Jessica A. Goldstein, University of Delaware, Department of Entomology and Wildlife Ecology Rebecca B. Simmons, University of North Dakota, Department of Biology A morphological revision of the tiger moth genus Syntomeida Harris. (Lepidoptera: Noctuoidea: Arctiidae: Arctiinae: Euchromiini). Abstract: Members of the family Arctiidae possess many intriguing life history traits, including the use of ultrasonic signals or pheromones for communication during courtship. Most arctiid genera exhibit either one or the other of these types of signals; however, both of these signals can be found in the same genus, Syntomeida Harris, making it an especially useful model for studying the evolution of these traits. The monophyly of Syntomeida has been uncertain historically. Here we present results supporting a new tree splitting Syntomeida into three different genera. Sixty-six Syntomeida and twenty-four outgroup specimens were dissected and their morphologies were examined. Twenty-three morphological characters provided the data matrix and a branch and bound search was performed. These results suggest that Syntomeida is not a real genus containing members of the genus Phoenicoprocta and a yet novel genus. This study shows how genitalia characteristics can be effective at distinguishing relationships at the species level. Also our data suggest that the use of ultrasound during courtship of arctiid moths evolved more than once. 2 trees 87 steps CI = 0.62 RI = 0.71 * Syntomeida vulcana Syntomeida syntomoides Syntomeida ipomoeae Syntomeida austera Syntomeida melanthus Eurota hermoine Agyrtidia uranophila Macrocneme cyanea Chrysoscale ignitia Histaea prosperina Syntomeida epilais Empyreuma pugione Empyreuma anassa Dycladia correbioides Chrysoscale prinicipalis Syntomeida joda Syntomeida hampsonii Phoenicoprocta chrysorrhoea Eurota picta Euchromia sperchia * * * * * trees 87 steps CI = 0.62 RI = 0.71 * Syntomeida vulcana Syntomeida syntomoides Syntomeida ipomoeaeSyntomeida austera Syntomeida melanthus Eurota hermoine Agyrtidia uranophila Macrocneme cyanea Chrysoscale ignitia Histaea prosperina Syntomeida epilais Empyreuma pugione Empyreuma anassa Dycladia correbioides Chrysoscale prinicipalis Syntomeida joda Syntomeida hampsonii Phoenicoprocta chrysorrhoea Eurota picta Euchromia sperchia * * * * * trees 87 steps CI = 0.62 RI = 0.71 * Syntomeida vulcana Syntomeida syntomoides Syntomeida ipomoeaeSyntomeida austera Syntomeida melanthus Eurota hermoine Agyrtidia uranophila Macrocneme cyanea Chrysoscale ignitia Histaea prosperina Syntomeida epilais Empyreuma pugione Empyreuma anassa Dycladia correbioides Chrysoscale prinicipalis Syntomeida joda Syntomeida hampsonii Phoenicoprocta chrysorrhoea Eurota picta Euchromia sperchia * * * * * Shared Characters: Superfamily Noctuoidea -tympanal organs in all ~50,000 species -hear ultrasound, which they use to avoid bats Family Arctiidae -sequester noxious secondary plant compounds -many wasp mimics -metathoracic tymbal organs produce ultrasound that disrupts bat echolocation Genus Syntomeida Harris. - 8 species of diurnal wasp mimics from Nearctics and Neotropics - genus established based on locality and external characteristics - S. syntomoides (Boisduval) S. ipomoeae (Harris) S. melanthus (Cramer)S. vulcana (Druce) S. austera (Dognin)S. hampsonii (Barnes) S. joda (Druce)S. epilais (Walker) Syntomeida appears to be made up of three separate lineages: 1. Core Syntomeida: S. syntomoides, S. ipomoeae, S. melanthus, S. vulcana, and S. austera All use chemical signals (pheromones) to locate a mate. They have tymbal organs, but do not use them during mating (Sanderford, pers. comm.) 2. Phoenicoprocta relatives S. hampsonii and S. joda Use pheromones, but have more striations on their tymbal organs than the previous species (pers. obs.). The presence of these striations suggests that they might use these tymbal organs in courtship, but no studies have examined this possibility. 3. Empyreuma relatives S. epilais Appears to have lost the ability to produce male mating pheromones and documented S. epilais courtship using tymbal organs (Sanderford & Conner, 1990). Pheromone Mating (Core Syntomeida and Phoenicoprocta relatives): -Female perches and releases long range pheromone -Male senses pheromone and flies upwind -Male finds female and releases short range pheromone -Female accepts male and copulation occurs Acoustic Mating (S. epilais): -Female perches and calls to male -Male hears calls and begins calling back meanwhile flying towards the female -Male finds female and continues to call Female accepts male and copulation occurs Objectives. 1.Does taxonomy reflects phylogeny and, accordingly, is the genus Syntomeida monophyletic? 2.If Syntomeida is not monophyletic, how are these species related? 