Group Title: BMC Biology
Title: Pair of lice lost or parasites regained: the evolutionary history of anthropoid primate lice
CITATION PDF VIEWER THUMBNAILS PAGE IMAGE ZOOMABLE
Full Citation
STANDARD VIEW MARC VIEW
Permanent Link: http://ufdc.ufl.edu/UF00099991/00001
 Material Information
Title: Pair of lice lost or parasites regained: the evolutionary history of anthropoid primate lice
Series Title: BMC Biology
Physical Description: Book
Language: English
Creator: Reed, David
Light, Jessica
Allen, Julie
Kirchman, Jeremy
Publication Date: 2007
 Notes
Abstract: BACKGROUND:The parasitic sucking lice of primates are known to have undergone at least 25 million years of coevolution with their hosts. For example, chimpanzee lice and human head/body lice last shared a common ancestor roughly six million years ago, a divergence that is contemporaneous with their hosts. In an assemblage where lice are often highly host specific, humans host two different genera of lice, one that is shared with chimpanzees and another that is shared with gorillas. In this study, we reconstruct the evolutionary history of primate lice and infer the historical events that explain the current distribution of these lice on their primate hosts.RESULTS:Phylogenetic and cophylogenetic analyses suggest that the louse genera Pediculus and Pthirus are each monophyletic, and are sister taxa to one another. The age of the most recent common ancestor of the two Pediculus species studied matches the age predicted by host divergence (ca. 6 million years), whereas the age of the ancestor of Pthirus does not. The two species of Pthirus (Pthirus gorillae and Pthirus pubis) last shared a common ancestor ca. 3–4 million years ago, which is considerably younger than the divergence between their hosts (gorillas and humans, respectively), of approximately 7 million years ago.CONCLUSION:Reconciliation analysis determines that there are two alternative explanations that account for the current distribution of anthropoid primate lice. The more parsimonious of the two solutions suggests that a Pthirus species switched from gorillas to humans. This analysis assumes that the divergence between Pediculus and Pthirus was contemporaneous with the split (i.e., a node of cospeciation) between gorillas and the lineage leading to chimpanzees and humans. Divergence date estimates, however, show that the nodes in the host and parasite trees are not contemporaneous. Rather, the shared coevolutionary history of the anthropoid primates and their lice contains a mixture of evolutionary events including cospeciation, parasite duplication, parasite extinction, and host switching. Based on these data, the coevolutionary history of primates and their lice has been anything but parsimonious.
General Note: Start page 7
General Note: M3: 10.1186/1741-7007-5-7
 Record Information
Bibliographic ID: UF00099991
Volume ID: VID00001
Source Institution: University of Florida
Holding Location: University of Florida
Rights Management: Open Access: http://www.biomedcentral.com/info/about/openaccess/
Resource Identifier: issn - 1741-7007
http://www.biomedcentral.com/1741-7007/5/7

Downloads

This item has the following downloads:

PDF ( PDF )


Full Text


0
BMC Biology ioMed Central



Research article

Pair of lice lost or parasites regained: the evolutionary history of
anthropoid primate lice
David L Reed*1, Jessica E Light', Julie M Allen1,2 and Jeremy J Kirchman1,2,3


Address: 'Florida Museum of Natural History, University of Florida, Gainesville, Florida 32611, USA, 2Department of Zoology, University of
Florida, Gainesville, Florida 32611, USA and 3New York State Museum, 3140 CEC, Albany, NY 12230, USA
Email: David L Reed* dreed@flmnh.ufl.edu; Jessica E Light jlight@flmnh.ufl.edu; Julie M Allen juliema@ufl.edu;
Jeremy J Kirchman jkirchma@mail.nysed.gov
* Corresponding author



Published: 7 March 2007 Received: 15 September 2006
BMC Biology 2007, 5:7 doi: 10.1 186/1741-7007-5-7 Accepted: 7 March 2007
This article is available from: http://www.biomedcentral.com/1741-7007/5/7
2007 Reed et al; licensee BioMed Central Ltd.
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0),
which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.



Abstract
Background: The parasitic sucking lice of primates are known to have undergone at least 25
million years of coevolution with their hosts. For example, chimpanzee lice and human head/body
lice last shared a common ancestor roughly six million years ago, a divergence that is
contemporaneous with their hosts. In an assemblage where lice are often highly host specific,
humans host two different genera of lice, one that is shared with chimpanzees and another that is
shared with gorillas. In this study, we reconstruct the evolutionary history of primate lice and infer
the historical events that explain the current distribution of these lice on their primate hosts.
Results: Phylogenetic and cophylogenetic analyses suggest that the louse genera Pediculus and
Pthirus are each monophyletic, and are sister taxa to one another. The age of the most recent
common ancestor of the two Pediculus species studied matches the age predicted by host
divergence (ca. 6 million years), whereas the age of the ancestor of Pthirus does not. The two
species of Pthirus (Pthirus gorillae and Pthirus pubis) last shared a common ancestor ca. 3-4 million
years ago, which is considerably younger than the divergence between their hosts (gorillas and
humans, respectively), of approximately 7 million years ago.
Conclusion: Reconciliation analysis determines that there are two alternative explanations that
account for the current distribution of anthropoid primate lice. The more parsimonious of the two
solutions suggests that a Pthirus species switched from gorillas to humans. This analysis assumes that
the divergence between Pediculus and Pthirus was contemporaneous with the split (i.e., a node of
cospeciation) between gorillas and the lineage leading to chimpanzees and humans. Divergence date
estimates, however, show that the nodes in the host and parasite trees are not contemporaneous.
Rather, the shared coevolutionary history of the anthropoid primates and their lice contains a
mixture of evolutionary events including cospeciation, parasite duplication, parasite extinction, and
host switching. Based on these data, the coevolutionary history of primates and their lice has been
anything but parsimonious.







