Download The origins and evolution of HIV

STUDENT’S GUIDE
Case Study
The origins and
evolution of HIV
Version 1.2
Anne Fischer
Formerly of the Max Planck
Institute for Evolutionary
Anthropology, Leipzig
Dean Madden [Ed.]
NCBE, University of Reading
Case Stu
Case Stud
the origins and evolution of hiv
PHOTO BY: Chris Jackson, Getty Images
Introduction
Prevalence of HIV
HIV (Human Immunodeficiency Virus) is the virus that can lead to AIDS
(Acquired Immune Deficiency Syndrome) in humans. AIDS is a disease
in which the immune system begins to fail, enabling other infections to
threaten the lives of patients. Since 1981, when it first began to spread
widely, HIV has caused the deaths of 25 million people worldwide.
According to current United Nations’ estimates, HIV will infect 90 million
people in Africa, leaving at least 18 million orphaned children there.
There are several different forms of HIV: evidence suggests that they
originated in Africa, but how are the different forms related to one another
and how did they enter the human population?
A boy sorts maize at the Reitutsire
orphanage in Maseru, Lesotho. The
orphanage is supported by Prince
Harry’s charity, Sentebale. Many adults
in Lesotho have been killed by AIDS
leaving a generation of over 380,000
orphans to fend for themselves.
IMAGE FROM: UN AIDS Global report, July 2008
This Case Study uses genetic sequence data from different types of HIV and
compares them with SIVs (simian immunodeficiency viruses), which are
found in wild chimpanzees and gorillas in Africa.
Estimated prevalence of HIV among
young adults (aged 15–49) by country in
2008.
Question
a. Study the map above. Describe the distribution of HIV. Which countries
have the greatest adult prevalence of HIV/AIDS? (Note: the map shows
the incidence of HIV/AIDS, not AIDS-related deaths.)
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PHOTO BY: Pascal Le Segretain, Getty Images
the origins and evolution of hiv
Transmission of HIV
HIV, the virus that causes AIDS, can be transmitted through:
•• unprotected sex (in semen or vaginal fluid);
•• blood (contact with contaminated material such as needles
and contaminated blood transfusions);
•• transmission from mother to child during pregnancy or at birth;
•• breast milk.
The virus was characterised in 1983 by a team from the Institut Pasteur
in France led by Luc Montagnier. In 2008, Montagnier and a colleague,
Françoise Barré-Sinoussi, were awarded the Nobel Prize in Physiology or
Medicine for their discovery of HIV.
An HIV particle (called a virion) is about 100 nm in diameter. This is about
1/20th of the length of a E. coli cell, and about 1/70th of the diameter of the
white blood cells that the virus infects.
Luc Montagnier, the co-discoverer
of HIV, photographed in 2008.
IMAGE FROM: Medical Art Service, Munich / Wellcome Images
The basic structure of the Human
Immunodeficiency Virus (HIV).
The virus’s spherical bilipid
membrane (yellow) is studded with
72 glycoproteins (green), made
from the proteins gp 120 and gp 41.
Beneath the membrane, a shell
made from the protein p 17 (pink)
surrounds the conical core or capsid
(yellow) made from p 24 protein.
The core contains two identical
single strands of RNA (ribonucleic
acid).
HIV has nine genes, compared to
about 25 000 genes in its human
host. These include sequences
encoding three enzymes required
for HIV replication: reverse
transcriptase, protease and
integrase (encoded by the pol gene).
A retrovirus
Like all viruses, HIV cannot reproduce by itself. To make new copies of
themselves, viruses must infect the cells of living organisms. HIV can only
replicate within human cells. HIV is a retrovirus. Retroviruses have RNA
(not DNA) as their genetic material. They use an enzyme called reverse
transcriptase to reverse-transcribe their RNA into DNA, which can then
be integrated into the host’s genome and replicated.
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the origins and evolution of hiv
DRAWING BY: Dean Madden, NCBE
New virus
leaves cell
New virus
assembled
HIV attaches to CD4
receptors on a T-cell
then fuses with the
host cell membrane
Viral RNA
transcribed
from DNA
Viral RNA (two
copies) and enzymes
enter the cell
Viral protease is
needed to process the
three viral proteins
DNA is transcribed
from viral RNA
Double-stranded
DNA is produced
Viral
integrase
DNA integrates
with the host
chromosome
HIV’s replication cycle
HIV is replicated in the human host’s cells as follows:
•• the virus binds to a protein called CD4 on the surface of the host’s
immune cells (e.g., lymphocytes). This allows the viral membrane to fuse
with the cell membrane, after which it releases the contents of the HIV
particle (virion) into the cell;
•• the viral RNA and three enzymes it encodes pass into the hosts’ cells. The
enzymes are a protease, a reverse transcriptase and an integrase, all of
which are needed for replication of the virus;
•• the viral RNA is reverse transcribed into DNA;
•• the viral DNA is then integrated into the genome of the host cell by the
integrase;
•• the DNA is transcribed back into RNA and translated into proteins that
form new viruses.
