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ReviewCite this article: Compton AA, Malik HS,

Emerman M. 2013 Host gene evolution traces

the evolutionary history of ancient

primate lentiviruses. Phil Trans R Soc B 368:

20120496.

http://dx.doi.org/10.1098/rstb.2012.0496

One contribution of 13 to a Theme Issue

‘Paleovirology: insights from the genomic fossil

record’.

Subject Areas:evolution, genetics, microbiology, immunology

Keywords:lentivirus, paleovirology, simian

immunodeficiency virus, human

immunodeficiency virus, primates

Author for correspondence:Michael Emerman

e-mail: memerman@fhcrc.org

& 2013 The Author(s) Published by the Royal Society. All rights reserved.

Host gene evolution traces theevolutionary history of ancientprimate lentiviruses

Alex A. Compton1,2, Harmit S. Malik3,4 and Michael Emerman2,3

1Molecular and Cellular Biology Graduate Program, University of Washington, Seattle, WA 98195, USA2Division of Human Biology, 3Division of Basic Sciences, and 4Howard Hughes Medical Institute,Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA

Simian immunodeficiency viruses (SIVs) have infected primate species long

before human immunodeficiency virus has infected humans. Dozens of

species-specific lentiviruses are found in African primate species, including

two strains that have repeatedly jumped into human populations within the

past century. Traditional phylogenetic approaches have grossly underesti-

mated the age of these primate lentiviruses. Instead, here we review how

selective pressures imposed by these viruses have fundamentally altered the

evolutionary trajectory of hosts genes and, even in cases where there now

remains no trace of the viruses themselves, these evolutionary signatures can

reveal the types of viruses that were once present. Examination of selection

by ancient viruses on the adaptive evolution of host genes has been used to

derive minimum age estimates for modern primate lentiviruses. This type of

data suggests that ancestors of modern SIV existed in simian primates more

than 10 Ma. Moreover, examples of host resistance and viral adaptation have

implications not only for estimating the age and host range of ancient primate

lentiviruses, but also the pathogenic potential of their modern counterparts.

1. Human lentivirus infections emerge from a brimmingreservoir with unclear boundaries

Simian immunodeficiency virus (SIV) is the collective name for a diverse group of

lentiviruses that naturally circulate among African non-human primate species.

With over 40 species-specific strains characterized to date and high seroprevalence

rates among reproductive adults, SIV amounts to an immense viral reservoir with

zoonotic potential (figure 1; [1]). In the early twentieth century, two new lentivirus

infections emerged in human populations. Human immunodeficiency virus type 1

(HIV-1) emerged via cross-species transmissions of SIVcpz from chimpanzees and

gorillas; four independent transmission events gave rise to HIV-1 groups M, N, O

and P. A separate lentiviral lineage, HIV-2, emerged in humans at least eight times

via cross-species transmissions from SIVsmm, found in sooty mangabeys (figure

1) [3]. Therefore, spillover from the pool of non-human lentiviruses can be both

recurrent and hazardous for human health. In fact, despite the species specificity

attributed to each SIV strain, transmissions of SIV between non-human primates

have been documented on numerous occasions and may lead to pathogenic out-

comes in these hosts as well (figure 1).

HIV-1 causes a chronic infection in humans. Nearly all infected individuals

progress to a state of life-threatening immunodeficiency (AIDS) if not treated

with antiretroviral therapy. A recent study spanning nearly 10 years showed

that chimpanzees infected with SIVcpz, the ancestor of HIV-1, also suffer from

decreased lifespan and immunopathologies characteristic of AIDS [4–6]. SIVcpz

is a recombinant lentivirus that arose following cross-species virus transmission(s)

from other ‘natural host’ species that comprise the SIV reservoir [7]. By contrast to

infected humans and chimps, the major predictors of disease progression (loss of

the CD4þ T-cell compartment and chronic immune activation) are notably absent

in natural hosts infected by SIV in the wild [8–11]. The mechanisms by which these

marmosets

tamarins

capuchins

squirrel monkeys

owl monkeys

sakis

howler monkeys

woolly monkeys

Macaques SIVmac, SIVstm

mangabeys SIVsm, SIVrcm

mandrills SIVmnd-1, SIVmnd-2

baboons SIVagm

AGM, guenons SIVagm, SIVmus, etc.

colobus monkeys SIVcol, SIVwrc, SIVolc

Proboscis, leaf monkeys

gibbons

orangutans

chimpanzees SIVcpz

bonobos

humans HIV-1, HIV-2

gorillas SIVgor

tarsiers

New Worldmonkeys

Old Worldmonkeys

hominoids

Colobinae

Cercopithecinae

lemurs

puffed lemurs

aye ayes

Indris, Sifaws

mouse, dwarf lemurs pSIV

bushbabies

lorises

prosimians

Figure 1. A representative phylogeny of primate species and their lentivirus associations. The evolutionary relationship between living primate species is shownwith branching orders reflecting the currently accepted species phylogeny [2]. The Old World Monkeys are subdivided into the Cercopithecinae and Colobinae sub-families. Lentiviruses naturally associated with a given species (green in the online version) include SIVsm/SIVrcm in mangabeys, SIVmnd-1/SIVmnd-2 in mandrills,SIVagm/SIVmus in AGM/guenon, and SIVcol/SIVwrc/SIVolc in colobus monkeys. Lentiviruses arising via cross-species transmission (red in the online version) includeSIVmac/SIVstm in macaques, SIVagm in baboons, SIVcpz in chimpanzees, HIV-1/HIV-2 in humans, and SIVgor in gorillas. Dashed lines depict cross-species virustransfer; lines originate from the donor species and terminate in an arrowhead at the recipient species. pSIV (blue in online version) is an endogenous primatelentivirus identified in the genomes of some lemur species. (Online version in colour.)

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primates remain AIDS-free despite lifelong infection and high

viremia have major implications for our understanding of

HIV-1 pathogenesis [11]. Furthermore, the lack of disease

progression is commonly attributed to coevolution between

the host and the virus, but the time-scale appropriate to describe

these relationships remains controversial.

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In some instances, closely related primate species also

harbour closely related, species-specific lentivirus strains.

Such cases support a scenario of host–virus cospeciation, in

which viral lineages diverge in parallel with diverging host

populations. However, in other instances, lentivirus phyl-

ogeny is discordant with that of their respective hosts.

These discordances reveal that host switching events (cross-

species transmission) can also give rise to lentivirus–host

relationships that must be younger by comparison [12,13].

Adding to the uncertain history of lentiviruses in primates is

the fact that current phylogenetic approaches employing a

molecular clock appear inadequate to reconstruct virus evol-

ution in deep time [14,15]. For example, previous attempts at

assigning a precise date of birth to the SIVsmm lineage have

yielded an age estimate of just centuries, orders of magnitude

less than what would be expected under conditions of host–

virus coevolution [16]. Reasons for this discrepancy likely

include the erosion of phylogenetic signal from viral sequences

by purifying selection [17] and by convergence on similar viral

genotypes (site saturation) over time [18].

