On quality of evidence in phylogeny reconstruction: a reply to Zrzavý's defence of the 'Ecdysozoa' hypothesis

Zrzavý’s arguments against the critical analyses of data supporting the Ecdysozoa hypothesis (Wägele et al., J. Zool. Syst. Evol. Res. 37, 211–223, 1999) are discussed. Zrzavý does not understand that the same basic principle of a priori weighting can be applied to sequence data as well as to morphological characters. Quality of evidence is the same as probability of homology, which is estimated from the number of discernible identical details. In sequences it is the number of identical nucleotides. Spectral analyses, dismissed by Zrzavý, visualize patterns of putative homologies present in alignments and also the number of positions supporting splits by chance alone. In cases in which old phylogenetic signals for a given monophylum are eroded in a gene, plesiomorphies and chance patterns will have strong influence on tree topologies and spectra. If plesiomorphies are a cause of errors, the addition of taxa that shorten internal branches is a remedy, although, in many cases such taxa may be extinct. The place of a priori estimations of data quality in a sequence of steps necessary for a phylogenetic analysis is shown. Morphological complexity is used as a proxy for a complex genetic basis and is used as a major criterion to compare characters of the Ecdysozoa and the Articulata. The details associated with the character ‘complex cuticle’ are discussed. Neither moulting nor the known components of the cuticle are novelties occurring only in Ecdysozoa. A published total evidence analysis is used to show that the number of coded characters does not necessarily reflect the quality of the data set. Zrzavý’s misunderstanding of the role of evolutionary scenarios is clarified and the importance of the use of additional biological data for plausibility arguments is explained. Plausibility arguments in favour of the Articulata hypothesis rely on facts found in functional morphology and in the fossil record. Zrzavý’s critique follows the actual mainstream but does not uncover logical mistakes or erroneous data analyses in the work of 86 . It is concluded that the Articulata hypothesis is a well‐founded alternative to the Ecdysozoa; it is based on much better morphological evidence and supported by plausibility arguments that currently do not exist for the Ecdysozoa.     

Annulonemertes (phylum Nemertea): when segments do not count

We estimated the phylogenetic position of the pseudosegmented ribbon worm Annulonemertes minusculus to test proposed evolutionary hypotheses to explain these body constrictions. The analysis is based on 18S rDNA sequences and shows that the species belongs to an apomorphic clade of hoplonemertean species. The segmentation has no phylogenetic bearing as previously discussed, but is a derived character probably coupled to the species' interstitial habitat.     

A phylogenetic analysis of the land plants

Phylogenetic systematics (cladistics) is a theory of phylogeny reconstruction and classification widely used in zoology. Taxa are grouped hierarchically by the sharing of derived (advanced) characters. The information is expressed in a cladogram, a best estimate of a phylogeny. Plant systematists generally use a phenetic system, grouping taxa on overall similarity which results in many groups being formed, at least in part, on the basis of shared primitive characters.
The methods of phylogenetic systematics are used to create a preliminary cladogram of land plants. The current classification of land plants is criticized for its inclusion of many groups which are not monophyletic.
Objections to the use of phylogenetic systematics in botany, apparent convergences within major groups and frequent hybridization, are shown to be invalid. It is concluded that cladistic analysis presents the best estimate of die natural hierarchy of organisms, and should be adopted by plant systematists in their assessment of plant interrelationships.     

Circularity,evolution, systematics … and circularity

Many have argued strongly that incorporation of evolutionary theory into systematics is dangerously circular, while others have maintained that such an integrated approach increases the accuracy of phylogenetic inference. Here, it is demonstrated that such blanket statements regarding exclusion or inclusion of evolutionary principles in systematics fail to distinguish between two very different types of principles. ‘Phylogeny-neutral’ evolutionary principles are those inferred without any recourse to specific phylogenetic hypotheses (e.g. via developmental genetics, biomechanics). In contrast, ‘phylogeny-dependent’ principles are those which can only be inferred on the basis of specific phylogenetic hypotheses (e.g. character associations detected via ‘comparative methods’). Inclusion of phylogeny-neutral principles in systematic studies as a priori assumptions can be justified, since these principles have (often strong) external empirical support from other spheres of investigation. However, inclusion of phylogeny-dependent principles in systematic studies is circular, since such principles have no external empirical support but are themselves derived from systematic studies. Advocating inclusion or exclusion of all (or as many as possible) evolutionary principles from phylogenetic analysis is therefore misguided. Rather, phylogeny-neutral principles are independently supported and can be included, while phylogeny-dependent principles are unjustified assumptions and should be excluded to avoid circularity. However, integration of complex phylogeny-neutral principles in systematics can create operational problems, even though there are no methodological reasons against their inclusion.     