Page 1 of 11
(page number not for citation purposes)







http://www.biomedcentral.com/1741-7007/5/7


Background
Sucking lice (Phthiraptera: Anoplura) are permanent and
obligate ectoparasites of eutherian mammals. These
highly specialized blood-sucking insects live in close asso-
ciation with their hosts and complete their entire life cycle
on the host [1]. Anoplurans have modified mouthparts
for feeding on host blood and because mammalian blood
differs widely among species in terms of its suitability for
louse nutrition [2], sucking lice can be highly host specific
[1,3]. Host specificity could also be reinforced by interac-
tions with the host's immune system. High host specificity
can arise from a long history of cospeciation between
hosts and their parasites. Cospeciation is speciation (or
cladogenesis) in a parasite lineage as a result of, or at the
same time as, host cladogenesis [4]. The current distribu-
tion of parasites on host taxa (the host-parasite associa-
tions) can be the result of cospeciation or various
historical events [5] such as host switching, sorting events
(extinction and lineage sorting), duplication events (par-
asites speciating on a single host lineage), and failure of
the parasite to speciate when the host speciates ('missing
the boat'; [6]). These historical processes can be detected
by comparing the phylogenies of hosts and their parasites
using methodologies such as reconciliation analysis [7].
Previous cophylogenetic studies of lice (both sucking lice
and chewing lice) have documented each of these histori-
cal events in various combinations (for a review, see [8]).

Humans (Homo sapiens) are parasitized by two genera of
sucking lice: one shared with chimpanzees (Pan spp.) and
the other shared with gorillas (Gorilla gorilla). Human
head and body lice, as well as chimpanzee lice, are mem-
bers of the genus Pediculus (Pediculus humanus and Pedicu-
lus schaeffi, respectively). There is no Pediculus species
known to parasitize gorillas. Human pubic lice and gorilla
lice belong to the genus Pthirus (Pthirus pubis and Pthirus
gorillae, respectively), and no Pthirus species is known to
parasitize chimpanzees. Pediculus and Pthirus are sister
taxa based on morphology and molecular data (Figure 1),
and primate lice are known to have cospeciated with their
hosts for at least 25 million years [9]. The curious distri-
bution of these two genera raises an interesting question
regarding the evolutionary history of primate lice. Why do
humans retain both genera, but chimpanzees and gorillas
have only one genus each?

Given what is already known about the coevolutionary
history of the lice and their hosts, we can speculate that
there are two mutually exclusive explanations that can
account for the current distribution of Pediculus and Pthi-
rus (Figure 2). The most parsimonious explanation (i.e.,
the explanation requiring the fewest number of steps) pre-
dicts perfect cospeciation between the primates and lice
with the addition of a single host switch. In this scenario,
the divergence between Pediculus humanus and Pediculus


schaeffi occurred at the same time as the split between
their hosts, humans and chimpanzees, ca. 6 million years
ago (MYA; [10]), and the split between Pediculus and Pthi-
rus occurred contemporaneously with the split between
gorillas and the lineage leading to chimpanzees and
humans (ca. 7 MYA; Figure 2A; [10]). These events were
followed some time later by one host switch of a Pthirus
species from gorillas to humans (Figure 2A). Host switch-
ing among lice is common in many groups of birds and
mammals [11-13]. This 'recent host switch' hypothesis
requires one evolutionary step and predicts that the diver-
gence between Pthirus pubis and Pthirus gorillae is more
recent than the chimpanzee/human split (Figure 2A).
How we might have acquired our pubic lice from gorillas
is not immediately apparent, however it would be inter-
esting to know whether the switch was very recent (say less
than 100,000 years old) or whether it was considerably
older.

The alternative hypothesis involves an ancient louse
duplication event that occurred on the ancestor of goril-
las, chimpanzees, and humans, which would have created
the lineages leading to the two extant genera, Pediculus
and Pthirus (Figure 2B). In this case, the timing of the
divergence between Pthirus pubis and Pthirus gorillae would
correspond to that of their hosts (ca. 7 MYA; [10]). In this
scenario, humans would have retained both genera, but
chimpanzees would have lost a Pthirus species and gorillas
would have lost a Pediculus species to extinction (Figure
2B). Such parasite duplications and extinctions are com-
mon in the lice of birds and mammals (e.g., [14]). This
parasite extinction or 'pair of lice lost' model is less parsi-
monious than the 'recent host switch' hypothesis listed
above because it requires at least three evolutionary steps
(a duplication of the parasite on a primate common
ancestor as well as the extinction of one Pediculus and one
Pthirus lineage, Figure 2B). Although the 'recent host
switch' hypothesis is more parsimonious, it is not neces-
sarily more likely than the 'pair of lice lost' hypothesis.
Each of the historical events (host switch, duplication,
and extinction) has some probability of occurrence, the
quantification of which is beyond the scope of this paper.
There are additional hypotheses that can be postulated
that have more evolutionary steps than the two presented
above, however there are no other hypotheses that have
an equal number of steps or fewer steps. For instance, one
might assume that the current distribution of lice resulted
from a host switch of Pthirus from humans to gorillas (the
opposite direction of the switch in the 'recent host switch'
model). However, this evolutionary scenario would
require at least five evolutionary steps.

The two hypotheses of parasite distributions ('pair of lice
lost' and 'recent host switch') are based on the premise of
maximizing the number of cospeciation events and mini-


Page 2 of 11
(page number not for citation purposes)


BMC Biology 2007, 5:7







http://www.biomedcentral.com/1741-7007/5/7


Figure I
Phylogenetic trees for primate lice and their vertebrate hosts redrawn from Reed et al. [9]. Trees are shown as
cladograms with no branch length information, and are based on molecular and morphological data. Dashed lines between
trees represent host-parasite associations. Humans are unique in being parasitized by two genera (Pediculus and Pthirus). Photo
credits: J. W. Demastes, T. Choe, and V. Smith.


mizing the number of events that deviate from cospecia-
tion, which are typical of analyses that attempt to
reconcile host and parasite associations that are based on
the concept of evolutionary parsimony (reconciliation
analysis as implemented in TreeMap; [7]). However, par-
simony-based reconciliation analyses do not take into
account branch lengths and divergence times informa-
tion that is essential to distinguish between the 'recent
host switch' and 'pair of lice lost' hypotheses. In the pres-
ence of significant cospeciation we can use the timing of
speciation events to differentiate among alternative
hypotheses of host-parasite associations [15 ].