Calibration of these time-scales is greatly aided by other

temporal markers. For instance, the discovery of SIV strains

in species endemic to Bioko, an island which became geo-

graphically isolated from the African mainland at least 10 000

years ago, provided a means to more accurately calibrate the

molecular clock phylogenetic analysis [19]. This led to the mini-

mum estimate of the time to most recent common ancestor of

SIV to be pushed back to 32 000 years, although it is likely

that this date still considerably underestimates the age of

the SIV lineage [19]. Because phylogenetic methods based

on viral sequences alone have failed to bridge the time gap

between ancestral viruses and their modern counterparts,

there is a keen interest in alternative approaches which estimate

the evolutionary age of virus families. By unearthing both

endogenous viral imprints in host genomes and instances of

virus-driven evolution of the host genome, we focus our

review on how such approaches have led to an appreciation

for the true antiquity of lentiviruses and their impact on

primate hosts.

2. Paleovirology: endogenous viruses andvirus-driven evolution of host immunity

Roughly 5–8% of the human genome consists of endogenous

retroviruses, the remnants of ancient, exogenous viruses that

integrated into the germline (reviewed in [20]). The availability

of sequenced genomes from a variety of organisms has led

to the discovery of additional endogenous viral elements

(EVEs), revealing that several virus families with relevance

to human health have far more distant origins than previ-

ously appreciated [21,22]. Dating of these EVEs does not rely

on using molecular clocks of virus evolution, but rather on

well-established dates of host species divergence. One such

endogenous lentiviral ‘fossil’ uncovered in the genomes of

Malagasy prosimians (mouse and dwarf lemurs, figure 1) pro-

vided the first genetic evidence of an early association between

primates and lentiviruses. This germline integration event

called prosimian SIV (pSIV), establishes that lentiviruses were

circulating among some primates at least 4 Ma [23,24]. pSIV is

a transitional viral form, sharing some aspects of genomic archi-

tecture with non-primate lentiviruses (such as dUTPase) while

lacking genes typically found in extant primate lentiviruses

(such as vpr) [23]. However, no trace of an endogenous

lentivirus-like entity has been identified in the genomes of

simian primates, which comprise the hosts for the known,

extant primate lentiviruses. Thus, despite its utility for other

viruses and for lentiviruses in other mammals [18,22], the

‘fossil’ record of viral imprints has been mostly uninformative

of the lentiviruses harboured by modern simian primates.

On the other hand, even in the absence of direct evidence

of past infections, ancient viruses can also be detected

indirectly by examining how they have driven adaptive

changes in antiviral immunity over time [25]. The discovery

of an extremely diverse set of innate immune genes with lenti-

virus-blocking potential, known as host restriction factors, has

provided a means to understand how lentiviruses impact

host evolution [26]. The rate at which non-synonymous

mutations accumulate between species is a classic measure of

adaptive evolution. Many restriction factors are evolving

under selection, and in fact it has been proposed that this is a

criterion by which they should be defined [26]. Furthermore,

because many of these cellular factors are directly antagonized

by the lentiviruses they have evolved to restrict, adaptive evo-

lution at the host–virus interface signals when lentivirus

infections likely occurred in the past [25]. Selective pressure

applied by virus-encoded antagonists will select adaptive

‘escape’ mutations in host factors at sites critical for the host–

virus interaction (figure 2a). Adaptive evolution of the host is

followed by virus counter-evolution in order to re-establish

the interaction, inciting a perpetual ‘genetic conflict’ character-

ized by host–virus antagonistic coevolution (figure 2a). (See

[27] for more comprehensive review of positive selection as a

result of virus–host interactions.)

Analysis of amino acid changes in restriction factor genes

isolated from a broad panel of extant primates, representing

tens of millions of years of evolution, provides a temporal

axis to determine when cases of virus-driven evolution

occurred in the past (figure 2b). Within the framework of the

host phylogeny, signatures of selection can be read like a gen-

etic ledger, documenting when a given host population

was exposed to a lentivirus threat. In order to gain specific

insights into the age and provenance of primate lentiviruses,

we focus only on restriction factors with antiviral specificity

for lentiviruses, which is assumed to reflect selection by ancient

lentiviral infections. Alternatively, even if the restriction factor

acts generally on a wider class of viruses or retroelements, the

positive selection of residues at known lentivirus-interacting

surfaces (figure 2c) permits deconvolution of the multiple

selective forces that may have acted on it and on the phylogeny

(figure 2d). These criteria allow us to discern positive selec-

tion signatures that are likely to have been driven exclusively

by lentiviruses.

As primate genomes encode a diverse arsenal of antiviral

effectors with lentivirus-blocking potential, SIV has left its

mark on the evolution of several host restriction factors that

block the viral life cycle at different stages. We discuss restriction

factors that serve as examples of lentivirus-driven evolution.

The analysis of the host restriction factor APOBEC3G

(A3G) has provided the most precise details in calculating

the age of ancient primate lentiviruses. A3G is an antiviral

cytidine deaminase that incorporates itself into virions and

mutates viral DNA produced during reverse transcription

[28]. While many retroviruses have evolved mechanisms to

evade or directly antagonize the activity of A3G [29–31],

the primate lentiviruses devote an entire accessory gene, vif,

time

restrictionfactor

(a) (b)

(c) (d )

virus-encodedantagonist

time

primate speciesphylogeny

lentiviralantagonist

non-lentiviralantagonists

time

***

** ***

***

***

***

Figure 2. Virus-driven evolution fuels a genetic conflict. (a) Depicted is a cycle of host – virus coevolution initiated when an antagonist encoded by a pathogenicvirus counteracts a host restriction factor. Host factor variants are drawn in solid black, and viral antagonists are drawn in white. Selection of host factor variants thatsuccessfully evade antagonism is followed by counter-selection of viral variants that re-establish the host – virus interaction, applying selective pressure on the hostonce again, resulting in a cycle of coevolution. Note that the ‘direction’ of the arms race can be reversed, such that host antiviral effectors ‘chase’ their viral targets.(b) The age of host genetic adaptations, and consequently of the causal pathogenic viruses, can be estimated when superimposed onto the primate species phy-logeny. For instance, the origins of positive selection on primate SAMHD1 appear to coincide with the birth of SAMHD1-degrading activity specific to somelentiviruses. (c) In cases where a particular restriction factor can be generally antagonized by a variety of different antagonists, information about binding sitescan nevertheless delineate those amino acid changes that occurred on account of lentiviruses rather than due to antagonists encoded by other viruses (grey shading).(d ) By tracing these lentiviral-specific changes on the primate phylogeny, one can infer whether one or several branches in primate phylogeny underwent selectionon account of pathogenic lentiviruses. For instance, despite being antagonized by several viruses, changes in APOBEC3G required to evade lentiviral Vif can be usedto calibrate Vif-escaping adaptive episodes in primate APOBEC3G. (Online version in colour.)

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to this purpose [32]. Vif binds A3G directly and links it to a

cellular E3 ubiquitin ligase to accelerate its turnover at the

proteasome [33–36]. Vif originated early during lentivirus

evolution and has been retained by every member of Lentivir-idae except equine infectious anaemia virus [18,37]. Therefore,

antagonism of primate A3G and its homologues in mammals

may be crucial for long-term lentivirus persistence, making it

an ideal candidate for studies of lentivirus-driven evolution.

An initial analysis of A3G evolution in primates exposed sig-

nals of selection throughout its length, suggesting that many

surfaces of the enzyme have been engaged by viral antagon-

ists of unknown origin [38]. However, a subsequent study of

A3G orthologues from a broad panel of Old World monkeys

(OWMs), including African green monkey (AGM) and sev-

eral other species comprising the SIV reservoir, revealed

recurrent selection at residues involved in the interaction

with the lentiviral Vif protein [39,40]. Single amino acid sub-

stitutions that allow evasion of Vif-mediated degradation

independently emerged on several occasions during primate

evolution, the oldest of which appeared at least 5 Ma in the

common ancestor of the guenon species. An even older Vif-

blocking adaptation in A3G, a multi-residue insertion event,

arose at least 12 Ma in a subset of OWMs (Colobinae) [39].

These findings suggest that, millions of years ago, lentiviruses

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left footprints of selection stamped in the A3G locus of simian

primates. Therefore, an examination of virus-driven evolution

of host immunity enables the inference of past lentivirus

infections in much deeper time-scales.

Similar inferences can be made from the evolution of the

newly discovered restriction factor SAMHD1, which imposes

a block to the viral reverse transcription in myeloid-derived

blood cells and resting CD4þ T cells [41]. Evolutionary analysis

of primate SAMHD1 revealed residues evolving under positive

selection that underlie interaction with and sensitivity to the

lentiviral-encoded antagonists Vpr and Vpx [42,43]. Interest-

ingly, mapping these signatures onto the primate phylogeny

found that the selection is most pronounced in members of

the Cercopithecinae lineage (figure 1), the OWM subfamily that

includes a large component of natural host species including

AGMs and sooty mangabeys. Taken together with functional

analyses demonstrating that SAMHD1-antagonizing activity

first arose in Vpr/Vpx from lentiviruses that infect the Cerco-pithecinae lineage, these results suggest that lentiviral pressure

led to both the origin and persistence of rapid evolution in pri-

mate SAMHD1. Dating this origin of positive selection using the

host species tree suggests that lentiviruses with the ability to

degrade SAMHD1 are at least 10 Myr old within primates.

Tetherin (also called BST-2) is a broadly acting antiviral

effector that ‘tethers’ virions to the cell surface to disallow

their release and spread. The non-specific antiviral mechan-

ism employed by Tetherin allows it to restrict a wide array

of enveloped virus families, including retroviruses, flavi-

viruses, herpesviruses and paramyxoviruses [26]. Despite

this apparent lack of specificity, a lentiviral-driven conflict

appears to have played a prominent role during the evo-

lutionary history of Tetherin in primates. Sequencing the

Tetherin locus from representatives of major primate clades

revealed that it is undergoing positive selection (adaptive

evolution) [44,45]. In particular, the most rapidly evolving

Tetherin residue in an analysis of OWM species maps pre-

cisely to one of the amino acids in the cytoplasmic tail of

Tetherin which confers species specificity to antagonism by

SIV Nef [45–48]. Nef downregulates Tetherin from the cell

surface [46], and presumably binds directly to these amino

acids in the cytoplasmic tail of Tetherin. This suggests that

the repeated amino acid substitutions at this single residue

were driven to allow escape from Nef antagonism [45]. How-

ever, because other viruses also encode Tetherin antagonists,

it is possible that another antagonist with the same specificity

as Nef was responsible for this host gene selection. Thus, a

caveat of using the host gene evolution to date paleoviruses

is that it may not be possible to determine which virus

family drove the ancient selection. However, when selection

converges on several antiviral factors inside the cell, as is

the case for the paleo-lentiviruses in primates, it increases

the confidence that an ancient virus that encoded Vif, Vpx/

Vpr and Nef was responsible for selection on multiple host

genes in a similar time period.

Finally, the analysis of the ‘birth’ of an antiviral gene

formed between the restriction factor TRIM5 and CypA,

called TRIM-CypA, supports the existence of an even older

lentivirus within primates [49]. Rhesus macaques (Macacamulatta) encode a TRIM-CypA fusion protein that emerged

in the ancestor of macaque species 5–6 Ma [50–52]. While

rhesus TRIM5a potently blocks HIV-1 replication, TRIM-

CypA exhibits specificity towards HIV-2 and SIVagm,

suggesting that it was selected millions of years ago in

response to a lentivirus with similar characteristics [53,54]. In

a remarkable demonstration of convergent evolution, a TRIM-

CypA fusion has evolved independently in owl monkeys

(Aotus), a genus endemic to South America, establishing that

the species of the New World may have been subjected to len-

tiviruses as well [55,56]. More recent analyses uncovered an

even older TRIM-CypA fusion protein, which was present in

the ancestor of all simian primates 43 Ma, and appears to

have been active for 10 Myr before undergoing decay and loss

of function [49]. If ‘birth’ of this older TRIM-CypA gene was

also driven by selective pressure by a lentivirus, then this

implies that lentiviruses have episodically infected simian pri-

mates for most of their evolutionary history. Thus, although

each method of detecting paleoviruses has its advantages and

shortcomings (reviewed in [15,21, 25,57]), the use of host gene

evolution to infer ancient viral infections has the potential to

identify past pathogenic challenges, where any direct traces of

the virus are long gone.

3. Host polymorphism as a readout for recentvirus-driven evolution

Most examples of lentivirus-driven selection we have discus-

sed so far examine interspecies divergence, presumably fixed

mutations that distinguish the antiviral gene of one species

from another. However, the selection can also manifest as vari-

ation (known as polymorphism) within a species or within very

closely related species, which provides a powerful means to

assess ongoing or very recent selective pressures. When applied

to the evolution of antiviral immunity, polymorphism may

result from relatively recent instances of virus-driven evolution

because the variation has not yet been purged or fixed within

the host population.

Analyses of gene diversity within AGMs (genus Chloro-cebus), a highly abundant natural host species of lentiviral

infections, have turned up polymorphisms in genes that

have relevance to ongoing and ancient virus–host evolution.

AGMs comprise four major subspecies known commonly as

vervet, tantalus, grivet and sabaeus monkeys, each occupying

distinct geographies in sub-Saharan Africa. Each AGM sub-

species harbours a distinct SIV strain (collectively referred

to as SIVagm). In two of those subspecies, single amino acid

changes in A3G emerged independently to allow evasion of

the lentiviral antagonist, Vif. Specifically, these adaptive

mutations prevent degradation by Vif from heterologous

SIVagm isolates (those found in the other populations),

suggesting that they are selected as a result of Vif-mediated

selective pressure [40].