Review of methodological problems of 'Computer cladistics' exemplified with a case study on isopod phylogeny (Crustacea: Isopoda)

Using a computerized phylogenetic analysis of the Isopoda (Crustacea: Peracarida) as source of typical errors and misunderstandings, problems that may occur in computer cladistics are reviewed. It is concluded that in addition to the errors that are possible in a conventional Hennigian analysis some specific methodological problems exist in computer cladistics. It is recommended that the OTU be replaced by the groundpattern concept. Tree statistics are not useful for comparing different competing hypotheses. Arguments ought to concentrate on the hypo-thetico-deductive steps of the analysis, i.e. on character analysis. The use of computers does not add objectivity to character analysis. Single outgroup taxa should not be used in assessing the character states of ingroups. Concerning isopod phylogeny, it is argued here that the tail fan of the Isopoda can probably be derived from the eumalacostracan groundpattern and did not evolve de novo within the Isopoda.     

Adolf Naef (1883–1949), systematic morphology and phylogenetics

Several authors have highlighted methodological similarities between Naef’s systematic morphology and Hennig’s phylogenetic systematics. Whereas this may indicate an influence of Naef on Hennig, the relevant issues – such as the principle of generality in character analysis and the threefold parallelism of classification, ontogeny and the Fossil Record – reach back beyond Naef and Hennig and were widely discussed in the German systematics literature of the late 19th and early 20th Century. The same is true of conceptual issues, such as the discussion of the principle of monophyly, which was first introduced by Haeckel in 1866 ( Rieppel 2011b , J Zool Syst Evol Res 49 :1). In spite of methodological and conceptual agreements, Naef’s systematic morphology differed fundamentally from Hennig’s phylogenetic systematics. Naef emphasized the role of unbiased observation and the immediate acquaintance of the investigator with the phenomena given in nature as the basis of natural science in general, and of his hierarchy of types in particular. From the hierarchy of types, Naef derived through conceptual‐logical analysis the natural system, which above the species level forms a nested hierarchy of intensionally defined classes, denoted by general names. The historical‐causal interpretation of the hierarchy of types in turn offers insight into the hypothetical reality of phylogeny. Hennig in contrast denied the possibility of theory‐free observation, indeed of assumption‐free science in general, and on that basis put metaphysical issues above epistemology. Tying individuality to spatiotemporal location, historicity and causality, Hennig took not only species (as did Naef) but also supraspecific monophyletic taxa as individuals, denoted by proper names. From the species up, the phylogenetic system thus becomes a nested hierarchy of complex wholes of increasing degrees of complexity. Diagnostic characters of species or higher taxa can then no longer define classes (as in Naef’s natural system) but are thought to indirectly indicate the phylogenetic relations on which alone the phylogenetic system is to be based.     

The threefold parallelism of Agassiz and Haeckel,and polarity determination in phylogenetic systematics

A parallel exists between the threefold parallelism of Agassiz and Haeckel and the three valid methods of polarity determination in phylogenetic systematics. The structural gradation among taxa within a linear hierarchy, ontogenetic recapitulation, and geological succession of the threefold parallelism resemble outgroup comparison, the ontogenetic method, and the paleontological method, respectively, which are methods of polarity determination in phylogenetic systematics. The parallel involves expected congruence among similar components of the distribution of character states among organisms. The threefold parallelism is a manifestation of a world view based on linear hierarchies, whereas polarity determination is part of the methodology of phylogenetic systematics which assumes that organisms are grouped into a nested hierarchy. The threefold parallelism facilitated the ranking of previously established taxa into linear hierarchies consisting mostly of paraphyletic groups. In contrast, methods of polarity determination identify apomorphies that determine and diagnose monophyletic taxa (clades) in the nested genealogical hierarchy. Taxa in linear hierarchies are defined by sets of character states, whereas clades are defined by common ancestry. Although the threefold parallelism was ostensibly abandoned with the rejection of Haeckel's biogenetic law, some of its components continue to facilitate the progressive scenarios that are common in evolutionary thought. Although a general view of progression in organismal history may be invalid, the progressive or directional sequence of character state changes that results in the characterization of a particular clade has considerable heuristic value. Agassiz's ostensibly nested hierarchy and other pre-Darwinian classifications do not provide support for the view that the natural system can be discovered without recourse to the principle of common descent.     

