Transcript 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. mesopora 40
100
80
T. mesopora 42
T. plumosa 11
100
T. plumosa 43
T. phyll ophora 39
76
T. fusci nervi s 53
100
T. fusci nervi s 54
T. ferr ugi nea 34
T. mel anosti gma 2
T. plebej a 13
100
100
T. robusta 16
T. sil vati ca 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