Earlier findings of polymorphism in the chemokine

receptor gene CCR5 of AGM provide further evidence of lenti-

virus-driven selection in this natural host species. The CCR5 is

used by HIV/SIV as a co-receptor to enter host cells. Non-

synonymous mutations in CCR5 of AGM subspecies cluster

in regions targeted by the viral envelope glycoprotein; this

variation inhibits SIVagm infection in vitro [58]. Interestingly,

the primary receptor for lentivirus entry, CD4, is also poly-

morphic in AGM [13] and is undergoing adaptive evolution

in primates at positions that govern virus entry [59]. Together,

signatures of recent selection in AGM suggest that both host

factors with antiviral functions and host factors that are essen-

tial for virus replication are subjected to ongoing evolutionary

change during the course of host–virus coevolution.

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Whereas the variation between species allows us to infer

the presence of ancient, pathogenic lentiviruses among pri-

mate ancestors, selection resolved to the population level

hints that contemporary lentiviruses other than SIVcpz may

also impact the fitness of their respective hosts. Indeed,

cases of AIDS, while rare, have been documented in SIV-

infected AGMs and sooty mangabeys in captivity [60,61].

Furthermore, cross-species transmission of apparently non-

pathogenic SIV strains, both in the laboratory and in the

wild, can induce disease in the non-natural host [62,63].

Even in the absence of full-blown AIDS, SIV infections may

have subtler, unrecognized consequences that, nevertheless,

seriously impact host fitness. For example, infection may

affect either reproductive rate, as observed in SIV-infected

chimpanzees [4], or social behaviours such as mate choice.

Long-term field studies of SIV-infected natural hosts are

needed to better understand the impact of wild lentivirus

infections on these and other parameters of host fitness.

20120496 4. Protective polymorphism: tripping up a viralopponent that cannot be outrun

The lentivirus genome has a mutation rate that is one million

times faster than that of its host [64]; so, primate species must

overcome a severe handicap in order to survive. Nonetheless,

it has been suggested that the presence of multiple host geno-

types may insulate populations from invading viruses by

slowing the rate of viral adaptation [65]. In fact, immune

polymorphism will not only benefit populations but also

individuals [66]. A potential protective role for heterozygos-

ity at the A3G gene is made possible by examining SIV

evolution following experimental infection of AGMs with

differing A3G genotypes.

As described earlier, the sabaeus population of AGM

encodes a unique, derived allele of A3G (D130H) that is

resistant to Vif from SIVagm strains infecting other AGM

populations. Using plasma samples isolated from three

sabaeus monkeys experimentally infected with SIVagm.Ver

(a virus naturally found in vervets) [67], we monitored the

sequence evolution in vif. Vif readily counteracted A3G in

individuals homozygous for the ancestral A3G allele. In ani-

mals that were homozygous for the A3G (D130H) allele that

was resistant to Vif from the initial virus inoculum, the vifgene evolved within the animal in order to counter this

A3G polymorphism [40]. However, in a sabaeus monkey

that was heterozygous, encoding an ancestral A3G allele

and a derived A3G (D130H) allele, SIVagm.Ver vif failed to

evolve the capacity to antagonize both variants in vivo(figure 3a) [40]. These findings suggest that the virus adap-

tation is dramatically impeded when two variants of a host

factor are present inside the cell.

Upon closer examination, the mechanism of constraint

imposed by A3G heterozygosity becomes clear. Adaptive

mutations selected in vivo enabled Vif to antagonize derived

A3G, but as a result, Vif lost activity against the ancestral

form (figure 3b, compare parental Vif with adapted Vif ).

Reversion of just one amino acid selected in Vif (Y84C)

resulted in a protein with the opposite specificity (figure 3b,

compare adapted Vif with adapted Vif þ C84Y), demon-

strates that this residue toggles the substrate specificity of

Vif. Therefore, virus adaptation to A3G is an evolutionary

trade-off, because evolving to target a variant of the host

factor compromises its ability to target the other. These data

support the idea that heterozygosity can provide an adaptive

advantage to the host and shift the balance of power in a

host–virus genetic conflict, at least temporarily. We find that

the Vif protein from SIVagm.sab, the lentivirus naturally circu-

lating among sabaeus monkeys in the wild, has overcome this

challenge and learned to counteract both derived and ancestral

variants of A3G [40]. However, this likely required repeated

transmission cycles and more extensive bouts of virus adap-

tation, because SIVagm.sab Vif shares less than 40% amino

acid identity with Vif from other SIVagm strains. Thus, hetero-

zygosity in restriction factors may have the capacity to severely

limit the ability of viruses to successfully counter-adapt during

the early stages of virus emergence.

5. Ebb and flow of the lentivirus reservoir: virusextinction and re-emergence

In addition to describing the immune adaptations of natural

host species that have resulted from, and possibly enabled,

long-term coexistence with lentiviruses, some features of pri-

mate immunity suggest that lentiviruses may have been

more widespread in the past than they are today. Species-

specific SIV strains have been defined in dozens of African

non-human primates, yet the relationship between lentiviruses

and some species is more ambiguous. Surveys of SIV in the

wild have identified ‘dead-end’ or ‘incidental’ hosts that are

not infected with unique versions of SIV but instead harbour

transient infections normally found in other species. Examples

include the yellow baboon, the chacma baboon and the white-

crowned mangabey, three species that, at low prevalence, carry

an SIVagm subtype usually confined to the autologous host

[68–70]. While clearly exposed to lentiviruses today, it is uncer-

tain whether such species were hosts to species-specific strains

in the past, although analyses of the genetics of their restriction

factors similar to that which has been done in AGMs [40] may

be able to discern between these two hypotheses.

Similarly, the first report of lentivirus infection in patas

monkeys, a species that is closely related to and sympatric

with sabaeus monkeys (AGM) in Senegal, showed that

approximately 5% of animals were cross infected with the

SIVagm.Sab subtype [71]. A later study showed that patas

monkeys share the immunophenotypic features of canonical

natural host species, namely the downregulation of the CD4

receptor from helper T cells, suggesting that a hypothetical

patas-specific SIV may have once existed [72]. This observa-

tion raises the possibility that adaptation of host immunity

(such as downregulation of lentivirus receptors or co-receptors)

may in fact mediate resistance to infection, thereby driving

the extinction of certain lentivirus strains. A similar conclusion

may be drawn for the white-crowned mangabey, which exhi-

bits a relatively low frequency of CD4þ T cells, a property

shared by other mangabey species that are known natural

hosts [73]. The prospect that genetic adaptations present in

these species may provide resistance to lentivirus infections

in the wild provides encouragement to gene therapeutic

approaches being developed to control the spread of HIV-1

in humans.

