Nuclear Intron Data Research Articles

Phylogeographical history of the Olive Woodpecker Dendropicos griseocephalus, a species widely distributed across Africa

Few studies have quantified the extent of genetic differentiation within widely distributed polytypic African bird species with disjunct ranges. Current knowledge indicates that high levels of genetic differentiation are found for such lineages but generalization of the pattern requires further comparisons with other co‐distributed taxa. We assessed the extent of phylogeographical structure across the range of the Olive Woodpecker Dendropicos griseocephalus using mitochondrial and nuclear intron data. The Olive Woodpecker occupies the forests of Central (Dendropicos griseocephalus ruwenzori) and Eastern (Dendropicos g. kilimensis) Africa, with a disjunct morphological lineage (Dendropicos g. griseocephalus) occurring in southern Africa. Each of the subspecies lineages can be diagnosed using morphology. Phylogenetic analyses of our sequence data recovered three monophyletic lineages with kilimensis sister to ruwenzori, and griseocephalus as sister to the clade uniting these two taxa. Molecular species delimitation methods and estimates of gene flow under the isolation‐with‐migration model suggest that the clade uniting the central and eastern subspecies may be recognized as distinct at the species level from the nominate subspecies, which is restricted to southern Africa. We conclude that D. griseocephalus (Boddaert, 1783) and D. ruwenzori (Sharpe, 1902) (including subspecies kilimensis) should be considered full species. The biogeographical pattern we uncover for the Olive Woodpecker differs from that of other co‐distributed widespread species both in terms of the order of sequence divergence of lineages occupying different areas of endemism in Africa, and in the timing of divergence, being younger (0.5–0.7 mya BP) than that recovered for the co‐distributed Square‐tailed Drongo Dicrurus ludwigii (0.9–1.6 mya BP).

Multilocus phylogeny of a cryptic radiation of Afrotropical long‐fingered bats (Chiroptera, Miniopteridae)

AbstractThe Old World bat family Miniopteridae comprises only the genus Miniopterus, which includes 20 currently recognized species from the Afrotropical realm and 15 species from Eurasia and Australasia. Since 2003, the number of recognized Miniopterus species has grown from 19 to 35, with most newly described species endemic to Madagascar and the Comoros Archipelago. We investigated genetic variation, phylogenetic relationships and clade membership in Miniopterus focusing on Afrotropical taxa. We generated mitochondrial cytochrome‐b (cyt‐b) and nuclear intron data (five genes) from 352 vouchered individuals collected at 78 georeferenced localities. Including 99 additional mitochondrial sequences from GenBank, we analysed a total of 25 recognized species. Mitochondrial genetic distances among cyt‐b‐supported clades averaged 9.3%, representing as many as five undescribed species. Multilocus coalescent delimitation strongly supported the genetic isolation of eight of nine tested unnamed clades. A large number of sampled clades in sub‐Saharan Africa are distributed wholly or partly in East Africa (nine of 13 clades), suggesting that Miniopterus diversity has been grossly underestimated. Although 25 of 27 cyt‐b and 23 of 25 nuclear gene tree lineages from the Afrotropics were strongly supported as monophyletic, a majority of deep nodes were poorly resolved in phylogenetic analyses. Long terminal branches subtending short backbone internodes in the phylogenetic analyses suggest a rapid radiation model of diversification. This hypothesis needs to be tested using more phylogenetically informative data.

Phylogeography and geographic variation in the Red-bellied Woodpecker (Melanerpes carolinus): characterization of mtDNA and plumage hybrid zones

ABSTRACT We used sequences of the mitochondrial ND2 gene, the nuclear TGFB-2 intron-5, and differences in plumage pattern to assay geographic variation in the Red-bellied Woodpecker (Melanerpes carolinus). Ten population samples from throughout its range indicated that ∼23% of the mitochondrial genetic variation was distributed among populations, largely due to the occurrence of a clade of haplotypes restricted to the Florida peninsula with a local frequency of 82%. In a hierarchical analysis, 37% of the mtDNA genetic variance was distributed between the Florida peninsula and elsewhere. A strong signal of population growth was detected in the non-peninsular portion of the range. Otherwise, geographic variation in haplotype frequencies was minor, with no evidence of isolation-by-distance. Nuclear intron data showed little variation among populations (Fst = 0.007). These results suggest recent secondary contact of 2 differentiated taxa following expansion from allopatric Pleistocene refugia. We modeled the ...