We have performed phylogenetic and cophylogenetic
analyses of two genes, the mitochondrial cytochrome c


oxidase subunit I (CoxI) gene and nuclear gene elonga-
tion factor 1 alpha (EF-lax) gene, to determine the shared
evolutionary history of primate lice and their hosts. We
also investigate the use of standard phylogenetic methods
for reconstructing coevolutionary histories when standard
cophylogenetic methods (e.g., reconciliation analysis)
cannot always find the solution that best fits the observed
data.

Results
Phylogenetic and cophylogenetic analyses
The partition homogeneity test determined that the Cox1
and EF-lax genes did not differ significantly (p = 0.94),
therefore a combined analysis was performed in addition
to analyses based on single genes. The best-fit model for


Page 3 of 11
(page number not for citation purposes)


WW W WW W w


Chimp


*****


Gorilla


w


OW Monkeys


Faehoti


BMC Biology 2007, 5:7


fteeftM ftW te


noaem






http://www.biomedcentral.com/1741-7007/5/7


A Host Switch
* Parasite Duplication
t Parasite Extinction


I


--Pediculus humanus
- Pthirus pubis

Pediculus schaeffi



Pthirus gorillae


Homo


Pan



Gorilla


Pedicinus sp. OW Monkeys


Pediculus humanus
Pthirus pubis


Pediculus schaeffi


t Pthirus sp.


- f Pediculus sp.


Pthirus gorillae


Homo


Pan



Gorilla


Pedicinus sp.


OW Monkeys


Figure 2
Cophylogenetic reconstructions with the host phylogeny for humans, chimpanzees, gorillas, and Old World
monkeys indicated by thick grey lines, and the louse phylogeny indicated by thin red and blue lines. (A) Recon-
struction showing perfect cospeciation between with hosts and parasites with the exception of a single host switch of Pthirus
sp. from gorillas to humans (marked by an arrow). (B) Cophylogenetic reconstruction showing an ancient duplication creating
two evolutionarily distinct lineages (Pediculus and Pthirus), each having cospeciated with gorillas, chimps, and humans with two
extinction events (marked with daggers). The reconciliation shown in panel A requires one evolutionary step (the host switch),
whereas reconciliation B requires three steps (one duplication and two extinctions).




Page 4 of 11
(page number not for citation purposes)


I


BMC Biology 2007, 5:7







http://www.biomedcentral.com/1741-7007/5/7


each individual gene and the combined-gene analysis of
lice was a transversion model (TrN+I+F model) that per-
mitted two nucleotide substitution rates for transversions,
one rate for transitions, unequal base frequencies, a rate
heterogeneity parameter (G), and a parameter for invari-
ant sites (I). Similarly, the best-fit model for the Cox1
gene from the primate host taxa was also TrN+I+F. Maxi-
mum likelihood (ML) analysis of the combined Cox1 and
EF-la dataset (as well as individual gene datasets) from
lice produced a single, well-supported phylogeny in agree-
ment with results from previous analyses (Figure 3; [9]).
When the same ML analysis was performed enforcing a
molecular clock, the resulting tree topology did not
change and the resulting tree score was not significantly
different than the unconstrained score (p = 0.7625). The
ML analysis of the host Cox1 data also resulted in a single
phylogeny in agreement with known relationships among
these primate taxa (Figure 3).

Cophylogenetic analyses using TreeMap [7] produced two
reconciliations of host and parasite phylogenies in agree-
ment with the 'recent host switch' and 'pair of lice lost'
hypotheses presented in Figure 2. The reconciliation con-
cordant with the 'recent host switch' hypothesis (Figure
2A) included five cospeciation events and one host switch
for a total cost of 1.0. The reconstruction concordant with
the 'pair of lice lost' hypothesis (Figure 2B) was less parsi-
monious. While this reconstruction included five cospeci-
ation events, there was also a single duplication event and
two losses resulting in a total cost of 3.0. Both reconcilia-
tions show significantly greater similarity between the
host and parasite trees than would be expected based on
chance alone (i.e., both reconciliations show significant
cospeciation, p < 0.05).

Divergence date estimation
Divergence date estimates differed little between r8s and
multidivtime analyses and between individual and com-
bined genes (Table 1). For convenience we will refer to
divergence date estimates based on the combined gene
tree used in multidivtime (Table 1 and Figure 3). Mean
divergence date estimates for the split between the chim-
panzee and human head/body lice (Pediculus schaeffi and
Pediculus humanus, respectively) averaged 6.39 MYA. The
divergence date estimates for the gorilla and human pubic
lice (Pthirus gorillae and Pthirus pubis, respectively) aver-
aged 3.32 MYA and are noticeably more recent than the
split between the two Pediculus species. The estimated
divergence date for the most recent common ancestor
(MRCA) of the two genera, Pthirus and Pediculus, was esti-
mated to be 12.95 MYA (Table 1 and Figure 3), noticeably
older than the MRCA of chimpanzees, humans, and goril-
las.


Discussion
Reconciliation analysis using TreeMap corroborates ear-
lier reports of significant cospeciation between primate
lice and their hosts [9]. These cophylogenetic analyses
also result in two reconciliations of the host and parasite
phylogenies where the most parsimonious reconstruction
favors the 'recent host switch' hypothesis (Figure 2A).
However, divergence date estimates conflict with the
results of the reconciliation analysis because the 'recent
host switch' hypothesis predicts that the divergence of
Pediculus and Pthirus would be roughly contemporaneous
with the split between gorillas and the lineage leading to
humans and chimpanzees. Our estimates of the MRCA of
Pediculus and Pthirus dates to roughly 13 MYA, not
remotely consistent with the MRCA of humans, chimpan-
zees, and gorillas (ca. 7 MYA; [10]). Given the much older
age of our MRCA of Pediculus and Pthirus, it is more appro-
priate, although less parsimonious, to assume that the ori-
gin of the two genera was the result of a parasite
duplication event rather than a cospeciation event ('pair
of lice lost' hypothesis; Figure 2B). It is curious that the
estimate of the MRCA of Pthirus and Pediculus (13 MYA) is
contemporaneous with the divergence of Orangutans
from other apes [10], however this is possibly coinciden-
tal. Lice do not parasitize orangutans; therefore, recon-
structing their role in the evolutionary history of primate
sucking lice will be difficult.