While it may be reasonable to suspect that most (if not all)

primate species in Africa have been infected by lentiviruses at

one time or another, primate habitats span the globe from the

New World to the Far East. Current surveillance efforts limit

10

100

A3G:

no Vif

0.1

derivedancestral

parental Vifadapted Vifadapted Vif + C84Y

infe

ctiv

ity (

%)

1

homozygous -ancestral

(a)

(b)

homozygous -derived

heterozygous

host A3Ggenotype:

viral vifadaptation status:

no adaptationnecessary

rapid adaptation toderived A3G variant

constraint imposed byancestral A3G impedes

adaptation to derived A3G

A3G Vif

Figure 3. Host heterozygosity may impede viral adaptation in the setting of cross-species transmission. (a) The outcome of experimental infection of three sabaeusAGMs that differ by A3G genotype is schematized. A3G variants are drawn as solid black bullets, whereas Vif variants are drawn as white crescents. The Vif-sensitive,ancestral A3G variant is readily antagonized by Vif. However, antagonism of the Vif-resistant, derived A3G variant requires counter-adaptation of the vif gene. Solidarrows signify in vivo evolution of vif in order to re-establish the interaction. In the ‘heterozygous’ monkey, expressing both the ancestral and derived A3G variant, vifcounter-evolution does not occur. (b) Single-round virus infections were performed using virions produced in the presence of Vif and A3G. Infectivity of viruses isreported as a percentage, relative to infectivity in the absence of A3G (100%). Error bars indicate standard deviation from the mean of three infection replicates.Residues found at the Vif-binding site of the ancestral and derived A3G variants are shown in grey boxes. Red bars correspond to the parental vif encoded by thevirus inoculum (SIVagm.Ver90). SIVagm.Ver90 Vif antagonizes the ancestral A3G variant, as illustrated by its ability to rescue virus infectivity (compare ‘no Vif ’ with‘parental Vif ’). By contrast, SIVagm.Ver90 cannot antagonize the derived A3G variant. Vif amplified following experimental infection of a monkey homozygous forthe derived A3G variant (‘adapted Vif ’) demonstrates elevated antagonism of derived A3G but a loss of activity against ancestral A3G (compare ‘adapted Vif ’ with‘parental Vif ’). Reversion of a cysteine residue selected in vivo (C84Y) results in Vif with the opposite specificity. (Online version in colour.)

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the range of SIV infections to the African continent, but a trail of

genetic evidence allows one to follow the spread of lentiviruses

out of Africa. For example, the Asian macaque species housed

in American primate centres recently acquired lentivirus infec-

tions via artificial cross-species transmission of SIV from sooty

mangabeys [74]. On the other hand, more recent hallmarks of

selection imprinted in other host factors strongly suggest the

possibility of a ‘natural’ rhesus-specific SIV strain in the

recent past. A high-frequency single nucleotide polymorphism

(SNP) in A3G of Indian-origin rhesus macaques, like those

identified in other members of the Cercopithecinae subfamily

of OWMs, encodes a unique charge-altering amino acid

change that affects susceptibility to the viral antagonist Vif

(figure 4) [39]. Likewise, there is an evidence of polymorphisms

in the TRIM5 gene in rhesus macaques [75], as well as the

presence of the TRIM5-CypA fusion gene in rhesus macaques

[50–52] that would provide protection against some lenti-

viruses. These polymorphisms may represent a vestigial

footprint of an extinct lentivirus that once circulated among

these primates. Thus, innovation in the innate immune

repertoire of the rhesus macaque may have been inspired by

encounters with lentiviruses in wild habitats.

Perhaps most striking of all, human evolutionary genetics

may allude to a lentiviral presence that preceded HIV-1. A

unique five amino acid deletion is fixed in the human ortho-

logue of Tetherin. As a broad acting antiviral effector, a

deletion in Tetherin could have been selected in response to

any number of enveloped viruses. However, this deletion

spans residues necessary for antagonism by the lentiviral

Nef protein used by many SIV strains, suggesting that it

may have been selected by a pathogenic, Nef-encoding lenti-

virus approximately 800 000 years ago (figure 4) [44,45,76].

Therefore, changes in innate immune genes of humans may

have been triggered by previous lentivirus infections in our

recent ancestors.

6. Looking backwards to move forwardThe evidence presented in this review for ancient lentiviral

infections of primate based on selections of host genes is

rhesus macaque – A3G D130N

humans –tetherin

deletion

lemurs – pSIV fossil

Colobinae – A3G insertion

guenons – A3G D130A

macaques – TRIM

-CypA

owl monkeys –

TRIM-CypA

time (ya)100 K1Myr10Myr40Myr 5Myr

AGM (sab) – A3G D130H

AGM (gri) – A3G K128E

moustached guenon – A3G K128E

simian ancestor

- TRIM

-CypA

SAMHD1-degrading activity

(Cercopithecinae)

Figure 4. An overview of host genetic adaptations used to infer past lentivirus infections in deep and shallow time. Adaptations are described by host species,followed by the gene name, protein fusion product, of lentivirus fossil. Those marked by a vertical flag are fixed adaptations and the minimum age is anchored inthe timeline. Mutations marked by a horizontal bracket are polymorphic in a single species of primate. The range of the bracket spans the minimum and maximumdate at which the mutation could have appeared. ya, years ago, Myr, million years, K, thousand years. (Online version in colour.)

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summarized in figure 4. An understanding of these virus origins

may allow us to better understand the host–virus interactions

that are pivotal to virus infection, transmission and long-term

persistence. While selected by virus infections of the past, the

strategies that primates have deployed to cope with lentiviruses,

and an understanding of how such adaptations constrain virus

fitness, may be exploited as novel therapeutic strategies to stem

modern pandemics in humans. For example, the introduction of

HIV-specific restriction factors into human cells is a burgeoning

avenue for antiviral gene therapy [77–79]. However, the vast

potential for viruses to counter-evolve in the face of genetic

innovations of their hosts means that a combinatorial approach

to gene therapy may prove most successful. That is, an enduring

blockade of host–virus interactions might only be achieved by

introducing multiple genetic changes in one or more antiviral

genes, like those naturally selected in hosts of SIV. Finally, an

appreciation for the size of the lentivirus reservoir, both past

and present, and for the pathogenic prowess of the many

species-specific virus strains contained therein, will help to

identify the human populations most at risk of zoonotic

infection and virus emergence.

Acknowledgements. We thank Lucie Etienne, Chelsea Spragg and PatrickMitchell for comments on the text.

Funding statement. A.A.C. was supported by a National Science Foun-dation Predoctoral Fellowship. This work was supported byNational Institutes of Health grant no. R01 AI30927 (to M.E.) and aNational Science Foundation CAREER award (to H.S.M.). H.S.M. isan Early Career Scientist of the Howard Hughes Medical Institute.