Evolutionary morphology of whip spiders: towards a phylogenetic system (Chelicerata: Arachnida: Amblypygi)*

The aim of this study is to present a cladogram and phylogenetic system and to use this to discuss the phylogeny and biogeography of the Amblypygi. A total of 29 morphological structures were studied, their plesiomorphic and apomorphic characters or character states were identified, and the resulting data matrix was analysed. As a result, the ‘old’Charontidae or Pulvillata emerge as a paraphyletic group; the genus Paracharon is the sister group of all other amblypygids, which are now termed Euamblypygi. The ‘new’Charontidae (sensu Quintero: the genera Stygophrynus and Charon) are the sister group of the Phrynida or Apulvillata; together they form the Neoamblypygi. The relationships of the genera of the Charinidae cannot be resolved with the available data. They may be a paraphyletic group. The genus Catageus is a possible candidate for being the sister group of the Neoamblypygi. The new system allows a discussion of the phylogeny and biogeography of whip spiders. It also points to unresolved taxa and thus indicates the questions future research should address.     

Heads or tails: a simple reliability check for multiple sequence alignments

The question of multiple sequence alignment quality has received much attention from developers of alignment methods. Less forthcoming, however, are practical measures for addressing alignment quality issues in real life settings. Here, we present a simple methodology to help identify and quantify the uncertainties in multiple sequence alignments and their effects on subsequent analyses. The proposed methodology is based upon the a priori expectation that sequence alignment results should be independent of the orientation of the input sequences. Thus, for totally unambiguous cases, reversing residue order prior to alignment should yield an exact reversed alignment of that obtained by using the unreversed sequences. Such "ideal" alignments, however, are the exception in real life settings, and the two alignments, which we term the heads and tails alignments, are usually different to a greater or lesser degree. The degree of agreement or discrepancy between these two alignments may be used to assess the reliability of the sequence alignment. Furthermore, any alignment dependent sequence analysis protocol can be carried out separately for each of the two alignments, and the two sets of results may be compared with each other, providing us with valuable information regarding the robustness of the whole analytical process. The heads-or-tails (HoT) methodology can be easily implemented for any choice of alignment method and for any subsequent analytical protocol. We demonstrate the utility of HoT for phylogenetic reconstruction for the case of 130 sequences belonging to the chemoreceptor superfamily in Drosophila melanogaster, and by analysis of the BaliBASE alignment database. Surprisingly, Neighbor-Joining methods of phylogenetic reconstruction turned out to be less affected by alignment errors than maximum likelihood and Bayesian methods.     

FLIPPERS AND PINNIPED PHYLOGENY: HAS THE PROBLEM OF CONVERGENCE BEEN OVERRATED?

The phylogenetic relationships of the major groups of pinnipeds are of longstanding controversy. According to prevailing paleontological opinion, pinnipeds comprise two distinct lineages of independent terrestrial carnivoran origin, A logical correlate of this view is that an enormous degree of convergence accounts for the striking resemblances of these groups.
It has been generally supposed that derived similarities common to pinnipeds are independently acquired adaptations to a highly constraining environment. This has not been adequately substantiated. Moreover, considerable evidence favors a contrary conclusion. Acceptance of convergence necessarily rests on demonstration that taxa possessing characters suspected to be convergent are in fact not mutal nearest allies. In the absence of such phylogenetic information, shared derived characters are indicative of common ancestry irrespective of whether they are deemed "functionally important." Given meager support for the proposition that pinnipeds are related to different terrestrial lineages, it is currently more judicious to regard the group's origin as monophyletic. In addition to the presumed prevalence of convergence, biogeography, the high degree of morphological divergence of phocids, and the aberrant nature of phocines have improperly contributed to acceptance of independent pinniped origins.     

