Walking stick (Insect: Phasmida) Phylogeny and the Evolution of Insect Flight Based on Molecular Data

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Taylor J. Maxwell and Dr. Michael F. Whiting, Zoology

The evolution and development of wings and flight has fascinated scientists for decades. From our research we happened upon an exciting discovery concerning flight in the phylogeny of walking sticks. A basic phylogeny was to be reconstructed from molecular data with the purpose of understanding the classification of the group. The phylogeny enabled us to discover the initial loss of wings and flight in phasmid history and the consequent gain of flight in later lineages. We are not aware of any other organism, which has lost and subsequently regained such a complex character.

Collaborators in Germany and Georgia have been using morphology to do a revision of the current classification. They sent us specimens representing almost all the currently recognized families and subfamilies (13 out of 19 subfamilies) from throughout the world. With 33 phasmid taxa and 10 outgroup taxa from various orders in Polyneoptera (the proposed sister taxa to order Phasmida), we sequenced the 28S (~2100 base pairs) and 18S (~1800 base pairs) ribosomal DNA genes.

Variable regions of the ribosomal genes, called expansion segments, were cut into blocks and the program POY (Wheeler and Gladstein, in press) was used to align them. The aligned pieces were but back together into a final alignment. The outgroup taxa were difficult to align in some regions, subsequently, they were excluded in those regions. This left only the aligned ingroup taxa (the walking sticks) in those regions. Only one area was excluded for both ingroup and outgroup taxa. Genes were analyzed both individually and in a combined data set. Both phylogenetic criterion, Parsimony and Maximum Likelihood, were employed to reconstruct phylogenetic trees using the program PAUP (Swofford, 1993). Parsimony was left unweighted, the model for Maximum Likelihood was GTR+I+G and it was obtained by using Modeltest 3.0 (Posada and Crandall, 1998). Bootstrap supports were obtained for all trees (500 replicates for Parsimony and 100 for Maximum Likelihood) and Bremer and partition Bremer supports were obtained for the parsimony trees. Bremer and bootstrap supports are intended to show relative support for different nodes of the resulting tree. Constraint trees were made to test against particular hypotheses of the tree and the Templeton (1983) signed rank test was used to compare the two hypotheses.

The resulting phylogenetic trees supported the monophyly of the order Phasmida relative to the outgroup taxa. The family Timematidae is the most basal family. Almost all of the proposed subfamilies are paraphyletic (incorrect groupings). This phylogeny disputes the former classifications by Gunther (1953) and Bradley and Galil (1977). With this phylogeny we were able to show the basal condition of winglessness in phasmids. It is undisputed that all proposed sister groups to Phasmida have wings and flight, therefore the primitive condition of the ancestor to walking sticks had flight. This condition was lost in Timematidae and remained so until it was regained by a few lineages much later in the tree. This is the first record of an insect, and any other organism for that matter, to have regained wings and flight after a loss. Many other insect groups have lost flight such as lice and fleas, yet they have never regained it once lost. It has been proposed that all winged insects had a single common ancestor which evolved flight once. The wings of the more derived phasmids appear to be homologous to the wings of other insects. This hints to some sort of developmental or regulatory gene control. It may be that some gene or genes involved in wing development were either not expressed or not recognized in the basal phasmid taxa that have no wings. The expression or recognition of these genes may have been turned back on in later phasmid lineages that do have wings. Developmentally, this is very exciting because this is a testable hypothesis. Walking sticks are easy to rear, easily obtained, and are large insects making it a very good lab organism.

References

  • Bradley, J.C., and B.S. Galil. 1977. The taxonomic arrangement of the Phasmatodea with keys to the subfamilies and tribes. Proc. Ent. Soc. Wash. 79: 176-208.
  • Gunther, K.,1953. Uber die taxonomische Gliederung und die geopgraphische Verbreitung der Insektenordnung der Phasmatodea. Beitr. Entomol. 3:541-563.
  • Posada, D. and Crandall. K.A. 1998. Modeltest: testing the model of DNA substitution. Bioinformatics 14(9): 817-818
  • Swofford, D.L., 1993. PAUP: Phylogenetic analysis using parsimony, version 3.1.1. Illinois Natural History Survey, Champaign.
  • Templeton, A.R. 1983. Phylogenetic inference from restriction endonuclease cleavage site maps with particular reverence to the evolution of humans and apes. Evolution 37:221-244.
  • Wheeler, W.C., and D.L. Gladstein. In Press. POY: Simultaneous alignment and tree reconstruction. Cladistics.