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Background  Most phylogenetic studies using current methods have focused on primary DNA sequence information. However, RNA secondary structures are particularly useful in systematics because they include characteristics, not found in the primary sequence, that give "morphological" information. Despite the number of recent molecular studies on octocorals, there is no consensus opinion about a region that carries enough phylogenetic resolution to solve intrageneric or close species relationships. Moreover, intrageneric morphological information by itself does not always produce accurate phylogenies; intra-species comparisons can reveal greater differences than intra-generic ones. The search for new phylogenetic approaches, such as by RNA secondary structure analysis, is therefore a priority in octocoral research.    Results  Initially, twelve predicted RNA secondary structures were reconstructed to provide the basic information for phylogenetic analyses; they accorded with the 6 helicoidal ring model, also present in other groups of corals and eukaryotes. We obtained three similar topologies for nine species of the Caribbean gorgonian genus  Eunicea  (candelabrum corals) with two sister taxa as outgroups (genera  Plexaura  and  Pseudoplexaura ) on the basis of molecular morphometrics of ITS2 RNA secondary structures only, traditional primary sequence analyses and maximum likelihood, and a Bayesian analysis of the combined data. The latter approach allowed us to include both primary sequence and RNA molecular morphometrics; each data partition was allowed to have a different evolution rate. In addition, each helix was partitioned as if it had evolved at a distinct rate.  Plexaura flexuosa  was found to group within  Eunicea ; this was best supported by both the molecular morphometrics and combined analyses. We suggest  Eunicea flexuosa  (Lamouroux, 1821) comb. nov., and we present a new species description including Scanning Electron Microscopy (SEM) images of morphological characteristics (sclerites).  Eunicea flexuosa ,  E. pallida ,  E. laxispica  and  E. mammosa  formed a separate clade in the molecular phylogenies, and were reciprocally monophyletic with respect to other  Eunicea  ( Euniceopsis  subgenus, e.g.  E. tourneforti  and  E. laciniata ) in the molecular morphometrics tree, with the exception of  E. fusca . Moreover, we suggest a new diagnostic character for  Eunicea , also present in  E. flexuosa : middle layer sclerites > 1 mm in length.    Conclusion  ITS2 was a reliable sequence for intrageneric studies in gorgonian octocorals because of the amount of phylogenetic signal, and was corroborated against morphological characters separating  Eunicea  from  Plexaura . The ITS2 RNA secondary structure approach to phylogeny presented here did not rely on alignment methods such as INDELS, but provided clearly homologous characters for partition analysis and RNA molecular morphometrics. These approaches support the divergence of  Eunicea flexuosa  comb. nov. from the outgroup  Plexaura , although it has been considered part of this outgroup for nearly two centuries because of morphological resemblance.

bmc evolutionary biology

Phylogenetic reconstruction using secondary structures of Internal Transcribed Spacer 2 (ITS2, rDNA): finding the molecular and morphological gap in Caribbean gorgonian corals