Taking phylogenetics beyond pattern analysis: can models of genome dynamics guide predictions about homoplasy in morphological and behavioral data sets?

Despite the considerable amount of interest in phylogeny reconstruction, patterns of homoplasy in morphological and behavioral data have received only limited attention to date, whereas the patterns of homoplasy in molecular data are relatively well understood. First, because the number of alternative molecular character states is strictly limited (particularly for nucleotide sequence data), higher rates of substitution generate higher levels of homoplasy. Second, depending on the relative proportions of constrained and unconstrained sites, each molecular data set has a time frame of applicability outside of which resolution becomes ambiguous. There is good evidence to suggest that numbers of alternative character states for morphological and even behavioral data may be similarly limited and that higher rates of evolution are often linked to higher rates of homoplasy. Like molecular data sets, morphological and behavioral data sets contain rapidly evolving characters as well as more conservative elements. Morphologies and behaviors related to sexual recognition and reproduction show low levels of intraspecific variation, but high levels of lability between species, making them crucial for species identification but often poor as markers of relationship at greater time depths. The organization theory of speciation derived by Carson is a model based on genome dynamics, and it predicts exactly this window of applicability for characters related to sexual reproduction. Nonsexual characters related to environmental adaptation should be applicable at greater phylogenetic depths. A better understanding of patterns of homoplasy enables a more sophisticated approach to the assessment of the relative reliabilities of alternative tree topologies.     

Homoplasy, homology, and the perceived special status of behavior in evolution

Evolutionary biologists tend to tread cautiously when considering how behavioral data might be incorporated into phylogenetic analyses, largely because of the preconception that behavior somehow constitutes a "special" set of characters that may be inherently more prone to homoplasy or subject to different selection regimes than those that operate on the morphological or genetic traits traditionally used in phylogenetic reconstruction. In this review, we first consider how the evolution of behavior has been treated historically, paying particular attention to why phylogenetic reconstruction has often failed to include behavioral traits. We then discuss, from a theoretical perspective, what reasons there are--if any--for assuming that behavioral traits should be more prone to homoplasy than other types of traits. In doing so, we review several empirical studies that tackle this issue head-on. Finally, we examine how behavioral features have been used to good effect in phylogenetic reconstruction. Our conclusion is that there seems to be little justification on theoretical grounds for assuming that behavior is in any way "special"--either particularly labile or particularly prone to exhibit high levels of homoplasy. Additionally, in reviewing historical perceptions of behavior and their links to conceptions of homology, we conclude that there is no compelling reason why behavior cannot be homologized or therefore why it should not prove phylogenetically informative. In subsequently considering several factors related to selection that influence the likelihood of homoplasy occurring in any trait system, we also found no clear trend predicting homoplasy disproportionately in behavioral systems. In fact, where studied, the degree of homoplasy seen in behavioral traits is comparable to that seen in other trait systems. Ultimately, there appear to be no grounds for dismissing behavior a priori from the class of phylogenetically informative characters.     

Reconstructing plumage evolution in orioles (Icterus): repeated convergence and reversal in patterns

Several empirical studies suggest that sexually selected characters, including bird plumage, may evolve rapidly and show high levels of convergence and other forms of homoplasy. However, the processes that might generate such convergence have not been explored theoretically. Furthermore, no studies have rigorously addressed this issue using a robust phylogeny and a large number of signal characters. We scored the appearance of 44 adult male plumage characters that varied across New World orioles (Icterus). We mapped the plumage characters onto a molecular phylogeny based on two mitochondrial genes. Reconstructing the evolution of these characters revealed evidence of convergence or reversal in 42 of the 44 plumage characters. No plumage character states are restricted to any groups of species higher than superspecies in the oriole phylogeny. The high frequency of convergence and reversal is reflected in the low overall retention index (RI = 0.66) and the low overall consistency index (CI = 0.28). We found similar results when we mapped plumage changes onto a total evidence tree. Our findings reveal that plumage patterns and colors are highly labile between species of orioles, but highly conserved within the oriole genus. Furthermore, there are at least two overall plumage types that have convergently evolved repeatedly in the three oriole clades. This overall convergence leads to significant conflict between the molecular and plumage data. It is not clear what evolutionary processes lead to this homoplasy in individual characters or convergence in overall pattern. However, evolutionary constraints such as developmental limitations and genetic correlations between characters are likely to play a role. Our results are consistent with the belief that avian plumage and other sexually selected characters may evolve rapidly and may exhibit high homoplasy. The overall convergence in oriole plumage patterns is an interesting evolutionary phenomenon, but it cautions against heavy reliance on plumage characters for constructing phylogenies.     