Historical relationships of three enigmatic phasianid genera (Aves: Galliformes) inferred using phylogenomic and mitogenomic data

The phylogeny of the Phasianidae (pheasants, partridges, and allies) has been studied extensively. However, these studies have largely ignored three enigmatic genera because of scarce DNA source material and limited overlapping phylogenetic data: blood pheasants (Ithaginis), snow partridges (Lerwa), and long-billed partridges (Rhizothera). Thus, phylogenetic positions of these three genera remain uncertain in what is otherwise a well-resolved phylogeny. Previous studies using different data types place Lerwa and Ithaginis in similar positions, but the absence of overlapping data means the relationship between them could not be inferred. Rhizothera was originally described in the genus Perdix (true partridges), although a partial cytochrome b (CYB) sequence suggests it is sister to Pucrasia (koklass pheasant). To identify robust relationships among Ithaginis, Lerwa, Rhizothera, and their phasianid relatives, we used 3692 ultra-conserved element (UCE) loci and complete mitogenomes from 19 species including previously hypothesized relatives of the three focal genera and representatives from all major phasianid clades. We used DNA extracted from historical specimen toepads for species that lacked fresh tissue in museum collections. Maximum likelihood and multispecies coalescent UCE analyses strongly supported Lerwa sister to a large clade which included Ithaginis at its base, and also including turkey, grouse, typical pheasants, tragopans, Pucrasia, and Perdix. Rhizothera was also in this clade, sister to a diverse group comprising Perdix, typical pheasants, Pucrasia, turkey and grouse. Mitogenomic genealogies differed from UCEs topologies, supporting a sister relationship between Ithaginis and Lerwa rather than a grade. The position of Rhizothera using mitogenomes depended on analytical choices. Unpartitioned and codon-based analyses placed Rhizothera sister to a tragopan clade, whereas a partitioned DNA model of the mitogenome was congruent with UCE results. In all mitogenome analyses, Pucrasia was sister to a clade including Perdix and the typical pheasants with high support, in contrast to UCEs and published nuclear intron data. Due to the strong support and consistent topology provided by all UCE analyses, we have identified phylogenetic relationships of these three enigmatic, poorly-studied, phasianid taxa.

Analysis of a Rapid Evolutionary Radiation Using Ultraconserved Elements: Evidence for a Bias in Some Multispecies Coalescent Methods

Rapid evolutionary radiations are expected to require large amounts of sequence data to resolve. To resolve these types of relationships many systematists believe that it will be necessary to collect data by next-generation sequencing (NGS) and use multispecies coalescent ("species tree") methods. Ultraconserved element (UCE) sequence capture is becoming a popular method to leverage the high throughput of NGS to address problems in vertebrate phylogenetics. Here we examine the performance of UCE data for gallopheasants (true pheasants and allies), a clade that underwent a rapid radiation 10-15 Ma. Relationships among gallopheasant genera have been difficult to establish. We used this rapid radiation to assess the performance of species tree methods, using ∼600 kilobases of DNA sequence data from ∼1500 UCEs. We also integrated information from traditional markers (nuclear intron data from 15 loci and three mitochondrial gene regions). Species tree methods exhibited troubling behavior. Two methods [Maximum Pseudolikelihood for Estimating Species Trees (MP-EST) and Accurate Species TRee ALgorithm (ASTRAL)] appeared to perform optimally when the set of input gene trees was limited to the most variable UCEs, though ASTRAL appeared to be more robust than MP-EST to input trees generated using less variable UCEs. In contrast, the rooted triplet consensus method implemented in Triplec performed better when the largest set of input gene trees was used. We also found that all three species tree methods exhibited a surprising degree of dependence on the program used to estimate input gene trees, suggesting that the details of likelihood calculations (e.g., numerical optimization) are important for loci with limited phylogenetic information. As an alternative to summary species tree methods we explored the performance of SuperMatrix Rooted Triple - Maximum Likelihood (SMRT-ML), a concatenation method that is consistent even when gene trees exhibit topological differences due to the multispecies coalescent. We found that SMRT-ML performed well for UCE data. Our results suggest that UCE data have excellent prospects for the resolution of difficult evolutionary radiations, though specific attention may need to be given to the details of the methods used to estimate species trees.

