Beyond the "Code": A Guide to the Description and Documentation of Biodiversity in Ciliated Protists (Alveolata, Ciliophora).

Recent advances in molecular technology have revolutionized research on all aspects of the biology of organisms, including ciliates, and created unprecedented opportunities for pursuing a more integrative approach to investigations of biodiversity. However, this goal is complicated by large gaps and inconsistencies that still exist in the foundation of basic information about biodiversity of ciliates. The present paper reviews issues relating to the taxonomy of ciliates and presents specific recommendations for best practice in the observation and documentation of their biodiversity. This effort stems from a workshop that explored ways to implement six Grand Challenges proposed by the International Research Coordination Network for Biodiversity of Ciliates (IRCN-BC). As part of its commitment to strengthening the knowledge base that supports research on biodiversity of ciliates, the IRCN-BC proposes to populate The Ciliate Guide, an online database, with biodiversity-related data and metadata to create a resource that will facilitate accurate taxonomic identifications and promote sharing of data.

A molecular and ultrastructural description of Spathidiopsis buddenbrocki and the phylogenetic position of the family Placidae (Ciliophora).

Spathidiopsis and Placus are the only two genera within the family Placidae. The family has been placed in the class Prostomatea and order Prorodontida because its members have somatic monokinetids with a radial transverse ribbon, a straight non-overlapping postciliary ribbon, and anteriorly directed non-overlapping kinetodesmal fibril, an apical cytostome lacking specialized oral cilia, a brosse, and toxicysts. To confirm the stability of this placement, ultrastructural morphology and small subunit rRNA gene sequences of Spathidiopsis socialis, Spathidiopsis buddenbrocki, and Placus striatus were determined. These data were combined with information from other ciliates, and phylogenetic trees were generated using maximum-likelihood and maximum-parsimony methods. The analyses confirmed the family Placidae to be a monophyletic group in the Prostomatea with the Placidae a sister group to a Cryptocaryon + Coleps + Prorodon clade.

The ultrastructure of Placus striatus and a revision of the family Placidae (Ciliophora).

The first ultrastructural description of the genus Placus is presented. Each somatic kinetosome has a cone-shaped axosomal plate, nonoverlapping postciliary microtubules, an anteriorly directed kinetodesmal fiber, and a radial ribbon of transverse microtubules, which extend laterally under the ciliary furrow and insert in the cortical ridge. A closed ring of paired kinetosomes encircles the cytostome. A brosse begins adjacent to the oral pairs and extends posteriorly for one-fourth to one-half the cell's length. Autapomorphies for Placus include bowling pin-shaped toxicysts extruded onto a distinct area of the cell surface immediately posterior to the brosse, and a brosse kinety consisting of a single row of paired cilia. Placus and its sister taxon Spathidiopsis both have spiraling kineties composed of single cilia inserted into the side of the ciliary furrow. Spathidiopsis can be distinguished from Placus because it has a brosse consisting of two kineties (i.e. one composed of paired cilia, the other of single cilia), a row of rod-shaped toxicyst-bearing palps extending around one side of the oral area and along the length of the brosse, and a mid-cell cortical inpocketing containing toxicysts and a segment of the brosse. A revised listing of species assigned to the family Placidae is given.

Phylogeny and systematic position of Zosterodasys (Ciliophora, Synhymeniida): a combined analysis of ciliate relationships using morphological and molecular data.

The Synhymeniida is characterized both by a band of somatic dikinetids, the synhymenium, extending across the surface of the cell and by a ventral cell mouth lacking specialized feeding cilia but subtended by a well-developed cyrtos. The synhymeniids have been hypothesized to be members of the class Nassophorea but our previous ultrastructural study of the synhymeniid genus Zosterodasys did not show any clear synapomorphies that would permit definitive placement in the Nassophorea or as a sister taxon to any of the other ciliate groups possessing a cyrtos. In the present study, simultaneous analysis of morphological and small subunit rDNA molecular data indicates that the Synhymeniida are sister to the class Phyllopharyngea and that this clade is, in turn, sister to the remaining Nassophorea, although this result is sensitive to dataset inclusion and alignment parameters. While this suggests that taxa with a ventral cyrtos might be united into a named taxon (e.g. resurrecting the Hypostomata), additional data are needed to reach a definitive conclusion.