Character evolution in Hydrozoa (phylum Cnidaria)

The diversity of hydrozoan life cycles, as manifested in the wide range of polyp, colony, and medusa morphologies, has been appreciated for centuries. Unraveling the complex history of characters involved in this diversity is critical for understanding the processes driving hydrozoan evolution. In this study, we use a phylogenetic approach to investigate the evolution of morphological characters in Hydrozoa. A molecular phylogeny is reconstructed using ribosomal DNA sequence data. Several characters involving polyp, colony, and medusa morphology are coded in the terminal taxa. These characters are mapped onto the phylogeny and then the ancestral character states are reconstructed. This study confirms the complex evolutionary history of hydrozoan morphological characters. Many of the characters involving polyp, colony, and medusa morphology appear as synapomorphies for major hydrozoan clades, yet homoplasy is commonplace.     

Weighting against homoplasy improves phylogenetic analysis of morphological data sets

The problem of character weighting in cladistic analysis is revisited. The finding that, in large molecular data sets, removal of third positions (with more homoplasy) decreases the number of well supported groups has been interpreted by some authors as indicating that weighting methods are unjustified. Two arguments against that interpretation are advanced. Characters that collectively determine few well‐supported groups may be highly reliable when taken individually (as shown by specific examples), so that inferring greater reliability for sets of characters that lead to an increase in jackknife frequencies may not always be warranted. But even if changes in jackknife frequencies can be used to infer reliability, we demonstrate that jackknife frequencies in large molecular data sets are actually improved when downweighting characters according to their homoplasy but using properly rescaled functions (instead of the very strong standard functions, or the extreme of inclusion/exclusion); this further weakens the argument that downweighting homoplastic characters is undesirable. Last, we show that downweighting characters according to their homoplasy (using standard homoplasy‐weighting methods) on 70 morphological data sets (with 50–170 taxa), produces clear increases in jackknife frequencies. The results obtained under homoplasy weighting also appear more stable than results under equal weights: adding either taxa or characters, when weighting against homoplasy, produced results more similar to original analyses (i.e., with larger numbers of groups that continue being supported after addition of taxa or characters), with similar or lower error rates (i.e., proportion of groups recovered that subsequently turn out to be incorrect). Therefore, the same argument that had been advanced against homoplasy weighting in the case of large molecular data sets is an argument in favor of such weighting in the case of morphological data sets. © The Willi Hennig Society 2008.     

Levels of Homoplasy in the Evolution of the Mammalian Skeleton

It is commonly believed that there are differences in the evolutionary lability of the crania, dentition, and postcrania of mammals, the latter two being more prone to homoplasy because of strong selective pressures for feeding and locomotion, respectively. Further, because of the fragmentary nature of fossils, phylogenetic analyses of extinct taxa often must utilize characters based on only one of these systems. In this paper the levels of homoplasy (as measured by the consistency index; CI) were compared in characters based on these three anatomical systems in therian mammals. No statistically significant differences were found in the overall CIs of 41 data sets based on dental, cranial, or postcranial characters. Differences in homoplasy within data sets with two or three kinds of data were not statistically significant. These findings suggest that dental, cranial, and postcranial characters can be equally prone to homoplasy and none should be automatically dismissed, disregarded, or systematically weighted in phylogenetic analyses. The level of homoplasy in characters derived from a given region of the skeleton may differ depending on the taxonomic level of the taxa considered. Dental, cranial, and postcranial characters may not constitute natural classes, yet examination of the phylogenetic signal of these subsets of data previous to a simultaneous analysis can shed light on significant aspects of the evolutionary process.     