Phylogeography of a Morphologically Cryptic Golden Mole Assemblage from South-Eastern Africa.

The Greater Maputaland-Pondoland-Albany (GMPA) region of southern Africa was recently designated as a centre of vertebrate endemism. The phylogeography of the vertebrate taxa occupying this region may provide insights into the evolution of faunal endemism in south-eastern Africa. Here we investigate the phylogeographic patterns of an understudied small mammal species assemblage (Amblysomus) endemic to the GMPA, to test for cryptic diversity within the genus, and to better understand diversification across the region. We sampled specimens from 50 sites across the distributional range of Amblysomus, with emphasis on the widespread A. hottentotus, to analyse geographic patterns of genetic diversity using mitochondrial DNA (mtDNA) and nuclear intron data. Molecular dating was used to elucidate the evolutionary and phylogeographic history of Amblysomus. Our phylogenetic reconstructions show that A. hottentotus comprises several distinct lineages, or evolutionarily significant units (ESUs), some with restricted geographic ranges and thus worthy of conservation attention. Divergence of the major lineages dated to the early Pliocene, with later radiations in the GMPA during the late-Pliocene to early-Pleistocene. Evolutionary diversification within Amblysomus may have been driven by uplift of the Great Escarpment c. 5–3 million years ago (Ma), habitat changes associated with intensification of the east-west rainfall gradient across South Africa and the influence of subsequent global climatic cycles. These drivers possibly facilitated geographic spread of ancestral lineages, local adaptation and vicariant isolation. Our study adds to growing empirical evidence identifying East and southern Africa as cradles of vertebrate diversity.

An empirical comparison of character‐based and coalescent‐based approaches to species delimitation in a young avian complex

The process of discovering species is a fundamental responsibility of systematics. Recently, there has been a growing interest in coalescent-based methods of species delimitation aimed at objectively identifying species early in the divergence process. However, few empirical studies have compared these new methods with character-based approaches for discovering species. In this study, we applied both a character-based and a coalescent-based approaches to delimit species in a closely related avian complex, the light-vented/Taiwan bulbul (Pycnonotus sinensis/Pycnonotus taivanus). Population aggregation analyses of plumage, mitochondrial and 13 nuclear intron character data sets produced conflicting species hypotheses with plumage data suggesting three species, mitochondrial data suggesting two species, and nuclear intron data suggesting one species. Such conflict is expected among recently diverged species, and by integrating all sources of data, we delimited three species verified with independently congruent character evidence as well as a more weakly supported fourth species identified by a single character. Attempts to validate species hypothesis using Bayesian Phylogenetics and Phylogeography (BPP), a coalescent-based method of species delimitation, revealed several issues that can seemingly affect statistical support for species recognition. We found that θ priors had a dramatic impact on speciation probabilities, with lower values consistently favouring splitting and higher values consistently favouring lumping. More resolved guide trees also resulted in overall higher speciation probabilities. Finally, we found suggestive evidence that BPP is sensitive to the divergent effects of nonrandom mating caused by intraspecific processes such as isolation-with-distance, and therefore, BPP may not be a conservative method for delimiting independently evolving population lineages. Based on these concerns, we questioned the reliability of BPP results and based our conclusions about species limits exclusively on character data.

High-latitude connectivity of the scleractinian coral Acropora tenuis in the south-western Indian Ocean, identified using nuclear intron and mitochondrial sequence data