Folliculinid ciliates: a new threat to Caribbean corals?

This is the first report of a putative pathogenic ciliate protozoan that has been associated with Caribbean corals. Previously, only 2 species of the phylum Ciliophora had been linked to coral diseases, and they were exclusive to the Indo-Pacific region. In this study, a ciliate of the genus Halofolliculina was found on 10 hard coral species at the National Parks of Los Roques and Morrocoy, Venezuela. The general morphology of this ciliate is very similar to that of Halofolliculina corallasia from the Indo-Pacific, which is known to cause skeletal eroding band. None of the other 31 genera in the family Folliculinidae are known to cause diseases in corals or in any other animal species. The presence of this ciliate, which shows a prevalence comparable to that of other epizootics in the Caribbean, suggests it could be a new threat to the coral reefs of this region.

A simple test: evaluating explanations for the relative simplicity of the Edwardsiidae (Cnidaria: Anthozoa).

Many members of the cnidarian subclass Zoantharia (sea anemones, corals, and their allies) pass through a larval stage with eight complete mesenteries and without posterior musculature. This larva is usually transient, developing into an adult with 12 or more mesenteries. The adults of one family of sea anemones, the Edwardsiidae, bear the larval number and arrangement of mesenteries and lack the pedal disc seen in other sea anemones. The morphology of the Edwardsiidae has been interpreted in a number of ways: (1) the Edwardsiidae are the most basal extant zoantharian, having diverged before the evolution of additional mesenteries and basal musculature; (2) they are relatively advanced sea anemones that have secondarily simplified because they burrow in sand or mud rather than attaching to a hard substrate; or (3) edwardsiids are derived anemones that have retained a juvenile morphology through paedomorphosis. Phylogenetic analyses of small subunit ribosomal gene sequences reveal that the Edwardsiidae are derived zoantharians, nested within sea anemones. None of the proposed explanations fully explain the edwardsiid's body plan; edwardsiid anatomy is a mosaic of retained primitive and derived features. The results of the present study provide insight into zoantharian phylogeny and illustrate how phylogenetic tests can be used to study the evolution of cnidarian body plans.

THE EUKARYOTIC KINGDOMS.

Abstract- Cladistic analysis of data from ultrastruetural and other cell biological studies indicates that neither the classical two kingdom nor the commonly accepted five kingdom classifications accurately represent the cellular diversity of eukaryotes. The resulting cladogram indicates instead that there are nine major groups of eukaryotes.

PARSIMONY, HOMOLOGY AND THE ANALYSIS OF MULTISTATE CHARACTERS.

Abstract- The order of states in a transformation series describes an internested set of synapomorphies. States adjacent to each other in the transformation series thus share a degree of homology not found in the other states. Whether the level of homology is relatively apomorphic is determined by rooting the order with outgroup comparison. The analysis of state order is a homology problem and is solved with a two-step process using similarity and congruence with other characters as criteria. Other methods that have been proposed (e.g. transformation series analysis, non-additive analysis, morphocline analysis, ontogenetic analysis) fail to apply both similarity and congruence, and thus cannot be used independently for determining character state order.

PARSIMONY AND THE CHOICE BETWEEN DIFFERENT TRANSFORMATIONS FOR THE SAME CHARACTER SET.