The 'pair of lice lost' hypothesis is also unsatisfactory
when compared to divergence date estimates. For the 'pair
of lice lost' hypothesis to be correct we must assume that
divergence between Pthirus pubis and Pthirus gorillae is
roughly contemporaneous with the split of gorilla from
the lineage leading to chimpanzees and humans (i.e., ca.
7 MYA). Our divergence date estimates of roughly 3-4
MYA (Table 1) is much younger than the host divergence
of 7 MYA [10] and is even younger than the divergence
between chimpanzees and humans (ca. 6 MYA).

The estimates of divergence dates argue for a more com-
plex evolutionary history than estimated by reconciliation
analysis. While reconciliation analysis serves to find the
most parsimonious reconstruction of host and parasite
evolutionary history by maximizing cospeciation events
and minimizing the cost of the reconstruction, it can only
identify possible scenarios describing the evolutionary
history between associated taxa. Incorporating branch
length data in other analyses is necessary to determine
which scenario best fits the observed data. For example,
post-hoc Mantel tests are commonly used to look for over-
all correlation in host and parasite data sets [16]. How-
ever, we have too few taxa to perform such an analysis,
and we have instead relied upon ad-hoc phylogenetic tests
to determine whether certain nodes of cospeciation were
contemporaneous.


Page 5 of 11
(page number not for citation purposes)


BMC Biology 2007, 5:7







http://www.biomedcentral.com/1741-7007/5/7


Fahrenholziapinnata - - ------------- -PJ'al'


12.95


PihPi i, ,' gorilla - Gorilla
2


Pilii ii,- pubis... Homo
-- Homo =


Pediculus humanus, -


2

- Pediculus schaeffi -- Pa"




Pedicinus hamadryas - - - - Papio ,


Pedicinus badii - - - -Procolobus


Figure 3
Maximum likelihood (ML) phylogeny of primate lice using the combined Cox I and EF- I a dataset with a best-
fit ML model of nucleotide substitution (left). Bootstrap values are indicated below the nodes and divergence estimates
are given above. Clade numbers used in Table I are provided to the right of each node. The ML phylogeny of the Cox gene
from host taxa is indicated on the right. Branch lengths are drawn to the same scale (substitutions/site), and are based on the
best-fit ML model of nucleotide substitution. Dashed lines connect hosts and their associated parasites.


To further examine the validity of the 'pair of lice lost'
hypothesis, we assessed whether the divergence of the two
Pthirus species was contemporaneous with the host diver-
gence of roughly 7 MYA. The branch length between the
two Pthirus species in the best-fit ML louse tree was artifi-
cially lengthened to approximate a branch that is the same
age as the branch between Pediculus schaeffi and Pediculus
humanus, thereby representing the split between chimp
and human lice. It is widely known that the gorilla, chim-
panzees, and human divergence times are very close in
age, so if our estimate is contemporaneous with the diver-
gence of chimpanzees and humans (the most conservative
expected age), then we cannot rule out true contempora-
neous times of divergence between the two Pthirus species
and their hosts. The likelihood score of this constrained


tree (with a branch length approximating a 6 million year
divergence between Pthirus gorillae and Pthirus pubis) was
significantly worse than the true louse tree (where the esti-
mated divergence was closer to 3-4 MYA; d.f. = 6, X2 =
12.91, p = 0.044). We can therefore reject contemporane-
ous divergence events and we can conclude that the split
between the human and gorilla species of Pthirus diverged
much more recently than the split between humans and
gorillas or even humans and chimpanzees. The divergence
within Pthirus is therefore the result of a host switch from
gorillas to humans (loosely defined) roughly 3-4 MYA, as
predicted in the 'recent host switch' hypothesis.

We can similarly examine the divergence date estimation
for the branch between Pthirus and Pediculus, which the


Page 6 of 11
(page number not for citation purposes)


BMC Biology 2007, 5:7


82 3







http://www.biomedcentral.com/1741-7007/5/7


Table I: Divergence date estimates.


EFIa Coxl and EFIla
(r8s, 20 MYA)


Coxl and EFlca Coxl Coxl
(r8s, 25 MYA) (r8s, 20 MYA) (r8s, 25 MYA)


Pedicinus (3) 10.63 (7.08-14.94) 10.23 (6.56-15.17) 10.91 (3.02-18.87)
Pthirus (1) 3.32 (1.84-5.61) 3.86 (2.05-7.49) 1.76 (0.05-6.75)
P. schaeffi and P. 6.39 (3.94-9.96) 6.87 (4.07-11.65) 6.65 (1.72-14.70)
humanus (2)
Pediculus and Pthirus (4) 12.95 (9.42-17.38) 13.03 (9.25-18.21) 14.66 (7.26-22.23)
OWM/Ape Calibration 22.50 (20.13-24.87) 22.39 (20.12-24.84) 22.48 (20.12-24.87)


Mean divergence date estimates (in millions of years) for the clades shown in Figure 3 (see Figure 3 for clade numbers). Divergence estimates using
a 20-25 calibration point for the split between Old World primate lice (Pedicinus) and Anthropoid primate lice (Pediculus and Pthirus) in multidivtime
are given in the first three columns (95% credibility intervals in parentheses). Mean divergence date estimates using 20 MYA and 25 MYA calibration
points for the split between Old World and Anthropoid primate lice using the Langley Fitch model in r8s are indicated in the final four columns for
the Coxl+EFlcI combined analysis and the Coxl gene alone.


'recent host switch' model would predict to be 7 MYA.
Our estimates were much older (Table 1), and shortening
the branch length artificially between Pthirus and Pediculus
to resemble a divergence near 7 MYA can be easily rejected
(p < 0.01). These analyses support an ancient duplication
at that node, consistent with the 'pair of lice lost' model.
Therefore, contrary to the results of the reconciliation
analysis, the divergence date estimates predict a much less
parsimonious explanation of current primate louse distri-
butions: a combination of the 'recent host switch' and
'pair of lice lost' hypotheses.