All scleractinian corals in southern Africa occupy the high-latitude margins of their biogeographical range, with some straddling multiple biogeographical regions. Connectivity among these populations is essential to maintain genetic diversity, and thus their conservation value. Therefore, the aim of this study was to elucidate the genetic diversity, connectivity and structure of Acropora sp. populations. Acropora sp. were sampled at four intertidal and subtidal sites over a distance of ~320 km, and assessed using primers that amplify the 3/550 nuclear intron found within the carbonic anhydrase gene and the mitochondrially encoded cytochrome oxidase I region of DNA. Results resolved the presence of four putative Acropora clades, genetically disparate according to 3/550 nuclear intron data. In contrast to this, cytochrome oxidase I data confirmed the presence of a single species: A. tenuis. Cytochrome oxidase I data were highly conserved at species level, supporting previous notions that this gene may not be suitable for species level delineation in acroporids. Genetic structuring and diversity of A. tenuis among the four sites revealed similar characteristics to Acropora austera of northern KwaZulu-Natal and Mozambique. Little genetic differentiation was found between all possible regions, suggesting no significant genetic differentiation across the biogeographical regions tested. A higher degree of isolation and lower differentiation and moderate to high connectivity indices suggested periods of lower-latitude colonisation and periods of population attrition may be a feature in the ecology of this species.

Phylogeography and postglacial recolonization of Europe by Rhinolophus hipposideros: evidence from multiple genetic markers

The demographic history of Rhinolophus hipposideros (lesser horseshoe bat) was reconstructed across its European, North African and Middle-Eastern distribution prior to, during and following the most recent glaciations by generating and analysing a multimarker data set. This data set consisted of an X-linked nuclear intron (Bgn; 543bp), mitochondrial DNA (cytb-tRNA-control region; 1630bp) and eight variable microsatellite loci for up to 373 individuals from 86 localities. Using this data set of diverse markers, it was possible to determine the species' demography at three temporal stages. Nuclear intron data revealed early colonization into Europe from the east, which pre-dates the Quaternary glaciations. The mtDNA data supported multiple glacial refugia across the Mediterranean, the largest of which were found in the Ibero-Maghreb region and an eastern location (Anatolia/Middle East)-that were used by R.hipposideros during the most recent glacial cycles. Finally, microsatellites provided the most recent information on these species' movements since the Last Glacial Maximum and suggested that lineages that had diverged into glacial refugia, such as in the Ibero-Maghreb region, have remained isolated. These findings should be used to inform future conservation management strategies for R.hipposideros and show the power of using a multimarker data set for phylogeographic studies.

First evaluation of mitochondrial DNA as a marker for phylogeographic studies of Calcarea: a case study from Leucetta chagosensis

In most animals mitochondrial DNA (mtDNA) evolves much faster than nuclear DNA. Therefore, and because of its shorter coalescent time, mitochondrial (mt) markers provide better resolution to trace more recent evolutionary events compared to nuclear DNA. But in contrast to most other Metazoa, previous studies suggested that in sponges mitochondrial sequence evolution is much slower, making mtDNA less suitable for studies at the intraspecific level. However, these observations were made in the class Demospongiae and so far no data exist for calcareous sponges (Class Calcarea). We here provide the first study that evaluates intraspecific mt sequence variation in Calcarea. We focus on arguably the best-studied species Leucetta chagosensis, for which three nuclear DNA marker datasets existed previously. We here sequenced the partial mitochondrial cytochrome oxidase subunit III gene (cox3). Our analyses reveal an unexpected variability of up to 8.5% in this mitochondrial marker. In contrast to other sponges where this marker evolves considerable slower than the nuclear internal transcribed spacer region (ITS), we found that cox3 in L. chagosensis evolves about five times as fast as ITS. The variability is similar to that of nuclear intron data of the species. The phylogeny inferred with cox3 is congruent with other markers, but separates earlier reported genetic groups much more distinctively than nuclear DNA. This provides further evidence for cryptic speciation in L. chagosensis. All these features make calcarean mtDNA exceptional among sponges and show its suitability for phylogeographic studies and potential as a species-specific (DNA barcoding) marker to distinguish morphologically identical cryptic species.

Inclusion of nuclear intron sequence data helps to identify the Asian sister group of New World pitvipers

Despite much effort towards resolving the molecular phylogenetic tree for pitvipers, some aspects remain unresolved. In particular, the sister group of the diverse New World radiation has remained impossible to identify with any certainty. In this study, which for the first time includes nuclear intron data from all major groups of Asian pitvipers as well as representatives of the New World radiation, Bayesian inference allows Gloydius to be identified as the most likely sister group to the New World radiation and sheds light on other ambiguous relationships among the Old World pitvipers. The sister group relationship of "Ovophis"okinavensis and "Trimeresurus"gracilis is confirmed by the addition of nuclear genes, and we hypothesise that they form a sister group to the Gloydius+New World clade, best supported when the phylogenetic signal from gaps is included in the form of a simple-coded matrix.

