PHYLOGENY OF NORTH AMERICAN APHAENOGASTER SPECIES
(HYMENOPTERA: FORMICIDAE) RECONSTRUCTED WITH MORPHOLOGICAL
AND DNA DATA
By
Bernice Bacon DeMarco

A DISSERTATION
Submitted to
Michigan State University
in partial fulfillment of the requirements
for the degree of
Entomology – Doctor of Philosophy
2015

ABSTRACT	
  
	
  
PHYLOGENY OF NORTH AMERICAN APHAENOGASTER SPECIES (HYMENOPTERA:
FORMICIDAE) RECONSTRUCTED WITH MORPHOLOGICAL AND DNA DATA
By
Bernice Bacon DeMarco
The ant genus Aphaenogaster Mayr is an ecologically diverse group that is common
throughout much of North America. Aphaenogaster has a complicated taxonomic history due to
variability of taxonomic characters. Novomessor Emery was previously synonymized with
Aphaenogaster, which was justified by the partial mesonotal suture observed in A. ensifera Forel.
Previous studies using Bayesian phylogenies with molecular data suggest Aphaenogaster is
polyphyletic. Convergent evolution and retention of ancestral similarities are two major factors
contributing to non-monophyly of Aphaenogaster. Based on 42 multi-state morphological
characters and five genes, we found Novomessor more closely related to Veromessor Forel and
that this clade is sister to Aphaenogaster. Our results confirm the validity of Novomessor stat. r.
as a separate genus and it is resurrected based on the combination of new DNA, morphological,
behavioral and ecological data.
Twenty-three Aphaenogaster species (Hymenoptera: Formicidae) occur in North
America. While morphology and ecology define most species, the species limits of a group in the
Eastern United States are unclear. In particular, the morphological and behavioral characters
once thought to define A. carolinensis, A. picea and A. rudis do not associate with their
hypothesized species limits. These observations suggest that these species are not monophyletic.
We therefore tested the monophyly of Aphaenogaster in the context of molecular phylogenetic
analyses. We used DNA data from five genes: CO1, CAD, EF1αF2, Long-wavelength
Rhodopsin and Wingless to reconstruct phylogenies for 44 Aphaenogaster and outgroup species.

In the resulting trees, reconstructed using parsimony and Bayesian inference, species boundaries
associate with well-supported monophyletic clades of individuals collected from multiple
locations. For example, A. carolinensis was monophyletic and a missing CAD intron was a
diagnostic trait for the clade. However, some clades were unresolved, and A. picea and A. rudis
were not monophyletic. Given the short branch lengths, these results suggest that these ants have
likely recently radiated, and lack of gene lineage sorting explains the non-monophyly of species.
Conversely, these results may indicate that clades of multiple species represent fewer but
morphologically varied species. Additional biological information concerning pre- and postmating barriers is needed before a complete revision of species boundaries for Aphaenogaster.
Aphaenogaster Mayr 1853, contains 227 species worldwide (Bolton 2006) with 23 valid
North American species, several species of which are hard to separate based on morphology
alone (Umphrey 1996). The difficulty in identifying some of these species is due to limited
diagnostic characters and to the lack of a comprehensive illustrated key. A recent analysis
returned three species from Aphaenogaster to Novomessor, thus making Aphaenogaster in North
America monophyletic (DeMarco and Cognato 2015). While many species have easily
identifiable morphological characters, some east coast species within the A. rudis clade in North
America are difficult to differentiate. Two of these species, A. carolinensis and A. miamiana, can
be diagnosed using DNA. The gene CAD was missing an intron in those taxa. Four additional
taxa, all identified morphologically as A. rudis, were found to be polyphyletic (DeMarco and
Cognato, in prep, or see Chapter 2).

	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
Copyright	
  by	
  
BERNICE	
  BACON	
  DEMARCO	
  
2015	
  

	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
	
  
This	
  work	
  is	
  dedicated	
  to	
  my	
  husband	
  for	
  his	
  unfailing	
  support	
  during	
  the	
  time	
  it	
  took	
  to	
  do	
  
my	
  research	
  and	
  complete	
  my	
  dissertation.	
  	
  I	
  will	
  be	
  forever	
  grateful.	
  

	
  

v	
  

ACKNOWLEDGEMENTS

This dissertation was possible with strong support from my major professor, Dr. Anthony
I. Cognato and from the rest of my graduate committee, Drs. Christina DiFonzo, L. Alan Prather
and Richard E. Triemer. All of you were willing to work with me, even though I chose a taxon
you were unfamiliar with. I am also indebted to Gary Parsons, Sarah Smith, and Rachel Olson
for help in the lab, reading papers and giving advice when needed.
Specimens were generously donated and loaned to this research by the following
individuals: Dr. Phil Ward (University of California, Davis), Dr. Jack Longino (University of
Utah, Dr. Corrie Moreau (The Field Museum, Chicago), Lloyd Davis, Dr. Gary Alpert, Stefan
Cover, Dr. David Lubertazzi and Dr. Aaron Ellison (Harvard University), Dr. Robert Johnson
(Arizona State University), Dr. Benoit Guenard (University of Hong Kong), Mark O’Brien
(University of Michigan Museum of Zoology), Dr. Richard Brown and Joe MacGown
(Mississippi Entomological Museum at Mississippi State University), Dr. Robin Verble-Pearson
(University of Arkansas at Little Rock) and Walter Tschinkel (Florida State University). This
work would not have been possible without the institutions housing these specimens and the
curators who maintain them. In addition, I received valuable advice from Drs. Moreau, Ward,
Alpert and Stefan Cover during the Ant Course and beyond. I am grateful for their support. I
am also grate to The University of Michigan for allowing me access to the Edwin S. Goerge
Reserve to collect speciemnes.
This work was made possible, in part, by funding provided by the Rhodes (Gene)
Thompson Memorial Fellowship and Michigan State University Entomology Department Hutson
Fund as well as travel grants from Systematics and Evolutionary Biology Division of

	
  

vi	
  

Entomological Society of America and the Harvard Ernst Mayr Grant. Additional funding was
provided by the following NSF Grant: 2012-2015, National Science Foundation. Dimensions in
Biodiversity: Collaborative Research: The climate cascade: functional and evolutionary
consequences of climatic change on species, trait, and genetic diversity in a temperate ant
community. PIs: Nate Sanders, Aaron Ellison, Nick Gotelli, Sara Helms Cahan, Bryan Ballif,
Rob Dunn.
Finally, I am indebted to my husband, children Paul and Stephanie, and to the rest of my
family for their patience and moral support throughout the entire degree program.
Please note that figures have been edited to conform to the Michigan State University
dissertation formatting guidelines. Please refer to resulting publications for the final version of
the figures.

	
  

vii	
  

TABLE	
  OF	
  CONTENTS	
  
	
  
LIST	
  OF	
  TABLES………………………………………………………………………………………………………..….…x	
  
	
  
LIST	
  OF	
  FIGURES…………………………………………………………………………………………………………...xi	
  	
  
	
  
CHAPTER	
  1	
  	
  
INTRODUCTION	
  TO	
  APHAENOGASTER	
  (HYMENOPTERA:	
  FORMICIDAE)…………………………..1	
  
Ecological	
  and	
  behavioral	
  diversity	
  of	
  Aphaenogaster…………………………………...…….....2	
  
Systematics of Aphaenogaster and preliminary cladistics analysis of morphology………5
CHAPTER	
  2	
  
	
  PHYLOGENETIC	
  ANALYSIS	
  OF	
  APHAENOGASTER	
  SUPPORTS	
  THE	
  RESURRECTION	
  OF	
  
NOVOMESSOR	
  (HYMENOPTERA:	
  FORMICIDAE)……………………………………………………………….7	
  
	
  
Abstract……………………………………………………………………………………………………………….8	
  
	
  
Introduction…………………………………………………………………………………………………...……8	
  
	
  
Materials	
  and	
  Methods………………………………………………………………………….……………10	
  
	
  
	
  
Morphological	
  characters………………………………………………………………………...11	
  
	
  
	
  
Molecular	
  characters……………………………………………………………………...………..14	
  	
  
	
  
Results………………………………………………………………………………………………………………16	
  
	
  
	
  
Identification	
  key	
  to	
  included	
  genera……………………………..………………...………17	
  
	
  
Discussion…………………………………………………………………………………………………..……..17	
  
	
  
	
  
Included	
  species……………………………………………………………………………………...21	
  
	
  
CHAPTER	
  3	
  	
  
A	
  MULTIPLE	
  GENE	
  PHYLOGENY	
  REVEALS	
  POLYPHYLY	
  AMONG	
  EASTERN	
  NORTH	
  
AMERICAN	
  APHAENOGASTER	
  SPECIES	
  (HYMENOPTERA:	
  FORMICIDAE)………………………..22	
  
	
  
Abstract…………………………………………………………………………………………..…………………23	
  
	
  
Introduction………………………………………………………………………………………………………23	
  
	
  
Materials	
  and	
  Methods……………………………………………………………………………………….26	
  
	
  
Results………………………………………………………………………………………………………………28	
  
	
  
Discussion………………………………………………………………………………………………………….30	
  
	
  
CHAPTER	
  4	
  	
  
APHAENOGASTER	
  (HYMENOPTERA:	
  FORMICIDAE)	
  OF	
  NORTH	
  AMERICA:	
  A	
  KEY	
  TO	
  
SPECIES	
  USING	
  MORPHOLOGY	
  AND	
  DNA………………………………………………………………………34	
  
	
  
Abstract…………………………………………………………………………………………..………………...35	
  
Introduction………………………………………………………………………………………....……………35	
  
	
  
Materials	
  and	
  Methods…………………………………………………………………….……...……….…37	
  
	
  
Glossary……………………………………………………………………………………………………...……..38	
  
	
  
Identification	
  key	
  to	
  Aphaenogaster	
  species……….……………….………………………...…….39	
  
	
  
Overview	
  of	
  Species……………………………………………………………………………………………43	
  
	
  
APPENDICES……………………………………………………………………………………………………………......55	
  
	
  

viii	
  

	
  
Appendix	
  A:	
  Tables	
  and	
  Figures	
  for	
  Chapter	
  2……………………………………………………..56	
  
	
  
Appendix	
  B:	
  Tables	
  and	
  Figures	
  for	
  Chapter	
  3……………………………………………………..68	
  
	
  
Appendix	
  C:	
  Tables	
  and	
  Figures	
  for	
  Chapter	
  4……………………………………….…………..109	
  
	
  
BIBLIOGRAPHY…………………………………………………………………………………………………………..138	
  
	
  

	
  

ix	
  

LIST	
  OF	
  TABLES	
  	
  
	
  
	
  
	
  
Table	
  1.1.	
  Specimens included in current analysis with associated localities and Genbank
numbers…………………………………………………………………………………………..57
Table 1.2. Morphological character state matrix for Aphaenogaster and outgroup species….…59
Table 1.3. PCR primers used for the amplification of gene loci……...……………………..…..63
Table	
  2.1.	
  Aphaenogaster and outgroup specimens with associated localities and Genbank
numbers………………………………………………………………………………………..…69
Table 2.2. Bootstrap and partitioned bremer support values which correspond to the label nodes
in the parsimony phylogeny (Fig. 2)…………………………………………………………….74
Table 3.1. A list of Aphaenogaster species known from North America……………...………110
	
  

	
  

x	
  

LIST	
  OF	
  FIGURES	
  
	
  
	
  
	
  
Figure 1.1. One of 5 most parsimonious trees reconstructed for 43 taxa with morphology data in
a TNT analysis. Bootstrap values are above the branches. Clades without bootstrap values were
not resolved in a strict consensus tree. A. = Aphaenogaster, C. = Camponotus, F. = Formica, M.
= Messor, My. = Myrmica, N. = Novomessor, So. = Solenopsis, St. = Stenamma, V. =
Veromessor…………………………………………………………………………………...….64
Figure 1.2. One most parsimonious tree reconstructed for 43 taxa with morphology and DNA
data (CO1, CAD, EF2, LWR, WG) of 43 in a TNT analysis. Bootstrap values/Bremer supports
are above the branches. A. = Aphaenogaster, C. = Camponotus, F. = Formica, M. = Messor,
My. = Myrmica, N. = Novomessor, So. = Solenopsis, St. = Stenamma, V. = Veromessor……...65
Figure 1.3. Bayesian majority rule consensus tree reconstructed for 43 taxa with morphology
and five genes (CO1, CAD, EF2, LWR, WG) in a Mr. Bayes analysis, Posterior probabilities
values greater than 90% are above the branches (* > 90%, **= 100%). Data were partitioned by
gene and codon position and and analyzed with a best-fit GTR + I + G model, 20 million
generations and a burn-in of 5,000,000 generations A. = Aphaenogaster, C. = Camponotus, F. =
Formica, M. = Messor, My. = Myrmica, N. = Novomessor, So. = Solenopsis, St. = Stenamma, V.
= Veromessor.……………..……………………………………………………………………..67
Figure 2.1. One of 64,525 MPT reconstructed for 123 taxa of Aphaenogaster and outgroups
with DNA data and analysis of 5 genes in PAUP*. Bootstrap values greater than 90% are above
the branches (* > 90%, **= 100%). A. = Aphaenogaster. Specimen numbers and
states/provinces where collected are displayed next to each sample. The names of nonmonophyletic species correspond to specific colors…………………………..….……………..77
Figure 2.2. One of 64,525 MPT shown as a cladogram reconstructed for 123 taxa of
Aphaenogaster and outgroups with DNA data and analysis of 5 genes in PAUP*. Node numbers
are above the branches. A. = Aphaenogaster. Specimen numbers and states/provinces where
collected are displayed next to each sample. The names of non-monophyletic species correspond
to specific colors…………………………………………………………………….………...…84
Figure 2.3. Maximum likelihood tree reconstructed for 123 taxa with DNA data and analysis of
5 genes in a RAxML analysis. Bootstrap values greater than 90% are above the branches (* >
90%, **= 100%). A. = Aphaenogaster. Specimen numbers and states/provinces where collected
are displayed next to each sample. The names of non-monophyletic species correspond to
specific
colors……………………………………………………………………………….……...……..91
Figure 2.4. Bayesian majority rule consensus tree reconstructed for 123 taxa with morphology
and five genes in a Mr. Bayes analysis, Posterior probabilities values greater than 90% are above
the branches (* > 90%, **= 100%). Data were partitioned by gene and codon position and
analyzed with a best-fit GTR + I + G model, 30 million generations and a burn-in of 7,500,000
	
  

xi	
  

generations. A. = Aphaenogaster. Specimen numbers and states/provinces where collected are
displayed next to each sample. The names of non-monophyletic species correspond to specific
colors………………………………………………………………………………….…...…..…99
Figure 3.1. Lateral view of Aphaenogaster mariae showing striae on first gastral
tergite…………………………………………………………………………………...……....111
Figure 3.2. Scape shapes for 4 species of Aphaenogaster…………………………………….112
Figure 3.3. Propodeal	
  spine	
  shape	
  and	
  angle	
  for	
  8	
  species	
  of	
  Aphaenogaster……………..113
Figure 3.4. Relative eye size for 4 species of Aphaenogaster………………………………...114
Figure 3.5. Lateral, head and dorsal views of Aphaenogaster ashmeadi (Emery)…………….115
Figure	
  3.7.	
  Lateral and head views of	
  Aphaenogaster boulderensis Smith…………………...116
Figure 3.6. Lateral, head and dorsal views of Aphaenogaster carolinensis Wheeler………….117
Figure 3.8. Lateral, head and dorsal views of Aphaenogaster flemingi Smith………………...118
Figure 3.9. Lateral, head and dorsal views of Aphaenogaster floridana Smith……………….119
Figure 3.10. Lateral, head and dorsal views of Aphaenogaster fulva Roger…………………..120
Figure 3.11. Lateral, head and dorsal views of Aphaenogaster huachucana Creighton………121
Figure 3.12. Lateral, head and dorsal views of Aphaenogaster lamellidens Mayr…………….122
Figure 3.13. Lateral, head and dorsal views of Aphaenogaster mariae Forel…………………123
Figure 3.14. Lateral, head and dorsal views of Aphaenogaster megommata Smith………...…124
Figure 3.15. Lateral, head and dorsal views of Aphaenogaster mexicana (Pergande)………...125
Figure 3.16. Lateral, head and dorsal views of Aphaenogaster miamiana Wheeler…….…….126
Figure 3.17. Lateral, head and dorsal views of Aphaenogaster mutica Pergande………….….127
Figure 3.18. Lateral, head and dorsal views of Aphaenogaster occidentalis (Emery)………...128
Figure 3.19. Lateral, head and dorsal views of Aphaenogaster patruelis Forel……………….129
Figure 3.20. Lateral, head and dorsal views of Aphaenogaster picea (Wheeler)……….……..130
Figure 3.21. Lateral, head and dorsal views of Aphaenogaster punctaticeps MacKay…..……131
	
  

xii	
  

Figure 3.22. Lateral, head and dorsal views of Aphaenogaster rudis Enzmann………...……..132
Figure 3.23. Lateral, head and dorsal views of Aphaenogaster tennesseensis (Mayr)………...133
Figure 3.24. Lateral, head and dorsal views of Aphaenogaster texana Wheeler………………134
Figure 3.25. Lateral, head and dorsal views of Aphaenogaster treatae Forel…………………135
Figure 3.26. Lateral, head and dorsal views of Aphaenogaster uinta Wheeler………………..136
Figure 3.27. Lateral, head and dorsal views of Aphaenogaster umphreyi Deyrup & Davis…..137

	
  

	
  

xiii	
  

CHAPTER 1
INTRODUCTION TO APHAENOGASTER (HYMENOPTERA: FORMICIDAE)

1	
  

Ecological and behavioral diversity of Aphaenogaster

Aphaenogaster Mayr 1853 contains 227 species worldwide (Bolton, 2006) with 23 valid
North American species, reduced from 31 original species descriptions. The North American
taxa have not been taxonomically reviewed in over 60 years (Creighton 1950). Umphrey (1996)
attempted to discriminate a complex group of ten sibling species of the Aphaenogaster fulvarudis-texana complex in northeastern US with karyotypes and morphology. He concluded that
karyotypes provided the best, but imperfect, means for species diagnosis. He acknowledged that
DNA would ultimately prove useful as a definitive method for separating these groups.
Aphaenogaster has been a popular genus for many studies including biology and natural
history (Lubertazzi 2012), tool use (Fellers and Fellers 1976), communication (Menzel and
Marquess 2008), interactions with other ant taxa (Bewick et al. 2014) and temperature tolerance
(Warren and Chick 2013).
In Connecticut, Lubertazzi (2012) found nesting sites for ants in the Aphaenogaster rudis
group in soil, in rotting wood, under rocks and in leaf litter. Nests in soil were shallow and had a
single entrance. Lubertazzi (2012) planted 25 artificial wooden nests at 3 different sites and
followed them through an entire year. Seventeen nests survived; all but one contained a queen.
Half the nests produced males, but only 3 produced female alates. The smallest nest contained
183 and the largest nest 1033 workers, with an average nest size of 613 individuals. He
measured foraging distances by placing baits randomly in a 10 m square area and following
workers back to the nests. The average foraging distance was 57 cm. Aphaenogaster behavior
was observed as timid around other ant species, and they did not defend foraging territories.
These ants laid a trail pheromone (Attygalle et al. 1998) using their poison gland to recruit nest
	
  

2	
  

mates to food items. They fed on small invertebrates including termites (Buczkowski and
Bennett 2007), eliaosome bearing seeds (Heithaus et al. 2005 and Clark and King 2012) and
even mushrooms (Carroll, et al. 1981). Luburtazzi (2012) also observed caste attributes. He
found that winged reproductives left the nest between late July and mid-August. Mated queens
and brood overwintered in the soil, and workers began foraging in early spring. They are some
of the earliest foragers observed in the forest. Larvae hatched from eggs in about 20 days, there
are four larval instars, with an average larval period of 28 days, and the pupal stage lasts 16 days.
Total time from egg to ecolsion averaged 64 days. Workers fed first instar larvae a liquid diet
from food stored in their crop, while later instars were able to ingest solid foods. Haskins (1960)
observed queens in Aphaenogaster picea able to survive 8-13 years.
Not all Aphaenogaster species nest in the same habitats. Aphaenogaster treatae nests in
open, sandy fields under clumps of grass or lichen (Talbot, 1954). Aphaenogaster megommata is
a nocturnal ant in the deserts of southwest US and nests in sand. Aphaenogaster tennesseensis is
a social parasite that enters the nests of Aphaenogaster rudis and A. fulva (Creighton, 1950). The
A. tennesseensis queen is attractive to the workers in the A. rudis and A. fulva nests, and they
unknowingly raise her eggs as their own (Creighton, 1950). Aphaenogaster tennesseensis have a
wide geographical range, from Virginia south to Florida, and east to Iowa and Nevada. They are
morphologically distinct in that they lack hairs on the mesosoma and gaster (Ellison, et al. 2012).
Aphaenogaster mariae Forel occurs in Virginia and Mississippi but is rarely collected. It is an
arboreal species with a starburst pattern of striae on the first gastral tergite (Ellison et al. 2012).
Aphaenogaster ashmeadi and A. treatae are identified by the size of a lobe at the base of the
scape (Creighton, 1950). Other NA Aphaenogaster do not have this lobe. Aphaenogaster
ashmeadi is found throughout the southeast, while A. treatae occurs from the southeast north into

