Download A Molecular Phylogeny of the Snail Killing Flies (Sciomyzidae

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 genericlevel 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*
71
80
54
63
95
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
T. arnaudi 23
100
T. arnaudi 24
T. kerteszi 46
100
T. kerteszi 47
T. mesopor a 40
100
80
T. mesopor a 42
T. plumosa 11
100
T. plumosa 43
T. phyl lophora 39
76
T. fusci nervi s 53
100
T. fusci nervi s 54
T. fer rug inea 34
T. mel anosti g ma 2
T. plebej a 13
100
100
T. robusta 16
T. sil vatica 35
T. montana 68
E. soli cita 5
E. soli cita 6
Bayesian Tree
E. soli cita 5
E. soli cita 6
T. montana 68
T. robusta 16
T. sil vatica 35
T. phyl lophora 39
T. mel anosti g ma 2
T. plebej a 13
T. fer rug inea 34
T. fusci nervi s 53
T. fusci nervi s 54
T. kerteszi 46
T. kerteszi 47
T. plumosa 11
T. plumosa 43
T. arnaudi 23
T. arnaudi 24
T. mesopor a 40
T. mesopor a 42
Timeline for completion of project
Specimens in hand for sequencing:
Atrichomelina
Coremacera
Dictya
Ectinocera
Elgiva
Euthycera
Hydromyia
Illione
Pherbina
1 (1)
1 (9)
2 (42)
1 (1)
1 (7)
1 (18)
1 (1)
1 (8)
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