Lack of congruence between morphological and molecular data in reconstructing the phylogeny of the galagonidae.

Published cladistic reconstructions of galagonid phylogeny based on morphological, behavioral, and genetic data have had few elements in common. A recent molecular study indicated that 2 of the 3 generic groupings derived from morphological data were not consistent with tree topologies constructed from the analysis of mitochondrial DNA sequences. In this study, we compiled and analyzed a data set based on craniodental morphology in 13 galagonid and 8 outgroup taxa, comprising 3 dwarf-lemur and 5 loris species, and subjected it to cladistic analysis. Our aim was not only to generate a new phylogenetic hypothesis based on these data, but also to investigate the conditions under which congruence could be achieved between these results and those obtained previously. The data set was found to be highly sensitive to the choice of outgroup, with the lorises showing high interspecific variability in cranial structure. Congruence between the craniodental and molecular trees could be achieved only if Arctocebus was used as the outgroup and two characters were preferentially weighted. Further progress in the reconstruction of galagonid phylogeny will require seeking consensus in a variety of other data sets, including postcranial morphology, behavior, and additional gene sequences. The effect of different outgroups on molecular analysis needs attention.     

MACROEVOLUTIONARY PATTERNS OF MORPHOLOGICAL DIVERSIFICATION AMONG PARASITIC FLATWORMS (PLATYHELMINTHES: CERCOMERIA)

Patterns of parasite morphological diversification were investigated using a morphological data base for the parasitic platyhelminths comprising 1,459 characters analyzed using phylogenetic systematic methods. Only 10.8% of the 1,882 character transformations are losses, casting doubt on views that parasites are secondarily simplified and exhibit degenerate evolution. Chi-squared analysis indicates that character loss in the Digenea and Monogenea occurs in proportion to total change and is disproportionately lower within the Eucestoda. In the Digenea fewer female characters and more male characters have been lost than expected by the total number of characters in that group, and more male and more nonreproductive characters have been lost in proportion to their distribution across groups. In the Monogenea fewer nonreproductive and more larval characters have been lost than expected within the group, and female character loss is high relative to other groups. In the Eucestoda fewer female and more larval characters have been lost than expected within the group, whereas loss of male and nonreproductive character is low, and loss of larval characters is high, compared to the other groups. Patterns of character loss result partially from characters that show repeated (homoplasious) loss in different groups. High consistency index and low homoplasy slope ratio values indicate that the parasitic platyhelminths show unusually low levels of homoplasy, casting doubt on views that parasite morphology is unusually adaptively plastic. Homoplasy within the monogeneans occurs in proportion to overall character change, is slightly higher than expected in the digeneans, and is much lower than expected within the eucestodes. Homoplasy occurs less often than expected in larval characters, and more often than expected in nonreproductive characters in the Digenea. Monogeneans show more homoplasy than expected for larval characters both within and among groups. Eucestodes show fewer homoplasious male and nonreproductive, and more homoplasious larval, characters than expected within the group, and higher homoplasy in larval characters and lower homoplasy in female and nonreproductive characters among groups.     

