,412 from seven different mitochondrial segments and 5,266 sites from four nuclear genes). We performed combinability tests to check conflicts in phylogenetic signals of separate genes and genomes. Phylogenetic analysis showed that the unrooted tree of ((curassows, horned guan) (guans, chachalacas)) was favored by most data partitions and that different data partitions provided support for different parts of the tree. In particular, the concatenated mitochondrial DNA (mtDNA) genes resolved shallower nodes, whereas the combined nuclear sequences resolved the basal connections among the major clades of curassows, horned guan, chachalacas, and guans. Therefore, we decided that for the Cracidae all data should be combined for phylogenetic analysis. Maximum parsimony (MP), maximum likelihood (ML), and Bayesian analyses of this large data set produced similar trees. The MP tree indicated that guans are the sister group to (horned guan, (curassows, chachalacas)), whereas the ML and Bayesian analysis recovered a tree where the horned guan is a sister clade to curassows, and these two clades had the chachalacas as a sister group. Parametric bootstrapping showed that alternative trees previously proposed for the cracid genera are significantly less likely than our estimate of their relationships. A likelihood ratio test of the hypothesis of a molecular clock for cracid mtDNA sequences using the optimal ML topology did not reject rate constancy of substitutions through time. We estimated cracids to have originated between 64 and 90 million years ago (MYA), with a mean estimate of 76 MYA. Diversification of the genera occurred approximately 41-3 MYA, corresponding with periods of global climate change and other Earth history events that likely promoted divergences of higher level taxa.

VICARIANT SPECIATION OF CURASSOWS (AVES, CRACIDAE): A HYPOTHESIS BASED ON MITOCHONDRIAL DNA PHYLOGENY

Abstract The curassows comprise 14 species of sedentary Neotropical birds classified in four genera (Crax, Nothocrax, Mitu, and Pauxi) in the family Cracidae. Congeneric species have a striking pattern of allopatric distributions that might be attributable to vicariance, dispersal, or a combination of the two. To test those biogeographic hypotheses, a strongly supported phylogeny was needed, so that existing problems of taxonomic rank could be solved and a better understanding of the groups evolutionary history attained. We therefore estimated the phylogenetic relationships of all 14 species, on the basis of 6,929 sites of six different mitochondrial DNA regions, and reassessed the status of the four genera. Sequences from the ND4 gene favored a tree that was highly incongruent with the tree recovered using the other five gene regions. However, when the ND4 sequences were concatenated with the sequences of the other genes, the optimal phylogeny was unchanged from that derived for the other genes. That combined tree was divided into two well-supported clades: one containing the seven species of Crax and the other containing the monospecific genus Nothocrax, as sister to a clade of the Mitu and Pauxi species. Mitu and Pauxi are not reciprocally monophyletic, which appears to be attributable to a distant hybridization event and a transfer of Mitu mtDNA into P. unicornis. We estimated divergence times; the diversification of curassow seems to have occurred from the Middle Miocene to the end of the Pliocene (9.5 to 1.6 Ma). Vicariance—following marine transgressions, the rise of the Andes, and subsequent changes in river basins in South America—seems to be the major mode of isolation that favored allopatric speciation in the group.

Mitochondrial genome organization and vertebrate phylogenetics

With the advent of DNA sequencing techniques the organization of the vertebrate mitochondrial genome shows variation between higher taxonomic levels. The most conserved gene order is found in placental mammals, turtles, fishes, some lizards and Xenopus. Birds, other species of lizards, crocodilians, marsupial mammals, snakes, tuatara, lamprey, and some other amphibians and one species of fish have gene orders that are less conserved. The most probable mechanism for new gene rearrangements seems to be tandem duplication and multiple deletion events, always associated with tRNA sequences. Some new rearrangements seem to be typical of monophyletic groups and the use of data from these groups may be useful for answering phylogenetic questions involving vertebrate higher taxonomic levels. Other features such as the secondary structure of tRNA, and the start and stop codons of protein-coding genes may also be useful in comparisons of vertebrate mitochondrial genomes.

DNA evidence for a Paleocene origin of the Alcidae (Aves: Charadriiformes) in the Pacific and multiple dispersals across northern oceans.

The Alcidae is a group of marine, wing-propelled diving birds known as auks that are distributed along the coasts of the northern oceans. It has been suggested that auks originated in the Pacific coastal shores as early as the Miocene, and dispersed to the Atlantic either through the Arctic coasts of Eurasia and North America (northern dispersal route), or through upwelling zones in the coastal areas of California to Florida (southern dispersal route), before the closure of the Isthmus of Panama in the Pliocene. These hypotheses have not been tested formally because proposed phylogenies failed to recover fully bifurcating, well-supported phylogenetic relationships among and within genera. We therefore constructed a large data set of mitochondrial and nuclear DNA sequences for 21 of the 23 species of extant auks. We also included sequences from two other extant and one extinct species retrieved from GenBank. Our analyses recovered a well-supported phylogenetic hypothesis among and within genera. Aethia is the only genus for which we could not obtain strong support for species relationships, probably due to incomplete lineage sorting. By applying a Bayesian method of molecular dating that allows for rate variation across lineages and genes, we showed that auks became an independent lineage in the Early Paleocene and radiated gradually from the Early Eocene to the Quaternary. Reconstruction of ancestral areas strongly suggests that auks originated in the Pacific during the Paleocene. The southern dispersal route seems to have favored the subsequent colonization of the northern Atlantic Ocean during the Eocene and Oligocene. The northern route across the Arctic Ocean was probably only used more recently after the opening of the Norwegian Sea in the Middle Miocene and the opening of the Bering Strait in the Late Miocene. We postulate that the ancestors of auks lived in a warmer world than that currently occupied by auks, and became gradually adapted to feeding in cool marine currents with high biomass productivity. Hence, warmer tropical waters are now a barrier for the dispersal of auks into the Southern Hemisphere, as it is for penguins in the opposite direction.