	
  

3	
  

Michigan. Aphaenogaster lamellidens is also morphologically distinct with a tooth or lobe on
the frontal carina that is rearward facing towards the back of the head (Creighton, 1950). They
nest in similar habitats as the rest of the northeastern Aphaenogaster species with nests in soil,
under rocks and in rotting pine and oak logs. They are found throughout the southeast.
Substrate vibration generating behavior has been observed in ants in the ant genera
Messor (Grasso et al. 1999), Novomessor (Markl and Holldobler 1978), Atta (Roces and
Holldobler 1996), and Solenopsis (Rauth and Vinson 2006). Ants in these genera use
stridulation to create sounds in response to the discovery of a food source. Menzel and Marquess
(2008) also observed substrate vibration generating behavior in Aphaenogaster. They described
this behavior and its causes in Aphaenogaster carolinensis. This ant produces vibrations by
striking, then dragging its mandible across a substrate surface. They concluded that this behavior
was not in response to food, but a reaction to the presence of non-nest mate conspecifics and to a
lesser extent, ants from other species.
Bewick et al. (2014) observed interactions between A. rudis and two other ant species,
Prenolepis imparis and Nylanderia faisonensis. The three species were chosen because of
different nest sizes and different feeding habits. Bewick et.al (2014) compared specific species
traits (food discovery rate, food clearance rate, body mass, dominance hierarchy and thermal
niche), the effect and interaction of interspecific competition and climate change on community
composition. Equalizing discovery rates, food clearance rates and dominance had a negative
effect on A. rudis and P. impairis, but a positive effect on N. faisonensis. Equalizing body mass
had the opposite effect. Compared to the other traits, loss of thermal niches had less of an effect
on the evenness of species distribution (but the most severe effect on local coexistence), with
increases for A. rudis and P. impairis and a decrease for N. faisonensis. The overall conclusion
	
  

4	
  

was that climate change would have a negative effect on P. impairis (known as the winter ant),
but also surprisingly on N. faisonensis, which is active during the summer months.
Aphaenogaster rudis faired the best and Bewick et al. (2014) predicted that the three community
species would decrease to A. rudis and P. impairis.
Warren and Chick (2013) examined data over a 38-year period of upward movement for
A. rudis and A. picea along the southern end of the Appalachian Mountain chain in Georgia. In
1974, 100% of Aphaenogaster ants at 900 m elevation were A. picea. By 2012, 25% of the
Aphaenogaster ants at 900 m were A. rudis and only 75% were A. picea. Warren and Chick
(2013) also tested thermal tolerance of individuals of both species. The absolute temperature
range for A. picea was a minimum of -0.5 °C and a maximum of 42.5°C. The absolute
temperature range for A. rudis was a minimum of 2 °C and a maximum of 43.5°C. These results
indicate possible changes in insect species as climates increase in temperature, due to the
differeing thermal tolerance levels for both species.
Systematics of Aphaenogaster and preliminary cladistics analysis of morphology
Aphaenogaster species systematics has been difficult for the lack of morphologically
diagnostic and phylogenetically informative characters. To demonstrate the need for additional
characters (i.e., DNA), I conducted a parsimony analysis using morphological characters gleaned
from previous studies (Creighton 1950, Ward 1985 and Coovert 2005). 42 morphological
characters were coded for 25 Aphaenogaster and 10 outgroup species (Chapter 1, Table 1). The
phylogenetic analysis resulted in 5 parsimonious trees; the strict consensus of the trees was
mostly unresolved (Chapter 1, Fig. 1). There was relatively high support for the clades
containing the outgroups, but no support for Aphaenogaster relationships.

	
  

5	
  

Given the lack of phylogenetically informative morphological characters, DNA was
utilized to potentially help resolve the phylogeny. Previous studies in ant systematics utilized a
number of genes to resolve relationships among ant taxa. We used one mitochondrial gene
(Cytochrome oxidase I), which was used for taxa within a genus (Branstetter 2012, Lucky 2011)
and four nuclear genes that have provided resolution at higher levels (Brady et al. 2006, Moreau
et al. 2013).
The purpose of this dissertation is to reconstruct Aphaenogaster phylogeny with
molecular characters to elucidate relationships within the genus. Chapter One incorporated a
small sample of 44 taxa to show that previously, Aphaenogaster was polyphyletic within North
America. Novomessor, which includes three species, was resurrected, making Aphaenogaster in
North America a monophyletic clade. Chapter Two examined a larger sample of 123 taxa and
showed a number of taxa as monophyletic, but some samples, identified as Aphaenogaster rudis,
were polyphyletic. Chapter Three provided a revised key to Aphaenogaster in North America
and included DNA data for definitive diagnosis of some species.

	
  

6	
  

CHAPTER 2
PHYLOGENETIC ANALYSIS OF APHAENOGASTER SUPPORTS THE RESURRECTION
OF NOVOMESSOR (HYMENOPTERA: FORMICIDAE)

	
  

7	
  

Abstract
The ant genus Aphaenogaster Mayr is an ecologically diverse group that is common
throughout much of North America. Aphaenogaster has a complicated taxonomic history due to
variability of taxonomic characters. Novomessor Emery was previously synonymized with
Aphaenogaster, which was justified by the partial mesonotal suture observed in A. ensifera Forel.
Previous studies using Bayesian phylogenies with molecular data suggest Aphaenogaster is
polyphyletic. Convergent evolution and retention of ancestral similarities are two major factors
contributing to non-monophyly of Aphaenogaster. Based on 42 multi-state morphological
characters and five genes, we found Novomessor more closely related to Veromessor Forel and
that this clade is sister to Aphaenogaster. Our results confirm the validity of Novomessor stat. r.
as a separate genus and it is resurrected based on the combination of new DNA, morphological,
behavioral and ecological data.
Introduction

The ant genus Aphaenogaster Mayr, 1853 is a speciose group, which has not been
taxonomically reviewed in over 60 years (Creighton 1950). Aphaenogaster contains 227
worldwide species (Bolton 2006) with 23 valid North American species reduced from 31 original
species descriptions. They are an ecologically diverse group that is common throughout much of
North America (Creighton 1950). They occur in deciduous forests, open grassy areas, pine
barrens and sand hills. Ecologically, they are general scavengers, feeding on a variety of
arthropods, and other small invertebrates; they are also keystone seed dispersers in mesic forests
of eastern North America (Lubertazzi 2012). Many species live in dead wood and promote
	
  

8	
  

decomposition and nutrient recycling (Warren and Bradford 2012).
Aphaenogaster has a complicated taxonomic history due to variability of taxonomic
characters. Mayr (1853) described the genus based on two new species from Italy, A. sardoa
Mayr, 1853 and A. senilis Mayr, 1853. Mayr (1863) later moved the genus into Atta (Fabricius,
1804) as a subgenus. Emery (1895) removed Aphaenogaster from Atta and placed it as a
subgenus of Stenamma Westwood, 1839. Emery (1908) decided it merited generic status.
Umphrey (1996) addressed the complicated Aphaenogaster fulva-rudis-texana complex using
morphometric characters and karyotypes to identify ten taxa that included six previously
recognized species and four undescribed species. He concluded that additional DNA data was
needed to define and diagnose these groups. Recent inclusion of DNA data in Bayesian
phylogenetic analyses resolved Aphaenogaster as polyphyletic, including Messor Forel, 1890
and Stenamma (Brady et al. 2006, Moreau and Bell 2013). Ward (2011) suggested that
convergent evolution and retention of ancestral similarities were two major factors contributing
to polyphyly of Aphaenogaster.
Aphaenogaster taxonomy was further complicated with the description of Novomessor
Emery, 1915 and Veromessor Forel, 1917. Brown (1974) synonymized Novomessor with
Aphaenogaster and returned two species, N. ensifera Forel, 1899 and N. manni Wheeler and
Creighton, 1934, to Aphaenogaster. Based on this synonymy, he reduced Novomessor to a
subgenus of Aphaenogaster. Hölldobler et al. (1976) resurrected Novomessor to generic status;
however, Bolton (1982 and 2003) considered Novomessor as a junior synonym of
Aphaenogaster and Veromessor as a junior synonym of Messor. The synonymy of Novomessor
with Aphaenogaster was justified by the partial mesonotal suture in A. ensifera (Brown 1974).
The Novomessor lineage of three species A. albisetosa Mayr, 1886, A. cockerelli André, 1893
	
  

9	
  

and A. ensifera, has several distinct morphological characters, as well as behaviors and habitat
preferences. Two of the species, A. albisetosa and A. cockerelli, do not have a mesonotal suture
and the three species differ from most other North American Aphaenogaster by their large size
(2x). They inhabit desert environments, but forage in the morning (Sanders and Gordon 2002), as
compared to other desert species of Aphaenogaster that forage at night (personal observation of
BBD). Aphaenogaster albisetosa and A. cockerelli also exhibit a stridulating behavior not
observed in other cogeners (Hölldobler et al. 1978). They drag their abdomen over sand to
recruit nestmates to help with prey. In addition, Hölldobler et al. (1976) suggested the
resurrection of Novomessor based on the presence of a new complex exocrine gland in the
Novomessor species. Ward et al. (2014) recently resurrected Veromessor based on DNA
evidence.
The preponderance of morphological, ecological and behavioral differences suggests the
validity of Novomessor. However, monophyly of Novomessor has not been tested, which is
necessary for the delimitation of a genus. The close relationship between Aphaenogaster,
Messor, Veromessor and Novomessor has made molecular tools crucial to understanding the
relationships between these taxa. In this study, we test the monophyly of Novomessor in
phylogenetic analyses using molecular, morphological, ecological and behavioral data from a
sample of North American Aphaenogaster species.
Materials and Methods
Specimens were collected at a number of North American localities, including wooded
areas of eastern and central US, and the western deserts. Additional specimens were borrowed
from colleagues and institutions (Table 1.1). Historically poorly collected areas were

	
  

10	
  

specifically targeted, such as the Michigan Upper Peninsula. Specimens were collected using an
aspirator and baits (peanut butter and pecan shortbread cookies) and stored in 100% ethanol at 80 °C. At least 12 workers per nest were collected at each site. Specimens were deposited at the
A.J. Cook Arthropod Research Collection, Michigan State University, East Lansing, Michigan.
The following 42 morphological characters/indices (Ward 1985, Bolton 1994, Lucky and Ward
2010, Brady and Ward 2005) were scored for the phylogenetic analysis (Table 1.2). All
multistate characters are unordered and each character is based on workers. Sculpture terms are
from Harris (1979).
Morphological characters
1. Cephalic index (head width / head length): (0) 0.99 mm or less; (1) 1 mm; (2) 1.01 mm +.
2. Frontal triangle: (0) shiny; (1) striated; (2) punctate; (3) finely punctate; (4) absent.
3. Lobe at base of scape: (0) lobe absent; (1) lobe flat and thin (as seen from side); length not
more than 1/5 scape; (2) lobe thick (as seen from side); length usually 1/4 scape or longer; (3)
small angled extension at scape base.
4. Sculpturing on mandible: (0) striated; (1) punctate.
5. Apical 4 segments paler in color than rest of antenna: (0) no; (1) yes (Coovert 2005).
6. Antennal segment number: (0) 10 segments; (1) 12 segments.
7. Antennal segment color: (0) same as head; (1) lighter than head; (2) darker than head.
8. Antennal club: (0) antennal funiculi without a differentiated club; (1) antennal funiculi
terminate in weak 4-segmented club; (2) antennal funiculi terminate in 3-segmented club; (3)
antennal funiculi terminate in 2-segmented club.
9. Psammophore: (0) no hairs under head; (1) psammophore present; (2) some long hairs under
head; but not a complete psammophore.
	
  

11	
  

10. Antennal scape index: (0) 0.99 or less; (1) 1; (2) 1.01 - 2.0; (3) 2.01 - 3.00; (4) 3.01+.
11. Palp formula indicates the number of maxillary and labral palp segments, respectively.
(0) 2,2; (1) 4,3; (2) 5,3; (3) 6,4.
12. Ocular index (eye length x eye width / head width): (0) 0.001 - 0.009; (1) 0.010 - 0.039; (2)
0.040 - 0.07 ;(3) 0.071 +.
13. Clypeal margin shape: (0) emarginated; (1) slightly emarginated; (2) straight; (3) emarginate
notched); (4) bicarinate without teeth; (5) bicarinate with teeth.
14. Sculpture pattern on head: (0) fine rugae; (1) long rugae; (2) long wavy rugae; (3) coarse
rugae; (4) coarse sculpturing; (5) shiny; (6) long to transverse rugae; (7) punctate; (8) finely
punctate.
15. Sculpture location on head: (0) to occiput; (1) to top of eyes; (2) to bottom of eyes; (3) none.
16. Posterior border of clypeus with deep; semicircular impression: (0) no; (1) yes.
17. Anterior edge of mesonotum rising abruptly above adjacent portion of pronotum: (0) no; (1)
yes.
18. Mesosoma lacking erect hairs: (0) no hairs present; (1) hairs present.
19. Sculpturing on pronotum: (0) punctate; (1) finely punctate; (2) coarsely punctate; (3) fine
rugae; (4) coarse rugae; (5) fine transverse rugae; (6) coarse transverse rugae; (7) coarse
sculpturing; (8) shiny.
20. Sculpturing on mesonotum: (0) punctate; (1) finely punctate; (2) coarsely punctate; (3) fine
rugae; (4) coarse rugae; (5) fine transverse rugae; (6) coarse transverse rugae; (7) coarse
sculpturing; (8) shiny.
21. Sculpturing on propodeum: (0) punctate; (1) finely punctate; (2) coarsely punctate; (3) fine
rugae; (4) coarse rugae; (5) fine transverse rugae; (6) coarse transverse rugae; (7) coarse

	
  

12	
  

sculpturing; (8) shiny.
22. Propodeal spines: (0) spines absent; (1) spines present; (2) spines present but small; (3)
spines present but very small; (4) spines present; but thin; (5) spines present, but small and
triangular.
23. Spine index (Spine width/Spine length): (0) = 0; (1) 0.01 - 0.85; (2) 0.86 - 1.2; (3) 1.21 - 3.0;
(4) 3.01 +.
24. Coxae sculpturing: (0) shiny; (1) finely punctate; (2) punctate; (3) 1st coxa finely punctate;
others shiny; (4) fine rugae.
25. Coxae color (compared to mesosoma): (0) same; (1) lighter; (2) darker.
26. Leg color (compared to mesosoma): (0) same; (1) lighter; (2) darker.
27. Weber’s length: (0) 0.75 - 0.99 mm; (1) 1.0 - 1.6 mm; (2) 1.61 - 2.0 mm; (3) 2.01 - 3.00 mm;
(4) 3.01 - 3.99 mm; (5) > 4.00 + mm.
28. Promesonotal suture: (0) no; (1) yes; (3) indistinct
29. Striae on first gastral tergite: (0) no striae; (1) Striae present.
30. Erect hairs on gastral tergite and sternite: (0) no; (1) yes.
31. Appressed hairs on gastral tergite and sternite: (0) no; (1) many; (2) sparse.
32. Metasoma color compared to mesosoma: (0) same; (1) lighter; (2) darker.
33. Gaster color compared to head: (0) same; (1) lighter; (2) darker.
34. Petiole/postpetiole: (0) petiole only; (1) Petiole and postpetiole.
35. Petiole index (petiole length/ petiole height): (0) 1.0 - 1.24;(1) 1.25 - 1.55; (2) 1.56 - 1.8; (3)

	
  

13	
  

1.81 +.
36. Postpetiole index (postpetiole length/ postpetiole height): (0) none; (1) 0.70 - 0.99 mm; (2)
1.00 - 1.25; (3) 1.26 +.
37. Hind femur length: (0) 0.8 - 0.99 mm; (1) 1.00 - 1.99 mm; (2) 2.00 - 2.99 mm; (3) 3.00 - 3.99
mm; (4) 4.00 mm +.
38. Outer face of frontal lobe bearing a flange which projects rearward in the form of a
tooth: (0) No tooth; (1) tooth present.
39. Mandible slender and triangular with outer margin not strongly curving toward midline: (0)
no; (1) yes.
40. Spine shape: (0) none; (1) angled back; (2) angled back; thin; (3) small right angle; (4)
angled up; (5) triangular; (6) angled back and curved in; (7) angled back; small; (8) angled up
and small; (9) curved back.
41. Spine angle: (0) 180°; (1) 120° +; (2) 130° +; (3) 140° +; (4) 150° +; (5) 160° +.
42. Petiole constricted with junction at gaster: (0) none; (1) slight constriction; (2) strong
constriction;
(3) no postpetiole.
Molecular characters
Molecular data were assembled for genetic loci, which were phylogenetically informative for ant
genera (Brady et al. 2006, Ward et al. 2010) (Table 1.3). DNA was extracted from ants
preserved in 100% ethanol using a silica-based spin column procedure (Qiamp, Qiagen Inc.,
Santa Clara, CA), following the manufacturer’s tissue protocol. Specific regions of

14	
  

mitochondrial (CO1, 650 base pairs or bp) and nuclear DNA [carbomoylphosphate synthase
(CAD, 816 bp), Elongation factor 1-alpha F2(EF2, 517 bp), Long Wavelength Rhodopsin(LWR,
560 bp) and Wingless(WG, 428 bp)] were amplified via polymerase chain reaction (PCR). The
total number of base pairs for all genes was 2972. For the mitochondrial gene CO1, the
annealing temperature was 50°C, and for the nuclear genes CAD and LWR were 54°C, for EF2,
53°C and for WG, 58°C. These loci were amplified following published protocols (Table 1.3).
After PCR, unincorporated deoxyribonucleotide triphosphates (dNTPs) and oligonucleotides
were removed from PCR reactions with Exo-SAP (http=//www.usbweb.com/) and directly
sequenced on an ABI 3700 automated sequencer using a BigDye (Applied Biosystems, Inc.,
Foster City, CA) fluorescent chemistry reaction, with both sense and anti-sense strands
sequenced for all individuals. Sequences were aligned using Sequencher® version 5.2 and
deposited in Genbank (Table 1.2). CO1 sequences were produced for all taxa. The following
taxa were missing sequences: CAD, Camponotus pennsylvanicus (DeGeer, 1773), Formica
glacialis Wheeler, 1908, Veromessor andrei (Mayr, 1886), Messor bouvieri Bondroit, 1918,
Myrmica latifrons Stärke,1927, Aphaenogaster balcanica (Emery, 1898), A. boulderensis Smith,
1941, A. floridana Smith, 1941, A. huachucana, A. mutica Pergande, 1896, A. patruelis Forel,
1886, A. tennesseensis (Mayr,1862), A. texana Wheeler 1915, A. treatae Forel, 1886 and A.
umphreyi Deyrup and Davis, 1998; EF2, V. andrei and A. huachucana Creighton, 1934; LWR,
Solenopsis aurea Wheeler, 1906, A. boulderensis, one individual of A. picea Wheeler, 1908. A.
texana, and A. uinta Wheeler, 1917; WG, one individual of A. ashmeadi (Emery, 1895), and A.
tennesseensis.
Phylogenetic analysis was performed using the computer software TNT (Goloboff et al.
2008). The analysis used the new technology search in TNT that included four search models:

15	
  

ratchet (Nixon 1999), sectorial searches, drifting and fusing. Default settings were used except
for ratchet, which was set at 10 perturbations and 200 iterations. Bootstrap analysis used
resampling, with 1000 replicates. Bremer support was preformed with the script Bremer.run
from the TNT wiki website (http://tnt.insectmuseum.org/index.php/Bremer_Support).
We also inferred a phylogeny with likelihood with RAxML (Stamatakis 2014) and
Bayesian analysis With Mr. Bayes via the CIPRES Gateway (Huelsenbeck and Ronquist 2001,
Miller et al. 2010). For both analyses, data were partitioned by gene, and codon position (Castoe
et al. 2004), with models of evolution applied independently to each partition (Nylander et al.
2004). We used MrModeltest 3.7 (Nylander 2004) for the selection of partition-specific
substitution models for the nucleotide data using the Akaike Information Criterion in order to
decrease the potential of over parameterizating the models although complex models often
perform as well or better than simpler models (Nylander et al. 2004). We followed guidelines to
make credible Bayesian inferences (Bollback 2002, Huelsenbeck and Ronquist 2001). The bestfit model for all genes was GTR + I + G.
Results
Using TNT (Goloboff et al. 2008) a morphological matrix was constructed with 42
characters and 43 taxa. Analysis of these data resulted in five most parsimonious trees. The
consensus of these trees was unresolved and showed no support except for the outgroups. The
three species of the Novomessor lineage, A. albisetosa, A. cockerelli and A. ensifera grouped
with Aphaenogaster (Figure 1.1). Veromessor was polyphyletic and Messor was within
Aphaenogaster. This illustrates the unreliability of using only morphological characters within
this group. However, morphological characters are diagnostic for these genera, including scape

16	
  

index and the amount of constriction between the postpetiole and the gaster (see Key).
Parsimony with morphology and DNA, in addition to maximum likelihood and Bayesian
analyses with DNA only, resolved a monophyletic Novomessor, which was sister to the
Veromessor species, and with Novomessor and Veromessor clades as sister to Aphaenogaster
(Figs. 2, 3). Since maximum likelihood and Bayesian analysis resulted in a nearly identical
topology, only the parsimony Bayesian analyses are shown. The European Messor species were
imbedded within the Aphaenogaster clade. The relationships among the Novomessor and
Veromessor species were well supported (Figures 1.2, 1.3). There was variable support for the
subclades within the Aphaenogaster clade (Figures 1.2, 1.3). These results confirm that
Novomessor stat. r. is monophyletic and is resurrected from synonomy under Aphaenogaster.
Identification	
  key	
  to	
  included	
  genera	
  
This	
  key	
  is	
  modified	
  from	
  Creighton	
  (1950)	
  for	
  the	
  following	
  4	
  genera.	
  
1a.	
  Scape	
  index	
  (scape	
  length/head	
  width)	
  is	
  less	
  than	
  1,	
  psammophore	
  often	
  
present………………………………………………………………………………………………….............2	
  
1b.	
  Scape	
  index	
  greater	
  than	
  1,	
  psammophore	
  absent…………………………….............3	
  
2a.	
  New	
  World	
  species	
  with	
  small	
  to	
  large	
  propodeal	
  spines………..…..Veromessor	
  
2b.	
  Old	
  World	
  species	
  with	
  no	
  or	
  small	
  propodeal	
  spines………………………..Messor	
  
3a.	
  Distinct	
  promesonotal	
  suture,	
  postpetiole	
  constricted	
  at	
  connection	
  to	
  
gaster…………………………………………..……………………………….……….…	
  Aphaenogaster	
  
3b.	
  Indistinct	
  promesonatal	
  suture,	
  postpetiole	
  not	
  constricted	
  at	
  connection	
  to	
  
gaster…………………….…………….……………………………………….……………….Novomessor	
  

Discussion
The lack of resolution of the morphology-based tree is not surprising because of the
limited number of variable characters found for Aphaenogaster. However, resolution was
17	
  

recovered where expected; for the outgroup species and Novomessor. The outgroup species have
several apomorphic characters that separate them from Aphaenogaster. Formica Linneaus 1758
and Camponotus Mayr, 1861 are in Formicinae, and have a petiole, but no post-petiole.
Myrmica Latreille, 1804 lacks a distinct peduncle, making the petiole shorter. Veromessor
species in this analysis have a complete psammophore, or a fringe of long hairs beneath the head
(V. andrei and V. julianus (Pergande, 1894)), while Messor species in this analysis have an
incomplete psammophore or few long hairs beneath the head, including M. bouvieri and M.
denticornis Forel, 1910. Stenamma has a bicarinate clypeus.
Along with morphology, several DNA, behavioral and habitat characters are diagnostic
for Novomessor. Novomessor albisetosa, N. cockerelli and N. ensifera are found in xeric
habitats, while most North American Aphaenogaster are found in woodland or field habitats.
Novomessor albisetosa and N. cockerelli are abundant at low mid-altitudes in arid habitats
(Wheeler and Creighton 1934). Both species form conspicuous nests with sloppy gravel craters,
and the workers are active from late afternoon into the night hours (Wheeler and Creighton
1934). They feed on seeds, plant material and dead or dying insects. Novomessor ensifera is
known only from Mexico, and nests in soil that consists of many large stones buried in coarse
sand (Kannowski 1954). Kannowski (1954) also did not observe plant material, but only dead
insects in their nests. Some Aphaenogaster occur in open, grassy habitats, pine barrens, and sand
hills. Most Eastern Aphaenogaster species build nests in soil, sand or under rocks, but in forest
habitats, nests may also be found in rotten logs, branches, stumps, and occasionally live trees.
Aphaenogaster texana, which is found in the southwest, occurs at a higher elevation, and is
found in dead logs or under rocks, which differs from those in the Novomessor lineage, but is
similar to many of the remaining North American Aphaenogaster species. Additionally, there are
18	
  

several desert dwelling species within Aphaenogaster including A. boulderensis, A. huachucana,
A. megommata Smith, 1963, and A. uinta. Morphologically, characters of the promesonatal	
  
suture,	
  and the postpetiole, are diagnostic for Novomessor. Reproductive characters such as the
forewing venation (Brown 1974) are potentially diagnostic but these characters need further
examination in a future study concerning Aphaenogaster.
This study provides another example of molecular phylogenies elucidating generic
boundaries for taxonomically challenging groups like Aphaenogaster. The
molecular/morphological-based phylogenies provide a strong justification for the delimitation
and recognition of Novomessor as for other ant genera in recent studies. Stenamma was shown
to form two separate clades (Holarctic and Middle American regions) using a ten gene
concatenated dataset (Branstetter 2012). Blaimer (2012) synonymized five of 13 former
subgenera of Crematogaster Lund, 1831 under C. (Orthocrema), and the remaining eight under
Crematogaster sensu stricto. Using five genes and morphology, Blaimer (2012) concluded that
there was a deep divergence event between Crematogaster and Orthocrema Santschi, 1918, and
provided a key to separate these subgenera based on morphology. Lucky and Ward (2010) and
Lucky (2011) also provided the first molecular and morphological phylogeny of Leptomyrmex
Mayr, 1862. They separated Leptomyrmex into two clades, “micro-“ and “macro-” Leptomyrmex.
The “macro” species have wingless queens and are found in Australia, New Caledonia and New
Guinea. The “micro-“ Leptomyrmex species are found only in southeast Australia. Additionally,
nine subspecies were elevated to species status (Lucky and Ward 2010). Given precedence set by
these studies, we resurrect Novomessor based on its monophyly, nucleotide differences, and
morphological diagnostic characters. Furthermore, our results (Figures 1.2, 1.3) and others
(Brady et al. 2006, Moreau and Bell 2013) indicate that Aphaenogaster is polyphyletic with
	
  

19	
  

inclusion of the European species of Messor which are sister to A. japonica (Figures 1.2, 1.3).
Additional phylogenetic study and subsequent generic revision are needed to resolve the
polyphyly of Aphaenogaster.
Genus Novomessor Emery, 1915
(Complete taxonomic references for Novomessor in Bolton 2006)
Diagnosis: Morphological characters that separate Novomessor from Aphaenogaster
include a head width and length each of greater than 2 mm, and a striated frontal triangle above
the clypeus. The intraocular distance is 1.4 mm or greater. The distance between the tips of the
spines is greater than 0.56 mm and the spine length is 1 mm or longer. The Weber’s length is 3
mm or greater, and the promesonotal suture is indistinct or absent. Characters that diagnose
Messor from Novomessor include a large metasternal process in Messor, which is smaller in
Novomessor and a quadrate head in Messor, which is elongate in Novomessor. In addition,
Novomessor has no constriction of the postpetiole as the gaster, Messor and Veromessor have a
slight constriction and Aphaenogaster has a strong constriction.
Description: Workers in Novomessor are 8-8.5 mm in length and reddish brown in color.
The head in all three species is longer than it is wide, the mesosoma has long transverse rugae
and the gaster is darker than the head. Novomessor albisetosa and N. cockerelli have long hairs
under the head resembling a psammophore, while Novomessor ensifera has only short hairs.
They have well-developed propodeal spines, averaging 1 mm in length. Novomessor albisetosa
and N. cockerelli can be difficult to distinguish from each other, but the long wavy rugae on the
head end at the top of the eyes in N. cockerelli and extend to the occiput in N. albisetosa.
Distribution: Novomessor albisetosa and N. cockerelli occur in southeastern Arizona,

20	
  

southern New Mexico, southwestern Texas and northern Mexico at elevations from 150 to 300
m. Novomessor cockerelli is found on the desert floor, with large, crater-like nest entrances
surrounded by coarse gravel. Novomessor albisetosa is found near N. cockerelli, but their nests
occur in the desert foothills, under flat rocks or stones, and surrounded by gravel. Novomessor
ensifera has only been found in Mexico, in sandy soil with large stones present (Kannowski
1954).
Biology: Both N. cockerelli and N. albisetosa forage late in the day and into evening, and
feed upon small insects, seeds and bits of plant tissue (Cook, 1953). Kannowski (1954)
describes a different foraging pattern for N. ensifera. He observed them foraging in the early
morning and late afternoon, and also only observed them feeding on insects, but no plant
material. He also found no plant pieces or seeds in their nests.
Included Species
Novomessor albisetosa (Mayr, 1886), new combination (restored status)
Novomessor cockerelli (André, 1983), new combination (restored status)
Novomessor ensifera (Forel, 1899), new combination (restored status)
Novomessor manni (Wheeler and Creighton, 1934) junior synonym of N. ensifera

	
  

21	
  

CHAPTER 3
A MULTIPLE GENE PHYLOGENY REVEALS POLYPHYLY AMONG EASTERN NORTH
AMERICAN APHAENOGASTER SPECIES (HYMENOPTERA: FORMICIDAE)

22	
  

Abstract
Twenty-three Aphaenogaster species (Hymenoptera: Formicidae) occur in North
America. While morphology and ecology define most species, the species limits of a group in the
Eastern United States are unclear. In particular, the morphological and behavioral characters of
A. carolinensis, A. picea and A. rudis overlap. These observations suggest that these three
species are not monophyletic. We therefore tested the monophyly of Aphaenogaster in the
context of molecular phylogenetic analyses. We used DNA data from five genes: CO1, CAD,
EF1αF2, Long-wavelength Rhodopsin and Wingless to reconstruct phylogenies for 44
Aphaenogaster and outgroup species. In the resulting trees, reconstructed using parsimony and
Bayesian inference, species boundaries associated with well-supported monophyletic clades of
individuals in most of the 23 North American Aphaenogaster collected from multiple locations.
However, some clades were unresolved, and both A. picea and A. rudis were not monophyletic.
Although this may indicate that clades of multiple species represent fewer but morphologically
varied species, given the short branch lengths, the lack of resolution may reflect the fact that
these ants have recently radiated, and a lack of gene lineage sorting explains the non-monophyly
of species. Additional biological information concerning pre- and post-mating barriers is needed
before a complete revision of species boundaries for Aphaenogaster.

Introduction
The number of recognized ant species worldwide increases each year. Hölldobler &
Wilson (1990) estimated that there were 8800 described species. By 2007, Fisher & Cover

23	
  

(2007) reported 12,000, and currently AntWeb (http://www.antweb.org) posts almost 16,000
valid species and subspecies. Ants are ubiquitous in many ecosystems, and ecologically
dominant as predators, scavengers and herbivores (Wilson & Hölldobler, 2005). Although ants
make up approximately 2 percent of the known global insect fauna, they comprise at least one
third of its biomass (Wilson & Hölldobler 2005). In the tropics, they can make up to 94% of the
biomass of tropical rainforest canopies (Davidson et al. 2003). Consequently, ants play an
important role in the environment and their study depends on a thorough understanding of their
diversity.
The woodland ant genus Aphaenogaster includes important seed dispersers in North
American forests, and has been the focus of a number of ecological and evolutionary studies
(Lubertazzi 2012, Warren & Chick 2013, Bewick et al. 2014). Previous systematic studies
focused on species descriptions (Kiran et al. 2008, Shattuck 2008, Longino & Cover 2004);
however, the magnitude of species diversity of Aphaenogaster is unclear due to few and
conserved distinguishing morphological characters.
Aphaenogaster have expanded frontal carinae that partially or wholly cover the antennal
insertions (Creighton 1950). All members of this genus have 12-segmented antennae with a long
scape, well-developed eyes, a two-segmented petiole, and (usually) distinct propodeal spines
(Coovert 2005). Approximately 18 morphological characters vary among 23 Aphaenogaster
species (Creighton 1950, Umphrey 1996, Coovert 2005). Ward (1985) replaced total body
length with Weber’s length. Recently, DeMarco & Cognato (2015) identified an additional 15
diagnostic characters, including the cephalic index, shape of the base of the antennal scape,
length of propodeal spines and sculpturing on the head and thorax. The base of the antennal
scape is particularly important, due to the wide range of shapes observed in different species.
24	
  

	
  
Current identification keys are based on the workers. Genitalia have not been described for most
species, except in Boudinot (2013), and original species descriptions have insufficient
information concerning queens and males.
The study of ant diversity has largely been based on morphological characters, although
in the last decade molecular characters provided crucial information for the determination of
generic and species limits (Branstetter 2012, LaPolla et al. 2010, Moreau & Bell 2013). Early
genetic studies revealed variability in chromosome number and enzymatic variation among
different ant species, which suggested potential taxonomic utility of molecular characters
(Whelden & Haskins 1953, Imai 1966, Tomaszewski et al. 1973, Pamilo et al. 1975). Indeed,
chromosomal and allozyme variation exist for Aphaenogaster species and among populations
(Crozier 1977). For example, A. rudis Enzmann, from the coastal plains of the US were n=20 and
nearly fixed for an esterase allele, while montane specimens were either n=18 or n=22 and had
variable allele frequencies (Crozier 1977).
Umphrey (1996) attempted to discriminate a complex group of ten sibling species of the
Aphaenogaster fulva-rudis-texana complex with karyotypes and morphology. Karyotyping of
223 colonies from 63 localities, mostly in Eastern North America, identified 10 genetic forms
including A. rudis, A. picea (Wheeler), A. miamiana Wheeler, A. carolinenesis Wheeler, A.
texana Wheeler, A. fulva Roger and four undescribed taxa. Chaetotaxy and a morphometric
analysis using 12 characters including characters such as head width, scape length, spine length,
and distance between the spines yielded little additional diagnostic information. While there was
some variability in size, shape and color, this variation was confounded by variation within a
colony or species. For example, A. rudis was morphologically similar to other species occurring
in the same habitat. Umphrey (1996) concluded that karyotypes provided the best, but imperfect,
	
  

25	
  

	
  
means for species diagnosis. He acknowledged that DNA would ultimately prove useful as a
definitive method for separating these groups.
A phylogeny based on DNA characters could define the relationships among
Aphaenogaster species and further diagnose North American species. A recent Bayesian
phylogeny testing the placement of the genus demonstrated non-monophyly of Aphaenogaster,
as a clade of Aphaenogaster species grouped with species in two other genera, Messor and
Stenamma (Brady et al. 2006, Branstetter 2012). Ward (2011) suggested that convergent
evolution and retention of ancestral similarities were two major factors contributing to this nonmonophyly. Aphaenogaster monophyly was resolved, in part, with the resurrection of
Novomessor (DeMarco & Cognato 2015). There are no published phylogenies based on DNA or
morphological data that focus on the species relationships within Aphaenogaster despite the
apparent need (Umphrey 1996, Lubertazzi 2012, Ward 2011, Ward et al. 2015), particularly for
the “fulva-rudis-texana” complex, as described by Umphrey (1996). In this study, we use
sequence data from five genes to reconstruct a phylogeny for 44 Aphaenogaster and outgroup
species. The resulting trees support some previously recognized groups, but also reveal
polyphyly among specimens identified as A. rudis.

Materials and Methods
Previous species concepts for Aphaenogaster were mainly morphological and genetic
(Crozier 1977, Umphrey 1996). Our phylogenetic approach necessitated a phylogenetic species
concept, founded in hypothesis testing (Hey 2006). Thus we tested the monophyly of the
currently recognized Aphaenogaster species. Non-monophyly of species suggested the need for
the revision of species boundaries.
	
  

26	
  

	
  
Ant collecting occurred in the eastern and central US forests and grasslands, and the
western forests and deserts. For hypothesis generating purposes, four additional samples were
included in the analysis from Costa Rica, Greece, Japan and Madagascar to begin to understand
the relationships between North American and worldwide Aphaenogaster. Specimens were
collected into 100% ethanol using an aspirator and baits (peanut butter and pecan shortbread
cookies) for analysis. GPS coordinates were recorded for all sites. At least 12 ants per nest were
collected, and 10 nests were sampled for within a 3 km radius to assess intraspecific variation at
a local level. Reproductive forms were collected when possible. Eight representatives from each
nest were pinned. Specimens collected were vouchered in the A.J. Cook Arthropod Research
Collection at Michigan State University (Table 1). Other individuals were stored in 100%
ethanol at -80 °C for future DNA analysis.
A molecular data set was assembled using genetic loci identified in a previous study of
ant phylogeny (Brady et al., 2006), including the nuclear protein coding genes wingless, longwavelength rhodopsin, elongation factor 1α F2 and the mitochondrial protein-coding gene COI.
The gene CAD was also used (Ward et al. 2010). DNA was extracted from 22 of 23 currently
recognized species of Aphaenogaster ants plus outgroups using a silica-based spin column
procedure (Qiamp, Qiagen Inc., Santa Clara, CA), following the manufacturer’s tissue protocol.
Specific regions of mitochondrial and nuclear DNA were amplified via polymerase chain
reaction (PCR). All PCR cocktails consisted of a total volume of 25µl and included 14.25-17.25
µl ddH20, 2.5µl 10X PCR buffer (Qiagen), 1.0µl 25mM MgCl2 (Qiagen), 0.5µl dNTP mix
(Qiagen), 2-5µl DNA template, 0.25µl HotStar Taq DNA polymerase (Qiagen). PCR reactions
were performed as specified by DeMarco and Cognato (2015). After PCR, unincorporated
deoxyribonucleotide triphosphates (dNTPs) and oligonucleotides were removed from PCR

	
  

27	
  

	
  
reactions with Exo-SAP (http://www.usbweb.com/category.asp?cat=pcr&id=78200) and directly
sequenced on an ABI 3700 automated sequencer using a BigDye (Applied Biosystems, Inc.,
Foster City, CA) fluorescent chemistry reaction. Both sense and anti-sense strands were
sequenced for all individuals.
Phylogenetic parsimony analysis was performed using the computer software PAUP*
(Swofford 2003). Bootstrap analysis used resampling, with 1000 replicates. Bremer support was
performed with TreeRoot v.2.0 (Sorenson 1999) with partition Bremer support for all genes. A
phylogeny was inferred with likelihood with RAxML (Stamatakis 2014) via the CIPRES
Gateway (Huelsenbeck & Ronquist 2001, Miller et al. 2010) with 1000 bootstrap replicates. A
phylogeny was also inferred with Bayesian analysis with Mr. Bayes via the CIPRES Gateway
(Huelsenbeck & Ronquist 2001, Miller et al. 2010). We followed guidelines to make credible
Bayesian inferences (Bollback 2002, Huelsenbeck & Ronquist 2001). Data were partitioned by
gene and codon position (Castoe et al. 2004), with models of evolution applied independently to
each partition (Nylander et al. 2004). We used MrModeltest 3.7 (Nylander 2004) for the
selection of partition-specific substitution models for the nucleotide data using the Akaike
Information Criterion in order to decrease the potential of over-parameterization of the models.
The best-fit model for all genes was GTR + I + G.
Results
All analyses recovered similar phylogenies and the Bayesian phylogeny was mostly
resolved. Most species represented by more than one individual were monophyletic and had
relatively high branch support, except A. rudis, A. carolinensis, A. picea, A. huachucana and A.
uinta (Figs. 1, 3 and 4). The parsimony tree differed compared to the likelihood and Bayesian
trees with a polyphyletic A. texana (Figs. 1, 3 and 4). The likelihood and Bayesian trees differed
	
  

28	
  

	
  
by the positions of A. carolinensis, and the A. ashmeadi (Emery) and A. treatae Forel clades
(Figs. 3 and 4). As indicated by the partition Bremer values, COI provided most of the support
followed by EF1α2 (Table 2, Fig. 2). The other genes (CAD, LWR and WG) provided little
support or conflicted with CO1 and EF1α2 as indicated by negative values. An intron was
missing from A. carolinensis and A. miamiana CAD sequences.
There was strong support for the outgroup taxa in the Formicinae with Camponotus and
Formica sister to the remaining taxa. This was also true for most of the Myrmicinae, including
Solenopsis, Stenamma, Myrmica, and Novomessor. Veromessor was sister to the European
Messor species and Aphaenogaster swammerdami. Aphaenogaster swammerdami was the only
species not within the Aphaenogaster clade. Aphaenogaster araneoides, from Costa Rica and an
undescribed species (JTL-001) from Mexico were sister to the other Aphaenogaster species.
Aphaenogaster japonica Forel, from Japan, was within the NA Aphaenogaster clade, as was A.
balcanica (Emery), from Greece.
Most of species collected west of the Rocky Mountains were grouped together near the
outgroup species. Aphaenogaster uinta Wheeler, like A. huachucana Creighton, was
polyphyletic. Aphaenogaster occidentalis (Emery) from Washington, Utah and Colorado formed
a monophyletic clade. There was strong support for the clade including A. tennesseensis (Mayr)
and A. mariae Forel. The clade containing A. fulva and A. umphreyi, is completely separate from
the A. rudis species complex.
Aphaenogaster floridana Smith and A. flemingi Smith were sister to the A. picea and A.
rudis clades. Aphaenogaster picea individuals were found two clades, one containing mostly
northern A. picea samples and the other individuals were in the A. rudis clade. Aphaenogaster

	
  

29	
  

	
  
rudis was not monophyletic, and appeared in 4 clades (Figs. 3,4). Taxa in the largest A. rudis
clade included A. rudis, A. carolinenesis and A. picea, in addition to A. miamiana, A. lamellidens
Mayr and A. texana, and the clade was sister to A. ashmeadi and A. treatae.
Discussion
We tested the monophyly of Aphaneogaster in the context of a multi-gene phylogenetic
analysis. In the resulting phylogenies, species boundaries associated with well-supported
monophyletic clades of individuals for 10 of 16 NA Aphaenogaster species. Many of these
monophyletic species contained morphological diagnostic characters discovered by previous
taxonomic studies. For example, A. tennesseensis lacks setae on the mesosoma and gaster and is
a nest parasite of A. rudis and A. fulva (Creighton 1950, Ellison et al. 2012). Aphaenogaster
mariae is an arboreal species with a starburst pattern of striae on the first gastral tergite (Ellison
et al. 2012). Aphaenogaster floridana is the only southeastern species lacking propodeal spines
and nests in sandy soil in pine forests in North Carolina and Florida (Creighton 1950).
Aphaenogaster flemingi is diagnosed by a shiny exoskeleton and thin propodeal spines
(Creighton 1950). Aphaenogaster fulva and A. umphreyi have upward pointing spines, and can
be separated from each other by the reduced eyes in A. umphreyi (Deyrup & Davis 1998). There
is no pattern to the type or the magnitude of difference among morphological characters that
diagnosis species; they can be obvious like the lack of spines or subtle like the pattern of striae.
Polyphyly of the remaining six species is an issue of concern because given the criteria of
monophyly, our phylogeny suggests the recognition of fewer species. The large clade of A. rudis
also includes A. ashmeadi, A. carolinensis, A. lamellidens, A. miamiana, A. texana and A.
treatae, and the placement of A. rudis individuals are scattered in six separate clades among

	
  

30	
  

	
  
these other species (Fig. 1). Other instances of paraphyly occur with A. fulva-A. umphreyi, A.
texana - A. huachucana, A. uinta and A. picea. It is tempting to synonymize these species in
order to preserve monophyly. However, many of the included species are well-supported
subclades with morphological and behavioral diagnostic characters. For example, the presence
and size of lobe at the base of the scape diagnoses A. ashmeadi and A. treatae, which are wellsupported monophyletic species (Creighton 1950). In other cases the diagnostic character is
minor, as with the smaller eye, which characterizes A. umphreyi from A. fulva (Deyrup and
Davis 1998). In addition, there are other potential molecular differences that could diagnose
species. For example, A. carolinensis and A. miamiana lack a 300 bp CAD intron as compared
to most other Aphaenogaster species. The remaining clades of individuals (e.g. A. rudis) may
represent unrecognized species that await the discovery of diagnostic characters. Morphology of
reproductive adults and nest architecture could provide these characters (Tschinkel 2011,
Boudinot 2013).
Moreover, there are a number of alternative reasons for the apparent polyphyly in the A.
rudis clade of NA Aphaenogaster, which would argue against abandoning existing nomenclature
without additional evidence. First, there may be an insufficient amount of phylogenetically
informative data for complete resolution. Although we sampled five genes known to resolve ant
phylogenies, only 1102 of 2967 characters were phylogenetically informative and most of the
phylogenetic support derived from COI and EF1αF2 (Table 2). The other genes gave little or
negative support, which is a pattern observed in other insect phylogenies (e.g., Damgaard &
Cognato 2003, Danforth et al. 2004). Doubling the number of sampled genes or increasing the
number of nucleotides to the 100,000’s via phylogenomic methods may help to resolve this
issue, as they have provided resolution for other taxa (Peterson, et al. 2012, Ward & Sumnicht

	
  

31	
  

	
  
2012).
It is also possible that recent species radiation could explain the low resolution due to a
lack of lineage sorting of gene lineages, as demonstrated with gallwasps (Rokas et al. 2003) and
Formica ants (Goropashnaya et al. 2004). Although the estimated age of this genus is 44 million
years (Moreau et al. 2006), the relatively short branches and minimal COI sequence variation
(mean 2.85%) observed for the A. rudis clade (not including the larger A. picea clade) suggest
more recent origins of the species. A possible Pleistocene origin of these species during the
expansion and contraction of glaciers could have contributed to the isolation of populations by
altitude and latitude in northeastern US, as has been shown for many other taxa (Cognato et al.
2003, Maroja et al. 2007, Lecocq et al. 2013). Potentially, pre- and post-mating isolating
mechanisms such as chromosomal	
  rearrangements may have developed in glacial refugia and
contributed to Aphaenogaster speciation. Potentially karyotype number may diagnose species
boundaries, because much chromosomal variation exists within subfamilies, genera and even
Aphaenogaster (Umphrey 1996, Menezes et al. 2013, Cardoso et al. 2014), and could be
important in the generating reproductive isolation (Lorite & Palomeque 2010). This is consistent
with the observation that distinct karyotypes associate with geographic distributions (Umphrey,
1996). For example, populations of western A. picea have n = 17, while eastern populations have
n = 18. Our specimens of A. picea occur in two clades (Fig. 2, 3, 4) but unfortunately, other than
one sample, A. rudis (# 43), we do not have associated karyotype numbers for our specimens. It
is unknown whether members of the A. rudis clade with different karyotypes produce viable
offspring or if other isolating mechanisms exist. Identification of these mechanisms and the
possibility of a speciation gene, such as those that cause hybrid male sterility in Drosophila,
could help resolve Aphaenogaster species relationships (Gomes & Civetta 2014). Obviously,
	
  

32	
  

	
  
more study is needed to resolve the non-monophyly of A. rudis and other species, provide
diagnostic characters, and to determine the existence of pre- or post- mating barriers among the
species. Thus, a revision of Aphaenogaster is premature.

	
  

33	
  

CHAPTER 4
APHAENOGASTER (HYMENOPTERA: FORMICIDAE) OF NORTH AMERICA: A KEY TO
SPECIES USING MORPHOLOGY AND DNA

	
  

34	
  

Abstract

Aphaenogaster Mayr 1853, contains 227 species worldwide (Bolton 2006) with 23 valid
North American species, several species of which are hard to separate based on morphology
alone (Umphrey 1996). The difficulty in identifying some of these species is due to limited
diagnostic characters and to the lack of a comprehensive illustrated key. A recent analysis
returned three species from Aphaenogaster to Novomessor, thus making Aphaenogaster in North
America monophyletic (DeMarco and Cognato 2015). While many species have easily
identifiable morphological characters, some east coast species within the A. rudis clade in North
America are difficult to differentiate. Two of these species, A. carolinensis and A. miamiana, can
be diagnosed using DNA. The gene CAD was missing an intron in those taxa. Four additional
taxa, all identified morphologically as A. rudis, were found to be polyphyletic (DeMarco and
Cognato, in prep, or see Chapter 2).

Introduction

Aphaenogaster Mayr 1853, contains 227 species worldwide (Bolton 2006) with
23 valid North American species, several species of which are hard to separate based on
morphology alone (Umphrey 1996). The difficulty in identifying some of these species is due to
limited diagnostic characters and to the lack of a comprehensive illustrated key. A recent
analysis returned three species from Aphaenogaster to Novomessor, thus making Aphaenogaster
in North America monophyletic (DeMarco and Cognato 2015). While many species have easily
identifiable morphological characters, some east coast species within the A. rudis clade in North
America are difficult to differentiate. Two of these species, A. carolinensis and A. miamiana, can
	
  

35	
  

be diagnosed using DNA. The gene CAD was missing an intron in those taxa. Four additional
taxa, all identified morphologically as A. rudis, were found to be polyphyletic (DeMarco and
Cognato, in prep, or see Chapter 2).
Aphaenogaster has been a popular genus for many studies including biology and natural
history (Lubertazzi 2012), tool use (Fellers and Fellers 1976), communication (Menzel and
Marquess 2008), interactions with other ant taxa (Bewick et al. 2014) and temperature tolerance
(Warren and Chick 2013). Aphaenogaster also have a variety of interesting behaviors. They laid
trail pheromones (Attygalle et al. 1998) using their poison glands to recruit nest mates to food
items. They fed on small invertebrates including termites (Buczkowski and Bennett 2007),
eliaosome bearing seeds (Heithaus et al. 2005 and Clark and King 2012) and even mushrooms
(Carroll et al. 1981). Haskins (1960) observed longevity in A. picea with queens able to survive
8-13 years. Menzel and Marquess (2008) observed substrate vibration generating behavior in A.
carolinensis. A worker would strike a substrate with its mandible and drag it across the surface.
This behavior was in response to the presence of non-nest mate conspecifics and ants from other
species.
Recently, Aphaenogaster species have become the focus of climate change studies.
Bewick et al. (2014), observed interactions among A. rudis and two other ant species, Prenolepis
imparis and Nylanderia faisonensis. They tested for the importance of different species traits
(food discovery rate, food clearance rate, body mass, dominance hierarchy and thermal niche),
how climate change affected community composition and how interspecific competition
mediated shifts in community composition in response to climate change. The overall conclusion
was that climate change would have a negative effect on P. impairis (known as the winter ant),
but also surprisingly on N. faisonensis, which is more active during the summer months.
	
  

36	
  

Aphaenogaster rudis faired the best and Bewick et al. (2014) predicted that the three community
species would decrease to two species including A. rudis and P. impairis. Nylanderia faisonensis
would be expatriated from its current range.
Warren and Chick (2013) examined data over a 38-year period of upward movement in
elevation for A. rudis and A. picea along the southern end of the Appalachian Mountain chain in
Georgia. They found 100% of Aphaenogaster ants at 900 m elevation in 1974 were A. picea. In
2012, 25% of the Aphaenogaster ants present were A. rudis and only 75% were A. picea. They
also tested thermal tolerance of individuals by increasing and decreasing temperatures. Their
results indicate possible changes in insect species as climates increase in temperature.
The studies described above, and others necessitate the ability to identify Aphaenogaster
species in research. It is thus timely to create a comprehensive identification key for NA
Aphaenogaster species, given their increased use as indicator species in climate change studies.

Materials and Methods

Collecting occurred in areas across North America, including the eastern and central US,
and the western deserts. Specimens were collected using an aspirator and baits (peanut butter and
shortbread cookies with pecans) to collect specimens. GPS coordinates were recorded for all
sites. Additional specimens were examined from the following museums: The Museum of
Comparative Zoology at Harvard University, The Smithsonian, The Field Museum in Chicago,
Mississippi State University, University of Michigan and the California Academy of Sciences.
Specimens were vouchered in the A.J. Cook Arthropod Research Collection at Michigan State
University as pinned and frozen samples.
	
  

37	
  

Most taxa can be identified using characters included in the key. Additional
morphological characters can be found in DeMarco and Cognato (2015). DNA data is required
to separate Aphaenogaster carolinensis and A. miamiana from A. rudis. Both of these taxa are
missing an intron in the gene CAD (carbomoylphosphate synthase). See an example in Genbank
(sample number KJ9205520). There are 545 base pairs in Aphaenogaster carolinensis and A.
miamiana. Aphaenogaster rudis contains 762 base pairs. Methods for sequencing this gene are
in DeMarco and Cognato (in prep).
Specimens were also photographed using a Canon EOS 5D Mark II camera with a Canon
Macro Pro lens (MP-E 65mm, 1-2.8, 1-5x). The images were taken using EOS Utility and
Zerene stacker in combination with Stackshot. The stacks were montaged using Helicon Focus.

Glossary

Many terms used in ant identification are unfamiliar to other entomologists and to nonentomologists needing to identify ant taxa. Therefore, a brief glossary of terms is included
(Bolton 1994, Fisher and Cover 2007).
Clypeus – the anterior sclerite of the head. The edge may be smooth, emarginate or notched.
Eye Size: variable (Figure 3.4)
Frontal carina – A pair of longitudinal ridges on the head, behind the clypeus.
Gaster – Abdominal segments four through seven, when a petiole and postpetiole are present.
Mesosoma – The second tagma of an ant’s body, including the thorax and propodeum.
Metasoma – The third tagma of an ant’s body, including the petiole, postpetiole (when present)
and gaster.

	
  

38	
  

Petiole – The second abdominal segment, reduced and isolated into a separate segment.
Piceous – Meaning pitch, or darkly colored.
Postpetiole - The third abdominal segment (in some ant taxa), reduced and isolated into a
separate segment.
Pronotum – The first tergite of the thorax.
Propodeal spines – spines present, extending from the propodeum. (Figure 3.3)
Propodeum – Morphologically, the first tergite of the abdomen, but forming the back of the
mesosoma.
Punctate – With numerous fine pits.
Rugae – wrinkled ridges, often forming parallel lines.
Rugose – containing rugae.
Scape – elongate basal section of antenna.
Spine shape – variable (Figure 3.2)
Spiracle – An orifice of the tracheal system. The propodeal spiracle is used to compare to the
propodeal spines.
Striae – fine lines.
Tergite – the upper sclerite of a segment.

Identification Key to Aphaenogaster species
Some characters are from Creighton (1950) and Coovert (2005), including striae, frontal carina
with rearward facing tooth, antennal scape shape, and color of last four antennal segments.
1

	
  

Striae at base of first gastral tergite,

39	
  

arboreal species (Figure 3.1)

Aphaenogaster mariae

1’

No striae on first gastral tergite, not arboreal

2

2 (1)

Propodeal spines absent

3

2’

Propodeal spines present (even if small)

4

3(2)

Gaster same color as head and mesosoma,
range = AL, FL, GA, MS, NC, SC

3’

Aphaenogaster floridana

Gaster darker than head and mesosoma
range = AZ, CA, CO, NM, MEX

4 (2)

Aphaenogaster boulderensis

No setae on mesosoma or metasoma,
spines strongly curved back,
wide range from NE to northern FL

4’

Aphaenogaster tennesseensis

Setae on mesosoma and metasoma, spines straight,
curved in or not strongly curved back

5

5(4)

Lobe at base of scape

6

5’

No lobe at base of scape

7

6(5)

Lobe one-fifth the length of the scape (Figure 3.3)
range = AL, FL, GA, LA, MS, NC, SC, TN

6’

Lobe one-fourth the length of scape (Fig. 3)
range = FL, IL, MI, MO, MS, NC, TN, VA

7(5)

Aphaenogaster megommata

Color varies from light brown to piceous,
eyes not large

	
  

Aphaenogaster treatae

Color light yellow in color, large eyes
range = AZ, CA, NV

7’

Aphaenogaster ashmeadi

8

40	
  

8(7)

Frontal carina with rearward-facing tooth
range = southeastern states

Aphaenogaster lamellidens

8’

Frontal carina without rearward-facing tooth

9

9(8)

Spines pointed upward from propodeum,
anterior edge of pronotum above mesonotum

9’

10

Spines angled back or reduced, anterior edge
of pronotum equal to or below mesonotum

11

10(9) Eyes reduced, reduced hind tibial spurs
RARE species (FL, AL)
10’

Aphaenogaster umphreyi

Eyes normal size, normal hind tibial spurs
range = AL, IL, MN, MS, NC, NJ, TN, VA, WS

Aphaenogaster fulva

11(9) Spines thin (Fig. 3) head and mesonotum shiny,
light brown in color, range = FL, LA, NC, MS
11’

Aphaenogaster flemingi

Spines variable, head and mesonotum not shiny,
color variable

12

12(11) Spine length less than or equal to diameter of

12’

propodeal spiracle

13

Spine length greater than diameter of spiracle.

15

13(12) Body unicolorous brown to black, with lighter legs

13’

range = southern CA, MEX (Baja Sur)

Aphaenogaster patruelis

Head and mesosoma light brown/tan, gaster dark

14

14(13) Head rounded (wider at occiput), clypeus notched
range = MEX (Baja Sur)

	
  

Aphaenogaster mutica

41	
  

14’

Head rectangular, clypeus emarginate
range = CA, ID, NV, UT

Aphaenogaster uinta

15(12) Spine shape triangular,
(See Fig. 2, like A. huachucana)
barely longer than width of propodeal spiracle
Scape with small triangular extension at base
range = AZ, NM
15’

Aphaenogaster huachucana

Spine shape not triangular,
longer than width of spiracle

16

16(14) Head narrowed posteriorly into neck, with collar

16’

RARE, only known from MEX

Aphaenogaster mexicana

Head not narrowed posteriorly

17

17(16) Last four antennal segments lighter in color
(except some forms in Canada) range = Northeast

17’

plus GA, NC,TN, and WV at higher altitudes

Aphaenogaster picea clade

Antenna unicolorous

18

18(17) Mesosoma with fine rugae
range = BC(Canada), CA, CO, OR, UT, WA, WY Aphaenogaster occidentalis
18’

Mesosoma punctate or coarsely rugose

19

19(18) Dorsum of head with coarse rugae

19’

range = AR, AZ, MO, NM, OK, TX

Aphaenogaster texana

Dorsum of head with fine rugae

20

20(19) Posterior border of head moderately pointed

42	
  

20’

RARE species, found only in New Mexico

Aphaenogaster punctaticeps

Posterior border of head rounded to flattened

21

21(20) Propodeal spines curved slightly inward
(dorsal view), coarse rugae on mesosoma,
Range = Al, FL, NC (CAD intron absent)
21’

Aphaenogaster miamiana

Propodeal spines straight, fine rugae
or punctate on mesosoma

22

22(21) Light to medium brown
range = NC to MS (CAD intron absent)
22’

Aphaenogaster carolinensis

Medium to dark brown
Widely distributed throughout East coast,
from Georgia to Massachusetts
and west to Minnesota (CAD intron present)

Aphaenogaster rudis clades

Overview of species

Aphaenogaster ashmeadi (Emery) (Figure 3.5.)
Taxonomic history:
Stenamma (Aphaenogaster) treatae var. ashmeadi Emery, C. 1895d: p. 302 (worker) U.S.A.
Combination in Aphaenogaster: (Wheeler 1913).
Combination in Aphaenogaster (Attomyrma), (Emery 1921).
Raised to species and senior synonym of A. hardeni: (Creighton 1950).
Type locality: Florida (holotype at USNM).

43	
  

Aphaenogaster ashmeadi is similar to A. treatae, but has a smaller lobe at the base of the scape.
The lobe is one-fifth the length of the scape. Specimens examined were collected from AL, FL,
GA, LA, MS, NC, SC and TN. They range in color from reddish brown to dark brown.

Aphaenogaster boulderensis Smith (Figure 3.6.)
Taxonomic history:
Aphaenogaster (Attomyrma) boulderensis Smith, M. R. 1941: p. 120 (worker) U.S.A.
Type locality: Arizona: Horseshoe Island in Mead Lake, beneath a lava rock. (holotype at
USNM).
Aphaenogaster boulderensis is one of the NA Aphaenogaster species without propodeal spines.
The head and mesosoma are light brown, and the gaster is dark brown. The antennal scapes pass
the occipital margin by one-third the length of the scape. Specimens examined were collected
from AZ, CA, NM, TX, UT, and Mexico.

Aphaenogaster carolinensis Wheeler (Figure 3.7.)
Taxonomic history:
Aphaenogaster texana var. carolinensis Wheeler, W. M. 1915: p. 414 (worker, queen)U.S.A.
Combination in Aphaenogaster (Attomyrma), (Emery 1921).
Subspecies of Aphaenogaster texana: (Creighton 1950).
Raised to species: (Umphrey 1996).
Type locality: North Carolina: Tyron, in open woods under stones (Holotype at Harvard MCZ).
Aphaenogaster carolinensis was described as similar to A. texana, but with shorter spines and
directed further backwards. This research finds overlapping morphological characters with both

44	
  

A. texana and A. rudis. DNA analysis is necessary to confirm identification by a missing intron
in the gene CAD (DeMarco and Cognato, in prep). Specimens examined were collected from NC
and MS.

Aphaenogaster flemingi Smith (Figure 3.8.)
Taxonomic history:
Aphaenogaster texana ssp. flemingi Smith, M. R. 1928: p. 275 (worker) U.S.A.
Raised to species: (Creighton 1950).
Senior synonym of Aphaenogaster macrospina (Smith 1958).
Type locality: Mississippi: at A and M College, in a stump (Holotype at USNM).
Aphaenogaster flemingi has slender, upward pointing propodeal spines, feeble sculpturing on the
mesosoma and an overall shiny appearance. Specimens examined were collected from FL, LA,
MS and NC.

Aphaenogaster floridana Smith (Figure 3.9.)
Taxonomic history:
Aphaenogaster (Attomyrma) floridana Smith, M. R. 1941: p. 118 (worker) U.S.A.
Type locality: Florida.
Aphaenogaster floridana is one of the NA Aphaenogaster species without propodeal spines. The
gaster is not significantly darker than the head and mesosoma (compared to A. boulderensis).
They are found in sandy pine scrub and mixed hardwood forest. Specimens examined were
collected from AL ,FL, GA and NC.

45	
  

Aphaenogaster fulva Roger (Figure 3.10.)
Taxonomic history:
Aphaenogaster fulva Roger, J. 1863: p. 190 (worker) U.S.A.
Combination in Stenamma (Aphaenogaster): (Emery 1895).
Combination in Aphaenogaster: (Wheeler 1913).
Combination in Aphaenogaster (Attomyrma), (Emery 1921).
Senior synonym of Aphaenogaster rubida (Brown 1949).
Current subspecies: nominal plus Aphaenogaster fulva azteca.
Type locality: “North America” (Holotype: unknown).
Aphaenogaster fulva is diagnosed by the spines pointing upward from propodeum and the
anterior edge of pronotum above mesonotum. They are similar to A. umphreyi, but have larger
eyes and larger hind tibial spurs. The last four antennal segments are lighter in color. Specimens
examined were collected from AL, IL, MN, MS, NC, NJ, TN, VA and WS.

Aphaenogaster huachucana Creighton (Figure 3.11.)
Taxonomic history:
Aphaenogaster (Attomyrma) huachucana Creighton, W. S. 1934: p. 189 (worker) U.S.A.
Current subspecies nominal plus crinimera.
Type locality: Arizona (Holotype missing from USNM, Syntype at Harvard MCZ).
Aphaenogaster huachucana is similar in appearance to A. texana, but is larger and found nesting
in rocky ledges as opposed to A. texana that can be found under logs and rocks. The antennal
scapes pass the occipital margin by one-third the length of the scape.Specimens examined were
collected from AZ and NM.

46	
  

Aphaenogaster lamellidens Mayr (Figure 3.12.)
Taxonomic history:
Aphaenogaster lamellidens Mayr, G. 1886: p. 444 (worker, queen, male) U.S.A.
Combination in Stenamma (Aphaenogaster) (Emery 1895).
Combination in Aphaenogaster (Wheeler 1913).
Combination in Aphaenogaster (Attomyrma) (Emery 1921).
Senior synonym of Aphaenogaster nigripes (Creighton 1950).
Type locality: Virginia (Holotype missing, syntype at Harvard MCZ).
Aphaenogaster lamellidens has dark legs compared to the rest of the body and the frontal carina
has a rearward-facing tooth. Specimens examined were collected from AL, AR, FL, LA, MO,
MS, NC, SC, TN and VA.

Aphaenogaster mariae Forel (Figure 3.13.)
Taxonomic history:
Aphaenogaster mariae Forel, A. 1886: p. 4 (worker) U.S.A.
Combination in Stenamma (Aphaenogaster): (Emery 1895).
Combination in Aphaenogaster (Attomyrma), (Emery 1921).
Type locality: Florida (Holotype at Naturhistorisches Museum in Vienna, Austria)
Aphaenogaster mariae has the diagnostic character of a starburst of striae (Ellison, et al. 2012) at
the base of the gaster. They also have very rugose sculpturing on the mesosoma. This species is
arboreal, and has only been collected on trunks of live trees at bait. One nest was observed in a
treehole. Specimens examined were collected from FL, MD, MS, NJ and TN.

47	
  

Aphaenogaster megommata Smith (Figure 3.14.)
Taxonomic history:
Aphaenogaster (Attomyrma) megommatus Smith, M. R. 1963: p. 244 (worker) U.S.A.
Type locality: Nevada: One mi N Camp Foster, Pyramid Lake, Washoe Co. (Holotype at
USNM).
Aphaenogaster megommata is a desert species that forages only at night. They have huge eyes
and miniscule propodeal spines. They are pale yellow in color. Specimens examined were
collected from AZ, CA and NV.

Aphaenogaster mexicana (Pergande) (Figure 3.15.)
Taxonomic history:
Ischnomyrmex mexicanum Pergande, T. 1896: p. 893 (worker) MEXICO.
Combination in Aphaenogaster (Ischnomyrmex): (Forel 1899).
Combination in Aphaenogaster (Deromyrma): (Emery 1915).
Type locality: Mexico: Tepic (Lectotype at CAS).
Aphaenogaster mexicana is rarely collected, but can be distinguished from other species in North
America due to the head being narrowed posteriorly into a neck with a collar. The antennal
scapes pass the occipital margin by one-half the length of the scape.

Aphaenogaster miamiana Wheeler (Figure 3.16.)
Taxonomic history:
Aphaenogaster (Attomyrma) texana miamiana Wheeler, W. M. 1932: p. 5 (worker, queen, male)
U.S.A.

48	
  

Raised to species: (Creighton 1950).
Type locality: Florida (Holotype at AMNH).
Aphaenogaster miamiana is within the A. rudis clade but can be distinguished by the more
rugose sculpturing on the head and mesosoma, and by a missing intron in the gene CAD
(DeMarco and Cognato, in prep.). Specimens examined were collected from FL and NC. This
species was previously only known from Florida.

Aphaenogaster mutica Pergande (Figure 3.17.)
Taxonomic history:
Aphaenogaster mutica Pergande, T. 1896: p. 891 (worker) MEXICO.
Combination in Aphaenogaster (Attomyrma), (Emery 1921).
Type locality: Mexico: Baja California Sur, Jose del Cabo (Holotype at CAS).
Aphaenogaster mutica has a head and mesosoma that are light brown, with a dark gaster.
The head is rounded (wider at occiput), with a notched clypeus. Specimens examined were
collected from Mexico.

Aphaenogaster occidentalis (Emery) (Figure 3.18.)
Taxonomic history:
Aphaenogaster occidentalis Emery, C. 1895d: p. 301 (worker) U.S.A.
Combination in Aphaenogaster (Attomyrma), (Emery 1921).
Senior synonym of A. borealis (Creighton, 1950).
Raised to species (Hunt and Snelling 1975).
Type locality: Washington: Pullman City (Holotype at Harvard MCZ).

	
  

49	
  

Aphaenogaster occidentalis has relatively short spines and scapes. This is the only species that
is a pest in Washington homes. Specimens examined were collected from WA, UT and CO.
The range extends further East than previously recorded.

Aphaenogaster patruelis Forel (Figure 3.19.)
Taxonomic history:
Aphaenogaster patruelis Forel, A. 1886: xli (worker) U.S.A.
Combination in Stenamma (Aphaenogaster): (Emery 1895).
Combination in Aphaenogaster (Attomyrma), (Emery 1921).
Subspecies of A. subterreanea (Emery 1895), Revived status as species (Wheeler 1904).
Senior synonym of A. willowsi (Creighton, 1950), of A. bakeri (Smith 1979).
Current subspecies: nominal plus carbonaria.
Type locality: Mexico: Guadelupe Island (Holotype at CAS).
Aphaenogaster patruelis ranges in color from dark brown to black, with lighter legs. The spines
are minute, less than the width of the propodeal spiracle. Specimens examined were collected
from CA and Mexico.

Aphaenogaster picea (Wheeler) (Figure 3.20.)
Taxonomic history:
Stenamma (Aphaenogaster) fulvum piceum Wheeler, W. M. 1908: p. 621 (worker, queen,
male)
(Emery, C. 1895 first available use of name)
Combination in Aphaenogaster (Attomyrma), (Emery 1921).

50	
  

Subspecies of A. fulva (Buren 1944), of A. rudis (Creighton, 1950), but Creighton incorrect as A.
picea is senior name.
Raised to species (Bolton 1995).
Type Locality: Connecticut ( Holotype unknown)
Aphaenogaster picea is diagnosed by the last four antennal segments being lighter in color than
the rest of the antenna, by its piceous color and northern ranges in North America. Species from
NC, TN, and WV occur at higher elevations. Other samples are from MI, MA, MN, OH, PA and
NY.

Aphaenogaster punctaticeps MacKay (Figure 3.21.)
Taxonomic history:
Aphaenogaster punctaticeps MacKay, W. P. 1989: p. 47 (worker) U.S.A.
Type Locality: New Mexico: Jornada Experimental Range, Dona Ana Co. (Holotype at USNM).
Aphaenogaster punctaticeps is similar to A. texana, but with the posterior border of head
moderately pointed as opposed to rounded. This is a rare species, found only in New Mexico.

Aphaenogaster rudis Enzmann (Figure 3.22.)
Taxonomic history:
Aphaenogaster fulva rudis Enzmann, J. 1947: p. 150 (worker, queen) U.S.A.
(Emery, 1895 first available use of name).
Raised to species (Creighton, 1950), but Creighton incorrect as A. picea is senior name.
Raised to species (Umphrey 1996).
Type locality: Virginia (Holotype unknown).

51	
  

This is the most commonly found Aphaenogaster species on the east coast. It is polyphyletic and
ranges in color from light to dark brown. The last four antennal segments are not lighter in color.
It cannot be distinguished from A. carolinensis without the gene CAD, which has an intron that
A. carolinensis is missing. (DeMarco and Cognato, in prep.) Specimens examined are from AR,
FL, GA, MI, MN, NC, NJ, OH, PA and VA.

Aphaenogaster tennesseensis (Mayr) (Figure 3.23.)
Taxonomic history:
Atta tennesseensis Mayr, G. 1862: p. 743 (worker) U.S.A.
Combination in Aphaenogaster (Roger 1863).
Combination in Stenamma (Aphaenogaster): (Emery 1895).
Combination in Aphaenogaster (Attomyrma), (Emery 1921).
Senior synonym of A. subrubra (Mayr 1886), of A. laevis (Mayr 1886) and of A. ecalcaritum
(Creighton 1950).
Type locality: Tennessee (Holotype at Naturhistorisches Museum, Vienna, Austria). This ant is
easily diagnosed by its lack of hair on the mesosoma and metasoma, and by the propodeal spines
that curve back towards the gaster. Specimens examined are from IA, MI, MN, OH, NE andVA.

Aphaenogaster texana Wheeler (Figure 3.24.)
Taxonomic history:
Aphaenogaster texana Wheeler, W.M. 1915: p. 306 (worker, queen, male) U.S.A.
(Emery, C. 1895 first available use of name).
Senior synonym of A. furvescens, A. silvestrii.

	
  

52	
  

Type locality: Texas (type unknown).
Aphaenogaster texana was described as similar to A. carolinensis, but with longer spines and
directed further upwards. This research finds overlapping morphological characters with both A.
texana and A. rudis. DNA data indicates that A. texana occurs west of the Mississippi River.
Specimens examined are from AR, AZ, MO and TX.

Aphaenogaster treatae Forel (Figure 3.25.)
Taxonomic history:
Aphaenogaster treatae Forel, A. 1886b: p. xl (worker, queen, male) U.S.A.
Combination in Stenamma (Aphaenogaster): (Emery 1895).
Combination in Aphaenogaster (Attomyrma), (Emery 1921).
Senior synonym of A. wheeleri (Creighton 1950).
Type locality: New Jersey: Vineland (Lectotype at CAS).
Aphaenogaster treatae is similar to A. ashmeadi, but has a larger lobe at the base of the scape.
The lobe is one-fourth the length of the scape. Specimens examined were collected from FL, IL,
MI, MO, MS, NC, TN and VA. They range in color from light to dark brown.

Aphaenogaster uinta Wheeler (Figure 3.26.)
Taxonomic history:
Aphaenogaster uinta Wheeler, W. M. 1917: p. 517 (worker, queen, male) U.S.A.
Combination in Aphaenogaster (Attomyrma), (Emery 1921).
Type locality: Utah: East Mill Creek, Salt Lake County (Holotype at Harvard MCZ).

53	
  

Aphaenogaster uinta is one of several Aphaenogaster species with a lighter head and mesosoma,
and darker gaster. They have large eyes, very short propodeal spines and scapes that extend just
beyond the occiput of the head. They live between thin layer of limestone in arid areas.
Specimens examined were collected from AZ, CA and NV.

Aphaenogaster umphreyi Deyrup & Davis (Figure 3.27.)
Taxonomic history:
Aphaenogaster umphreyi Deyrup, M.; Davis, L. 1998: p. 88 (worker) U.S.A.
Type locality: Florida: Putnam County, Sandhill habitat (Holotype at Harvard MCZ).
Aphaenogaster umphreyi is diagnosed by the spines pointing upward from propodeum and the
anterior edge of pronotum above mesonotum. They are similar to A. fulva, but have smaller eyes
and smaller hind tibial spurs. The last four antennal segments are not lighter in color.
Specimens examined were collected from FL.

54	
  

APPENDICES	
  

55	
  

APPENDIX	
  A	
  
	
  
	
  
Tables	
  and	
  Figures	
  for	
  Chapter	
  2	
  

	
  

56	
  

Table 1.1. Specimens included in current analysis with associated localities and Genbank numbers.
DNA
Taxon name
Collection locations
Genbank
Genbank Genbank
extraction
codes
CO1 #
CAD #
EF2 #
A. araneoides
Aphara01
CR: La Selva
KJ920514 KJ920549 KJ920579
A. ashmeadi 7
Aphash07
USA: North Carolina
KJ920515 KJ920550 KJ920581
A. ashmeadi 12
Aphash12
USA: North Carolina
KJ920516 KJ920551 KJ920580
A. balcanica
Aphbal01
Greece: Kefalonia
KJ920517 N/A
KJ920582
A. boulderensis
Aphbou01
USA: California
KJ920518 N/A
KJ920583
A. carolinenesis
Aphcar01
USA: Mississippi
KJ920519 KJ920552 KJ920584
A. flemingi
Aphfle01
USA: Florida
KJ920522 KJ920555 KJ920587
A. floridana
Aphflo01
USA: Florida
KJ920523 KJ920556 KJ920588
A. fulva 5
Aphful05
USA: North Carolina
KJ920524 KJ920557 KJ920590
A. fulva 7
Aphful07
USA: North Carolina
KJ920525 KJ920558 KJ920589
A. huachucana
Aphhua01
USA: Arizona
KJ920526 N/A
N/A
A. japonica
Aphjap01
Japan: Chugoku
KJ920527 KJ920559 KJ920591
A. lamellidens
Aphlam01
USA: Virginia
KJ920528 KJ920560 KJ920592
A. mariae
Aphmar01
USA: Virginia
KJ920529 KJ920561 KJ920593
A. megommata
Aphmeg01
USA: Nevada
KJ920530 KJ920562 KJ920594
A. miamiana
Aphmia01
USA: Florida
KJ920531 KJ920563 KJ920595
A. mutica
Aphmut01
Mexico: Baja CA Sur
KJ920532 N/A
KJ920596
A. occidentalis
Aphocc01
USA: Washington
KJ920533 KJ920564 KJ920597
A. patruelis
Aphpat01
USA: California
KJ920534 N/A
KJ920598
A. picea 1
Aphpic01
USA: Michigan
KJ920535 KJ920565 KJ920599
A. picea 6
Aphpic06
USA: Minnesota
KJ920536 KJ920566 KJ920600
A. picea 39
Aphpic39
USA: Massachusetts
KJ920537 KJ920567 KJ920601
A. rudis 2
Aphrud02
USA: Michigan
KJ920538 KJ920568 KJ920602
A. rudis 4
Aphrud04
USA: Ohio
KJ920539 KJ920569 KJ920603
A. rudis 8
Aphrud08
USA: New Jersey
KJ920540 KJ920570 KJ920604

57	
  

Genbank
LWR #
KJ920615
KJ920616
KJ920617
KJ920618
N/A
KJ920619
KJ920622
KJ920623
KJ920624
KJ920625
KJ920626
KJ920627
KJ920628
KJ920629
KJ920630
KJ920631
KJ920632
KJ920633
KJ920634
KJ920635
KJ920636
N/A
KJ920637
KJ920638
KJ920639

Genbank
WG #
KJ920650
N/A
KJ920651
KJ920652
KJ920653
KJ920654
KJ920657
KJ920658
KJ920660
KJ920659
KJ920661
KJ920662
KJ920663
KJ920664
KJ920665
KJ920666
KJ920667
KJ920668
KJ920669
KJ920670
KJ920671
KJ920672
KJ920673
KJ920674
KJ920675

Table 1.1. (cont’d).
DNA
Taxon name
extraction
codes
A. swammerdami
A. tennesseensis
A. texana
A. treatae
A. uinta
A. umphreyi
C.pennsylvanicus
F. glacialis
Me. bouvieri
Me. denticornis
My. latifrons
N. albisetosa
N. cockerelli
N. ensifera
So. aurea
St. diecki
V. andrei
V. juliana

A = Aphaenogaster
C = Camponotus
F = Formica

Aphswa01
Aphten01
Aphtex01
Aphtre01
Aphuin01
Aphump01
Campen01
Forgla01
Mesbou01
Mesden01
Myrlat01
Novalb01
Novcoc01
Novens01
Solaur01
Stedie01
Verand01
Mesjul01

Collection locations

Genbank
CO1 #

Genbank
CAD #

Genbank
EF2 #

Genbank
LWR #

Genbank
WG #

Madagascar: Antsiranana
USA: Virginia
USA: Arizona
USA: Michigan
USA: California
USA: Florida
USA: Michigan
USA: Michigan
Spain: Mallorca
S Africa: Western Cape
USA: Massachusetts
USA: Arizona
USA: Arizona
Mexico: Guerrero
USA: Arizona
USA: Minnesota
USA: California
Mexico: Baja CA Sur

JQ742635
KJ920541
KJ920542
KJ920543
KJ920544
KJ920545
KJ920508
KJ920509
JQ742637
JQ742636
KJ920510
KJ920513
KJ920520
KJ920521
KJ920512
JQ742647
DQ074325
KJ920511

JQ742579
N/A
KJ920571
N/A
KJ920572
N/A
N/A
N/A
JQ742581
JQ742580
N/A
KJ920548
KJ920553
KJ920554
KJ920547
JQ742591
N/A
KJ920546

EF013388
KJ920605
KJ920606
KJ920607
KJ920608
KJ920609
KJ920573
KJ920574
EF013447
EF013446
KJ920576
KJ920578
KJ920585
KJ920586
KJ920577
JQ742693
N/A
KJ920575

EF013546
KJ920640
KJ920641
KJ920642
N/A
KJ920643
KJ920610
KJ920611
EF013590
EF013589
KJ920613
KJ920614
KJ920620
KJ920621
N/A
JQ742738
HE963100
KJ920612

JQ742891
N/A
KJ920676
KJ920677
KJ920678
KJ920679
KJ920644
KJ920645
JQ742893
JQ742892
KJ920647
KJ920649
KJ920655
KJ920656
KJ920648
JQ742903
HE963097
KJ920646