Bayesian inference of the metazoan phylogeny; a combined molecular and morphological approach

Metazoan phylogeny remains one of evolutionary biology's major unsolved problems. Molecular and morphological data, as well as different analytical approaches, have produced highly conflicting results due to homoplasy resulting from more than 570 million years of evolution. To date, parsimony has been the only feasible combined approach but is highly sensitive to long-branch attraction. Recent development of stochastic models for discrete morphological characters and computationally efficient methods for Bayesian inference has enabled combined molecular and morphological data analysis with rigorous statistical approaches less prone to such inconsistencies. We present the first statistically founded analysis of a metazoan data set based on a combination of morphological and molecular data and compare the results with a traditional parsimony analysis. Interestingly, the Bayesian analyses demonstrate a high degree of congruence between morphological and molecular data, and both data sets contribute to the result of the combined analysis. Additionally, they resolve several irregularities obtained in previous studies and show high credibility values for controversial groups such as the ecdysozoans and lophotrochozoans. Parsimony, on the contrary, shows conflicting results, with morphology being congruent to the Bayesian results and the molecular data set producing peculiarities that are largely reflected in the combined analysis.     

Correlation with a limited set of behavioral niches explains the convergence of somatic morphology in mygalomorph spiders

Understanding the drivers of morphological convergence requires investigation into its relationship with behavior and niche space, and such investigations in turn provide insights into evolutionary dynamics, functional morphology, and life history. Mygalomorph spiders (trapdoor spiders and their kin) have long been associated with high levels of morphological homoplasy, and many convergent features can be intuitively associated with different behavioral niches. Using genus-level phylogenies based on recent genomic studies and a newly assembled matrix of discrete behavioral and somatic morphological characters, we reconstruct the evolution of burrowing behavior in the Mygalomorphae, compare the influence of behavior and evolutionary history on somatic morphology, and test hypotheses of correlated evolution between specific morphological features and behavior. Our results reveal the simplicity of the mygalomorph adaptive landscape, with opportunistic, web-building taxa at one end, and burrowing/nesting taxa with structurally modified burrow entrances (e.g., a trapdoor) at the other. Shifts in behavioral niche, in both directions, are common across the evolutionary history of the Mygalomorphae, and several major clades include taxa inhabiting both behavioral extremes. Somatic morphology is heavily influenced by behavior, with taxa inhabiting the same behavioral niche often more similar morphologically than more closely related but behaviorally divergent taxa, and we were able to identify a suite of 11 somatic features that show significant correlation with particular behaviors. We discuss these findings in light of the function of particular morphological features, niche dynamics within the Mygalomorphae, and constraints on the mygalomorph adaptive landscape relative to other spiders.     

PATTERNS OF VARIATION IN LEVELS OF HOMOPLASY

Patterns of variation in levels of homoplasy were explored through statistical analyses of standardized consistency indexes. Data were obtained from 60 recent cladistic analyses of a wide variety of organisms based on several different kinds of characters. Consistency index is highly correlated with the number of taxa included in an analysis, with homoplasy increasing as the number of taxa increases. This observation is compatible with a simple model of character evolution in which 1) the probability of character-state change increases with the total number of branches in a tree and 2) the number of possible states of a character is limited. Consistency index does not show a significant relationship to the number of characters utilized in an analysis or to the taxonomic rank of the terminal taxa. When the relationship between consistency index and number of taxa is taken into account, there is no significant difference between plant and animal data sets in the amount of homoplasy. Likewise, the level of homoplasy in morphological and molecular data sets does not appear to differ significantly, although there are still too few molecular studies to be confident of this result. Future comparisons of consistency indexes, including studies along the lines established here, must take into account the influence of the number of taxa on homoplasy.     

PENGUINS,PETRELS, AND PARSIMONY: DOES CLADISTIC ANALYSIS OF BEHAVIOR REFLECT SEABIRD PHYLOGENY?

