The evolutionary history of holometabolous insects inferred from transcriptome-based phylogeny and comprehensive morphological data

Abstract

Background: Despite considerable progress in systematics, a comprehensive scenario of the evolution of phenotypic characters in the mega-diverse Holometabola based on a solid phylogenetic hypothesis was still missing. We addressed this issue by de novo sequencing transcriptome libraries of representatives of all orders of holometabolan insects (13 species in total) and by using a previously published extensive morphological dataset. We tested competing phylogenetic hypotheses by analyzing various specifically designed sets of amino acid sequence data, using maximum likelihood (ML) based tree inference and Four-cluster Likelihood Mapping (FcLM). By maximum parsimony-based mapping of the morphological data on the phylogenetic relationships we traced evolutionary transformations at the phenotypic level and reconstructed the groundplan of Holometabola and of selected subgroups.

Results: In our analysis of the amino acid sequence data of 1,343 single-copy orthologous genes, Hymenoptera are placed as sister group to all remaining holometabolan orders, i.e., to a clade Aparaglossata, comprising two monophyletic subunits Mecopterida (Amphiesmenoptera + Antliophora) and Neuropteroidea (Neuropterida + Coleopterida). The monophyly of Coleopterida (Coleoptera and Strepsiptera) remains ambiguous in the analyses of the transcriptome data, but appears likely based on the morphological data. Highly supported relationships within Neuropterida and Antliophora are Raphidioptera + (Neuroptera + monophyletic Megaloptera), and Diptera + (Siphonaptera + Mecoptera). ML tree inference and FcLM yielded largely congruent results. However, FcLM, which was applied here for the first time to large phylogenomic supermatrices, displayed additional signal in the datasets that was not identified in the ML trees.

Conclusions: Our phylogenetic results imply that an orthognathous larva belongs to the groundplan of Holometabola, with compound eyes and well-developed thoracic legs, externally feeding on plants or fungi. Ancestral larvae of Aparaglossata were prognathous, equipped with single larval eyes (stemmata), and possibly agile and predacious. Ancestral holometabolan adults likely resembled in their morphology the groundplan of adult neopteran insects. Within Aparaglossata, the adult's flight apparatus and ovipositor underwent strong modifications. We show that the combination of well-resolved phylogenies obtained by phylogenomic analyses and well-documented extensive morphological datasets is an appropriate basis for reconstructing complex morphological transformations and for the inference of evolutionary histories.

Figure 1

Combined and simplified cladogramm of holometabolan insect relationships, with selected autapomorphies for the clades addressed in this study. The topology is taken from the ML tree inferred from dataset 1 (i.e., the complete datamatrix). (1) Bootstrap support (BS) (bottom, black) is derived from 72 bootstrap replicates (MRE-based bootstopping criterion) of dataset 1. (2) BS values for the specific phylogenetic relationship (bottom, red) are derived from ML tree inferences from the seven specific decisive datasets 1 to 7. (3) relative support [%] values for the specific phylogenetic relationship (top) are derived from the Four-cluster Likelihood Mapping (FcLM) with the seven specific decisive datasets. Apomorphies are selected from the full lists of reconstructed groundplan characters (see Additional file 4, Chapter 5).

Figure 2

Illustration of reconstructed groundplan larva of Holometabola. The putative groundplan larva was orthognathous, and equipped with simplified but distinctly developed compound eyes, and well developed thoracic legs. Abdominal prolegs and cerci were absent. For a list of larval and adult groundplan characters of Holometabola, see Table 4. ce: compound eye. fro: frons. ant: antenna. cl: clypeus. lbr: labrum. md: mandible. mx: maxille. lb: labium. t1: tergite of first thoracic segment. pl1: pleurite of first thoracic segment. spi2: spiracle of second thoracic segment. plr: pleural ridge. cx: coxa. tr: trochanter. fe: femur. tib: tibia. ta: tarsus. cla: claw. spiI: spiracle of first abdominal segment. sV: sternite of fifth abdominal segment. spiVIII: spiracle of eighth abdominal segment. tX: tergite of tenth abdominal segment.

Figure 3

Three holometabolan adult thorax states. A) A thorax with approximately equally sized pterothoracic segments is possibly ancestral for Aparaglossata (Figure shows thorax of Nannochorista neotropica (Mecoptera, Nannochoristidae); prothorax not shown.). B) shows a thorax of taxa with anteromotorism, i.e., flight with mainly the fore wings (e.g., Hymenoptera, Trichoptera, “higher” Lepidoptera, and Diptera; figure shows Ptychoptera sp. (Diptera, Ptychopteridae)). This state is possibly ancestral for Holometabola. However, the reconstruction of the ancestral state of this character in the formal analysis remained ambiguous for Holometabola and Aparaglossata. C) shows a thorax of taxa with posteromotorism, i.e., flight with the hind wings (Coleoptera and Strepsiptera; figure shows Mengenilla moldrzyki (Strepsiptera, Mengenillidae)). red: muscles. blue: sceleton. green: gut. yellow: nerves. Numerals refer to thoracic segments. th: thorax segment. g: ganglion. dlm: dorsal longitudinal muscle. dvm: dorso-ventral muscle. vlm: ventral longitudinal muscle (not visible in A and B). A 3D version of this figure can be found as Additional file 5 (Click on image to activate animation).