3.How did the use of acoustic signals during courtship evolve in Euchrominii? Materials and Methods. -Dissected 20 species from 10 genera -The ingroup was made up of the 8 species from Syntomeida -The outgroup was made up of 12 species from 9 genera -We used 23 characters 17 of which were genitalia characters -We used MacClade 4.06 (Maddison & Maddison 2000) and PAUP (Swofford 2000) -Transformation series analysis (TSA) was performed resulting in 2 trees with a length of 87 steps Discussion. We had originally hypothesized S. epilais would be found to be closely related to the genus Empyreuma and therefore the use of acoustic signals during mating evolved once (Figure 2). This assertion was not supported by our analysis. Behavioral studies need to be done on the taxa nested in the same clade as S. epilais: Macrocneme, Histaea, Empyreuma, Dycladia and Chrysoscale to see whether or not these species also use their tymbal organs in mating (Figure 3). The behaviors between S. epilais and Empyreuma are not homologous. In S. epilais, both males and females call to each other. In Empyreuma, only males produce sound during courtship. Hence the use of sound in courtship evolved two different ways. This observation is logical because the structures needed to produce these sounds were already present in all tiger moths. The use of these structures in courtship instead of defense probably involved only a simple behavioral change. Our tree supported the other preliminary observations that S. joda and S. hampsonii would be assimilated into the genus Phoenicoprocta, which was clear from their similar morphology. This group has well developed tymbal organs with many striations, which may mean that, like S. epilais, they also use their tymbal organs for courtship. A system where moths use both pheromones and acoustic signals during courtship has not been seen, but it would be interesting to observe the courtship behavior of these two species. At the very least it suggests that members of this genus have relatively advanced defenses against bat predation. Future Directions. Now that the relationships of the genus Syntomeida have been deduced, future research should focus on the systematic placement of “S. epilais”, using molecular and morphological characters. Once the sister genus of S. epilais have been found, then studies examining the evolution of intraspecific communication in arctiids and the extent of this communication can be conducted. Results. There is especially strong support for moving S. hampsonii and S. joda to Phoenicoprocta with a decay index value of 9 (Figure 1). The decay index does offer further support that S. hampsonii and S. joda are each other’s closest relatives by grouping them together with a decay index value of 3. Syntomeida epilais is also separated from the rest of the clade Syntomeida (S. syntomoides, S. ipomoeae, S. melanthus, S. vulcana, and S. austera). The genus Empyreuma is supported, but S. epilais is not included in it, as we had expected. The grouping of the two Empyreuma species E. anassa and E. pugione without S. epilais is strongly supported by the decay index and has a decay value of 5. The genus Syntomeida, as we hypothesized, was also supported by a decay index of 2 as well as the taxon jackknifing. The K-H test (Kashino & Hasegawa, 1989) found that our tree is significantly better than the 1000 random trees as well as the traditional tree and our hypothesized tree (p<0.05 for each comparison). Aknowledgements. Specimens: USNM, CMNH, BMNH, LACM Advice: Jordan Price (SMCM), Julian Dohanue (LACM), Bill Conner (WFU) Figure 1. Observed phylogenetic tree of genus Syntomeida and outgroups used in study. The asterix (*) are branches supported 100% by taxon jackknifing and the numbers are decay indices above 1. Figure 2. Possible evolution of acoustic signals during mating if this character evolved twice in this clade. Figure 3. Possible evolution of acoustic signals during mating if this character evolved once in this clade. image adapted from Works cited. Barnes, W New species of North American Lepidoptera. The Canadian Entomologist 36:165. Boisduval, J. A Glaucopsis syntomoides. Spec. Gen. Lepid. 1: pl. 16, (fig. 4.) Bremmer, K Branch support and tree stability. Cladistics 10: Cramer, J Sphinx melanthus. Papillons Exot. 3: pl. 248 C. Dognin, P Syntomeida austera. Ann. Soc. Entomol. Belg. 46:228. Druce, H Mr. H. Druce on new Species of Lepidoptera. Annals Mag. Nat. Hist. 4:83. Druce, H Supplement. Biol. Centr.-Amer., Lepidoptera Heterocera 2: Harris, T. W Catalogue of North American Sphinges. American Journal of Science 36: Kashino, H. and M. Hasegawa Evaluation of the maximum likelihood estimate of the evolutionary tree topologies from DNA sequence data, and the branching order in Hominoidea. Journal of Molecular Evolution 29: Maddison, D. R. and W. P. Maddison Macclade: Analysis of phylogeny and character evolution, version Sinauer Associates, Sunderland, Massachusets. Mickevich, M. F Transformation series analysis. Systematic Zoology 31: Sanderford, M. V. and W. E. Conner Courtship sounds of the polka-dot wasp moth, Syntomeida epilais. Naturwissenschaften 77: Swofford, D. L PAUP: Phylogenetic Analysis Using Parsimony (*And Other Methods), version 4.0b10. Sinauer Associates, Sunderland, Massachusetts. Walker, F List of the specimens of lepidopterous insects in the collection of the British Museum.1: