The Controversy of the Phylogyeny of Corbiculate Bees and what could cause it'

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The Controversy of the Phylogyeny of Corbiculate
Beesand what could cause it.

• Lisa Booth

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Outline

• Background
• About Corbiculate Bees
• Behavioral Phylogeny
• Morphological Phylogeny
• Molecular Phylogeny
• Methods and Results
• Alignment
• Outgroup
• Gene
• Conclusions

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Corbiculate Bees Why are they studied?

• Include economically important species
• Form complex eusocial societies

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Corbiculate Bees What are they?

• Family Apidae
• Subfamily Apinae
• Monophyletic group of Long-tongued bees
• Comprised of 4 monophyletic extant tribes
• Euglossini Orchid bee
• Bombini Bumble bee
• Apini Honey bee
• Meliponini Stingless bee

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Euglossini Orchid bee

• Pollinate orchids by collecting chemicals on
  their legs
• Do not possess eusocial behavior
• Most are solitary

www.ufv.br/dbg/bee/euglossa.htm
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Bombini Bumble bee

• Important for crop and wildflower pollination
• Are social
• Form small colonies

www.sankey.ws/bombini.jpg
www.malawicichlidhomepage.com/macro_nature/
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Apini Honey bee
www.durhamsbeefarm.com/photobee.htm

• Only 6-11 species
• Developed social behavior and structure 30
  million years ago
• Maintain the same society today
• Highly eusocial

www.sphoto.com/medium/wkbeecells76.jpg
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Meliponini Stingless bee
www.ecuador-images.net/insect-bee18.jpg

• Bite, but do not sting
• Some produce honey, but not enough to harvest in
  most cases
• Are highly eusocial

encarta.msn.com/.../Stingless_Bee_in_Amber.html
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The Behavioral Phylogeny

• Nolls Research
• 42 Behavioral characteristics
• Characters are based on
• Nest construction
• Food storage
• Worker reproduction
• Female-female interaction
• Food provisioning

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The Behavioral Phylogeny
Orchid
Bumble
Honey
Stingless
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The Behavioral Phylogeny

• Characters that separate the bumble bees from the
  stingless bees.
• 6 Hygienic Habits
• 13 Brood Cell Construction
• 16 Brood Cell Construction
• 18 Cell Construction and Oviposition
• 20 Cell Provisioning
• 24 Brood Food Origin
• 28 Foraging Behavior
• 29 Division of Non-reproductive behavior
• 30 Food exchange among adults
• 31 Nectar exchange among the individuals before
  being stored
• 36 Colony foundation
• 37 Queen foraging
• 40 Dominant female or queen disappearance
• 41 Morphological differences between castes

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The Morphological Phylogeny

• Serraos Research
• Proventricular structure
• Widened Apex
• Triangular Apex
• Spine-like hairs
• Long columnar folds

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The Morphological Phylogeny
Stingless
Honey
Bumble
Orchid
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The Molecular Phylogeny

• Camerons research
• Used Molecular data from 4 genes
• Nuclear DNA
• 28S
• Opsin
• Mitochondrial DNA
• 16S
• Cytochrome b

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The Molecular Phylogeny
Honey
Orchid
Bumble
Stingless
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The Molecular Phylogeny
Orchid
Bumble
Stingless
Honey
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The Molecular Phylogeny
Honey
Orchid
Bumble
Stingless
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The Molecular Phylogeny
Honey
Bumble
Stingless
Orchid
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The Molecular Phylogeny
Honey
Orchid
Bumble
Stingless
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Controversy

• Both Nolls data and Serraos data support the
  single origin of eusocial bees
• Camerons data supports the dual origin of
  eusocial bees

www.entomology.cornell.edu
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Methods Repeat Nolls Analysis

• Copy data matrix from Nolls paper into a NEXUS
  format
• Use PAUP 4.0 beta for a Parsimony analysis
• Use PAUP 4.0 beta to find Bootstrap values
• Looked at each character individually

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Results Behavioral Analysis
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Results Behavioral Analysis

• Single Origin Characteristics
• 4 Nest Coating
• 6 Hygienic Habits
• 9 Cell Walls
• 10 Brood Cell Architecture
• 13 Brood Cell Construction
• 16 Brood Cell Construction
• 18 Cell Construction and ovipositition
• 20 Cell Provisioning
• 24 Brood Food Origin
• 28 Foraging Behavior
• 29 Division of Nonreproductive behavior
• 30 Food exchange among adults
• 31 Nectar exchange among the individuals before
  being stored
• 40 Dominant female or queen disappearance
• 41 Morphological differences between castes