The logical priority of the tree over characters and some of its consequences for taxonomy

The aim of the present paper is to explore the role of the character in phylogenetic systematics. I argue that too much emphasis is put on particular characters rather than congruence both in the choice of phylogenetic hypotheses and in taxonomic decisions. This means that the logical priority of the tree over the characters is neglected. To a large extent, this is a result of not paying enough attention to the individuality thesis which states that clades are historical individuals and hence contingent in nature.     

Phylogenetically structured variance in felid bite force: the role of phylogeny in the evolution of biting performance

A key question in evolution is the degree to which morphofunctional complexes are constrained by phylogeny. We investigated the role of phylogeny in the evolution of biting performance, quantified as bite forces, using phylogenetic eigenvector regression. Results indicate that there are strong phylogenetic signals in both absolute and size‐adjusted bite forces, although it is weaker in the latter. This indicates that elimination of size influences reduces the level of phylogenetic inertia and that the majority of the phylogenetic constraint is a result of size. Tracing the evolution of bite force through phylogeny by character optimization also supports this notion, in that relative bite force is randomly distributed across phylogeny whereas absolute bite force diverges according to clade. The nonphylogenetically structured variance in bite force could not be sufficiently explained by species‐unique morphology or by ecology. This study demonstrates the difficulties in identifying causes of nonphylogenetically structured variance in morphofunctional character complexes.     

Utility of rbcS gene as a novel target DNA region for brown algal molecular systematics

The usefulness of molecular phylogenetic studies has increased remarkably as the quantity and quality of available DNA sequences has increased. When compared with the progress that has occurred in angiosperms and animals, there have been relatively few target DNA regions identified for use in taxonomic studies of brown algae. Therefore, in this study, we developed a new set of primers to amplify Rubisco small subunit (rbcS) gene sequences and determined the rbcS gene sequences of various species of brown algae including those belonging to Dictyotales, Ectocarpales, Fucales and Sphacelariales. The level of sequence variations in the rbcS gene varied according to the brown algal lineages. When focusing on the relationship of species within the genus Sargassum, the rbcS gene sequences provided useful information regarding the phylogenetic relationship among sections of the subgenus Bactrophycus. Based on the broad applicability and phylogenetic utility of the rbcS gene, we suggest that the sequence be used as a new target region for the molecular systematics of brown algae.     

On the information content of characters in comparative morphology and molecular systematics

A review of the fundamental difference between single molecular-sequence positions, or numerical characters, and complex morphological characters is the subject of this study. It has been found that transformation series of single complex structures contain enough information to allow a priori determination of character order and that rooting of a dendrogram is possible without out-group comparison, while trees based on less-informative characters can usually only be rooted with out-group comparison. Furthermore, the quality of total information used is decisive in discriminating between hypotheses of relationships. Numerical methods for the inference of phylogenies have been found to be useful for high numbers of characters that have only a low information content, while the Hennigian procedure seems to be preferable for complex characters.     

Old trees, new trees--is there any progress?

The amount of comparative data for phylogenetic analyses is constantly increasing. Data come from different directions such as morphology, molecular genetics, developmental biology and paleontology. With the increasing diversity of data and of analytical tools, the number of competing hypotheses on phylogenetic relationships rises, too. The choice of the phylogenetic tree as a basis for the interpretation of new data is important, because different trees will support different evolutionary interpretations of the data investigated. I argue here that, although many problematic aspects exist, there are several phylogenetic relationships that are supported by the majority of analyses and may be regarded as something like a robust backbone. This accounts, for example, for the monophyly of Metazoa, Bilateria, Deuterostomia, Protostomia (= Gastroneuralia), Gnathifera, Spiralia, Trochozoa and Arthropoda and probably also for the branching order of diploblastic taxa (“Porifera”, Trichoplax adhaerens, Cnidaria and Ctenophora). Along this “backbone”, there are several problematic regions, where either monophyly is questionable and/or where taxa “rotate” in narrow regions of the tree. This is illustrated exemplified by the probable paraphyly of Porifera and the phylogenetic relationships of basal spiralian taxa. Two problems span wider regions of the tree: the position of Arthropoda either as the sister taxon of Annelida (= Articulata) or of Cycloneuralia (= Ecdysozoa) and the position of tentaculate taxa either as sister taxa of Deuterostomia (= Radialia) or within the taxon Spiralia. The backbone makes it possible to develop a basic understanding of the evolution of genes, molecules and structures in metazoan animals.     