Given our estimates of divergence dates, the most likely
evolutionary history is that Pthirus and Pediculus diverged
on an ancestor of chimpanzee, human, and gorilla
roughly 13 MYA (a duplication event), with each genus
then having the potential to cospeciate with descendent
hosts (Figure 4). However, only the gorillas retained Pthi-
rus with an extinction of Pthirus on the branch leading to
both humans and chimpanzees (Figure 4). Pediculus was
maintained on the lineage leading to humans and chim-
panzees but lost from the gorilla lineage, and the two
resulting species (Pediculus schaeffi and Pediculus humanus)
diverged in tandem with their primate hosts roughly six
million years ago (Figure 4). Approximately 3-4 MYA, a
Pthirus species switched from the gorilla lineage to the lin-
eage leading to modem humans. It is important to note
that this happened after the divergence of chimpanzees
and humans and that these data suggest humans acquired
their pubic louse from gorillas not recently, but rather 3-
4 million years ago. In total, this coevolutionary scenario
requires four evolutionary steps (one duplication, two
losses, and one host switch), and is a combination of both
the 'recent host switch' and the 'pair of lice lost' hypothe-
ses.

Conclusion
Evidence suggests that Pthirus pubis has been associated
with humans for several million years, and likely arrived
on humans via a host switch from gorillas. Despite the fact


that human pubic lice are primarily transmitted via sexual
contact, such contact is not required to explain the host
switch. Parasites often switch from a given species to a
predator of that species [17], and are sometimes found to
switch to unrelated hosts in communally used areas, such
as roosting or nesting sites [18]. The host switch in ques-
tion could have resulted from any form of contact
between archaic humans and gorillas including, but not
limited to, feeding on or living among gorillas. Regardless
of how the transfer occurred, suitable habitat had to be
available on the new human host for the host switch to be
successful. For example, it is possible that the switch of
Pthirus from gorillas to humans coincides with a change in
available niche space in humans, such as the loss of body
hair. Further study, however, is required to test such a
hypothesis.

Because Pthirus has been associated with humans for sev-
eral million years, this taxon can be examined in the same
way that Pediculus humanus has to study the evolutionary
history of its human host [9,19,20]. Pthirus pubis repre-
sents an independent, ecological replicate that went
through the same evolutionary history on humans as their
head/body lice, and can be used to test predictions made
from Pediculus humanus. Pediculus humanus shows genetic
evidence of population expansion out of Africa roughly
100,000 years ago, which is concordant with host evolu-
tionary history [9]. However, in contrast to the shallow
mitochondrial DNA (mtDNA) gene history of humans
(human mtDNA coalesce to a common ancestor within
200,000 years, [21-23]), Pediculus humanus has three
deeply divergent mtDNA lineages that share a MRCA ca. 2
million years ago, which is far older than the age of their
modem human hosts [9]. Perhaps a worldwide sample of
Pthirus pubis will mirror that of Pediculus humanus, and
show both the population expansion 100,000 years ago
and the three deeply divergent mtDNA lineages. Under-
standing human evolutionary history from the perspec-
tive of its parasites may provide useful insight into a brief
period of history that is not fully recorded in the host fos-


Page 7 of 11
(page number not for citation purposes)


MRCA
(clade number)


Coxi and EFlac


BMC Biology 2007, 5:7







BMC Biology 2007, 5:7



A Host Switch
0 Parasite Duplication
t Parasite Extinction


http://www.biomedcentral.com/1741-7007/5/7


3-4 MY


Pediculus humanus
-- Pthirus pubis


Homo


Pediculus schaeffi Pan


Pthirus gorillae


Gorilla


Pedicinus sp. OW Monkeys


Figure 4
Coevolutionary reconstruction of primate lice and their hosts based on reconciliation analysis and divergence
date estimation. Thick grey lines represent the host phylogeny for humans, chimpanzees, gorillas, and Old World monkeys.
Thin black lines (solid and dashed) represent the louse lineages. This evolutionary scenario depicts a parasite duplication ca. 13
MYA leading to the extant genera Pediculus (solid lines) and Pthirus (dashed lines). One species from each lineage is depicted as
having gone extinct (dagger), and a single host switch ca. 3-4 MYA is shown by an arrow within the Pthirus lineage. The diver-
gence of the chimpanzee and human lice (Pediculus spp.) are shown as having diverged in tandem with their hosts.


sil record or in host DNA [24]. However, if parasites are to
provide much clarity, it will likely be only after many
human parasites have been examined.

The advent of parsimony-based reconciliation analysis
has permitted many researchers to assess phylogenetic
congruence in a wide array of host-parasite assemblages.
However, this method is more limited than Bayesian
approaches [25] to studying cophylogenetics, which eval-
uate not only topological congruence but also the com-
parative timing of host and parasite divergences. It is
imperative that we continue to put into practice the theo-
retical work that has propelled systematists forward in
recent years. Only then can we hope to uncover the more
complex interactions between hosts and parasites.

Methods
Specimen collection and preparation
Samples of Pthirus gorillae (from gorillas), Pthirus pubis
(from humans), Pediculus humanus (from humans), Pedic-
ulus schaeffi (from chimpanzees), Pedicinus hamadryas
(from baboons), Pedicinus badii (from red colobus mon-
keys), and one outgroup species (Fahrenholzia reducta)
were collected for this study (Table 2). Lice in the genus
Pedicinus parasitize only Cercopithecoid monkeys (Old
World Monkeys; OWM) whereas the genera Pediculus and
Pthirus parasitize only the Anthropoid primates (apes). All
lice were preserved in 95% EtOH and stored at -80C.
DNA was extracted from louse specimens using the tech-
nique of Johnson and Clayton [26] and Reed et al. [9],


which enabled extraction of whole genomic DNA from
each louse while retaining the entire louse body as a
voucher specimen. The Qiagen DNeasy Tissue Kit (QIA-
GEN Inc., Valencia, California) was used to isolate
genomic DNA from the body of each louse according to
louse-specific protocols [9,26,27]. After DNA extraction,
lice were mounted on slides and retained as vouchers.
Voucher specimens will be deposited in the Price Institute
for Phthirapteran Research collection (University of
Utah).

PCR and sequencing
PCR amplification and sequencing of a portion of the
mitochondrial cytochrome c oxidase subunit I gene
(CoxI; 858 bp) was performed using the primers
LCO1718 [9] and H7005 [16]. PCR amplification and
sequencing of 345 bp of the nuclear elongation factor 1
alpha (EF-lax) gene were performed using the primers
For3 and Chol0 [28]. Double-stranded PCR amplifica-
tions for both Cox1 and EF-la( were performed in 25 ptl
reaction volumes using 10 ptl of Eppendorf HotMaster
PCR Mix (Fisher Scientific), 1 ptl of each primer (at 10
mM), and 2 ptl of DNA template. The amplification proto-
col required an initial denaturation step of 94C for 10
min, followed by 5 cycles of 94 C (1 min), 48 C (1 min),
and 65 C (2 min), then 30 cycles of 940C (1 min), 52C
(1 min), and 65 C (2 min) and a final extension of 650C
for 10 minutes. Amplified fragments were purified using
ExoSAP-IT (USB Corporation) and sequenced in both
directions. Sequences were edited using Sequencher v.


Page 8 of 11
(page number not for citation purposes)








http://www.biomedcentral.com/1741-7007/5/7


Table 2: Specimens examined. Louse taxa included in phylogenetic and cophylogenetic analyses.


Genbank Accession Numbers


Louse Species


Host Species Voucher ID Collection Locality


Host Identification


CoxI


Pediculus humanus
Pediculus schaeffi
Pthirus pubis
Pthirus gorillae
Pedicinus hamadryas
Pedicinus badii
Fahrenholzia pinnata


Pdcap9.20.05.25
Pdsch5.23.05
Ptpub 1.19.06.3
Ptgor8.1.06.6
Qnham2.4.01.2
Qnbad7.24.06.9
Fzpin I 63


USA, Florida, West Palm Beach
Uganda
UK, Scotland, Glasgow
Uganda
Captive (SW Found for Biomed. Res.)
Uganda
USA, Nevada, Tonopah


Homo sapiens (WP007)
Pan troglodytes
Homo sapiens (GLA 140)
Gorilla gorilla (051122CAWBB00 I)
Papio hamadryas
Procolobus badii
Perognathus longimembris (MLZ 2039)


Abbreviations are as follows: Moore Laboratory of Zoology (MLZ), Page Lab, University of Glasgow (GLA), Lice Solutions, West Palm (WP), and
Maryland Gorilla Veterinary Project (MGVP).


4.2.2 (Gene Codes Corporation, Ann Arbor, Michigan)
and aligned by eye using Se-Al v2.0all http-zl
evolve.zps.ox.ac.uk/Se-Al/Se-Al.html. Primer sequences
were removed and sequences were trimmed in reference
to the translated protein sequence using Se-AL v2.01all
and MacClade 4.0 [29]. All sequences were submitted to
[Genbank: EF152552-EF152564] and alignments to
[TreeBase: #SN3269]. Sequences of the Cox1 gene from
the primate host taxa were downloaded [Genbank:
NC001807, NC001643, NC001645, NC001992,
NC0082191. EF-lax sequences were not available for sev-
eral primate taxa, and were therefore not examined.

Phylogenetic and cophylogenetic analyses
The partition homogeneity test [30] in PAUP *4.0b 10 [31]1
was used to evaluate phylogenetic congruence of the louse
Cox1 and EF-la data sets. One thousand partition repli-
cates were analyzed by maximum parsimony (heuristic
search option with random addition replicates and tree
bisection-reconnection branch swapping). Modeltest [32]
was used to determine the best-fit ML model for the
molecular data. Phylogenetic analyses were conducted on
host and parasite data sets using maximum likelihood
(ML) with branch and bound searches using the best-fit
model in PAUP* 4. Ob 10 [311. Nonparametric bootstraps
(100 replicates) were performed to assess nodal support
for the louse phylogeny. ML searches were performed with
and without the 'enforce clock' constraint in order to test
the hypothesis of a molecular clock in the Cox1 and EF- 1
louse datasets. The resulting ML host and parasite trees
with branch lengths estimated from the best-fit ML model
were then used in cophylogenetic analyses.

TreeMap (v. 2.0.2; [7]) was used to determine whether
host and parasite trees were more similar to one another
than would be expected by chance. Default costs for evo-
lutionary events (codivergence = 0, host switching = 1,
duplication = 1, and loss = 1) were used. Significance val-
ues were calculated from a sample of 1,000 randomly gen-
erated trees.


Divergence date estimation
Because the lice and their primate hosts showed signifi-
cant codivergence and because molecular data did not dif-
fer significantly from clocklike behavior, divergence dates
were estimated using methods that both adhere to and
relax the molecular clock. Divergence dates were esti-
mated in the program r8s [33], using the Langley and
Fitch (LF) model which assumes a molecular clock. Dates
were also estimated using a parametric Bayesian approach
[34] in the program multidivtime. This method relaxes the
molecular clock and allows rate variation among genes
and lineages, and it is therefore appropriate for datasets
that utilize more than one molecular marker. The topol-
ogy resulting from ML analysis of the combined 2-gene
data set was used in multidivtime. The Cox1 best-fit ML tree
and the combined 2-gene topologies were used in r8s.
Divergence dates were estimated using each individual
gene (with branch lengths optimized on the best ML tree)
as well as a combined 2-gene dataset.