• Dual Origin Characteristics
• 7 Wax envelope covering the brood area
• 8 Materials employed in cell construction
• 12 Reuse of cells or space where larvae are
  reared
• 25 Brood food composition
• 27 Food storage
• 35 Removal of cell walls after cocoon spinning
• 38 Social regulation

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Methods Repeat Camerons Analysis

• Found Camerons aligned data on-line
• Realigned data using ClustalX
• Gap penalty of 10/0.05
• Aligned Ingroup species first
• Aligned Outgroup sequences to the Ingroup
• Used PAUP 4.0 beta for Parsimony and Likelihood
  analyses
• Used Mr. Bayes for a Bayesian analysis

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Results Repeat Camerons Analysis
Bumble
Stingless
Orchid
Honey
28S
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Results Repeat Camerons Analysis
Honey
Bumble
Orchid
Opsin
Stingless
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Results Repeat Camerons Analysis
Bumble
Orchid
Honey
Stingless
16S
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Results Repeat Camerons Analysis
Honey
Orchid
Bumble
Cytochrome b
Stingless
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Results Repeat Camerons Analysis
Honey
Orchid
Bumble
Concatenated
Stingless
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Methods Look at another gene

• Found a new gene on GenBank, Elongation Factor
  1-alpha (nuclear)
• Aligned DNA sequences with ClustalX
• Gap penalty of 10/0.05
• Aligned Ingroup first
• Aligned Outgroup sequences to Ingroup
• Used PAUP for Parsimony and Likelihood analyses
• Used Mr. Bayes for a Bayesian analysis

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Results Look at new gene
Bumble
Stingless
Honey
Elongation Factor
Orchid
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Methods Investigate Outgroups

• Redefined the Outgroup using Camerons outgroup
  species
• Melissodes
• Centris
• Anthophora
• Habropoda
• Xylocopa
• Found new outgroup species for the Cytochrome b
  and Elongation Factor genes
• Allodape
• Braunsapis
• Ceratina

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Results Investigate Outgroups (Parsimony
Trees)
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Results Investigate Outgroups (Likelihood
Trees)
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Results Investigate Outgroups (Bayesian Trees)
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Conclusions Alignment

• Not able to repeat Camerons alignment for all
  genes.
• Calls into question Camerons alignments
• Alignment does affect the tree
• Concatenated Tree
• 16S Tree

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Conclusions Gene Choice

• The different genes do not agree.
• 28S and 16S have similar tree topologies of
  (AE)(BM).
• Both are ribosomal non-coding
• Opsin and Cytochrome b have similar tree
  topologies of A(E(BM)).
• Both are genes for enzymes
• EF has a topology of (AE)(BM).
• Enzyme gene but does not match other enzyme genes.

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Conclusions Outgroups

• Outgroup choice is very important
• Xylocopa and Centris seem to affect the tree the
  most
• Braunsapis, Allodape and Ceratina seem to be good
  outgroups, but I wasnt able to find much data to
  test them fully
• Camerons outgroups didnt group together, they
  mixed with the ingroup which makes them a poor
  outgroup

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Conclusions Model of Analysis

• Slight differences between the different tree
  options (parsimony, likelihood, and Bayesian)
• 16S and Opsin
• were the most
• sensitive to
• the Model of
• Analysis

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Conclusions Controversy

• Not an easily solved problem
• Based on majority it seems that the most likely
  topology is A(E(BM)).
• The Red blocks in the tables
• Least sensitive to all the variants
• Alignment
• Gene
• Outgroup

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Acknowledgements

• Thanks to Dennis Pearl and John Wenzel for all
  their suggestions and help.
• Thanks also to Jeff Pan for his help with
  computer programs.

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References

• Danforth, B.N., J. Fang, S. Sipes, S.G. Brady
  E. Almeida (2004). Phylogeny and molecular
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  Cornell University, Ithaca, NY http//www.entomolo
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• Winston, Mark L. and Charles D. Michener. "Dual
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  Proc. National Academy of Science 74.3 (Mar
  1977) 1135-1137.
• Cameron, Sydney A. and Patrick Mardulyn.
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  Origins of Highly Eusocial Behavior in Bees
  (HymenopteraApinae)." Systematic Biology 50.2
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• Serrao, J.E. A comparative study of the
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  (Hymenoptera, Apidae). Micron 32 (2001)
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