We inferred secondary structure models of the internal transcribed spacers (ITS) 1 and 2 of bush crickets using a combined comparative and thermodynamic approach. The inferred secondary structure models were used to account for interdependency of interacting nucleotides in a phylogenetic analysis of the bush cricket genus Poecilimon. Our analysis indicates that the two previously reported conformational structures (i.e., hairpin and ring) of ITS2 are likely to fold in bush crickets as well and that both predicted structures are similar to those proposed for other eukaryotes. Comparing predicted ITS1 secondary structure models proved to be difficult because of substantial variation in their nucleotide sequence length. Our study revealed that the phylogenetic signal of ITS1 and ITS2 is largely congruent with that preserved in the mitochondrial genes 16S rRNA, tRNA‐Val and 12S rRNA. The phylogenetic signal in both the nuclear and the mitochondrial genome question the monophyly of the genus Poecilimon: species of the genera Poecilimonella, Parapoecilimon, Polysarcus and Phonochorion consistently cluster within Poecilimon.

The application of mixed nucleotide ⁄ doublet substitution models has recently received attention in RNA-based phylogenetics. Within a Bayesian approach, it was shown that mixed models outperformed analyses relying on simple nucleotide models. We analysed an mt RNA data set of dragonflies representing all major lineages of Anisoptera plus outgroups, using a mixed model in a Bayesian and parsimony (MP) approach. We used a published mt 16S rRNA secondary consensus structure model and inferred consensus models for the mt 12S rRNA and tRNA valine. Secondary structure information was used to set data partitions for paired and unpaired sites on which doublet or nucleotide models were applied, respectively. Several different doublet models are currently available of which we chose the most appropriate one by a Bayes factor test. The MP reconstructions relied on recoded data for paired sites in order to account for character covariance and an application of the ratchet strategy to find most parsimonious trees. Bayesian and parsimony reconstructions are partly differently resolved, indicating sensitivity of the reconstructions to model specification. Our analyses depict a tree in which the damselfly family Lestidae is sister group to a monophyletic clade Epiophlebia + Anisoptera, contradicting recent morphological and molecular work. In Bayesian analyses, we found a deep split between Libelluloidea and a clade ÔAeshnoideaÕ within Anisoptera largely congruent with TillyardÕs early ideas of anisopteran evolution, which had been based on evidently plesiomorphic character states. However, parsimony analysis did not support a clade ÔAeshnoideaÕ, but instead, placed Gomphidae as sister taxon to Libelluloidea. Monophyly of Libelluloidea is only modestly supported, and many inter-family relationships within Libelluloidea do not receive substantial support in Bayesian and parsimony analyses. We checked whether high Bayesian node support was inflated owing to either: (i) wrong secondary consensus structures; (ii) under-sampling of the MCMC process, thereby missing other local maxima; or (iii) unrealistic prior assumptions on topologies or branch lengths. We found that different consensus structure models exert strong influence on the reconstruction, which demonstrates the importance of taxon-specific realistic secondary structure models in RNA phylogenetics.