Combined Analysis of Extant Rhynchonellida (Brachiopoda) using Morphological and Molecular Data

Abstract

Independent molecular and morphological phylogenetic analyses have often produced discordant results for certain groups which, for fossil-rich groups, raises the possibility that morphological data might mislead in those groups for which we depend upon morphology the most. Rhynchonellide brachiopods, with more than 500 extinct genera but only 19 extant genera represented today, provide an opportunity to explore the factors that produce contentious phylogenetic signal across datasets, as previous phylogenetic hypotheses generated from molecular sequence data bear little agreement with those constructed using morphological characters. Using a revised matrix of 66 morphological characters, and published ribosomal DNA sequences, we performed a series of combined phylogenetic analyses to identify conflicting phylogenetic signals. We completed a series of parsimony-based and Bayesian analyses, varying the data used, the taxa included, and the models used in the Bayesian analyses. We also performed simulation-based sensitivity analyses to assess whether the small size of the morphological data partition relative to the molecular data influenced the results of the combined analyses. In order to compare and contrast a large number of phylogenetic analyses and their resulting summary trees, we developed a measure for the incongruence between two topologies and simultaneously ignore any differences in phylogenetic resolution. Phylogenetic hypotheses generated using only morphological characters differed among each other, and with previous analyses, whereas molecular-only and combined Bayesian analyses produced extremely similar topologies. Characters historically associated with traditional classification in the Rhynchonellida have very low consistency indices on the topology preferred by the combined Bayesian analyses. Overall, this casts doubt on the use of morphological systematics to resolve relationships among the crown rhynchonellide brachiopods. However, expanding our dataset to a larger number of extinct taxa with intermediate morphologies is necessary to exclude the possibility that the morphology of extant taxa is not dominated by convergence along long branches.

Keywords: Brachiopoda; combined analyses; morphology; paleontology; phylogenetics.

Figure 1

Comparison of published morphological a) and molecular b) phylogenies for the Rhynchonellida. One way to display disagreement between two phylogenetic topologies is a graphical “tangle-gram,” where nodes are optimally rotated so that matches between tip taxon labels in topologies positioned horizontally are maximized. Lines are drawn between identical taxon pairs, such that crossing lines now indicate incongruent relationships. The tree on the left a) is the majority-rule consensus from Schreiber et al. (2013)’s morphological analysis under maximum parsimony, with characters reweighted relative to their consistency indices (as depicted in their fig. 4). The phylogeny on the right b) is the half-compatibility Bayesian posterior phylogeny from Cohen and Bitner (2013a, their Fig. 3), based on SSU and LSU rDNA. This “tangle-gram” figure was constructed using the R library “phytools,” v0.5-20 (Revell 2012). Relationships have been collapsed to the generic level. Superfamilies are not labeled on these phylogenies, due to the high incongruence of those higher taxa with any existing phylogenetic hypothesis (Cohen and Bitner 2013a; Schreiber et al. 2013).

Figure 2

Flowchart illustrating the workflow for the simulation-based sensitivity analyses. In the first step, a combined dataset composed of the original molecular data and a simulated morphological matrix is generated, using the majority-rule consensus summary from analysis Morph-Pars-18t as the basis for simulating the morphological data. This artificial dataset is then analyzed in MrBayes, using the same settings as analysis Comb-BMaxI-18t. The entire process is then replicated 10 times, and the output is summarized and compared with the summary topologies obtained from the empirical analyses.

Figure 3

Conceptual diagram of pairwise tree comparisons, illustrating how the typically used Robinson–Foulds metric (RF; Robinson and Foulds 1981) contrasts with the CD measure introduced in this study. Notably, RF penalizes trees for differences due to lack of resolution, but CD simultaneously ignores differences due to lack of resolution, making it ideal for comparing summary topologies. Note that, unlike RF, CD values are scaled to be between 0 and 1.

Figure 4

Phylogenies inferred from the revised morphological data only, including only those taxa for which molecular data are also available (Cohen and Bitner 2013a). a) Majority-rule consensus from a maximum parsimony analysis (analysis Morph-Pars-18t). Upper left node labels are bootstrap percentages; values not shown for nodes with bootstraps less than 50%. Center right node labels are the percentage of most parsimonious trees a clade was observed in, with nodes observed 100% indicated by an asterisk (*). Lower left node labels are Bremer support values for nodes with Bremer support values greater than zero. b) Half-compatibility tree from a Bayesian analysis (analysis Morph-BMaxI-18t) with nodes labeled by their posterior probabilities, rounded to two significant digits. Probabilities are not shown when effectively equal to 1.00.

Figure 5

Half-compatibility summaries from Bayesian analyses appliedto a combined dataset of the molecular and revised morphologicaldata, under strictest model and prior settings for the morphologicalcharacters. a) Analysis Comb-BMaxI-26t, containing all taxa in thedataset including extant taxa with no molecular data. Rhynchonellidegenera are labeled by their current superfamily membership(Savage et al. 2002): respectively, Dimerelloidea (D), andHemithiridoidea, (H) Norelloidea (N), and Pugnacoidea (P). b)Analysis Comb-BMaxI-18t, containing only those taxa that have bothknown morphological and molecular data. Nodes are labeled with theirposterior probabilities, rounded to two significant digits.Probabilities are not shown when effectively equal to1.00.

Figure 6

Majority rule consensus trees from maximum parsimony analyses applied to a combined dataset of the molecular and revised morphological data. These are respectively a) Analysis Comb-Pars-26t, with all taxa considered, and b) Analysis Comb-Pars-18t, containing only those taxa that have both known morphological and molecular data. For both cladograms, two node labels are shown. Upper left node labels are bootstrap percentages; values not shown for nodes with bootstraps less than 50% and values at 100% indicated by an asterisk (*). Lower left node labels are the percentage of most parsimonious trees in which a clade was observed, with nodes observed 100% indicated by an asterisk (*).

Figure 7

half-compatibility summary from one iteration of the sensitivity analysis, where the molecular data were combined with simulated “perfect” morphological data. The latter is a matrix of binary characters, the same size as our revised matrix, with state transitions evenly placed without homoplasy on nodes of the majority rule consensus topology from analysis Morph-Pars-26t (displayed in Fig. 4a). Nodes are labeled with their posterior probabilities, rounded to two significant digits. Probabilities are not shown when effectively equal to 1.00. All iterations of the sensitivity analyses produced half-compatibility summaries with identical topologies (see Supplementary Material available on Dryad).

Figure 8

The morphological characters most consistent with the combined analysis are not typically used within the Rhynchonellida. a–c) are histograms of consistency indices calculated using the half-compatibility topology from analysis Comb-BMaxI-18t (combined dataset), for three morphological character subsets. a) Those characters often considered to be useful for systematic purposes in the Rhynchonellida (characters 1, 3, 6, 7, 8, 11, 41, 47). b) Those characters used for distinguishing articulated brachiopod lineages from inarticulated out-groups, which do not vary among the Rhynchonellida (characters 59, 61, 62, 65, 66). c) Remaining characters. The remainder with high consistency (12, 23, 26, 28, 30, 44, 53) have high consistency indices due to rhynchonellide taxa for which that character is unknown or inapplicable.