Me = Messor
My = Myrmica
N = Novomessor

So = Solenopsis
St = Stenamma
V = Veromessor
58

Table 1.2. Morphological character state matrix for Aphaenogaster and outgroup species
Character
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22
Species
A. araneoides
A. ashmeadi 7
A. ashmeadi 12
A. balcanica
A. boulderensis
A. carolinensis
A. flemingi
A. floridana
A. fulva 5
A. fulva 7
A. huachucana
A. japonica
A. lamellidens
A. mariae
A. megommata
A. miamiana
A. mutica
A. occidentalis
A. patruelis
A. picea 1
A. picea 6
A. picea 39
A. rudis 2

0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
1
0
0
0
0

2
0
1
1
1
0
0
0
1
0
1
0
0
0
1
1
1
1
0
0
0
0
0

0
1
0
3
0
0
3
3
0
0
3
0
0
0
3
0
0
3
0
0
0
0
0

0
0
0
0
0
0
0
0
0
0
0
0
0
2
0
0
0
0
0
0
0
0
0

0
0
1
1
0
0
0
0
1
1
0
0
0
0
0
0
0
0
0
1
1
1
0

1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1

0
1
2
0
0
0
0
0
2
2
0
0
0
0
2
0
0
0
0
0
0
0
1

0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0

0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0

4
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
0
0
0
2
2
2
2

2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2

3
2
1
1
1
1
1
2
1
1
1
1
2
1
3
1
1
1
1
1
1
1
1
59

1
1
0
0
0
0
1
1
0
0
0
0
0
0
3
0
3
0
0
0
0
0
0

2
0
0
0
0
2
7
0
0
0
0
0
0
4
0
4
0
2
0
0
0
0
0

0
0
0
0
1
0
0
0
0
0
0
0
0
0
1
0
1
1
2
2
2
1
2

0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0

0
0
1
0
0
0
0
0
1
1
0
0
0
0
0
0
0
0
0
0
0
0
0

1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1

3
0
0
3
1
0
1
1
0
0
0
8
0
7
8
2
1
3
1
1
1
1
0

3
0
0
3
3
0
1
1
0
0
0
0
0
7
1
2
3
3
1
1
0
1
0

0
0
0
3
3
0
1
1
0
0
0
8
0
7
5
2
1
0
1
5
0
1
0

2
1
1
1
2
1
1
2
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1

Table 1.2 (cont’d).
Character

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22

Species
A. tennesseensis
A. texana
A. treatae
A. uinta
A. umphreyi
C. pennsylvanicus
F. glacialis
Me. bouvieri
Me. denticornis
My. latifrons
N. albesitosa
N. cockerelli
N. ensifera
So. aureus
St. diecki
V. andrei
V. julianus

0
1
0
2
0
1
0
2
2
0
1
0
0
0
0
2
0

0
0
0
1
0
3
2
3
3
0
2
2
2
0
0
1
1

0
3
2
0
0
0
0
0
0
0
0
0
0
0
0
3
0

0
0
0
0
0
3
0
0
0
0
0
0
0
0
0
0
0

1
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0

1
1
1
1
1
1
1
1
1
1
1
1
1
0
1
1
1

0
0
0
0
0
0
0
0
0
0
0
0
0
1
1
0
0

0
0
0
0
0
0
0
2
2
1
0
0
0
3
1
2
1

0
0
0
0
0
0
0
1
2
0
0
0
0
0
0
1
1

2
2
2
2
2
1
1
0
0
0
2
0
2
0
0
0
0

2
2
2
2
2
3
3
2
2
2
2
2
2
0
2
2
2

1
1
2
2
1
3
2
1
3
1
2
2
2
0
0
1
1

60

1
0
0
0
0
0
2
1
1
2
1
2
2
5
4
1
1

0
0
0
2
3
8
8
0
0
1
2
2
6
5
1
1
1

2
2
2
1
0
0
0
0
0
0
0
1
1
3
0
0
0

0
0
0
0
0
0
0
0
0
1
0
0
0
0
2
0
0

0
0
0
0
1
0
0
0
0
0
0
0
0
0
0
0
0

0
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1

0
0
5
1
4
1
1
3
3
4
1
1
5
8
6
4
4

0
0
5
3
0
1
1
4
4
4
4
4
5
8
6
4
6

4
0
5
3
4
1
1
4
4
4
4
4
5
8
6
4
6

1
1
1
1
1
0
0
2
2
1
1
1
1
0
1
1
1

Table 1.2 (cont’d).
Character
Species
A. araneoides
A. ashmeadi 7
A. ashmeadi 12
A. balcanica
A. boulderensis
A. carolinensis
A. flemingi
A. floridana
A. fulva 5
A. fulva 7
A. huachucana
A. japonica
A. lamellidens
A. mariae
A. megommata
A. miamiana
A. mutica
A. occidentalis
A. patruelis
A. picea 1
A. picea 6
A. picea 39
A. rudis 2
A. rudis 4
A. rudis 8
A. swammerdami
A. tennesseensis

23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42

1
2
2
2
0
2
2
0
2
2
2
3
2
3
2
2
1
2
3
2
2
2
2
3
2
1
3

2
1
4
1
1
1
1
0
4
4
0
0
1
0
0
1
1
1
1
1
0
0
0
0
1
9
1

0
0
1
3
0
1
1
0
1
1
0
1
1
1
0
1
0
0
0
1
1
1
1
1
1
3
1

0
0
0
1
0
1
1
0
0
1
0
1
1
0
0
1
0
0
0
1
1
1
1
1
1
0
0

5
3
2
0
3
2
3
3
2
2
3
3
3
2
3
2
3
2
2
2
2
2
2
3
2
0
3

1
1
1
0
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
0
1

0
0
0
3
0
0
0
0
0
0
0
0
0
1
0
0
0
0
0
0
0
0
0
0
0
5
0

1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
0

1
0
2
0
0
1
2
0
2
2
2
2
1
2
2
0
2
2
0
1
1
1
2
2
2
0
1

0
0
2
1
2
0
2
0
2
2
0
0
1
1
0
0
0
2
0
0
0
0
0
0
0
1
1
61	
  

0
0
2
1
2
0
0
0
2
2
0
2
1
1
0
0
0
2
0
0
0
0
0
0
0
1
1

1
1
1
0
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
0
1

1
1
2
0
1
1
1
3
2
1
2
1
1
0
2
1
1
1
1
1
1
1
1
1
1
0
1

2
2
3
1
1
3
1
3
2
2
3
2
3
2
1
3
2
1
1
2
2
1
2
2
2
2
1

4
2
1
1
2
1
4
2
1
1
2
2
1
1
2
1
2
1
1
1
1
1
1
1
1
1
1

0
0
0
2
0
0
0
0
0
0
0
0
1
0
0
0
0
0
0
0
0
0
0
0
0
4
0

1
1
1
2
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
4
1

3
1
1
0
3
1
2
3
4
4
5
5
1
1
5
6
3
7
7
1
1
1
1
1
1
0
9

1
4
2
1
1
6
4
1
2
2
3
3
3
6
1
3
1
3
2
5
5
5
4
4
4
1
5

2
2
2
2
2
2
0
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2

Table 1.2 (cont’d).
Character
Species
A. texana
A. treatae
A. uinta
A. umphreyi
C. pennsylvanicus
F. glacialis
Me. bouvieri
Me. denticornis
My. latifrons
N. albesitosa
N. cockerelli
N. ensifera
So. aureus
St. diecki
V. andrei
V. julianus

A = Aphaenogaster
C = Camponotus
F = Formica

23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42

3
2
2
1
0
0
3
2
2
1
3
1
0
3
2
1

1
1
0
1
5
1
3
3
1
2
1
1
0
0
1
1

0
1
0
0
0
0
1
1
0
0
0
0
0
1
3
1

0
1
0
0
0
0
1
1
0
2
0
0
0
1
1
0

3
3
3
3
4
3
0
0
2
4
5
5
0
1
0
3

1
1
1
1
0
0
0
0
0
0
0
0
0
0
0
3

0
0
0
0
0
0
3
4
0
0
0
0
0
0
3
0

1
1
1
1
1
1
1
1
1
1
1
1
1
1
3
1

1
1
2
2
1
1
0
0
1
0
0
0
1
1
0
1

0
0
2
0
0
0
1
1
0
2
2
2
0
0
1
2

Me = Messor
My = Myrmica
N = Novomessor

0
0
2
0
0
0
1
0
0
2
2
2
2
0
0
2

1
1
1
1
0
0
0
0
1
1
1
1
1
1
0
1

2
0
1
1
3
3
0
0
0
2
1
1
1
1
0
2

3
2
3
2
0
0
1
1
1
1
3
1
1
2
1
3

2
2
1
1
3
3
1
1
1
3
4
4
0
0
1
2

0
0
0
0
0
0
2
2
0
0
0
0
0
0
1
0

So = Solenopsis
St = Stenamma
V = Veromessor

62

1
1
1
1
2
2
2
3
2
1
1
1
2
2
2
2

7
1
5
4
0
0
0
0
1
1
1
1
0
8
0
4

3
4
1
2
0
0
2
2
3
6
6
4
0
2
2
2

2
2
2
2
3
3
2
2
2
0
1
0
2
2
1
1

Table 1.3. PCR primers used for the amplification of gene loci.
Gene
CO1

Primer
LCO
HCO

Sequence
GGT CAA CAA ATC ATA AAG ATA TTG G
TAA ACT TCA GGT GAC CAA AAA ATC A

Source
Folmer, et al. (1994)
Folmer, et al. (1994)

CAD

CD892F
CD1491R

5'- GGYACCGGRCGTTGYTAYATGAC -3'
5'- GCCGCARTTNAGRGCRGTYTGYCC -3'

Ward, et al. (2010)
Ward, et al. (2010)

EF1-alpha F2

F2-557F
F2-1118R

5'- GAACGTGAACGTGGTATYACSAT -3'
5'- TTACCTGAAGGGGAAGACGRAG -3'

Brady, et al. (2006)
Brady, et al. (2006)

LW Rhod

LR143F
LR639ER

5'- GACAAAGTKCCACCRGARATGCT -3'
5'- YTTACCGRTTCCATCCRAACA -3'

Ward & Downie (2005)
Ward & Downie (2005)

Wingless

Wg578F
Wg1032R

5'- TGCACNGTGAARACYTGCTGGATGCG -3'
5'- ACYTCGCAGCACCARTGGAA -3'

Ward & Downie (2005)
Ward & Downie (2005)

63

A. occidentalis
A. uinta
A. boulderensis
A. mutica
A. megommata
A. flemingi
A. floridana
N. cockerelli
N. ensifera Novomessor
N. albisetosa
A. araneoides
A. ashmeadi 7
A. treatae
A. japonica
A. huachucana
A. texana
A. lamellidens
A. miamiana
A. carolinenesis
M. bouvieri
M. denticornis
V. andrei
A. balcanica
A. swammerdami
A. ashmeadi 1
A. mariae
A. fulva 5
A. fulva 7
Aphaenogaster
A. umphreyi
A. rudis 2
A. rudis 4
A. rudis 8
A. picea 6
A. picea 1
A. picea 39
A. tennesseensis
A. patruelis
My. latifrons
98
V. julianus
St. diecki
100
Outgroups
So. aurea
F. glacialis
C. pennsylvanicus
3.0
Figure 1.1. One of 5 most parsimonious trees reconstructed for 43 taxa with morphology data in
a TNT analysis. Bootstrap values are above the branches. Clades without bootstrap values were
not resolved in a strict consensus tree. A. = Aphaenogaster, C. = Camponotus, F. = Formica, M.
= Messor, My. = Myrmica, N. = Novomessor, So. = Solenopsis, St. = Stenamma, V. =
Veromessor.

64

97/11
87/8

A. carolinensis
A. miamiana
A. rudis 8
A. treatae
A. huachucana
94/6
A. texana
A. lamellidens
100/17
A. picea 6
99/15
A. picea 39

A. rudis 2
A. rudis 4
A. picea 1
99/13
A. ashmeadi 1
A. ashmeadi 7
A. flemingi
A. floridana
A. megommata
A. mutica
A. boulderensis
A. patruelis
99/2
A. uinta
A. occidentalis
A. fulva 5
98/4
A.
fulva 7
99/2
A. araneoides
Aphaenogaster
A. umphreyi
91/10
A. mariae
A. japonica A. tennesseensis
99/11
M. bouvieri
M. denticornis
A. balcanica
N. albisetosa
97/17
100/18 97/11
N. cockerelli
Novomessor
100/23
N. ensifera
V. andrei
100/71
V. julianus
So. aurea
St. diecki
A. swammerdami
Outgroups
My. latifrons
F. glacialis
C. pennsylvanicus
100/14

3.0

Figure 1.2. One most parsimonious tree reconstructed for 43 taxa with morphology
and DNA data (CO1, CAD, EF2, LWR, WG) of 43 in a TNT analysis. Bootstrap
values/Bremer support are above the branches. A. = Aphaenogaster, C. =
Camponotus, F. = Formica, M. = Messor, My. = Myrmica, N. = Novomessor, So. =
Solenopsis, St. = Stenamma, V. = Veromessor.

65

*
**
**

**

**
**

Novomessor albisetosa
Novomessor cockerelli
**
Novomessor ensifera
Veromessor andrei
*
Veromessor julianus
Stenamma diecki
Solenopsis aurea
Myrmica latifrons

Camponotus pennsylvanicus
Formica glacialis

0.2
Figure 1.3. Bayesian majority rule consensus tree reconstructed for 43 taxa with morphology
and five genes (CO1, CAD, EF2, LWR, WG) in a Mr. Bayes analysis, Posterior probabilities
values greater than 90% are above the branches (* > 90%, **= 100%). Data were partitioned by
gene and codon position and and analyzed with a best-fit GTR + I + G model, 20 million
generations and a burn-in of 5,000,000 generations Novomessor and outgroups.

66

Figure 1.3. (cont'd).

**
**
**
**

**
**
**

**

Aphaenogaster carolinenesis
Aphaenogaster miamiana
Aphaenogaster rudis 8
Aphaenogaster treatre
Aphaenogaster huachucana
Aphaenogaster texana
Aphaenogaster lamellidens
rudis 2
* ** Aphaenogaster
Aphaenogaster rudis 4
Aphaenogaster picea 6
Aphaenogaster picea 39
** **
Aphaenogaster picea 1
** Aphaenogaster ashmeadi 1
**
Aphaenogaster ashmeadi 7
Aphaenogaster flemingi
**Aphaenogaster floridana
** Messor bouvieri
Messor denticornis
** Aphaenogaster japonica
Aphaenogaster boulderensis
Aphaenogaster uinta
Aphaenogaster
megommata
*
**Aphaenogaster mutica
Aphaenogaster patruelis
Aphaenogaster occidentalis
Aphaenogaster mariae
**Aphaenogaster tennesseensis
Aphaenogaster balcanica
Aphaenogaster fulva 5
Aphaenogaster fulva 7
Aphaenogaster umphreyi
Aphaenogaster araneoides
Aphaenogaster swammerdami

67

APPENDIX	
  B	
  
Tables	
  and	
  Figures	
  for	
  Chapter	
  3	
  

68	
  

Table 2.1. Aphaenogaster and outgroup specimens with associated localities and Genbank numbers
Taxon name, author and specimen number
Aphaenogaster araneoides Emery
Aphaenogaster ashmeadi(Emery) 7
Aphaenogaster ashmeadi 10
Aphaenogaster ashmeadi 12
Aphaenogaster balcanica (Emery)
Aphaenogaster boulderensis Smith
Aphaenogaster carolinenesis Wheeler 1
Aphaenogaster carolinenesis 2
Aphaenogaster carolinenesis 3
Aphaenogaster carolinenesis 12
Aphaenogaster carolinenesis 16
Aphaenogaster flemingi Smith
Aphaenogaster floridana Smith 1
Aphaenogaster floridana 6
Aphaenogaster fulva 1
Aphaenogaster fulva 4
Aphaenogaster fulva 5
Aphaenogaster fulva 6
Aphaenogaster fulva 7
Aphaenogaster fulva 10
Aphaenogaster fulva 11
Aphaenogaster fulva 12
Aphaenogaster fulva 13
Aphaenogaster fulva 17
Aphaenogaster fulva 18
Aphaenogaster huachucana Creighton 1
Aphaenogaster huachucana 4

Genbank
COI#
KJ920514
KJ920515
KP730068
KJ920516
KJ920517
KJ920518
KJ920519
KP730069
KP730070
KP730071
KP730072
KJ920522
KJ920523
KP730073
KP730074
KP730075
KJ920524
KP730076
KJ920525
KP730077
KP730078
KP730079
KP730080
KP730081
KP730082
KJ920526
KP730083
69

Genbank
CAD #
KJ920549
KJ920550
N/A
KJ920551
N/A
N/A
KJ920552
KP860427
KP860428
KP860429
KP860430
KJ920555
KJ920556
N/A
KP860431
KP860432
KJ920557
KP860433
KJ920558
KP860434
KP860435
KP860436
KP860437
KP860438
KP860439
N/A
KP860440

Genbank
EF1αF2 #
KJ920579
KJ920581
KP730150
KJ920580
KJ920582
KJ920583
KJ920584
KP730151
KP730152
KP730153
KP730154
KJ920587
KJ920588
KP730155
KP730156
KP730157
KJ920590
KP730158
KJ920589
KP730159
KP730160
KP730161
KP730162
KP730163
KP730164
N/A
KP730165

Genbank
LWR #
KJ920615
KJ920616
KP860353
KJ920617
KJ920618
N/A
KJ920619
KP860354
KP860355
KP860356
KP860357
KJ920622
KJ920623
KP860358
KP860359
KP860360
KJ920624
KP860361
KJ920625
KP860362
KP860363
KP860364
KP860365
KP860366
KP860367
KJ920626
KP860368

Genbank
WG #
KJ920650
N/A
N/A
KJ920651
KJ920652
KJ920653
KJ920654
KP730229
KP730230
KP730231
KP730232
KJ920657
KJ920658
KP730233
KP730234
KP730235
KJ920660
KP730236
KJ920659
KP730237
KP730238
KP730239
KP730240
KP730241
KP730242
KJ920661
KP730243

Table 2.1. (cont’d).
Taxon name, author and specimen number
Aphaenogaster japonica Forel
Aphaenogaster lamellidens Mayr 1
Aphaenogaster lamellidens 2
Aphaenogaster mariae Forel 1
Aphaenogaster mariae 2
Aphaenogaster megommata Smith
Aphaenogaster miamiana Wheeler 1
Aphaenogaster miamiana 2
Aphaenogaster miamiana 3
Aphaenogaster miamiana 5
Aphaenogaster mutica Pergande
Aphaenogaster occidentalis (Emery) 1
Aphaenogaster occidentalis 4
Aphaenogaster occidentalis 5
Aphaenogaster patruelis Forel
Aphaenogaster picea (Wheeler) 1
Aphaenogaster picea 2
Aphaenogaster picea 3
Aphaenogaster picea 4
Aphaenogaster picea 6
Aphaenogaster picea 15
Aphaenogaster picea 16
Aphaenogaster picea 19
Aphaenogaster picea 21
Aphaenogaster picea 23
Aphaenogaster picea 26
Aphaenogaster picea 27
Aphaenogaster picea 28

Genbank
COI#
KJ920527
KJ920528
KP730084
KJ920529
KP730085
KJ920530
KJ920531
KP730086
KP730087
KP730088
KJ920532
KJ920533
KP730089
KP730090
KJ920534
KJ920535
KP730091
KP730092
KP730093
KJ920536
KP730094
KP730095
KP730096
KP730097
KP730098
KP730099
KP730100
KP730101
70

Genbank
CAD #
KJ920559
KJ920560
KP860441
KJ920561
KP860442
KJ920562
KJ920563
KP860443
KP860444
KP860445
N/A
KJ920564
KP860446
KP860447
N/A
KJ920565
KP860448
KP860449
KP860450
KJ920566
KP860451
KP860452
KP860453
KP860454
KP860455
KP860456
KP860457
KP860458

Genbank
EF1αF2 #
KJ920591
KJ920592
KP730166
KJ920593
KP730167
KJ920594
KJ920595
KP730168
KP730169
KP730170
KJ920596
KJ920597
KP730171
KP730172
KJ920598
KJ920599
KP730173
KP730174
KP730175
KJ920600
KP730176
KP730177
KP730178
KP730179
KP730180
KP730181
KP730182
KP730183

Genbank
LWR #
KJ920627
KJ920628
KP860369
KJ920629
KP860370
KJ920630
KJ920631
KP860371
KP860372
KP860373
KJ920632
KJ920633
KP860374
KP860375
KJ920634
KJ920635
KP860376
KP860377
KP860378
KJ920636
KP860379
KP860380
KP860381
KP860382
KP860383
KP860384
KP860385
KP860386

