Toward reconstructing the evolution of advanced moths and butterflies (Lepidoptera: Ditrysia): an initial molecular study

Abstract

Background: In the mega-diverse insect order Lepidoptera (butterflies and moths; 165,000 described species), deeper relationships are little understood within the clade Ditrysia, to which 98% of the species belong. To begin addressing this problem, we tested the ability of five protein-coding nuclear genes (6.7 kb total), and character subsets therein, to resolve relationships among 123 species representing 27 (of 33) superfamilies and 55 (of 100) families of Ditrysia under maximum likelihood analysis.

Results: Our trees show broad concordance with previous morphological hypotheses of ditrysian phylogeny, although most relationships among superfamilies are weakly supported. There are also notable surprises, such as a consistently closer relationship of Pyraloidea than of butterflies to most Macrolepidoptera. Monophyly is significantly rejected by one or more character sets for the putative clades Macrolepidoptera as currently defined (P < 0.05) and Macrolepidoptera excluding Noctuoidea and Bombycoidea sensu lato (P < or = 0.005), and nearly so for the superfamily Drepanoidea as currently defined (P < 0.08). Superfamilies are typically recovered or nearly so, but usually without strong support. Relationships within superfamilies and families, however, are often robustly resolved. We provide some of the first strong molecular evidence on deeper splits within Pyraloidea, Tortricoidea, Geometroidea, Noctuoidea and others.Separate analyses of mostly synonymous versus non-synonymous character sets revealed notable differences (though not strong conflict), including a marked influence of compositional heterogeneity on apparent signal in the third codon position (nt3). As available model partitioning methods cannot correct for this variation, we assessed overall phylogeny resolution through separate examination of trees from each character set. Exploration of "tree space" with GARLI, using grid computing, showed that hundreds of searches are typically needed to find the best-feasible phylogeny estimate for these data.

Conclusion: Our results (a) corroborate the broad outlines of the current working phylogenetic hypothesis for Ditrysia, (b) demonstrate that some prominent features of that hypothesis, including the position of the butterflies, need revision, and (c) resolve the majority of family and subfamily relationships within superfamilies as thus far sampled. Much further gene and taxon sampling will be needed, however, to strongly resolve individual deeper nodes.

Figure 1

Characterization of tree populations resulting from very-extensive heuristic ML searches (10,000 GARLI runs) on four character sets. For each character set there are two side-by-side graphs: A, E for noLRall2 + nt2; B, F for nt12; C, G for nt123; D, H for nt3. The left-hand graph plots the number of searches resulting in a tree with given topological difference score (Y axis), against the topological difference itself (X axis), where "topological difference" for each search result tree defined as the number of nodes collapsed in the strict consensus between that tree and the overall best tree ("best" tree). A dashed vertical line marks the median topological difference for each data set. The right-hand graph plots topological difference from the overall best tree (Y axis) against difference in ln L from the overall best tree (X axis), expressed as a percent of the best score, for all trees with likelihood scores within 0.005% of the best.

Figure 2

Relationships among sampled families of Ditrysia, comparing working hypothesis to results of five-gene analyses. Fig. 2A. Relationships among sampled families (only) according to composite working hypothesis [5]. Figs. 2B-F: Family relationships according to five-gene analyses, condensed from corresponding 123-taxon trees. Black triangles represent multiple exemplars. Bootstrap values (ML analyses) or posterior probabilities (Bayesian analysis) ≥50% are shown above branches. The corresponding 123- taxon trees, with support levels, are given in Additional files 3, 4, 5, 6 and 7. Fig. 2B. Nt123 ML analysis. Fig. 2C. Nt123 Majority rule consensus tree from Bayesian analysis; "§' symbol marks differences from nt123 ML tree. Fig. 2D. NoLRall2 + nt2 ML analysis. Fig. 2E. Nt12 ML analysis. Fig. 2F. Nt3 ML analysis.

Figure 3

Best 123-taxon ML tree found for nt123. The ML nt123 topology is shown, with bootstrap values (BP) above branches (cladogram on left) separately calculated for ML nt123, ML nt12, and NoLRall2 + nt2, posterior probabilities from Bayesian nt123 analysis below branches. Dashes denote BP < 50%; brackets around BP or posterior probability mean group not recovered in the best ML tree for that character set and analysis. Branch lengths of the phylogram (right side) are proportional to total nucleotide change in ML nt123 tree. Major clade assignment (column to right of taxon names) according to working hypothesis (Fig. 2A): M = Macrolepidoptera; O-M = non-macrolepidopteran Obtectomera; A-O = non-obtectomeran Apoditrysia; D-A= non-apoditrysian Ditrysia.