Phylogenetics of barbets (Aves: Piciformes) based on nuclear and mitochondrial DNA sequence data

A combination of nuclear (β-fibrinogen intron 7; 938 bp) and mitochondrial (cytochrome b; 1045 bp) DNA sequence data analyzed with model-based phylogenetic methods yields a hypothesis of barbet and toucan relationships supported by high Bayesian posterior probabilities and several synapomorphic indels in the nuclear intron data. The basal topology differs from previous morphology and mitochondrial DNA sequence based hypotheses, but is consistent with DNA–DNA hybridization results. The mitochondrial DNA sequence data provide resolution at the tips of the tree, but when analyzed alone, yield a different basal topology than the combined data. However, the basal nodes in the mitochondrial-based tree have little statistical support. Combined data analysis produced strong statistical support at basal nodes and a relatively simple geographic structure. Barbets from each of the three tropical regions are monophyletic, but the Old World barbets are paraphyletic. The African and New World clades are sister taxa, and the Asian clade is basal. This geographic structure indicates that similarities in plumage, voice, and behavior between Gymnobucco and Calorhamphus are convergent. The data are not conclusive, but suggest paraphyly of the New World barbets. Only 4% of the Bayesian posterior distribution unites Semnornis with the other New World barbets.

NUCLEAR INTRON SEQUENCES FOR PHYLOGENETICS OF CLOSELY RELATED MAMMALS: AN EXAMPLE USING THE PHYLOGENY OFMUS

DNA sequence data from 2 nuclear introns (mitochondrial malate dehydrogenase intron 7 and acid phosphatase type V intron 2) were collected for 9 taxonomic units of rodents, 8 of which were in the genus Mus. Data were used to infer phylogenetic history of the 9 taxa, which was then compared with a consensus phylogeny taken from previous literature. In general, the nuclear intron data provided strong support (bootstrap proportions > 80) for relationships accepted in the consensus tree. The intron data disagreed with previous mitochondrial data in that 2 M. m. domesticus sequences were paraphyletic with respect to M. m. castaneus and M. m. musculus. This paraphyly, if real, likely reflects the longer lineage-sorting time of the nuclear genome compared with the mitochondrial genome. Introns seem to provide an attractive source of nuclear DNA sequence data for phylogenetic analysis, but mitochondrial data are likely to be superior for very recent divergences.

Predicting nuclear gene coalescence from mitochondrial data: the three-times rule.

Coalescence theory predicts when genetic drift at nuclear loci will result in fixation of sequence differences to produce monophyletic gene trees. However, the theory is difficult to apply to particular taxa because it hinges on genetically effective population size, which is generally unknown. Neutral theory also predicts that evolution of monophyly will be four times slower in nuclear than in mitochondrial genes primarily because genetic drift is slower at nuclear loci. Variation in mitochondrial DNA (mtDNA) within and between species has been studied extensively, but can these mtDNA data be used to predict coalescence in nuclear loci? Comparison of neutral theories of coalescence of mitochondrial and nuclear loci suggests a simple rule of thumb. The "three-times rule" states that, on average, most nuclear loci will be monophyletic when the branch length leading to the mtDNA sequences of a species is three times longer than the average mtDNA sequence diversity observed within that species. A test using mitochondrial and nuclear intron data from seven species of whales and dolphins suggests general agreement with predictions of the three-times rule. We define the coalescence ratio as the mitochondrial branch length for a species divided by intraspecific mtDNA diversity. We show that species with high coalescence ratios show nuclear monophyly, whereas species with low ratios have polyphyletic nuclear gene trees. As expected, species with intermediate coalescence ratios show a variety of patterns. Especially at very high or low coalescence ratios, the three-times rule predicts nuclear gene patterns that can help detect the action of selection. The three-times rule may be useful as an empirical benchmark for evaluating evolutionary processes occurring at multiple loci.