Whether or not behavior accurately reflects evolutionary relationships (phylogeny) has been hotly debated by ethologists and comparative psychologists. Previous studies attempting to resolve this question have generally lacked a quantitative, phylogenetic approach. In this study we used behavior and life-history (BLH) information (72 characters) to generate phylogenetic trees for 18 seabird species (albatrosses, petrels, and penguins). We compared these trees with trees obtained from isozyme electrophoretic analysis of blood proteins (15 loci and 98 electromorphs) and partial mitochondrial 12S ribosomal DNA sequences (381 base pairs). Cladistic analysis of the BLH data set generated three MP trees (tree length = 243, CI = 0.52, RI = 0.57) with significant cladistic structure. The BLH characters were classified into four types (foraging, agonistic, reproductive, and life history) and levels of homoplasy for each type were measured. No significant differences were found among these categories. The BLH trees were shown to be significantly more congruent with the electrophoretic and 12S sequence trees than expected by chance. This indicates that seabird BLH data contains phylogenetic signal. Areas of incongruence between BLH trees and a phylogeny generated by combining the data sets were predicted to result from ecological constraints that did not covary with phylogeny. These predictions were supported by the results of a concentrated changes test. This study found that this BLH data set was no more homoplasious than molecular data and that BLH trees were significantly congruent with molecular trees.     

The Phylogeny of the Gasterosteidae: Combining Behavioral and Morphological Data Sets

Genealogical relationships among the five genera of stickleback (Gasterosteidae: Smegmomorpha) fishes were reconstructed based upon 47 behavioral and 89 morphological characters. Phylogenetic systematic analysis of the combined data set produced a single MPT (consistency index of 82.54%) structured ( Spinachia ( Apeltes-((Pungitius + Culaea )( Gasterosteus aculeatus + G. wheatlandi )))). When analyzed separately, both behavioral and morphological data sets produced the same tree topology, although the consistency index (excluding uninformative characters) was higher for the behavioral (90.70%) than for the morphological (68.13%) characters. This total evidence analysis provides a highly robust estimate of phylogeny for the sticklebacks and reiterates the importance of discarding a priori biases about behavioral evolution when selecting characters for phylogenetic analysis.     

The lacrimal/ectethmoid region of waterfowl (Aves,Anseriformes): Phylogenetic signal and major evolutionary patterns

Waterfowl (Aves, Anseriformes) constitute an ancient global radiation, and understanding the pattern and timing of their evolution requires a well-corroborated phylogeny including extant species and fossils. Following the molecular advances in avian systematics, however, morphology has often been held as misleading, yet congruence with molecular data has been shown to vary considerably among different skeletal parts. Here, we explore phylogenetic signal in discrete characters of the lacrimal/ectethmoid region of waterfowl, which is highly variable among species and constitutes a rich source of data. We do so by combining cladistic and multivariate approaches, and using phylogenetic comparative methods. We quantitatively recognize three major morphological types among lacrimal bones, and discuss homoplasy and potential synapomorphies of major clades using a molecular backbone tree. Our results clearly indicate that the lacrimal bone carries substantial phylogenetic signal and could be of systematic value at different levels of the phylogeny of waterfowl, feeding the exploration of other regions of the skull with this combined approach.     

Stabilizing selection on behavior and morphology masks positive selection on the signal in a salamander pheromone signaling complex

Natural selection maintains the integration and coordination of sets of phenotypic characters that collectively perform a task. In functional complexes in which characters span molecular to behavioral levels of organization, we might then expect similar modes of selection to produce similar patterns in evolutionary divergence at each level. To test this expectation, we diagnosed selection at behavioral, morphological, and molecular levels for courtship pheromone signaling by plethodontid salamanders. At the levels of morphology and behavior tens of millions of years of stasis (stabilizing selection) occur on each side of a transition from vaccination to olfactory delivery modes. As a proxy for the molecular level, we used plethodontid receptivity factor (PRF), a protein that is an active component of the pheromone. We cloned PRF from 12 Plethodon spp. spanning the delivery transition and obtained multiple alleles from each individual surveyed. Analyses of 61 alleles for PRF identified elevated nonsynonymous over synonymous substitution rates along lineages in a molecular phylogeny, and at 8% of sites in the protein, indicating that positive (directional) selection has acted on this vertebrate pheromone gene. Structural models showed PRF is in a family of cytokines characterized by a four-alpha-helix bundle. Positive selection in PRF was associated with receptor binding sites that are under purifying selection in other cytokines of that family. The evolutionary dynamics of the plethodontid pheromone delivery complex consists of stabilizing selection on morphological and behavioral aspects of signal delivery but positive selection on the signal mediated by receptors. Thus, different selection modes prevail at different levels in this reproductive functional complex. Evolutionary studies of integrated sets of characters therefore require separate analyses of selective action at each level.     

