Phylogeny of the order Corallimorpharia (Cnidaria: Hexacorallia) Hi, Good Afternoon~ My name is Ha-Rim Cha and I am a graduate student of department of EEB. My doctoral research focuses on systematics of the order Corallimorpharia and today I would like to talk about the phylogenetic study of this taxon. Harim Cha Department of Ecology and Evolutionary Biology Natural History Museum & Biodiversity Research Center The University of Kansas harim@ku.edu

Corallimorpharia Coral-like sea anemones One of six orders of Hexacorallia Coral-like sea anemones Solitary or clonal polyps without skeleton Current Classification four families, 12 genera, and 50 species Widely distributed from tropical to polar areas and from shallow to deep water Adorian Ardelean George Miller Corallimorpharia is one of six orders of Zoantharia, and because of intermediate characters between stony corals and sea anemones, this groups is often called “coral-like sea anemones.” Members of Corallimorpharia are solitary or colonial polyps without skeleton, and the size of polyp is from 10 mm to 500 mm in width of oral disc. According to the current classification, four families, 14 nominal genera, and 50 species are included. Corallimorpharians distribute very widely in the seas. From tropical to polar area and from shallow to deep water. The horizontal and vertical wide range of distribution allow the significance of the global scale research of this group. Also, these animals are one of the beauty of underwater biota and also in aquariums. In spite of its beauty, the order Corallimorpharia is one of questionable taxon in scientific perspectives. Within the order, although there is literature inventorying corallimorpharians, not only the species composition of this order but also the number of valid genera have been unclear because of incomplete diagnosis among genera and the inconsistency in usage among orders. Also, as I mentioned earlier of this slides, corallimorpharians have attributes of both orders Actiniaria and Scleractinia. So, the systematic position of this taxon has been disputed and most of precious hypotheses based on morphology differ between authors depending on the characters focused. Vincent B. Hargreaves

Corallimorpharia Questions about relationships within the order Cylindrical body (CO I) Family Sideractiidae Family Corallimorphidae Plate-like body (CO II) Family Ricordeidae Family Discosomatidae The first question is the relationship within the order. The relationship between two different morphological groups in the order, the one is cylindrical body group which has capitate or digiform tentacles, and the other is plate-like body group that can be characterized by low column and numerous reduced tentacles, is still questionable. Vincent B. Hargreaves Vincent B. Hargreaves

Corallimorpharia Questions about relationship between Corallimorpharia and orders Actiniaria (sea anemones) and Scleractinia (stony corals) Hypothesis 1 Hypothesis 2 SC CO AC SC CO AC The second question is for the relationship among orders. Based on the different two group of morphology within corallimorpharians, the order Corallimorpharia may not be a monophyletic group. This hypothesis has been supported by several molecular studies. More expanding study is the relationship between corallimorpharia and two closest orders, stony corals and sea anemones. There are three hypothesis for the relationship. My animals are different but may be close to stony corals rather than with sea anemones. OR, three orders may have evolved independently, OR Corallimorpharia may not an order and a part of two other orders. Molecular studies? 28S rDNA (Chen et al. 1995) - polyphyly 16S mtDNA (Romane and Cairns 200) - paraphyly 18S rDNA (Bernston et al. 1999) - paraphyly Immonulogical study? Fautin and Lowenstein 1992 - non monophyly Hypothesis 3 Hypothesis 4 SC CO AC SC CO CO AC

Phylogenetic hypothesis based on molecular data a. Fautin & Lowenstein (1994): radioimmunoassay b. Chen et al. (1995): 28S rRNA c. France et al. (1996): 16S rRNA d. Daly et al. (2003): Morphology, 16S, 18S, 28S e. Medina et al. (2006): mtDNA

Phylogenetic analysis using morphological characters Major characters used for analysis: Calcareous exoskeleton Polyp shape / size Tentacle shape / number / arrangement Nature of mesenteries and mesenterial arrangement Muscle development Nematocyst composition Life style – solitary or clonal Habitat Zooxanthellae Methods: 32 morphological characters 12 genera of Corallimorpharia, 6 genera of Scleractinia, and 11 genera of Actiniaria Outgroup: genus Cerianthus Maximum parsimony criterion, 1000 bootstrap replicates