References

1. Hahn BH, Shaw GM, De Cock KM, Sharp PM. 2000AIDS as a zoonosis: scientific and public healthimplications. Science 287, 607 – 614. (doi:10.1126/science.287.5453.607)

2. Perelman P et al. 2011 A molecular phylogeny ofliving primates. PLoS Genet. 7, e1001342. (doi:10.1371/journal.pgen.1001342)

3. Sharp PM, Hahn BH. 2010 The evolution of HIV-1and the origin of AIDS. Phil. Trans. R. Soc. B 365,2487 – 2494. (doi:10.1098/rstb.2010.0031)

4. Keele BF et al. 2009 Increased mortality and AIDS-like immunopathology in wild chimpanzees infectedwith SIVcpz. Nature 460, 515 – 519. (doi:10.1038/nature08200)

5. Rudicell RS et al. 2010 Impact of simianimmunodeficiency virus infection on chimpanzee

population dynamics. PLoS Pathog. 6, e1001116.(doi:10.1371/journal.ppat.1001116)

6. Etienne L et al. 2011 Characterization of a newsimian immunodeficiency virus strain in anaturally infected Pan troglodytes troglodyteschimpanzee with AIDS-related symptoms.Retrovirology 8. (doi:10.1186/1742-4690-8-4)

7. Bailes E, Gao F, Bibollet-Ruche F, Courgnaud V,Peeters M, Marx PA, Hahn BH, Sharp PM. 2003Hybrid origin of SIV in chimpanzees. Science 300,1713. (doi:10.1126/science.1080657)

8. Rey-Cuille MA, Berthier JL, Bomsel-Demontoy MC,Chaduc Y, Montagnier L, Hovanessian AG,Chakrabarti LA 1998 Simian immunodeficiency virusreplicates to high levels in sooty mangabeyswithout inducing disease. J. Virol. 72, 3872 – 3886.

9. Broussard SR, Staprans SI, White R, Whitehead EM,Feinberg MB, Allan JS. 2001 Simian immunodeficiencyvirus replicates to high levels in naturally infectedAfrican green monkeys without inducing immunologicor neurologic disease. J. Virol. 75, 2262 – 2275.(doi:10.1128/JVI.75.5.2262-2275.2001)

10. Cichutek K, Norley S. 1993 Lack of immunesuppression in SIV-infected natural hosts. AIDS7(Suppl. 1), S25 – S35. (doi:10.1097/00002030-199301001-00004)

11. Chahroudi A, Bosinger S, Vanderford T, Paiardini M,Silvestri G. 2012 Natural SIV hosts: showing AIDSthe door. Science 335, 1188 – 1193. (doi:10.1126/science.1217550)

12. Charleston MA, Robertson DL. 2002 Preferential hostswitching by primate lentiviruses can account for

rstb.royalsocietypublishing.orgPhilTransR

SocB368:20120496

9

on August 13, 2013rstb.royalsocietypublishing.orgDownloaded from

phylogenetic similarity with the primate phylogeny.Syst. Biol. 51, 528 – 535. (doi:10.1080/10635150290069940)

13. Wertheim JO, Worobey M. 2007 A challenge to theancient origin of SIVagm based on African greenmonkey mitochondrial genomes. PLoS Pathog. 3,e95. (doi:10.1371/journal.ppat.0030095)

14. Sharp PM, Bailes E, Gao F, Beer BE, Hirsch VM, HahnBH. 2000 Origins and evolution of AIDS viruses:estimating the time-scale. Biochem. Soc. Trans. 28,275 – 282.

15. Holmes EC. 2003 Molecular clocks and the puzzle ofRNA virus origins. J. Virol. 77, 3893 – 3897. (doi:10.1128/JVI.77.7.3893-3897.2003)

16. Wertheim JO, Worobey M. 2009 Dating the age ofthe SIV lineages that gave rise to HIV-1 and HIV-2.PLoS Comput. Biol. 5, e1000377. (doi:10.1371/journal.pcbi.1000377)

17. Wertheim JO, Kosakovsky Pond SL. 2011 Purifyingselection can obscure the ancient age of virallineages. Mol. Biol. Evol. 28, 3355 – 3365. (doi:10.1093/molbev/msr170)

18. Gifford RJ. 2012 Viral evolution in deep time:lentiviruses and mammals. Trends Genet. 28,89 – 100. (doi:10.1016/j.tig.2011.11.003)

19. Worobey M et al. 2010 Island biogeography revealsthe deep history of SIV. Science 329, 1487. (doi:10.1126/science.1193550)

20. Katzourakis A, Rambaut A, Pybus OG. 2005 Theevolutionary dynamics of endogenous retroviruses.Trends Microbiol. 13, 463 – 468. (doi:10.1016/j.tim.2005.08.004)

21. Holmes EC. 2011 The evolution of endogenous viralelements. Cell Host Microbe 10, 368 – 377. (doi:10.1016/j.chom.2011.09.002)

22. Katzourakis A, Gifford RJ. 2010 endogenous viralelements in animal genomes. PLoS Genet. 6,e1001191. (doi:10.1371/journal.pgen.1001191)

23. Gifford RJ, Katzourakis A, Tristem M, Pybus OG,Winters M, Shafer RW. 2008 A transitionalendogenous lentivirus from the genome of a basalprimate and implications for lentivirus evolution.Proc. Natl Acad. Sci. USA 105, 20 362 – 20 367.(doi:10.1073/pnas.0807873105)

24. Gilbert C, Maxfield D, Goodmn S, Feschotte C. 2009Parallel germline infiltration of a lentivirus in twoMalagasy lemurs. PLOS Genet. 5, e1000425. (doi:10.1371/journal.pgen.1000425)

25. Emerman M, Malik HS. 2010 Paleovirology: modernconsequences of ancient viruses. PLoS Biol. 8,e1000301. (doi:10.1371/journal.pbio.1000301)

26. Duggal NK, Emerman M. 2012 Evolutionary conflictsbetween viruses and restriction factors shapeimmunity. Nat. Rev. Immunol. 12, 687 – 695.(doi:10.1038/nri3295)

27. Daugherty MD, Malik HS. 2012 Rules ofengagement: molecular insights from host – virusarms races. Annu. Rev. Genet. 46, 677 – 700.(doi:10.1146/annurev-genet-110711-155522)

28. Malim MH, Bieniasz PD. 2012 HIV restriction factorsand mechanisms of evasion. Cold Spring Harb.Perspect. Med. 2, a006940. (doi:10.1101/cshperspect.a006940)

29. Russell RA, Wiegand HL, Moore MD, Schafer A,McClure MO, Cullen BR. 2005 Foamy virus Betproteins function as novel inhibitors of the APOBEC3family of innate antiretroviral defense factors.J. Virol. 79, 8724 – 8731. (doi:10.1128/JVI.79.14.8724-8731.2005)

30. Derse D, Hill SA, Princler G, Lloyd P, Heidecker G.2007 Resistance of human T cell leukemia virus type1 to APOBEC3G restriction is mediated by elementsin nucleocapsid. Proc. Natl Acad. Sci. USA 104,2915 – 2920. (doi:10.1073/pnas.0609444104)

31. Kolokithas A, Rosenke K, Malik F, Hendrick D,Swanson L, Santiago ML, Portis JL, Hasenkrug KJ,Evans LH. 2010 The glycosylated Gag protein of amurine leukemia virus inhibits the antiretroviralfunction of APOBEC3. J. Virol. 84, 10 933 – 10 936.(doi:10.1128/JVI.01023-10)