A phylogenetic hot spot for evolutionary novelty in Middle American treefrogs

Among the various types of evolutionary changes in morphology, the origin of novel structures may be the most rare and intriguing. Here we show statistically that the origins of different novel structures may be correlated and phylogenetically clustered into "hot spots" of evolutionary novelty, in a case study involving skull elements in treefrogs. We reconstruct phylogenetic relationships within a clade of Middle American treefrogs based on data from 10 nuclear and four mitochondrial genes and then analyze morphological evolution across this tree. New cranial elements are rare among anurans and tetrapods in general, but three novel elements have evolved within this clade, with a 40% increase in the number of skull roof elements in some species. Two of these elements also evolved in a related clade of treefrogs, and these two novel elements may have each evolved repeatedly within one or both clades. The molecular phylogeny suggests striking homoplasy in cranial morphology and shows that parsimony and Bayesian analyses of the morphological data have produced misleading results with strong statistical support. The origins of the novel elements are associated with an overall increase in the ossification of dermal skull roof elements (suggesting peramorphosis) and with the evolution of a novel adaptive behavior. Our study may be the first to statistically document significant phylogenetic clustering and correlation in the origins of novel structures, and to demonstrate the strongly misleading effects of peramorphosis on phylogenetic analysis.     

A phylogenomic perspective on gene tree conflict and character evolution in Caprifoliaceae using target enrichment data,with Zabelioideae recognized as a new subfamily

The use of diverse data sets in phylogenetic studies aiming for understanding evolutionary histories of species can yield conflicting inference. Phylogenetic conflicts observed in animal and plant systems have often been explained by hybridization, incomplete lineage sorting (ILS), or horizontal gene transfer. Here, we used target enrichment data, species tree, and species network approaches to infer the backbone phylogeny of the family Caprifoliaceae, while distinguishing among sources of incongruence. We used 713 nuclear loci and 46 complete plastome sequence data from 43 samples representing 38 species from all major clades to reconstruct the phylogeny of the family using concatenation and coalescence approaches. We found significant nuclear gene tree conflict as well as cytonuclear discordance. Additionally, coalescent simulations and phylogenetic species network analyses suggested putative ancient hybridization among subfamilies of Caprifoliaceae, which seems to be the main source of phylogenetic discordance. Ancestral state reconstruction of six morphological characters revealed some homoplasy for each character examined. By dating the branching events, we inferred the origin of Caprifoliaceae at approximately 66.65 Ma in the late Cretaceous. By integrating evidence from molecular phylogeny, divergence times, and morphology, we here recognize Zabelioideae as a new subfamily in Caprifoliaceae. This work shows the necessity of using a combination of multiple approaches to identify the sources of gene tree discordance. Our study also highlights the importance of using data from both nuclear and plastid genomes to reconstruct deep and shallow phylogenies of plants.     

PHYLOGENETIC HYPOTHESES UNDER THE ASSUMPTION OF NEUTRAL QUANTITATIVE-GENETIC VARIATION

There are many situations in which the only available characters for reconstructing phylogenies are morphological. Those traits that are subject only to the forces of mutation and random genetic drift can be used to obtain unbiased estimates of phylogenetic relationships. However, the accurate recovery of a phylogeny from information on neutral characters requires the procurement of data for a large number of independent traits, individuals, and populations. Phylogenetic trees fit to data from more than five species will almost always contain topological errors, even with very large data sets. The population-genetic consequences of the neutral model are reviewed, and some statistical methods for testing whether the diversification of a phylogeny is compatible with such a model are outlined. The theory is then applied to a very large data set on cranial morphology in modern man. The results are consistent with the hypothesis that interracial differences in human skull dimensions are a simple consequence of random drift and mutation.