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Gene-tree-estimation error is a major concern for coalescent methods of phylogenetic inference. We sampled eight empirical studies of ancient lineages with diverse numbers of taxa and genes for which the original authors applied one or more coalescent methods. We found that the average pairwise congruence among gene trees varied greatly both between studies and also often within a study. We recommend that presenting plots of pairwise congruence among gene trees in a dataset be treated as a standard practice for empirical coalescent studies so that readers can readily assess the extent and distribution of incongruence among gene trees. ASTRAL-based coalescent analyses generally outperformed MP-EST and STAR with respect to both internal consistency (congruence between analyses of subsamples of genes with the complete dataset of all genes) and congruence with the concatenation-based topology. We evaluated the approach of subsampling gene trees that are, on average, more congruent with other gene trees as a method to reduce artifacts caused by gene-tree-estimation errors on coalescent analyses. We suggest that this method is well suited to testing whether gene-tree-estimation error is a primary cause of incongruence between concatenation- and coalescent-based results, to reconciling conflicting phylogenetic results based on different coalescent methods, and to identifying genes affected by artifacts that may then be targeted for reciprocal illumination. We provide scripts that automate the process of calculating pairwise gene-tree incongruence and subsampling trees while accounting for differential taxon sampling among genes. Finally, we assert that multiple tree-search replicates should be implemented as a standard practice for empirical coalescent studies that apply MP-EST. [ABSTRACT FROM AUTHOR]

It has recently been concluded that phylogenomic data from 310 nuclear genes support the clade of (Amborellales, Nymphaeales) as sister to the remaining angiosperms and that shortcut coalescent phylogenetic methods outperformed concatenation for these data. We falsify both of those conclusions here by demonstrating that discrepant results between the coalescent and concatenation analyses are primarily caused by the coalescent methods applied (MP-EST and STAR) not being robust to the highly divergent and often mis-rooted gene trees that were used. This result reinforces the expectation that low amounts of phylogenetic signal and methodological artifacts in gene-tree reconstruction can be more problematic for shortcut coalescent methods than is the assumption of a single hierarchy for all genes by concatenation methods when these approaches are applied to ancient divergences in empirical studies. We also demonstrate that a third coalescent method, ASTRAL, is more robust to mis-rooted gene trees than MP-EST or STAR, and that both Observed Variability (OV) and Tree Independent Generation of Evolutionary Rates (TIGER), which are two character subsampling procedures, are biased in favor of characters with highly asymmetrical distributions of character states when applied to this dataset. We conclude that enthusiastic application of novel tools is not a substitute for rigorous application of first principles, and that trending methods (e.g., shortcut coalescent methods applied to ancient divergences, tree-independent character subsampling), may be novel sources of previously under-appreciated, systematic errors. [ABSTRACT FROM AUTHOR]

[Display omitted] • Summary coalescent methods are not robust to gene-tree misrooting errors. • Summary coalescent methods are not robust to homology errors. • Summary coalescent methods are not robust to differential sampling of taxa. • Additional conflicting retroelement insertions are revealed. • 20,850 loci and 4345 retroelements do not robustly resolve palaeognath phylogeny. Phylogenomic analyses of ancient rapid radiations can produce conflicting results that are driven by differential sampling of taxa and characters as well as the limitations of alternative analytical methods. We re-examine basal relationships of palaeognath birds (ratites and tinamous) using recently published datasets of nucleotide characters from 20,850 loci as well as 4301 retroelement insertions. The original studies attributed conflicting resolutions of rheas in their inferred coalescent and concatenation trees to concatenation failing in the anomaly zone. By contrast, we find that the coalescent-based resolution of rheas is premised upon extensive gene-tree estimation errors. Furthermore, retroelement insertions contain much more conflict than originally reported and multiple insertion loci support the basal position of rheas found in concatenation trees, while none were reported in the original publication. We demonstrate how even remarkable congruence in phylogenomic studies may be driven by long-branch misplacement of a divergent outgroup, highly incongruent gene trees, differential taxon sampling that can result in gene-tree misrooting errors that bias species-tree inference, and gross homology errors. What was previously interpreted as broad, robustly supported corroboration for a single resolution in coalescent analyses may instead indicate a common bias that taints phylogenomic results across multiple genome-scale datasets. The updated retroelement dataset now supports a species tree with branch lengths that suggest an ancient anomaly zone, and both concatenation and coalescent analyses of the huge nucleotide datasets fail to yield coherent, reliable results in this challenging phylogenetic context. [ABSTRACT FROM AUTHOR]