32. Malim MH, Emerman M. 2008 HIV-1 accessoryproteins: ensuring viral survival in a hostileenvironment. Cell Host Microbe 3, 388 – 398.(doi:10.1016/j.chom.2008.04.008)

33. Conticello SG, Harris R, Neuberger MS. 2003 The Vifprotein of HIV triggers degradation of thehuman antiretroviral DNA deaminase APOBEC3G.Curr. Biol. 13, 2009 – 2013. (doi:10.1016/j.cub.2003.10.034)

34. Yu X. 2003 Induction of APOBEC3G ubiquitinationand degradation by an HIV-1 Vif-Cul5-SCF complex.Science 302, 1056 – 1060. (doi:10.1126/science.1089591)

35. Mehle A, Goncalves J, Santa-Marta M, McPike M,Gabuzda D. 2004 Phosphorylation of a novel SOCS-box regulates assembly of the HIV-1 Vif-Cul5complex that promotes APOBEC3G degradation.Genes Dev. 18, 2861 – 2866. (doi:10.1101/gad.1249904)

36. Yu Y, Xiao Z, Ehrlich ES, Yu X, Yu XF. 2004 Selectiveassembly of HIV-1 Vif-Cul5-ElonginB-ElonginC E3ubiquitin ligase complex through a novel SOCSbox and upstream cysteines. Genes Dev. 18,2867 – 2872. (doi:10.1101/gad.1250204)

37. Han GZ, Worobey M. 2012 Endogenous lentiviralelements in the weasel family (Mustelidae). Mol.Biol. Evol. 29, 2905 – 2908. (doi:10.1093/molbev/mss126)

38. Sawyer SL, Emerman M, Malik HS. 2004 Ancientadaptive evolution of the primate antiviral DNA-editing enzyme APOBEC3G. PLoS Biol. 2, E275.(doi:10.1371/journal.pbio.0020275)

39. Compton AA, Emerman M. 2013 Convergence anddivergence in the evolution of the APOBEC3G-Vifinteraction reveal ancient origins of simianimmunodeficiency viruses. PLoS Pathog. 9,e1003135. (doi:10.1371/journal.ppat.1003135)

40. Compton AA, Hirsch V, Emerman M. 2012 The hostrestriction factor APOBEC3G and retroviral Vif proteincoevolve due to ongoing genetic conflict. CellHost Microbe 11, 91 – 98. (doi:10.1016/j.chom.2011.11.010)

41. Ayinde D, Casartelli N, Schwartz O. 2012 RestrictingHIV the SAMHD1 way: through nucleotidestarvation. Nat. Rev. Microbiol. 10, 675 – 680.(doi:10.1038/nrmicro2862)

42. Laguette N et al. 2012 Evolutionary and functionalanalyses of the interaction between the myeloidrestriction factor SAMHD1 and the lentiviral Vpxprotein. Cell Host Microbe 11, 205 – 217. (doi:10.1016/j.chom.2012.01.007)

43. Lim ES, Fregoso OI, McCoy CO, Matsen FA, Malik HS,Emerman M. 2012 The ability of primatelentiviruses to degrade the monocyte restrictionfactor SAMHD1 preceded the birth of the viralaccessory protein Vpx. Cell Host Microbe 11, 194 –204. (doi:10.1016/j.chom.2012.01.004)

44. Sauter D et al. 2009 Tetherin-driven adaptationof Vpu and Nef function and the evolution ofpandemic and nonpandemic HIV-1 strains. CellHost Microbe 6, 409 – 421. (doi:10.1016/j.chom.2009.10.004)

45. Lim ES, Malik HS, Emerman M. 2010 Ancientadaptive evolution of tetherin shaped the functionsof Vpu and Nef in human immunodeficiency virusand primate lentiviruses. J. Virol. 84, 7124 – 7134.(doi:10.1128/JVI.00468-10)

46. Jia B et al. 2009 Species-specific activity of SIV Nefand HIV-1 Vpu in overcoming restriction bytetherin/BST2. PLoS Pathog. 5, e1000429. (doi:10.1371/journal.ppat.1000429)

47. Zhang F et al. 2009 Nef proteins from simianimmunodeficiency viruses are tetherin antagonists.Cell Host Microbe 6, 54 – 67. (doi:10.1016/j.chom.2009.05.008)

48. Zhang F, Landford WN, Ng M, McNatt MW,Bieniasz PD, Hatziioannou T, Desrosiers RC. 2011SIV Nef proteins recruit the AP-2 complex toantagonize Tetherin and facilitate virion release.PLoS Pathog. 7, e1002039. (doi:10.1371/journal.ppat.1002039)

49. Malfavon-Borja R, Wu LI, Emerman M, Malik HS.2013 Birth, decay, and reconstruction of an ancientTRIMCyp gene fusion in primate genomes. Proc.Natl Acad. Sci. USA 110, E583 – E592. (doi:10.1073/pnas.1216542110)

50. Dietrich EA, Jones-Engel L, Hu SL. 2010 Evolution ofthe antiretroviral restriction factor TRIMCyp in OldWorld primates. PLoS ONE 5, e14019. (doi:10.1371/journal.pone.0014019)

51. Wilson SJ, Webb BL, Ylinen LM, Verschoor E,Heeney JL, Towers GJ. 2008 Independent evolutionof an antiviral TRIMCyp in rhesus macaques. Proc.Natl Acad. Sci. USA 105, 3557 – 3562. (doi:10.1073/pnas.0709003105)

52. Newman RM, Hall L, Kirmaier A, Pozzi LA, Pery E,Farzan M, O’Neil SP, Johnson W, Luban J. 2008Evolution of a TRIM5-CypA splice isoform in OldWorld monkeys. PLoS Pathog. 4, e1000003. (doi:10.1371/journal.ppat.1000003)

53. Ylinen LM, Price AJ, Rasaiyaah J, Hue S, Rose NJ,,Marzetta F, James LC, Towers GJ, Luban J. 2010Conformational adaptation of Asian macaqueTRIMCyp directs lineage specific antiviral activity.PLoS Pathog. 6, e1001062. (doi:10.1371/journal.ppat.1001062)

54. Dietrich EA, Brennan G, Ferguson B, Wiseman RW,O’Connor D, Hu S-L. 2011 Variable prevalence andfunctional diversity of the antiretroviral restriction

rstb.royalsocietypublishing.orgPhilTransR

SocB368:20120496

10

on August 13, 2013rstb.royalsocietypublishing.orgDownloaded from

factor TRIMCyp in Macaca fascicularis. J. Virol. 85,9956 – 9963. (doi:10.1128/JVI.00097-11)

55. Sayah DM, Sokolskaja E, Berthoux L, Luban J.2004 Cyclophilin A retrotransposition into TRIM5explains owl monkey resistance to HIV-1. Nature430, 569 – 573. (doi:10.1038/nature02777)