Maximum Parsimony analysis (CI = 0.58, RI = 0.69) Corallimorpharia (CO) Scleractinia (SC) Actiniaria (AC) Ceriantharia (CE) CO I CO II Maximum Parsimony analysis (CI = 0.58, RI = 0.69) Strict consensus tree of 12 equally parsimonious trees Number of parsimony informative characters = 30

Phylogenetic analysis using molecular characters Genes sequenced: 16S mtDNA (1233 bp), 28S rDNA (345 bp), 18S rDNA (1836 bp) Taxon sampling: Corallimorpharia - more than one genus of each family except family Sideractidae - collected from Oman and Caribbean - borrowed from the Field Museum Scleractinia - Robust and Complex groups Actiniaria - at least one taxon of each tribe Outgroup: Genus Cerianthus MacClade 4.0 and PAUP 3.3.1 Maximum likelihood Maximum parsimony – 2000 bootstrap replicates So, I did phylogenetic analysis using morphological characters of corallimorpharian genera and three genera of Actiniaria and Scleractinia. I used 32 characters that are collected from my own observations using type species and non-type species of each genus. Three genera of each Scleractinia and Actiniaria were added for analysis. Genus Cerianthus was chosen for the outgroup. For the analysis, MacClade and PAUP were used. Each transformation series has equal weight. And all characters unordered when analyzed. The heuristic method and Branch-and Bound search were used for searching the most parsimonious tree because of the number of taxa we consider. Accelerated transformation (ACCTRAN) was used for optimization of character states.

Results 16S mtDNA Maximum Likelihood Maximum Parsimony analysis Robust Scleractinia (RS) Maximum Parsimony analysis Strict consensus of 9 equally parsimonious trees Number of parsimony-informative characters: 498 Complex Scleractinia (CS) Ceriantharia (CE) Robust Scleractinia (RS) As a result, I got a 50% majority-rule strict consensus tree was computed based on 16 equally parsimonious trees. Corallimorpharia (CO) CO II Corallimorpharia (CO) CO I Complex Scleractinia (CS) Actiniaria (AC) Actiniaria (AC) Ceriantharia (CE)

28S rDNA maximum likelihood analysis CO CS AC CE RS CO II CO I CE CO AC Scleractinia CO II CO I

Summary of results Phylogeny based on morphological characters 1) Plate-like corallimorpharians are more closely related to deep-sea cylindrical corallimorpharians (genera Corallimorphus, Sideractis, Nectactis, and Sphincteractis) than to shallow-water ones. 2) Corallimorpharia is a monophyletic group 3) Corallimorpharia is a sister taxon of Scleractinia Each gene gives a different signal on phylogenetic relationships of Corallimorpharia 1) 16S mtDNA: Corallimorpharia is monophyletic and a sister taxon of the Complex group of Scleractinia 2) 28S rDNA: Corallimorpharia is monophyletic and a sister taxon of Scleractinia 3) 18S rDNA: Corallimorpharia is within Scleractinia Based on the result, the order Corallimorpharia may not be a monophyletic group. Second, two families of Corallimorpharia are clustered to actiniarian genera and two families are clustered to scleractinian genera. This result supports one of hypothesis about the relationship which claims a part of corallimorpharians may belong to Actiniaria and the others may belong to Scleractinia. And the tree show clear clustering pattern of two different morphological groups within the order.

Further study Better molecular markers are required to resolve higher level phylogeny of Hexacorallia Extensive morphological data including polyp anatomy of scleractinians may help to clarify the position of corallimorpharians Data from deep-sea corallimorpharians (family Sideractidae) will complete sampling of corallimorpharians and may be able to provide new insights on corallimorpharian phylogeny

Acknowledgments Dr. Daphne Fautin and all lab people Dr. Adorian Ardelean (WHOI) Dr. Paulyn Cartwright (KU) Dr. Rachel Collin (STRI) Dr. Meg Daly (OSU) Mr. Matthew Kost (KU) Ms. Abby Reft Dr. Sandra Romano (UVI) NSF grant 9978106 in the program Partnerships to Enhance Expertise in Taxonomy (PEET) to Dr. Daphne Fautin KU Natural History Museum and Biodiversity Research Center Department of Ecology and Evolutionary Biology, KU