For the parametric Bayesian analysis, model parameters
for the F84+F model were estimated for each gene sepa-
rately using the baseml program in PAMLv3.14 [35]1. These
parameters were then used in the program estbranches
[34,36] to estimate the ML and the variance-covariance
matrix of the branch length estimates for each gene.
Lastly, the program multidivtime [34,36], utilizing the out-
put files from estbranches and implementing Markov chain
Monte Carlo (MCMC) sampling, was used to estimate
prior and posterior distribution of rates and divergence
time estimates among lineages. The prior assumption for
the mean and standard deviation of the time of the
ingroup root node (rttm) was set to 3.0 time units, where
1 time unit represents 10 million years. This value corre-
sponds to the upper limit of the split between hominoid
and cercopithecoid primates. The mean and standard
deviation for the prior distribution of the rate of evolution
at the ingroup node (rtrate and rtratesd) was determined
following the protocol of Jansa et al. (rttm; [37]). To avoid
violation of the definition of the prior, rtratesd was set to



Page 9 of 11
(page number not for citation purposes)


EF152552
EF152553
EF152554
EF152555
AY696007
EF152556
EF152557


EF152558
EF152559
EF152560
EF152561
EF152562
EF152563
EF152564


BMC Biology 2007, 5:7








http://www.biomedcentral.com/1741-7007/5/7


its maximum value (equal to rtrate). The Markov chain
was initialized by randomly selecting the initial parameter
value and each Markov chain was sampled every 100
cycles for 1,000,000 generations with a burn in of
100,000 cycles.

A calibration point of 22.5 + 2.5 MYA was used for the
split between Pedicinus and Pediculus+Pthirus. This diver-
gence of 20-25 MYA corresponds to the split between
OWM and apes [38-41]. Since lice and their primate hosts
show significant cospeciation, we can use this well-estab-
lished host calibration based on fossil data to calibrate the
louse phylogenetic trees. It is preferable to use more than
one calibration point when estimating divergence dates
[42,43], however the small number of nodes in our trees
make that impossible. Furthermore, Reed et al. [9] showed
that the calibration point of 20-25 MYA yielded esti-
mated clade ages that were very similar to those estimated
from a calibration of 5-7 million years between the
human and chimpanzee lice (P. humanus and P. schaeffi,
respectively).

Authors' contributions
JEL and JMA collected specimens. JEL, JMA and JJK per-
formed molecular lab work. DLR and JEL analyzed data
and wrote the manuscript. All authors provided com-
ments on initial and final drafts of the manuscript.

Acknowledgements
We wish to thank K. Shepherd at Lice Solutions and D. Clayton at the Uni-
versity of Utah for samples of head lice, Debbie Cox (Chimpanzee Sanctu-
ary, Wildlife Conservation Trust and the Jane Goodall Institute) for samples
of chimpanzee lice, D. Dean (Salt Lake City Public Health Center) for pubic
lice, C. Whittier and M. Cranfield (Maryland Zoo Gorilla Veterinary
Project) for gorilla lice, and K. Rice (Southwest Foundation for Biomedical
Research) for baboon lice. We thank F. K. Barker for assistance with multi-
divtime and two anonymous reviewers for the helpful comments and sug-
gestions. This work was funded in part by The National Science Foundation
(DBI 0102112, DBI 0445712, and DEB 0555024) and the University of Flor-
ida Research Opportunity Fund to DLR.

References
I. Marshall AG: The Ecology of Ectoparasitic Insects. London:
Academic Press; 1981.
2. Murray MD, Nicholls DG: Studies on the ectoparasites of seals
and penguins: 1. The ecology of the louse Lepidophthirus mac-
rorhini Enderlein on the southern elephant seal, Mirounga
leonina (L.). AusJZoolog 1965, 13:437-454.
3. Kim KC, Pratt HD, Stojanovich CJ: The Sucking Lice of North
America. Pennsylvania: Pennsylvania State University; 1986.
4. Brooks DR: Testing the context and extent of host-parasite
coevolution. Syst Zoolog 1979, 28:299-307.
5. Clayton DH, AI-Tamimi S, Johnson KP: The ecological basis of
coevolutionary history. In Tangled Trees: Phylogeny, Cospeciation
and Coevolution Edited by: Page R. Chicago: University of Chicago;
2003:310-341.
6. Paterson AM, Gray RD: Host-parasite cospeciation, host
switching and missing the boat. In Host-Parasite Evolution: General
Principles and Avian Models Edited by: Clayton DH, Moore J. Oxford:
Oxford Univ. Press; 1997:236-250.
7. Charleston MA, Page RDM: TreeMap. v. 2.0.2. Software distributed
by authors 2002.