• Up to 86% of internal gene-tree branches are dubiously or arbitrarily resolved. • Collapsing branches increased species-tree coalescent branch lengths by up to 455%. • Severe and clearly justified collapsing method for likelihood: 0% SH-like aLRT. • Severe and clearly justified collapsing method for parsimony: strict consensus. • Collapsing can improve congruence between coalescent and concatenation analyses. In two-step coalescent analyses of phylogenomic data, gene-tree topologies are treated as fixed prior to species-tree inference. Although all gene-tree conflict is assumed to be caused by lineage sorting when applying these methods, in empirical datasets much of the conflict can be caused by estimation error. Weakly supported and even arbitrarily resolved clades are important sources of this estimation error for gene trees inferred from few informative characters relative to the number of sampled terminals, and the resulting extraneous conflict among gene trees can negatively impact species-tree inference. In this study, we quantified the relative severity of alternative methods for collapsing gene-tree branches for seven empirical datasets and quantified their effects on species-tree inference. The branch-collapsing methods that we employed were based on the strict consensus of optimal topologies, various bootstrap thresholds, and 0% approximate likelihood ratio test (SH-like aLRT) support. Up to 86% of internal gene-tree branches are dubiously or arbitrarily resolved in reanalyses of these published phylogenomic datasets, and collapsing these branches increased inferred species-tree coalescent branch lengths by up to 455%. For two datasets, the longer inferred branch lengths sometimes impacted inference of anomaly-zone conditions. Although branch-collapsing methods did not consistently affect the species-tree topology, they often increased branch support. The more severe and clearly justified gene-tree branch-collapsing methods, which we recommend be broadly applied for two-step coalescent analyses, are use of the strict consensus in parsimony analyses and the collapse clades with 0% SH-like aLRT support in likelihood analyses. Collapsing dubiously or arbitrarily resolved branches in gene trees sometimes improved congruence between coalescent-based results and concatenation trees. In such cases, we contend that the resolution provided by concatenation should be preferred and that incomplete lineage sorting is a poor explanation for the initial conflict between phylogenetic approaches. [ABSTRACT FROM AUTHOR]

• Partitioned coalescence support (PCS) is automated for genome-scale datasets. • PCS summarizes the support and conflict among gene trees in coalescence analyses. • PCS detects biases in phylogenomic support provided by outlier gene trees. • PCS quantifies the effects of missing taxa in gene trees. • PCS reveals the instability of clades with high bootstrap or Bayesian support. Genomic datasets sometimes support conflicting phylogenetic relationships when different tree-building methods are applied. Coherent interpretations of such results are enabled by partitioning support for controversial relationships among the constituent genes of a phylogenomic dataset. For the supermatrix (=concatenation) approach, several methods that measure the distribution of support and conflict among loci were introduced over 15 years ago. More recently, partitioned coalescence support (PCS) was developed for phylogenetic coalescence methods that account for incomplete lineage sorting and use the summed fits of gene trees to estimate the species tree. Here, we automate computation of PCS to permit application of this index to genome-scale matrices that include hundreds of loci. Reanalyses of four phylogenomic datasets for amniotes, land plants, skinks, and angiosperms demonstrate how PCS scores can be used to: (1) compare conflicting results favored by alternative coalescence methods, (2) identify outlier gene trees that have a disproportionate influence on the resolution of contentious relationships, (3) assess the effects of missing data in species-tree analysis, and (4) clarify biases in commonly-implemented coalescence methods and support indices. We show that key phylogenomic conclusions from these analyses often hinge on just a few gene trees and that results can be driven by specific biases of a particular coalescence method and/or the differential weight placed on gene trees with high versus low taxon sampling. The attribution of exceptionally high weight to some gene trees and very low weight to other gene trees counters the basic logic of phylogenomic coalescence analysis; even clades in species trees with high support according to commonly used indices (likelihood-ratio test, bootstrap, Bayesian local posterior probability) can be unstable to the removal of only one or two gene trees with high PCS. Computer simulations cannot adequately describe all of the contingencies and complexities of empirical genetic data. PCS scores complement simulation work by providing specific insights into a particular dataset given the assumptions of the phylogenetic coalescence method that is applied. In combination with standard measures of nodal support, PCS provides a more complete understanding of the overall genomic evidence for contested evolutionary relationships in species trees. [ABSTRACT FROM AUTHOR]