Genbank
WG #
KJ920662
KJ920663
KP730244
KJ920664
KP730245
KJ920665
KJ920666
KP730246
KP730247
KP730248
KJ920667
KJ920668
KP730249
KP730250
KJ920669
KJ920670
KP730251
KP730252
KP730253
KJ920671
KP730254
KP730255
KP730256
KP730257
KP730258
KP730259
KP730260
KP730261

Table 2.1. (cont’d).
Taxon name, author and specimen number
Aphaenogaster picea 30
Aphaenogaster picea 31
Aphaenogaster picea 32
Aphaenogaster picea 36
Aphaenogaster picea 37
Aphaenogaster picea 39
Aphaenogaster rudis Enzmann 2
Aphaenogaster rudis 4
Aphaenogaster rudis 6
Aphaenogaster rudis 8
Aphaenogaster rudis 10
Aphaenogaster rudis 12
Aphaenogaster rudis 14
Aphaenogaster rudis 15
Aphaenogaster rudis 16
Aphaenogaster rudis 17
Aphaenogaster rudis 19
Aphaenogaster rudis 28
Aphaenogaster rudis 29
Aphaenogaster rudis 32
Aphaenogaster rudis 33
Aphaenogaster rudis 34
Aphaenogaster rudis 41
Aphaenogaster rudis 43
Aphaenogaster rudis 44
Aphaenogaster rudis 45
Aphaenogaster rudis 46
Aphaenogaster rudis 47

Genbank
COI#
KP730102
KP730103
KP730104
KP730105
KP730106
KJ920537
KJ920538
KJ920539
KP730107
KJ920540
KP730108
KP730109
KP730110
KP730111
KP730112
KP730113
KP730114
KP730115
KP730116
KP730117
KP730118
KP730119
KP730120
KP730121
KP730122
KP730123
KP730124
KP730125
71

Genbank
CAD #
KP860459
KP860460
KP860461
KP860462
KP860463
KJ920567
KJ920568
KJ920569
KP860464
KJ920570
KP860465
KP860466
KP860467
KP860468
KP860469
KP860470
KP860471
KP860472
KP860473
KP860474
KP860475
KP860476
KP860477
KP860478
KP860479
KP860480
KP860481
KP860482

Genbank
EF1αF2 #
KP730184
KP730185
KP730186
KP730187
KP730188
KJ920601
KJ920602
KJ920603
KP730189
KJ920604
KP730190
KP730191
KP730192
KP730193
KP730194
KP730195
KP730196
KP730197
KP730198
N/A
N/A
KP730199
KP730200
KP730201
KP730202
KP730203
KP730204
KP730205

Genbank
LWR #
KP860387
KP860388
KP860389
KP860390
KP860391
N/A
KJ920637
KJ920638
KP860392
KJ920639
KP860393
KP860394
KP860395
KP860396
KP860397
N/A
KP860398
KP860399
KP860400
KP860401
KP860402
KP860403
KP860404
KP860405
N/A
KP860406
KP860407
KP860408

Genbank
WG #
KP730262
KP730263
KP730264
KP730265
KP730266
KJ920672
KJ920673
KJ920674
KP730267
KJ920675
N/A
KP730268
KP730269
N/A
KP730270
KP730271
KP730272
KP730273
KP730274
KP730275
KP730276
N/A
KP730277
KP730278
KP730279
N/A
KP730280
KP730281

Table 2.1. (cont’d).
Taxon name, author and specimen number
Aphaenogaster rudis 48
Aphaenogaster rudis 49
Aphaenogaster rudis 50
Aphaenogaster rudis 51
Aphaenogaster swammerdami Forel
Aphaenogaster tennesseensis (Mayr) 1
Aphaenogaster tennesseensis 2
Aphaenogaster tennesseensis 5
Aphaenogaster tennesseensis 6
Aphaenogaster texana Wheeler 1
Aphaenogaster texana 3
Aphaenogaster texana 4
Aphaenogaster texana 5
Aphaenogaster texana 6
Aphaenogaster texana 7
Aphaenogaster treatae Forel 1
Aphaenogaster treatae 4
Aphaenogaster uinta Wheeler 1
Aphaenogaster uinta 2
Aphaenogaster umphreyi Deyrup & Davis 1
Aphaenogaster umphreyi 2
Camponotus castanaeus (Latreille)
Camponotus nearcticus Emery
Camponotus pennsylvanicus (DeGeer)
Formica glacialis Wheeler
Formica subintegra Wheeler
Formica subsericea Say
Messor bouvieri Bondroit

Genbank
COI#
KP730126
KP730127
KP730128
KP730129
JQ742635
KJ920541
KP730130
KP730131
KP730132
KP730133
KJ920542
KP730135
KP730136
KP730137
KP730138
KJ920543
KP730139
KJ920544
KP730140
KJ920545
KP730141
KP730061
KP730062
KJ920508
KJ920509
KP730063
KP730064
JQ742637
72

Genbank
CAD #
KP860483
KP860484
KP860485
KP860486
JQ742579
N/A
N/A
N/A
KP860487
N/A
KJ920571
KP860488
KP860489
KP860490
KP860491
N/A
N/A
KJ920572
N/A
N/A
KP860492
N/A
N/A
N/A
N/A
N/A
N/A
JQ742581

Genbank
EF1αF2 #
KP730206
KP730207
KP730208
KP730209
EF013388
KJ920605
KP730210
KP730211
KP730212
KP730213
KJ920606
KP730215
KP730216
KP730217
KP730218
KJ920607
KP730219
KJ920608
KP730220
KJ920609
KP730221
KP730143
KP730144
KJ920573
KJ920574
KP730145
KP730146
EF013447

Genbank
LWR #
KP860409
KP860410
KP860411
KP860412
EF013546
KJ920640
KP860413
KP860414
KP860415
N/A
KJ920641
KP860416
KP860417
KP860418
KP860419
KJ920642
KP860420
N/A
KP860421
KJ920643
KP860422
KP860349
KP860350
KJ920610
KJ920611
N/A
KP860351
EF013590

Genbank
WG #
KP730282
KP730283
KP730284
KP730285
JQ742891
N/A
KP730286
KP730287
KP730288
KP730289
KJ920676
KP730291
KP730292
KP730293
KP730294
KJ920677
KP730295
KJ920678
KP730296
KJ920679
KP730297
KP730223
N/A
KJ920644
KJ920645
KP730224
KP730225
JQ742893

Table 2.1. (cont’d).
Taxon name, author and specimen number
Messor denticornis Forel
Myrmica incompleta Provancher
Myrmica latifrons Stärcke
Novomessor albisetosa (Mayr)
Novomessor cockerelli André 1
Novomessor cockerelli André 2
Novomessor ensifera Forel
Solenopsis aurea Wheeler
Solenopsis invicta Buren
Stenamma diecki Emery
Veromessor andrei (Mayr)
Veromessor julianus (Pergande)
JTL-001 (undescribed species)

Genbank
COI#
JQ742636
KP730065
KJ920510
KJ920513
KJ920520
KP730066
KJ920521
KJ920512
KP730067
JQ742647
DQ074325
KJ920511
KP730142

73	
  

Genbank
CAD #
JQ742580
N/A
N/A
KJ920548
KJ920553
KP860424
KJ920554
KJ920547
KP860425
JQ742591
N/A
KJ920546
N/A

Genbank
EF1αF2 #
EF013446
KP730147
KJ920576
KJ920578
KJ920585
KP730148
KJ920586
KJ920577
KP730149
JQ742693
N/A
KJ920575
KP730222

Genbank
LWR #
EF013589
N/A
KJ920613
KJ920614
KJ920620
KP860352
KJ920621
N/A
N/A
JQ742738
HE963100
KJ920612
KP860423

Genbank
WG #
JQ742892
N/A
KJ920647
KJ920649
KJ920655
KP730226
KJ920656
KJ920648
KP730228
JQ742903
HE963097
KJ920646
KP730298

Table 2.2. Bootstrap and partitioned bremer support values that
correspond to the label nodes in the parsimony phylogeny (Fig. 2).

Node 1
Node 2
Node 3
Node 4
Node 5
Node 6
Node 7
Node 8
Node 9
Node 10
Node 11
Node 12
Node 13
Node 14
Node 15
Node 16
Node 17
Node 18
Node 19
Node 20
Node 21
Node 22
Node 23
Node 24
Node 25
Node 26
Node 27
Node 28
Node 29
Node 30
Node 31
Node 32
Node 33
Node 34
Node 35
Node 36
Node 37
Node 38

Bootstrap
62
51
62
<50
64
67
85
61
60
84
97
59
85
98
<50
<50
<50
<50
<50
<50
63
<50
58
<50
55
55
86
85
74
<50
91
81
100
100
96
54
52
61

COI CAD EF2 LWR Wg
-0.1 0.0
0.1
0.0 0.1
0.1
0.0
0.0
0.0 -0.1
0.0 -2.0 2.9
0.0 0.0
0.1
0.0
0.1
0.0 -0.1
0.9
0.0 -0.1 0.0 0.1
0.6
0.0
0.1
0.0 0.3
2.8
0.0
0.0
0.0 0.1
1.0
0.0 -0.1 0.0 0.1
0.0
0.0 -0.1 0.0 1.1
1.7
0.0
0.1
0.0 0.1
7.8
0.0 -0.5 -0.4 0.1
-0.6 0.0
0.0
2.0 -0.3
4.0
0.0
0.0
0.0 0.0
3.4
0.0
0.0
1.0 0.7
-0.6 0.7
0.0
0.8 0.1
-1.4 1.0
0.1
1.6 -0.3
-0.9 0.9 -0.1 1.2 -0.1
-0.4 0.8 -0.1 0.5 0.2
0.3
0.0
0.0
0.0 -0.3
-0.7 0.0
0.0
0.0 0.7
0.7
0.0
0.0
0.0 0.3
-0.5 0.0
0.0
0.0 0.6
1.2
0.0 -0.1 0.0 -0.1
-0.6 0.0
0.0
2.0 -0.3
-0.2 1.5
1.4
0.0 0.3
-0.2 1.6
1.2 -0.2 0.6
2.8
0.0
0.0
0.0 0.2
0.0
2.0
0.9
0.0 0.1
0.2
0.0 -0.1 0.9 -0.1
-0.1 0.0
0.1
0.0 0.1
13.2 -5.5 0.2
0.0 0.2
0.0
1.0 -0.1 0.0 0.1
11.4 0.1
0.3
0.0 0.2
11.0 0.0 -0.4 1.0 0.4
6.0
0.1
0.9
0.0 0.0
1.9
0.0 -1.1 0.0 0.2
-0.8 1.5
1.7 -0.4 1.1
-0.5 0.0
0.4
0.0 0.1
74

Sum
0.0
0.0
1.0
0.0
1.0
1.0
3.0
1.0
1.0
2.0
7.0
1.0
4.0
5.0
1.0
1.0
1.0
1.0
0.0
0.0
1.0
0.0
1.0
1.0
3.0
3.0
3.0
3.0
1.0
0.0
8.0
1.0
12.0
12.0
7.0
1.0
3.0
0.0

Table 2.2 (cont’d).
Bootstrap
Node 39
Node 41
Node 42
Node 43
Node 44
Node 45
Node 46
Node 47
Node 48
Node 49
Node 50
Node 51
Node 52
Node 53
Node 54
Node 55
Node 56
Node 57
Node 58
Node 59
Node 60
Node 61
Node 62
Node 63
Node 64
Node 65
Node 66
Node 67
Node 68
Node 69
Node 70
Node 71
Node 72
Node 73
Node 74
Node 75
Node 76
Node 77
Node 78

<50
52
<50
<50
<50
<50
93
<50
<50
<50
<50
54
66
98
<50
<50
90
87
88
<50
100
<50
100
100
<50
62
<50
<50
52
<50
<50
<50
<50
80
73
100
100
94
84

COI CAD EF2 LWR Wg

Sum

-0.3 0.0
2.0
0.0
0.2 -0.5
0.1
0.2
0.2
1.4
0.0
0.4
-0.1 1.8
0.0
1.2
0.1
0.0
0.0
0.0
1.2
0.0
0.1
0.0
-0.1 1.0
0.0
8.7
6.8 -3.0
0.3
0.0
21.6 -5.9
41.8 -17.0
1.4
0.0
1.5
0.0
9.1
0.0
6.7
0.1
7.0
0.0
16.1 13.0
0.6
0.0
3.3
0.0
-0.4 0.0
0.2
0.0
2.6
0.0
0.1
0.0
-0.9 0.0
0.4
0.0
-1.1 0.0
0.9
0.0
1.4 -1.0
40.2 -7.1
13.8 1.0
8.2
0.0
34.4 -16.0

0.0
1.0
1.0
1.0
1.0
1.0
3.0
1.0
0.0
0.0
1.0
1.0
1.0
10.0
4.0
1.0
12.0
8.0
1.0
1.0
13.0
12.0
8.0
31.0
2.0
4.0
2.0
2.0
4.0
5.0
1.0
1.0
1.0
1.0
1.0
26.0
17.0
10.0
2.0

0.2
-0.2
0.0
0.1
0.0
0.5
0.6
-0.5
0.0
0.0
-0.1
-0.1
-0.1
-0.1
0.2
0.8
-1.1
-5.6
-0.5
-0.2
3.0
2.4
0.2
-0.1
0.8
-0.1
1.0
0.9
0.6
2.9
0.9
-0.1
0.9
0.2
0.2
-2.2
1.9
-0.2
-5.2
75	
  

0.0
-1.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
-0.9
-3.1
0.0
0.0
0.0
2.0
1.0
1.0
0.0
0.0
0.0
0.0
0.0
1.0
0.0
0.0
0.0
0.0
0.0
-1.3
0.0
1.0
-3.0

0.1
0.2
1.3
0.6
-0.6
0.2
0.7
0.3
0.0
0.0
-0.1
1.0
0.1
1.4
0.0
-0.1
-1.7
-8.1
0.1
-0.3
0.9
0.8
-0.1
1.0
0.7
0.8
1.4
0.9
0.8
1.0
1.0
0.6
1.2
0.0
0.4
-3.5
0.3
0.9
-8.2

Table 2.2 (cont’d).
Node 79

Bootstrap
63

Node 80

61

1.4

0.0

-0.1

0.0

-0.3

1.0

Node 81

64

2.0

0.0

-1.1

0.0

0.1

1.0

Node 82

69

1.6

0.0

-0.6

0.0

0.0

1.0

Node 83
Node 84
Node 85
Node 86
Node 87
Node 88
Node 89
Node 90
Node 91
Node 92
Node 93
Node 94
Node 95
Node 96
Node 97
Node 98
Node 99
Node 100
Node 101
Node 102
Node 103
Node 104
Node 105
Node 106

75
92
<50
<50
<50
<50
99
89
100
65
100
98
100
89
97
68
90
100
84
100
81
100
100
100

1.9
1.3
-4.0
34.3
34.7
8.2
27.5
22.2
24.2
-0.1
47.3
12.2
9.0
21.7
42.0
6.7
34.3
12.8
2.9
29.9
2.4
25.7
16.0
21.0

0.0
0.9
6.0
-16.0
-16.0
0.0
2.5
4.0
-6.0
2.0
8.0
-3.0
4.0
0.2
-16.5
0.0
-16.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0

-1.1
2.8
8.9
-5.1
-5.5
5.9
-3.4
-3.3
7.0
0.1
3.9
0.0
1.0
-2.3
13.7
1.9
-5.0
17.0
3.0
11.0
0.9
10.3
54.0
22.9

0.0 0.1
1.0
0.0 0.0
5.0
0.7 2.5 14.0
-3.0 -8.2 2.0
-3.0 -8.2 2.0
1.0 7.9 23.0
-1.5 -2.1 23.0
-7.0 -4.0 12.0
0.0 1.8 27.0
0.0 0.0
2.0
0.0 -0.2 59.0
3.0 1.9 14.0
-1.0 8.0 21.0
-2.8 -4.8 12.0
-4.1 -15.0 20.0
1.0 -0.7 9.0
-3.0 -8.3 2.0
0.0 0.2 30.0
0.0 0.1
6.0
8.0 0.1 49.0
0.0 0.7
4.0
0.0 13.9 50.0
0.0 0.0 70.0
1.0 8.1 53.0

Totals

COI CAD EF2 LWR Wg
-0.1 0.0 -0.1 0.0 1.2

Sum
1.0

751.1 -73.1 142.9 -5.7

76

-6.1 809.0

Messor bouvieri
Messor denticornis
*
A. swammerdami
*
Veromssor andrei
Veromessor julianus
** Novomessor cockerelli 1
Novomessor cockerelli 2
*
**
**
Novomessor albisetosa
*
Novomessor ensifera
Solenopsis aurea
*
Solenopsis invicta
**
Stenamma diecki
** Myrmica latifrons
Myrmica incompleta
Camponotus castanaeus
Camponotus pennsylvanicus
**
Camponotus nearcticus
Formica subintegra
Formica glacialis
** Formica subsericea

*

90.0

Figure 2.1. One of 64,525 MPT reconstructed for 123 taxa of
Aphaenogaster and outgroups with DNA data and analysis of 5 genes in
PAUP*. Bootstrap values greater than 90% are above the branches (* >
90%, **= 100%). A. = Aphaenogaster. Specimen numbers and states/
provinces where collected are displayed next to each sample. The names
of non-monophyletic species correspond to specific colors.

77

Figure 2.1. (cont'd).

** A. mariae 2 MS
A. fulva 1 VA
A. fulva 7 NC
*
A. fulva 4 ON
A. fulva 6 NC
A. fulva 10 MS
A. fulva 11 MA
A. fulva 12 IN
A. fulva 17 ON
A. fulva 5 MS
A. fulva 13 IL
A. fulva 18 GA
A. umphreyi 1 FL
A. umphreyi 2 AL
A. aeraneoides CR
JTL-001 undescribed

78

Figure 2.1. (cont'd).

*

79

A. occidentalis 4 CO
** A. occidentalis 5 UT
A. occidentalis 1 WA
A. uinta 1 CA
A. boulderensis CA
A. mutica MX
A. patruelis CA
A. uinta 2 NV
A, megommata NV
A. japonica JA
A. balcanica GR
A. tennesseensis 1 VA
** A. tennesseensis 2 MN
A. tennesseensis 5 IA
A. tennesseensis 6 NE
A. mariae 1 VA
** A. mariae 2 MS
A. fulva 1 VA
A. fulva 7 NC

Figure 2.1. (cont'd).

A. picea 28 MA
A. picea 36 MA
A. picea 19 MA
A. picea 16 NJ
A. picea 37 MA
A. picea 30 IN
A. picea 31 ON
A. rudis 14 (picea) ON
A. picea 23 MI
A. picea 15 MI
A. rudis 29 (picea) ON
A. picea 2 MI
A. picea 4 NY
** A. picea 3 OH
A. picea 6 MN
A. picea 27 VA
A. picea 39 MA
A. huachucana 4 AZ
A. flemingi FL
A. floridana 6 GA
** A. floridana 7 FL

80

Figure 2.1. (cont'd).

A, rudis 16 OH
A. rudis 43 MO
A. rudis 2 MI
* A. rudis 4 OH
A. rudis 12 NJ
A. rudis 34 PA
A. ashmeadi 7 NC
*
** A. ashmeadi 12 NC
* A. ashmeadi 10 NC
A. treatae 1 MI
** A. treatae 4 AR
A. picea 21 MA
A. picea 26 NC
* A. picea 1 MI
A. picea 32 ON
A. picea 28 MA

81

Figure 2.1. (cont'd).

A. texana 3 AZ
A. texana 4 AZ
A. huachucana 3 AZ
A. lamellidens 1 VA
A. lemellidens 2 MS
A. rudis 10 MN
A. rudis 45 IN
A. texana 7 MO
A. texana 6 AR
A. rudis 44 TN
A. rudis 17 NC
A. rudis 32 GA
A. rudis 33 GA
A. rudis 41 GA
A. texana 5 MO

82

Figure 2.1. (cont'd).