Abstract- When phylogeneticists choose among alternative hypotheses, they choose the one that requires the fewest ad hoc assumptions, i.e. the one that is the most parsimonious. For some systematists, choosing among alternative transformation series for the same set of taxa is equivalent to attaining trees with shorter length and minimal homoplasy. Homoplasy is shown to be composed of hierarchical discordance and scattering, which are recognized and described for the first time. Neither the consistency nor retention indices can be used in assessing different theories of multistate character transformation because both are affected by the shape of the transformation series rather than the character state distribution on a tree. Fits of transformations to a tree are better assessed by comparing the transformation to the cladogram character and the nearest neighbor network. Nearest neighbor networks are graphical representations of the nearest neighbor matrix. Transformations with the closest greatest number of matches between cladogram characters and the least complexity in the nearest neighbor network are preferred. These transformations are shown to make the fewest number of ad hoc statements and hence to be the most parsimonious. A means for obtaining cladogram characters and nearest neighbor matrices using a widely distributed microcomputer program is presented.

Character Coding and Inapplicable Data.

Inapplicable character states occur when character complexes are absent or reduced in some of the taxa. Several approaches have been proposed for representing such states in a character matrix so that the inapplicable condition has no effect on the placement of taxa and/or the applicable states are independent and not redundant. Here we examine each of these approaches and demonstrate that all have shortcomings. Coding inapplicables as "?" (reductive coding), although flawed, is currently the best way to analyze data sets that contain inapplicable character states.

Support, Ribosomal Sequences and the Phylogeny Of The Eukaryotes.

Sequences of the small subunit (SSU) ribosomal RNA are considered useful for reconstructing the tree of life because this molecule is found in all organisms and is large enough not to have become saturated with multiple mutations. However, these data sets are large, difficult to align, and have extreme biases in base compositions which makes their phylogenetic signal ambiguous. Large ambiguous data sets may have many most-parsimonious trees, and finding them all may be impossible using convential phylogenetic methods. To examine the reliability of the number and relationships of eukaryotic kingdoms proposed by previous analyses of the SSU, we calculated trees from aligned sequences from eukaryotes in the Ribosomal Database Project using parsimony jackknifing which uses a resampling procedure to rapidly search large data sets for the branches that are strongly supported and eliminates poorly supported groups. Two separate analyses were carried out: an analysis in which all bases were equally weighted, and one in which transversions only were used. The parsimony jackknife procedure was able to efficiently find trees in which most major groups of eukaryotes were supported and in which some evolutionary hypotheses proposed by previous workers were tested. The relationships of these major groups to each other were largely unresolved, indicating that the SSU data, as represented in this database, is insufficient for answering questions about these deep branches. Interestingly, the analysis of transitions differs from the results of the entire data set, primarily being less resolved. This indicates that transversional mutations are important contributors to the resolved structure of the tree.

PARSIMONY JACKKNIFING OUTPERFORMS NEIGHBOR-JOINING.

Abstract- Because they are designed to produced just one tree, neighbor-joining programs can obscure ambiguities in data. Ambiguities can be uncovered by resampling, but existing neighbor-joining programs may give misleading bootstrap frequencies because they do not suppress zero-length branches and/or are sensitive to the order of terminals in the data. A new procedure, parsimony jackknifing, overcomes these problems while running hundreds of times faster than existing programs for neighbor-joining bootstrapping. For analysis of large matrices, parsimony jackknifing is hundreds of thousands of times faster than extensive branch-swapping, yet is better able to screen out poorly-supported groups.

The misleading effects of composite taxa in supermatrices.

With the amount of available sequence data rapidly increasing, supermatrices are at the forefront of systematic studies. As an alternative to supertrees, supermatrices utilize a total evidence approach where different genes and other lines of data are merged into a single data matrix, which is then analyzed in an attempt to obtain the phylogeny that best explains the data. However, questions may arise when combining data sets in which one or more taxa do not have sequences available for each individual gene. Two possible solutions to this situation are to either leave all taxa separate and code unavailable sequences as missing, or to combine taxa at a level for which monophyly is assumed a priori. By reanalyzing the previous work of, we show that combining taxa may yield misleading results, i.e., hypotheses of relationships that are not supported by the underlying data.