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Finding the nearest relatives of
Nasonia (Hymenoptera: Pteromalidae)
Roger Burks
University of California, Riverside
Department of Entomology
What is Nasonia?
• Gregarious puparial parasitoids of
calyptrate flies in bird nests and
refuse
• Model system, better known than any
other species of Chalcidoidea—
genome project ongoing
• Three species, each infected by two
unique strains of Wolbachia
The three species of Nasonia
• Females almost identical (Darling & Werren 1990)
• Males differ in degree of wing reduction
• Nasonia vitripennis worldwide, synanthropic
• N. giraulti in eastern North America,
N. longicornis in western North America
– specialized on flies in bird nests
Wolbachia basic background
• Bacteria infecting arthropods and filarial nematodes
• Transmitted vertically from mother to offspring (Binnington &
Hoffmann 1989)
• Cause crossing incompatibility in Nasonia (Breeuwer & Werren 1990)
• Phylogenetic congruence between bacteria and host
usually absent– horizontal transmission?
• May cause rapid speciation in arthropods (Laven 1959, 1967; Breeuwer
& Werren 1990)
How Wolbachia affects Nasonia
• Cytoplasmic Incompatibility (Breeuwer & Werren 1990)
– Causes death of offspring of mothers that do not have
same Wolbachia strains as the father
• Incompatible crosses:
– Uninfected female x infected male
– Infected female x male infected by at least one different
strain
• Infection rate near 100% in wild Nasonia
– “Cured” colonies used to study Wolbachia effects in lab
Why Nasonia’s relationships still
need studying
• Nasonia is a model system for evolutionary
biology studies, yet…
• Ancestral states cannot be inferred with only
three analyzed species!
• No agreement in classification of wasps in its
family (Pteromalidae)
• Needed: means to reject some pteromalids as
close Nasonia relatives
Pteromalidae is a scary taxon
•
•
•
•
587 genera in 31 subfamilies
Pteromalinae with only 283 genera
Parasitoids of various terrestrial arthropods
No previous phylogenetic analysis using
more than 10 pteromaline genera
• Previous analyses with either morphology
only or 28S ribosomal sequences only
Pteromalinae molecular vs.
morphological rates of evolution
• 283 genera of Pteromalinae, but...
• 28S D2 sequence divergence equal to that
of the genus Aphelinus (Heraty 2004)
• Rapid morphological evolution or
ribosomal constraints?
• Rapid evolution due to Wolbachia?
Tools for the search
• Morphology
– 105 morphological characters (work in progress)
• 28S D2-D5 ribosomal DNA, Wingless
– Secondary structure alignment for 28S (Gillespie et al. 2005)
to be compared with POY results
• Analysis with parsimony (PAUP, TNT, POY),
maximum likelihood, Mr. Bayes
• Hypothesis testing with ML using CONSEL
Outgroup selection
• Based on Heraty lab matrix of Chalcidoidea
– 28S D2-D5, 18S E17-E35 ribosomal DNA
– 471 taxa (including outgroups)
– All families, 84 total subfamilies represented
• Subfamilies Diparinae, Ormocerinae are
legitimate outgroups for Pteromalinae
Combined 28S and Wingless molecular results, Parsimony (PAUP)
black = Pteromalinae
red = other Pteromalids
* = Wolbachia positive
Numbers indicate
bootstrap support
(1000 replicates)
Agrees with simple POY
run in topology
1176 steps in PAUP
rci = 0.209
ri = 0.403
Combined 28S and Wingless molecular results, Mr. Bayes 3.1
black = Pteromalinae
red = other Pteromalids
* = Wolbachia positive
6 parameters, 4 chains, partitioned
by gene region, 1 million
generations
Numbers indicate
posterior probability
Combined 28S and Wingless molecular results, Likelihood
black = Pteromalinae
red = other Pteromalids
* = Wolbachia positive
model: GTR+I+G
program: PAUP
Testing hypotheses not present in the optimum
maximum likelihood tree (500 total sampled trees for test)
constraint tree with:
au test p value
sh test p value
Nasonia+ Urolepis clade
0.81
1.00
Nasonia + Trichmalopsis
+ Urolepis paraphyly
0.50
0.85
monophyletic
Pteromalinae
0.38
0.80
Nasonia +
Trichomalopsis clade
0.23
0.79
monophyletic
Trichomalopsis
0.07
0.70
0.01**
0.50
paraphyletic Nasonia
au = approximately unbiased test (Shimodaira 2002)
sh = Shimodaira-Hasegawa test (Shimodaira & Hasegawa 1999)
Problem: Not enough variation to have statistical power
Solution: Add a more rapidly evolving gene
Candidates:
Long-wavelength Rhodopsin—multiple copies?
Pten—contains intron, but short
Cytochrome Oxidase I & II—AT richness
Perspective
• Trichomalopsis sarcophagae 28S sequence (>1100
base pairs) differs from that of Nasonia vitripennis
by only 1 base pair
• Sampling remains incomplete
– Nasonia not well-surveyed in Palearctic region
– Trichomalopsis with 54 species!
Trichomalopsis microptera male
They differ by only one base pair in 28S??
Trichomalopsis sarcophagae
Nasonia vitripennis
Further goals
• Sequence from more species of
Trichomalopsis, other genera near Nasonia
(>120 specimens to be sequenced)
• Finish morphological analysis
• Wolbachia survey across Pteromalinae,
comparing bacteria and wasp phylogenies
Acknowledgments
Advisory committee:
John Heraty
Richard Stouthamer
Bob Luck
Cheryl Hayashi
Jack Werren
Matt Yoder
Doug Yanega
Serguei Triapitsyn
Lara Baldo
James Russell
Genet Tulgetske
Danel Vickerman
Heraty lab:
Dave Hawks
Johan Liljeblad
James Munro
Jeremiah George
Jason Mottern
Chrissy Romero
Adena Why
Jutta Burger
Matt Buffington
Funded by: NSF FIBR: 0328363