A. rudis 8 NJ
A. rudis 15 NC
A. rudis 46 NC
A. rudis 47 NC
A. rudis 48 NC
A. rudis 19 VA
A. rudis 28 VA
A. carolinensis 2 NC
A. carolinensis 3 NC
A. carolinensis 12 NC
A. carolinensis 1 MS
A. carolinensis 16 MS
A. rudis 49 VA
A. rudis 50 GA
* A. rudis 51 GA
A. rudis 6 VA
A. miamiana 1 FL
A. miamiana 3 FL
A. miamiana 2 FL
A. miamiana 5 NC

83

JTL-001 undescribed
89 Messor bouvieri
90 Messor denticornis
91 A. swammerdami
92

96
106

93
94
95

99
97
98

100

101
102
105
103
104

Veromssor andrei
Veromessor julianus
Novomessor cockerelli 1
Novomessor cockerelli 2
Novomessor albisetosa
Novomessor ensifera
Solenopsis aurea
Solenopsis invicta
Stenamma diecki
Myrmica incompleta
Myrmica latifrons
Camponotus castanaeus
C. pennsylvanicus
Camponotus nearcticus
Formica subintegra
Formica glacialis
Formica subsericea

20.0

Figure 2.2. One of 64,525 MPT shown as a cladogram reconstructed for
123 taxa of Aphaenogaster and outgroups with DNA data and analysis of
5 genes in PAUP*. Node numbers are above the branches. A. =
Aphaenogaster. Specimen numbers and states/provinces where collected
are displayed next to each sample. The names of non-monophyletic
species correspond to specific colors.

84

Figure 2.2. (cont'd).

79
86

80

81

87

82
84
88
83

85

A. floridana 6 GA
A. floridana 7 FL
A. occidentalis 4 CO
A. occidentalis 5 UT
A. occidentalis 1 WA
A. uinta 1 CA
A. boulderensis CA
A. mutica MX
A. patruelis CA
A. uinta 2 NV
A, megommata NV
A. japonica JA
A. balcanica GR
A. tennesseensis 1 VA
A. tennesseensis 2 MN
A. tennesseensis 5 IA
A. tennesseensis 6 NE
A. mariae 1 VA
A. mariae 2 MS
A. fulva 1 VA
A. fulva 7 NC
A. fulva 4 ON
A. fulva 6 NC
A. fulva 10 MS
A. fulva 11 MA
A. fulva 12 IN
A. fulva 17 ON
A. fulva 5 MS
A. fulva 13 IL
A. fulva 18 GA
A. umphreyi 1 FL
A. umphreyi 2 AL
A. aeraneoides CR
JTL-001 undescribed
Messor bouvieri
Messor denticornis
A. swammerdami MAD

Figure 2.2. (cont'd).

A. huac hucana 4 AZ
A. flemingi FL
A. floridana 6 GA
A. floridana 7 FL
62 A. occidentalis 4 CO
A. occidentalis 5 UT
67 A. occidentalis 1 WA
A. uinta 1 CA
68
64 A. boulderensis CA
69
A. mutica MX
66
A. patruelis CA
70 66
A. uinta 2 NV
71
A, megommata NV
A. japonica JA
A. balcanica GR
73 A. tennesseensis 1 VA
75 A. tennesseensis 2 MN
A. tennesseensis 5 IA
74
77
A. tennesseensis 6 NE
A. mariae 1 VA
76
A. mariae 2 MS
A. fulva 1 VA

72

78

85

86

Figure 2.2. (cont'd).

43
44

57

45
42
46
49 47
48
50
58
54
51
59

53

56

52
55

61

60

87

A. picea 32 ON
A. picea 28 MA
A. picea 36 MA
A. picea 19 MA
A. picea 16 NJ
A. picea 37 MA
A. picea 30 IN
A. picea 31 ON
A. rudis 14 (picea) ON
A. picea 23 MI
A. picea 15 MI
A. rudis 29 (picea) ON
A. picea 2 MI
A. picea 4 NY
A. picea 3 OH
A. picea 6 MN
A. picea 27 VA
A. picea 39 MA
A. huachucana 4 AZ
A. flemingi FL
A. floridana 6 GA
A. floridana 7 FL
A. occidentalis 4 CO
A. occidentalis 5 UT
A. occidentalis 1 WA
A. uinta 1 CA
A. boulderensis CA
A. mutica MX

Figure 2.2. (cont'd).

37

41
36

88

A. texana 5 MO
27 A, rudis 16 OH
28 A. rudis 43 MO
30 A. rudis 2 MI
A. rudis 4 OH
31
29A. rudis 12 NJ
A. rudis 34 PA
32 A. ashmeadi 7
33 A. ashmeadi 12
35 A. ashmeadi 10
A. treatae 1
34 A. treatae 4
38 A. picea 21 MA
39 A. picea 26 NC
40 A. picea 1 MI
A. picea 32 ON
A. picea 28 MA
A. picea 36 MA
A. picea 19 MA

Figure 2.2. (cont'd).

A. texana 3 AZ
A. texana 4 AZ
14
A. huachucana 3 AZ
A. lamellidens 1
15
A. lamellidens 2
16
A. rudis 10 MN
17
A. rudis 45 IN
18
19 A. texana 7 MO
20 A. texana 6 AR
22 A. rudis 44 TN
24
A. rudis 17 NC
21 A. rudis 32 GA
25
23 A. rudis 33 GA
A. rudis 41 GA
A. texana 5 MO
A, rudis 16 OH
A. rudis 43 MO
A. rudis 2 MI
A. rudis 4 OH
A. rudis 12 NJ
A. rudis 34 PA
A. ashmeadi 7
89
A. ashmeadi 12
12
13

26

Figure 2.2. (cont'd).

1
2
3
4

5
7
8

6

10

9

90

A. rudis 8 NJ
A. rudis 15 NC
A. rudis 46 NC
A. rudis 47 NC
A. rudis 48 NC
A. rudis 19 VA
A. rudis 28 VA
A. carolinensis 2 NC
A. carolinensis 3 NC
A. carolinensis 12 NC
A. carolinensis 1 MS
A. carolinensis 16 MS
A. rudis 49 VA
A. rudis 50 GA
A. rudis 51 GA
A. rudis 6 VA
A. miamiana 1 FL
A. miamiana 3 FL
A. miamiana 2 FL
A. miamiana 5 NC

**
*

**

Myrmica incompleta

Formica subintegra
** Formica glacialis
Formica subsericea
Camponotus nearcticus
Camponotus castanaeus
Camponotus pennsylvanicus
0.2

Figure 2.3. Maximum likelihood tree reconstructed for 123 taxa with DNA data and
analysis of 5 genes in a RAxML analysis. Bootstrap values greater than 90% are above the
branches (* > 90%, **= 100%). A. = Aphaenogaster. Specimen numbers and states/
provinces where collected are displayed next to each sample. The names of nonmonophyletic species correspond to specific colors.

91

Figure 2.3. (cont'd).

Messor denticornis
Messor bouvieri
**
A. swammerdami
Veromssor andrei
Veromessor julianus
** Novomessor cockerelli 2
** Novomessor cockerelli 1
**
Novomessor albisetosa
Novomessor ensifera
Stenamma diecki
Solenopsis aurea
**
Solenopsis invicta
Myrmica incompleta
Myrmica latifrons

*

**

**

**

92

Figure 2.3. (cont'd).

A. balcanica GR
A. umphreyi 1 FL
A. umphreyi 2 AL
A. fulva 18 GA
A. fulva 11 MA
A. fulva 4 ON
A. fulva 12 IN
A. fulva 10 MS
A. fulva 1 VA
A. fulva 7 NC
A. fulva 6 NC
A. fulva 17 ON
A. fulva 5 MS
A. fulva 13 IL
A. aeraneoides CR
JTL-001 undescribed

93

Figure 2.3. (cont'd).

A. occidentalis 5 UT
** A. occidentalis 4 CO
A. occidentalis 1 WA
A. patruelis CA
A. boulderensis CA
A. mutica MX
A. uinta 2 NV
A. uinta 1 CA
A, megommata NV
A. tennesseensis 1 VA
** A. tennesseensis 2 MN
A. tennesseensis 5 IA
**A. tennesseensis 6 NE
A. mariae 2 MS
** A. mariae 1 VA
A. balcanica GR
A. umphreyi 1 FL

94

Figure 2.3. (cont'd).

A. treatae 1
A. picea 28 MA
A. picea 36 MA
A. picea 19 MA
** A. picea 16 NJ
A. picea 37 MA
A. picea 31 ON
A. picea 30 IN
A. rudis 14 (picea) ON
A. picea 23 MI
A. picea 15 MI
* A. rudis 29 (picea) ON
A. picea 4 NY
A. picea 2 MI
* A. picea 3 OH
A. picea 6 MN
A. picea 27 VA
A. picea 39 MA
A. picea 26 NC
A. picea 21 MA
A. picea 32 ON
A. picea 1 MI
A. floridana 6 GA
A. floridana 7 FL
A. flemingi FL
A. huachucana 4 AZ
A. japonica JA

95

Figure 2.3. (cont'd).

A, rudis 16 OH
A. rudis 43 MO
A. rudis 2 MI
A. rudis 34 PA
A. rudis 4 OH
A. rudis 12
A. ashmeadi 12
** A. ashmeadi 7
* A. ashmeadi 10
A. treatae 4
**A. treatae 1

96

Figure 2.3. (cont'd).

A. rudis 48 NC
A. rudis 28 VA
A. rudis 19 VA
A. miamiana 3 FL
A. miamiana 1 FL
A. miamiana 2 FL
A. miamiana 5 NC
A. huachucana 3 AZ
A. texana 3 AZ
A. texana 4 AZ
A. texana 6 AR
A. texana 5 MO
A. texana 7 MO
A. rudis 17 NC
A. rudis 32 GA
A. rudis 10 MN
A. rudis 44 TN
A. rudis 45 IN
A. rudis 33 GA
A. rudis 41 GA
A. lamellidens 2
**
A. lamellidens 1
A, rudis 16 OH

97

Figure 2.3. (cont'd).

A. caro linensis 3 NC
A. carolinensis 2 NC
A. carolinensis 12 NC
A. carolinensis 1 MS
A. carolinensis 16 MS
A. rudis 51 GA
A. rudis 50 GA
A. rudis 49 VA
A. rudis 6 VA
A. rudis 8 NJ
A. rudis 46 NC
A. rudis 15 NC
A. rudis 47 NC
A. rudis 48 NC
* A. rudis 28 VA
A. rudis 19 VA
** A. miamiana 3 FL
A. miamiana 1 FL
A. miamiana 2 FL
A. miamiana 5 NC

98

**
**
Formica subintegra
** Formica glacialis
Formica subsericea
Camponotus nearcticus
Camponotus pennsylvanicus
Camponotus castanaeus

0.8

Figure 2.4. Bayesian majority rule consensus tree reconstructed for 123 taxa with
morphology and five genes in a Mr. Bayes analysis, Posterior probabilities values greater
than 90% are above the branches (* > 90%, **= 100%). Data were partitioned by gene and
codon position and analyzed with a best-fit GTR + I + G model, 30 million generations and a
burn-in of 7,500,000 generations. A. = Aphaenogaster. Specimen numbers and states/
provinces where collected are displayed next to each sample. The names of nonmonophyletic species correspond to specific colors.

99

Figure 2.4. (cont'd).

Messor bouvieri
Messor denticornis
A. swammerdami
**
Veromssor andrei
Veromessor julianus
*
Novomessor cockerelli 1
** Novomessor cockerelli 2
**
Novomessor albisetosa
*
Novomessor ensifera
Stenamma diecki
Solenopsis aurea
**
Solenopsis invicta
Myrmica incompleta
Myrmica latifrons

100

Figure 2.4. (cont'd).

A. boulderensis CA
A. mutica MX
A. patruelis CA
A. occidentalis 4 CO
*
A. occidentalis 5 UT
** A. occidentalis 1 WA
*
A. uinta 1 CA
A. aeraneoides CR
** JTL-001 undescribed MX

101

Figure 2.4. (cont'd).

A. umphreyi 1 FL
A. umphreyi 2 AL
A. fulva 18 GA
A. fulva 1 VA
A. fulva 7 NC
A. fulva 4 ON
A. fulva 6 NC
A. fulva 10 MS
A. fulva 11 MA
A. fulva 12 IN
**
A. fulva 17 ON
A. fulva 5 MS
A. fulva 13 IL
A. balcanica GR
A. tennesseensis 1 VA
A. tennesseensis 2 MN
** A. tennesseensis 5 IA
A. tennesseensis 6 NE
**
A. mariae 1 VA
** A. mariae 2 MS
A. japonica JA
A, megommata NV
A. uinta 2 NV

102

Figure 2.4. (cont'd).

A. picea 21 MA
* A. picea 2 MI
A. picea 4 NY
A. picea 3 OH
A. picea 6 MN
A. picea 28 MA
A. picea 36 MA
A. picea 19 MA
A. picea 30 IN
A. picea 31 ON
A. rudis 14 (picea) ON
A. picea 16 NJ
**
A. picea 37 MA
A. picea 27 VA
A. picea 39 MA
A. picea 15 MI
A. picea 23 MI
A. rudis 29 (picea) ON
A. floridana 6 GA
**
A. floridana 7 FL
A. flemingi FL
A. huachucana 4 AZ

103

Figure 2.4. (cont'd).

A. rudis 49 VA
A. rudis 50 GA
A. rudis 51 GA
A. rudis 6 VA
A, rudis 16 OH
A. rudis 43 MO
A. rudis 2 MI
A. rudis 34 PA
A. rudis 4 OH
A. rudis 12 NJ
A. picea 26 NC
A. picea 32 ON
** A. picea 1 MI
A. picea 21 MA

104

Figure 2.4. (cont'd).

A. rudis 49 VA
A. rudis 50 GA
A. rudis 51 GA
A. rudis 6 VA
A, rudis 16 OH
A. rudis 43 MO
A. rudis 2 MI
A. rudis 34 PA
A. rudis 4 OH
A. rudis 12 NJ
A. picea 26 NC
A. picea 32 ON
** A. picea 1 MI
A. picea 21 MA

105

Figure 2.4. (cont'd).

A. rudis 45 IN
A. rudis 10 MN
A. rudis 33 GA
A. rudis 46 NC
A. rudis 8 NJ
A. rudis 15 NC
A. rudis 47 NC
A. rudis 48 NC
A. rudis 19 VA
A. rudis 28 VA
A. carolinensis 2 NC
A. carolinensis 3 NC
A. carolinensis 12 NC
* A. carolinensis 1 MS
A. carolinensis 16 MS
A. miamiana 1 FL
A. miamiana 3 FL
** A. miamiana 2 FL
A. miamiana 5 NC

106

Figure 2.4. (cont'd).

A. rudis 45 IN
A. rudis 10 MN
A. rudis 33 GA
A. rudis 46 NC
A. rudis 8 NJ
A. rudis 15 NC
A. rudis 47 NC
A. rudis 48 NC
A. rudis 19 VA
A. rudis 28 VA
A. carolinensis 2 NC
A. carolinensis 3 NC
A. carolinensis 12 NC
* A. carolinensis 1 MS
A. carolinensis 16 MS
A. miamiana 1 FL
A. miamiana 3 FL
** A. miamiana 2 FL
A. miamiana 5 NC

107

Figure 2.4 (cont'd).

A. ashmeadi 7 NC
** A. ashmeadi 12 NC
** A. ashmeadi 10 NC
A. treatae 1 MI
**
A. treatae 4 AR
A. rudis 41 GA
A. huachucana 1 AZ
A. texana 3 AZ
** A. texana 4 AZ
A. texana 6 AR
A. texana 5 MO
A. texana 7 MO
A. lamellidens 1 VA
A. lamellidens 2 MS
A. rudis 17 NC
A. rudis 32 GA
A. rudis 44 TN
A. rudis 45 IN
A. rudis 10 MN
A. rudis 33 GA
A. rudis 46 NC

108

APPENDIX	
  C:	
  
Tables	
  and	
  Figures	
  for	
  Chapter	
  4	
  

109	
  

Table 3.1. A list of Aphaenogaster species known from North America
Genus Aphaenogaster
Aphaenogaster ashmeadi (Emery 1895)
Aphaenogaster boulderensis Smith 1941
Aphaenogaster carolinensis Wheeler 1915
Aphaenogaster flemingi Smith 1928
Aphaenogaster floridana Smith 1941
Aphaenogaster fulva Roger 1863
Aphaenogaster huachucana Creighton 1934
Aphaenogaster lamellidens Mayr 1886
Aphaenogaster mariae Forel 1886
Aphaenogaster megommata Smith 1963
Aphaenogaster mexicana (Pergande 1896)
Aphaenogaster miamiana Wheeler 1932
Aphaenogaster mutica Pergande 1896
Aphaenogaster occidentalis (Emery 1895)
Aphaenogaster patruelis Forel 1886
Aphaenogaster picea (Wheeler 1908)
Aphaenogaster punctaticeps MacKay 1989
Aphaenogaster rudis Enzmann 1947
Aphaenogaster tennesseensis (Mayr 1862)
Aphaenogaster texana Wheeler 1915
Aphaenogaster treatae Forel 1886
Aphaenogaster uinta Wheeler 1917
Aphaenogaster umphreyi Deyrup & Davis 1998

110	
  

Figure 3.1. Lateral view of Aphaenogaster mariae
showing striae on first gastral tergite.

111

Aphaenogaster rudis

Aphaenogaster huachucana

Aphaenogaster treatae

Aphaenogaster ashmeadi

Figure 3.2. Scape shapes for 4 species of Aphaenogaster.

112

Aphaenogaster mariae

Aphaenogaster fulva

Aphaenogaster tennesseensis

Aphaenogaster rudis

Aphaenogaster boulderensis

Aphaenogaster texana

Aphaenogaster flemingi

Aphaenogaster huachucana

Figure 3.3. Propodeal spine shape and angle for 8 Aphaenogaster species.
113

Aphaenogaster fulva

Aphaenogaster umphreyi

Aphaenogaster rudis

Aphaenogaster megommata

Figure 3.4. Relative eye size for 4 species of Aphaenogaster.
114

Figure 3.5. Lateral, head and dorsal views of Aphaenogaster ashmeadi (Emery).

115	
  

Figure	
  3.6.	
  Lateral and head views of	
  Aphaenogaster boulderensis Smith.

116	
  

Figure 3.7. Lateral, head and dorsal views of Aphaenogaster carolinensis
Wheeler.

117	
  

Figure 3.8. Lateral, head and dorsal views of Aphaenogaster flemingi Smith.
118	
  

Figure 3.9. Lateral, head and dorsal views of Aphaenogaster floridana Smith.

119	
  

Figure 3.10. Lateral, head and dorsal views of Aphaenogaster fulva Roger.

120	
  

Figure 3.11. Lateral, head and dorsal views of Aphaenogaster huachucana
Creighton.

121	
  

Figure 3.12. Lateral, head and dorsal views of Aphaenogaster lamellidens Mayr.

122	
  

Figure 3.13. Lateral, head and dorsal views of Aphaenogaster mariae Forel.

123	
  

Figure 3.14. Lateral, head and dorsal views of Aphaenogaster megommata Smith.

124	
  

(photos from Antweb)	
  

Figure 3.15. Lateral, head and dorsal views of Aphaenogaster mexicana
(Pergande).

125	
  

Figure 3.16. Lateral, head and dorsal views of Aphaenogaster miamiana Wheeler.

126	
  

Figure 3.17. Lateral, head and dorsal views of Aphaenogaster mutica Pergande.

127	
  

Figure 3.18. Lateral, head and dorsal views of Aphaenogaster occidentalis
(Emery).

128	
  

Figure 3.19. Lateral, head and dorsal views of Aphaenogaster patruelis Forel.

129	
  

Figure 3.20. Lateral, head and dorsal views of Aphaenogaster picea (Wheeler).

130	
  

(photo from AntWeb, photo by Jen Fogarty)

Figure 3.21. Lateral, head and dorsal views of Aphaenogaster punctaticeps
MacKay.

131	
  

Figure 3.22. Lateral, head and dorsal views of Aphaenogaster rudis Enzmann.

132	
  

(Mayr).

Figure 3.23. Lateral, head and dorsal views of Aphaenogaster tennesseensis

133	
  

Figure 3.24. Lateral, head and dorsal views of Aphaenogaster texana Wheeler.

134	
  

Figure 3.25. Lateral, head and dorsal views of Aphaenogaster treatae Forel.

135	
  

Figure 3.26. Lateral, head and dorsal views of Aphaenogaster uinta Wheeler.

136	
  

Figure 3.27. Lateral, head and dorsal views of Aphaenogaster umphreyi
Deyrup & Davis.

137	
  

BIBLIOGRAPHY	
  

138	
  

BIBLIOGRAPHY

Attygalle, A.B., Kern, F., Huang, Q., Meinwald, J. (1998). Trail pheromone of the myrmicine
ant Aphaenogaster rudis (Hymenoptera: Formicidae). Naturwissenschaften 85: 38–41.
Bewick, S., Stuble, K.L., Lessard, J.P., Dunn, R.R., Adler, F.R., Sanders, N.J. (2014).
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