Department of Biological Sciences Systematics of the snail-killing flies in the tribe Tetanocerini (Diptera: Sciomyzidae): Evolution of larval feeding strategies in the genus Tetanocera Eric G. Chapman Department of Biological Sciences Kent State University Ph. D. Prospectus

Sciomyzid Biology • Larvae are predators, parasitoids, or saprophages of: - Terrestrial Snails - Semi-aquatic and aquatic non-operculate snails - Operculate aquatic snails - Semi-terrestrial succineid snails - Slugs - Snail eggs - Fingernail clams - Freshwater oligochaete worms • Members of the family were grouped into 17 distinct feeding guilds by Knutson & Vala (2002).

Sciomyzid Relationships Maximum Parsimony cladogram of generic-level relationships with 1000 bootstrap replicates (values greater than 50 shown). Data from Marinoni & Mathis (2000).

Tetanocera Biology • All species are predators/parasitoids of snails as larvae • Eggs are laid in the snails’ habitat • Newly hatched larvae are “free living” - Search for a suitable host snail - Attach to the host and begin feeding • Once the snail dies, the larvae continue feeding until all suitable tissue is gone • Larvae then search for another host snail • Pupation occurs either while floating or on land

Tetanocera Biology • Species grouped into 6 feeding guilds by Knutson & Vala (2002) according to the type of prey and habitat • Aquatic Pulmonate Snails • Shoreline Pulmonate Aquatic Snails • Succiniid Snails • Slugs • Terrestrial Snails - Six feeding guilds is 2 more that any other genus of Tetanocerini

Tetanocera plebeja

Tetanocera plumosa

Past Tetanocera Research • 40 species worldwide have been described (29 North American species) • Life cycles of about 2/3 of the N. American species have been discovered by Dr. Benjamin Foote. • A morphological phylogenetic analysis of the N. Am. species done by Dr. Maryanne Shemory as part of her dissertation - 29 taxa & 22 characters did not paint a clear picture of the evolutionary relationships between the species

Tetanocera Relationships Strict consensus of 2 MP trees - bootstrap values > 50 shown (1000 replicates). Data from Shemory (1997) re-analyzed using PAUP*

Materials & Methods • Genes sequenced thus far (all mitochondrial): - COI (Cytochrome Oxidase I)…….1449 bp - COII (Cytochrome Oxidase II)……731 bp - 16s…………………………………428 bp Total = 2608 bp • Genes to be tried: - 28s (nuclear) - 12s (mitochondrial) - Other nuclear markers…

Results • The 16s, COI, & COII genes of 1-4 individuals of 12 Tetanocera species and 1 outgroup (Elgiva) • A total of 19 specimens were sequenced • Parsimony/bootstrap analysis shows that there is enough variability in these genes to be useful in phylogenetic analysis

Neighbor-Joining Dendogram

Bayesian Tree

Timeline for completion of project Specimens in hand for sequencing: Atrichomelina 1 (1) Coremacera 1 (9) Dictya 2 (42) Ectinocera 1 (1) Elgiva 1 (7) Euthycera 1 (18) Hydromyia 1 (1) Illione 1 (8) Pherbina 1 (4) Pherbellia 1 (92) Poecilographa 1 (1) Pteromicra 1 (18) Renocera 3 (7)? Sciomyza 1 (5) Sepodon 3 (72) Tetanocera 20 (40) Tripetoptera 1 (2)

Timeline for completion of project • Collecting trips: - Summer 2003: collect across Canada -Goal: get remainder of N. Am. Tetanocera - Winter 2003-2004: - Trip to Argentina & Chile - June 2004: Trip to Mexico & Guatemala -Goal: get members of the 15 sciomyzid genera endemic to the Neotropics

Materials & Methods • Fresh specimens preserved in 100% EtOH • Thorax removed and total DNA isolated using Qiagen DNeasy Animal Tissue Kit • 16s Ribosomal gene fragment (550 bp) amplified using 16Sar-L and 16Sbr-H primers • CO1 Ribosomal gene fragment (650 bp) amplified using LCO 22-me and HCO 700-dy primers

Materials & Methods • Amplified DNA purified electrophoretically overnight in a 1.5% NuSeive gel, and cleaned with the Wizard Gel Extraction kit • Sequencing reactions on purified 16s & CO1 DNA (both light & heavy strands) done with Amplicycle sequencing kit • DNA sequenced on a LiCor 4200S-2 gel scanner using KBplus 5.5% acrylamide gel • Bases called using LiCor Base ImagIR image analysis program

Materials & Methods • Light & heavy strand reads aligned using GeneJockeyII and discrepancies were cleared up by examining the gel images • 16s & CO1 sequences of all available taxa were aligned with GeneJockeyII (saved as text) • Text file converted to “nexus” file using Microsoft Word • Data matrix (nexus file) “cleaned up” using MacClade • Trees generated with PAUP* for Parsimony & Neighbor-Joining analysis, and with Mr. Bayes for Bayesian analysis