56. Ribeiro IP, Menezes AN, Moreira MA, Bonvicino CR,Seuanez HN, Soares MA. 2005 Evolution ofcyclophilin A and TRIMCyp retrotransposition in NewWorld primates. J. Virol. 79, 14 998 – 15 003.(doi:10.1128/JVI.79.23.14998-15003.2005)

57. Patel MR, Emerman M, Malik HS. 2011Paleovirology—ghosts and gifts of viruses past.Curr. Opin. Virol. 1, 304 – 309. (doi:10.1016/j.coviro.2011.06.007)

58. Kuhmann SE, Madani N, Diop OM, Platt EJ, MorvanJ, Muller-Trutwin MC, Barre-Sinoussi F, Kabat D.2001 Frequent substitution polymorphisms inAfrican green monkey CCR5 cluster at critical sitesfor infections by simian immunodeficiency virusSIVagm, implying ancient virus – host coevolution.J. Virol. 75, 8449 – 8460. (doi:10.1128/JVI.75.18.8449-8460.2001)

59. Zhang ZD, Weinstock G, Gerstein M. 2008 Rapidevolution by positive Darwinian selection in T-cellantigen CD4 in primates. J. Mol. Evol. 66, 446 – 456.(doi:10.1007/s00239-008-9097-1)

60. Ling B, Apetrei C, Pandrea I, Veazey RS, Lackner AA,Gormus B, Marx PA. 2004 Classic AIDS in a sootymangabey after an 18-year natural infection.J. Virol. 78, 8902 – 8908. (doi:10.1128/JVI.78.16.8902-8908.2004)

61. Traina-Dorge V, Blanchard J, Martin L, Murphey-Corb M. 1992 Immunodeficiency andlymphoproliferative disease in an African greenmonkey dually infected with SIV and STLV-I. AIDSRes. Hum. Retroviruses 8, 97 – 100. (doi:10.1089/aid.1992.8.97)

62. Hirsch VM, Dapolito G, Johnson PR, Elkins WR,London WT, Montali RJ, Goldstein S, Brown C.1995 Induction of AIDS by simian immunodeficiencyvirus from an African green monkey: species-specific variation in pathogenicity correlates

with the extent of in vivo replication. J. Virol. 69,955 – 967.

63. Beer BE, Brown CR, Whitted S, Goldstein S, GoekenR, Plishka R, Buckler-White A, Hirsch VM. 2005Immunodeficiency in the absence of high viral loadin pig-tailed macaques infected with simianimmunodeficiency virus SIVsun or SIVlhoest. J. Virol.79, 14 044 – 14 056. (doi:10.1128/JVI.79.22.14044-14056.2005)

64. Lemey P, Rambaut A, Pybus OG. 2006 HIVevolutionary dynamics within and among hosts.AIDS Rev. 8, 125 – 140.

65. Meyerson NR, Sawyer SL. 2011 Two-steppingthrough time: mammals and viruses. TrendsMicrobiol. 19, 286 – 294. (doi:10.1016/j.tim.2011.03.006)

66. Carrington M et al. 1999 HLA and HIV-1:heterozygote advantage and B*35-Cw*04disadvantage. Science 283, 1748 – 1752. (doi:10.1126/science.283.5408.1748)

67. Goldstein S et al. 2006 Comparison of simianimmunodeficiency virus SIVagmVer replication andCD4þ T-cell dynamics in vervet and sabaeus Africangreen monkeys. J. Virol. 80, 4868 – 4877. (doi:10.1128/JVI.80.10.4868-4877.2006)

68. van R, Engelbrecht S, Mwenda J, Laten JD, RobsonBA, Stander T, Chege GK 1998 Simianimmunodeficiency viruses (SIVs) from eastern andsouthern Africa: detection of a SIVagm variant froma chacma baboon. J. Gen. Virol. 79, 1809 – 1814.

69. Jin MJ, Rogers J, Phillips-Conroy JE, Allan JS,Desrosiers RC, Shaw GM, Sharp PM, Hahn BH. 1994Infection of a yellow baboon with simianimmunodeficiency virus from African greenmonkeys: evidence for cross-species transmission inthe wild. J. Virol. 68, 8454 – 8460.

70. Tomonaga K et al. 1993 Isolation andcharacterization of simian immunodeficiency virusfrom African white-crowned mangabey monkeys(Cercocebus torquatus lunulatus). Arch. Virol. 129,77 – 92. (doi:10.1007/BF01316886)

71. Bibollet-Ruche F, Galat-Luong A, Cuny G, Sarni-Manchado P, Galat G, Durand J-P, Pourrut X, Veas F.

1996 Simian immunodeficiency virus infection in apatas monkey (Erythrocebus patas): evidence forcross-species transmission from African greenmonkeys (Cercopithecus aethiops sabaeus) in thewild. J. Gen. Virol. 77, 773 – 781. (doi:10.1099/0022-1317-77-4-773)

72. Apetrei C, Gaufin T, Gautam R, Vinton C, Hirsch V,Lewis M, Brenchley J, Pandrea I. 2010 Pattern ofSIVagm infection in patas monkeys suggests thathost adaptation to simian immunodeficiency virusinfection may result in resistance to infection andvirus extinction. J. Infect. Dis. 202(Suppl. 3),S371 – 376. (doi:10.1086/655970)

73. Vinton C et al. 2011 CD4-Like immunologicalfunction by CD42 T cells in multiple natural hostsof simian immunodeficiency virus. J. Virol. 85,8702 – 8708. (doi:10.1128/JVI.00332-11)

74. Apetrei C et al. 2006 Kuru experiments triggered theemergence of pathogenic SIVmac. AIDS 20, 317 –321. (doi:10.1097/01.aids.0000206498.71041.0e)

75. Newman RM et al. 2006 Balancing selection andthe evolution of functional polymorphism in OldWorld monkey TRIM5a. Proc. Natl Acad. Sci. USA103, 19 134 – 19 139. (doi:10.1073/pnas.0605838103)

76. Sauter D, Vogl M, Kirchhoff F. 2011 Ancient origin ofa deletion in human BST2/Tetherin that confersprotection against viral zoonoses. Hum. Mutat. 32,1243 – 1245. (doi:10.1002/humu.21571)

77. Sakuma R, Noser JA, Ohmine S, Ikeda Y. 2007Inhibition of HIV-1 replication by simian restrictionfactors, TRIM5a and APOBEC3G. Gene Ther. 14,185 – 189.

78. Voit RA, McMahon MA, Sawyer SL, Porteus MH.2013 Generation of an HIV resistant T-cell line bytargeted ‘stacking’ of restriction factors. Mol. Ther.21, 786 – 795. (doi:10.1038/mt.2012.284)

79. Chan E, Schaller T, Eddaoudi A, Zhan H, Tan CP,Jacobsen M, Thrasher AJ, Towers GJ, Qasim W2012 Lentiviral gene therapy against humanimmunodeficiency virus type 1, using a novel humanTRIM21-cyclophilin A restriction factor. Hum. GeneTher. 23, 1176 – 1185. (doi:10.1089/hum.2012.083)