8. Page RDM, (ed.): Tangled Trees: Phylogenies, Cospeciation,
and Coevolution. Chicago: University of Chicago Press; 2003.
9. Reed DL, Smith VS, Hammond SL, Rogers AR, Clayton DH: Genetic
analysis of lice supports direct contact between modern and
archaic humans. PLoS Biol 2004, 2(I I):e304.
10. Stauffer RL, Walker A, Ryder OA, Lyons-Weiler M, Hedges SB:
Human and ape molecular clocks and constraints on paleon-
tological hypotheses. J Hered 2001, 92(6):469-474.
I I. Barker SC, Close RL: Zoogeography and Host associations of
the Heterodoxus octoseriatus group and H. ampullatus
(Phthiraptera: Boopiidae) from rock wallabies (Marsupialia:
Petrogale). Int J Parasitol 1990, 20(8): 1081-1087.
12. Weckstein JD: Biogeography explains cophylogenetic patterns
in toucan chewing lice. Syst Biol 2004, 53(1):154-164.
13. Johnson KP, Adams RJ, Clayton DH: The phylogeny of the louse
genus Brueelia does not reflect host phylogeny. BiolJ Linnean
Soc 2002, 77(2):233-247.
14. Clayton DH, Price RD: Taxonomy of New World Columbicola
(Phthiraptera: Philopteridae) from the Columbiformes
(Aves), with descriptions of five new species. Ann Ent Soc Am
1999, 92(5):675-685.
15. Percy DM, Page RDM, Cronk QCB: Plant-insect interactions:
double-dating associated insect and plant lineages reveals
asynchronous radiations. Sys Biol 2004, 53(1):120-127.
16. Hafner MS, Sudman PD, Villablanca FX, Spradling TA, Demastes JW,
Nadler SA: Disparate rates of molecular evolution in cospeci-
ating hosts and parasites. Science 1994, 265(5175): 1087-1090.
17. Whiteman NK, Santiago-Alarcon D, Johnson KP, Parker PG: Differ-
ences in straggling rates between two genera of dove lice
(Insecta: Phthiraptera) reinforce population genetic and
cophylogenetic patterns. IntJ Parasitol 2004, 34:1 I I113-1 I 119.
18. Johnson KP, Weckstein JD, Witt CC, Faucett RC, Moyle RG: The
perils of using host relationships in parasite taxonomy: phyl-
ogeny of the Degeeriella complex. Mol Phylog Evol 2002,
23(2):150-157.
19. Kittler R, Kayser M, Stoneking M: Molecular evolution of Pedicu-
lus humanus and the origin of clothing. Curr Biol 2003,
13:1414-1417.
20. Ashford RW: Parasites as indicators of human biology and
evolution. J Med Microb 2000, 49(9):771-772.
21. Cann RL, Stoneking M, Wilson AC: Mitochondrial DNA and
human evolution. Nature 1987, 325:31-36.
22. Vigilant L, Stoneking M, Harpending H, Hawkes K, Wilson AC: Afri-
can populations and the evolution of human mitochondrial
DNA. Science 1991, 253:1503-1507.
23. Ingman M, Kaessmann H, Paabo S, Gyllensten U: Mitochondrial
genome variation and the origin of modern humans. Nature
2000, 408:708-713.
24. Whiteman NK, Parker PG: Using parasites to infer host popula-
tion history: a new rationale for parasite conservation. Anim
Conserv 2005, 8:175-18 1.
25. Huelsenbeck JP, Rannala B, Larget B: A Bayesian framework for
the analysis of cospeciation. Evolution 2000, 54(2):352-364.
26. Johnson KP, Clayton DH: Coevolutionary history of ecological
replicates: comparing phylogenies of wing and body lice to
columbiform hosts. In Tangled Trees: Phylogeny, Cospeciation and
Coevolution Edited by: Page R. Chicago: University of Chicago;
2003:262-286.
27. Cruickshank RH, Johnson KP, Smith VS, Adams RJ, Clayton DH, Page
R: Phylogenetic analysis of partial sequences of elongation
factor I-alpha identifies major groups of lice (Insecta:
Phthiraptera). Mol Phylog Evol 2001, I 9(2):202-215.
28. Danforth BN,Ji S: Elongation factor-I a occurs as two copies in
bees: implications for phylogenetic analysis of EFIa
sequences in insects. Mol Biol Evol 1998, 15:225-235.
29. Maddison WP, Maddison DR: MacClade: Analysis of phylogeny
and character evolution. v. 4.05. Sunderland, MA: Sinauer; 2002.
30. Farris JS, Kallersjo M, Kluge AG, Bult C: Testing significance of
congruence. Cladistics 1994, 10:315-320.
31. Swofford DL: PAUP*. Phylogenetic Analysis Using Parsimony
(*and Other Methods). v. 4bl0. Sunderland, Massachusetts: Sin-
auer Associates; 2002.
32. Posada D, Crandall KA: MODELTEST: Testing the model of
DNA substitution. Bioinformatics 1998, 14:817-818.





Page 10 of 11
(page number not for citation purposes)


BMC Biology 2007, 5:7








http://www.biomedcentral.com/1741-7007/5/7


33. Sanderson MJ: r8s: inferring absolute rates of molecular evolu-
tion and divergence times in the absence of a molecular
clock. Bioinformatics 2003, 1 9(2):301-302.
34. Thorne JL, Kishino H: Divergence time and evolutionary rate
estimation with multilocus data. Systc Biof 2002, 51 (5):689-702.
35. Yang ZH: PAML: a program package for phylogenetic analysis
by maximum likelihood. Comp App Biosc 1997, 13(5):555-556.
36. Kishino H, Thorne JL, Bruno WJ: Performance of a divergence
time estimation method under a probabilistic model of rate
evolution. Mol Biof Evol 2001, 18(3):352-36 1.
37. Jansa SA, Barker FK, Heaney LR: The pattern and timing of diver-
sification of Philippine endemic rodents: Evidence from
mitochondrial and nuclear gene sequences. Syst Biol 2006,
55(1):73-88.
38. Young NM, MacLatchy L: The phylogenetic position of Moroto-
pithecus. j Hum Evo 2004, 46(2):163.
39. Maclatchy L: The oldest ape. EvolAnthropo 2004, 13(3):90-103.
40. Steiper ME, Young NM, Sukarna TY: Genomic data support the
hominoid slowdown and an Early Oligocene estimate for the
hominoid-cercopithecoid divergence. Proc Not! Acad Sci USA
2004, 101(49):17021-17026.
41. Kumar S, Filipski A, Swarna V, Walker A, Hedges SB: Placing confi-
dence limits on the molecular age of the human-chimpanzee
divergence. Proc Not! Acad Sci USA 2005, 102(52):18842-18847.
42. Porter ML, Perez-Losada M, Crandall KA: Model-based multi-
locus estimation of decapod phylogeny and divergence
times. Mol Phylogen Evol 2005, 37(2):355-369.
43. Lee MSY: Molecular clock calibrations and metazoan diver-
gence dates. j Mol Evol 1999, 49(3):385-391.


Page 11 of 11
(page number not for citation purposes)


Publish with BioMed Central and every
scientist can read your work free of charge
"BioMed Central will be the most significant development for
disseminating the results of biomedical research in our lifetime."
Sir Paul Nurse, Cancer Research UK
Your research papers will be:
available free of charge to the entire biomedical community
peer reviewed and published immediately upon acceptance
cited in PubMed and archived on PubMed Central
yours you keep the copyright
Submit your manuscript here: BioMedcentral
http://www.biomedcentral.com/info/publishing adv.asp


BMC Biology 2007, 5:7




University of Florida Home Page
© 2004 - 2010 University of Florida George A. Smathers Libraries.
All rights reserved.

Acceptable Use, Copyright, and Disclaimer Statement
Last updated October 10, 2010 - - mvs