Predicting protein functional sites with phylogenetic motifs

In this report, we demonstrate that phylogenetic motifs, sequence regions conserving the overall familial phylogeny, represent a promising approach to protein functional site prediction. Across our structurally and functionally heterogeneous data set, phylogenetic motifs consistently correspond to functional sites defined by both surface loops and active site clefts. Additionally, the partially buried prosthetic group regions of cytochrome P450 and succinate dehydrogenase are identified as phylogenetic motifs. In nearly all instances, phylogenetic motifs are structurally clustered, despite little overall sequence proximity, around key functional site features. Based on calculated false-positive expectations and standard motif identification methods, we show that phylogenetic motifs are generally conserved in sequence. This result implies that they can be considered motifs in the traditional sense as well. However, there are instances where phylogenetic motifs are not (overall) well conserved in sequence. This point is enticing, because it implies that phylogenetic motifs are able to identify key sequence regions that traditional motif-based approaches would not. Further, phylogenetic motif results are also shown to be consistent with evolutionary trace results, and bootstrapping is used to demonstrate tree significance.     

Evaluating evolutionary pressures and phylogenetic signal in earthworms: a case study – the number of typhlosole lamellae in Hormogastridae (Annelida,Oligochaeta)

Rarely have phylogenetic comparative methods been used to study the correlation between phenotypic traits and environmental variables in invertebrates. With the widespread convergence and conservativeness of the morphological characters used in earthworms, these comparative methods could be useful to improve our understanding of their evolution and systematics. One of the most prominent morphological characters in the family Hormogastridae, endemic to Mediterranean areas, is their multilamellar typhlosole, traditionally thought to be an adaptation to soils poor in nutrients. We tested the correlation of body size and soil characteristics with the number of typhlosole lamellae through a phylogenetic generalized least squares (PGLS) analysis. An ultrametric phylogenetic hypothesis was built with a 2580‐bp DNA sequence from 90 populations, used in combination with three morphological and 11 soil variables. The best‐supported model, based on the Akaike information criterion, was obtained by optimizing the parameters lambda (λ), kappa (κ), and delta (δ). The phylogenetic signal was strong for the number of typhlosole lamellae and average body weight, and was lower for soil variables. Increasing body weight appeared to be the main evolutionary pressure behind the increase in the number of typhlosole lamellae, with soil texture and soil richness having a weaker but significant effect. Information on the evolutionary rate of the number of typhlosole lamellae suggested that the early evolution of this character could have strongly shaped its variability, as is found in an adaptive radiation. This work highlights the importance of implementing the phylogenetic comparative method to test evolutionary hypotheses in invertebrate taxa.     

Phylogenetic analysis of the Amphitritinae (Polycnaeta: Terebellidae)

The phylogenetic relationships of the Amphitritinae (Polychaeta: Terebellidae) were studied using parsimony analysis of 22 external morphological characters. To choose outgroups to polarize the characters, I carried out a preliminary analysis of the relationships of the four terebellid subfamilies and the Trichobranchidae. The single most parsimonious tree from the analysis supports monophyly of the Terebellidae by the presence of ventral glandular shields. However, this character is homoplasious within the Terebellomorpha, and further evaluation of the Terebellidae is recommended. Artacama and Thelepus were chosen as outgroups for the analysis of amphitritine genera. The generic level analysis yielded seven equally parsimonious trees, which are consistent in their topologies except for the relationships among seven genera in one large clade. In all trees, Artacama is the sister taxon to a large clade within the Amphitritinae; the Artacaminae is therefore synonymized with the Amphitritinae, which is diagnosed by the presence of double rows of uncini. Within the Amphitritinae, the status of several monotypic genera is questioned; plesiomorphic character states indicated by the analysis are discussed. The results presented are offered as working hypotheses of the relationships among amphitritine genera. The large number of homoplasies indicated by the analysis emphasizes the need to further evaluate these hypotheses using additional characters. With a robust phylogenetic hypothesis of amphitritine relationships, a re-classification of the group based on apomorphic character states can be undertaken, and questions regarding the evolution of morphological characters, reproductive modes, or biogeographical patterns can be properly addressed.