Using this mechanism, up to ten billion new viruses can be produced every
day. This rapid replication, coupled with a high mutation rate, contributes
to HIV’s variability and evolutionary success.
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the origins and evolution of hiv
HIV is placed within a subgroup
of the retroviruses, called
lentiviruses (lenti is Latin for
slow, and lentiviruses have a long
incubation period). This diagram
shows the relationship between
various lentiviruses.
Notice that there are two main
forms of HIV: HIV-1 and HIV-2.
Other lentiviruses include SIV
(simian immunodeficiency virus),
BIV (bovine immunodeficiency)
virus and FIV (feline
immunodeficiency virus) which
infect apes and monkeys, cows and
cats, respectively.
Two types of HIV
Two types of HIV infect humans: HIV-1 and HIV-2. HIV-1 is easilytransmitted. It is virulent and is the cause of the majority of HIV
infections globally. HIV-1 can be divided into three subgroups: HIV-1-M,
HIV-1-N and HIV-1-O, of which HIV-1-M is the most prevalent and has
spread around the world.
HIV-2 is less virulent than HIV-1 and is not transmitted as easily. It is largely
confined to West Africa. The effects of both viruses on humans are similar:
HIV-1 and HIV-2 are therefore distinguished by their genomes.
HIV denialism
Some people, including scientists who are not experts on HIV, have
suggested that HIV is not the cause of AIDS. They therefore question the
validity of HIV testing and treatment for AIDS. The mainstream scientific
community has rejected these claims. Unfortunately, some governments,
particularly those in South Africa, have until very recently supported
AIDS denialism and encouraged the use of ineffective ‘treatments’ such as
vitamin supplements. This has contributed to the failure of South Africa’s
response to its AIDS epidemic, although the situation is improving now.
Question
b. Now you know something about HIV, look once more at the map on
page 2. Suggest several different explanations for the distribution of
HIV/AIDS shown on the map.
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the origins and evolution of hiv
Where did HIV originate?
Since HIV was discovered in early 1980s, there has been considerable
speculation about its origin. One hypothesis suggests that HIV was
transmitted to humans from other primates. The close genetic relationship
between humans and primates makes it likely that viruses could be
transmitted between these species. Non-human primates (e.g., monkeys
and apes) carry HIV-like viruses, called SIVs (simian immunodeficiency
viruses). Unlike HIV, the viruses that non-human primates carry rarely
cause any disease in their hosts. These animals are called asymptomatic
carriers (which means that they display no disease symptoms).
PHOTO BY: Irwin Bernstein, University of Georgia.
Studying the evolutionary relationships between strains of HIV and related
SIVs from African primates has made it possible to discover more about the
likely origin of HIV. By comparing the genetic sequences of HIV and HIVlike viruses from non-human primates, one can identify which species is
most likely to have transmitted the virus to humans.
Clues from genes and geography
Scientists were able to show in 1989 that the RNA sequence of SIV from
both captive and wild Sooty mangabeys was very similar to that of HIV-2.
This supports the idea that HIV-2 originated in non-human primates. Other
evidence supporting the idea of cross-species transmission is the overlapping
geographical distribution in west Africa of SIV-infected Sooty mangabeys
and humans infected with HIV-2.
Sooty mangabeys are West African
primates that carry an SIV which is
thought to be the origin of HIV-2.
Historical range of the sooty mangabey in West Africa (shown in green). The
geographical range of these monkeys corresponds closely with the occurance of HIV-2
in humans. From: Santiago, M. L. et al. (2005) Journal of Virology 79 (19) 12515–12527.
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The origin of HIV-1
The origin of HIV-1 was unclear for many years. Chimpanzees and gorillas
are a potential source, but they are endangered species and it is not easy
to get blood samples from living animals in the wild. A breakthrough
came when Beatrice Hahn and her colleagues in Brimingham, Alabama,
developed a method of isolating DNA and RNA from faecal samples collected
from the forest floor.
Analysis of faeces from wild chimpanzees and gorillas has revealed the
presence of SIV in these species. Comparisons of chimpanzee and gorilla
SIV and human HIV-1 sequences were made by a team led by Paul Sharp at
the University of Nottingham, in co-operation with Hahn and many other
researchers. The work showed that these viruses are very similar. The
natural habitat of chimpanzees and gorillas coincides with the epicentres
of HIV-1 epidemics. Furthermore, the central African region encompassing
Gabon, Cameroon, Equatorial Guinea and the Republic of Congo, is the only
place where all three subgroups of HIV-1 (M, N and O) are found.
Sequences of SIVcpz (the SIV that infects chimpanzees) and SIVgor (the
SIV that infects gorillas) that resemble HIV-1 sequences most closely have
been found in chimpanzees and gorillas inhabiting the same geographical
region. How could transmission between species occur?
PHOTO BY: Mila Zinkova, Wikimedia Commons
PHOTO BY: Thomas Lersch, Wikimedia Commons
Chimpanzees and gorillas are the closest living relatives of humans. These
species, as other primate species, are commonly hunted for food (bushmeat)
and orphan chimps are sometimes kept as pets. Such pets are a natural
reservoir for the disease and they could transmit SIVcpz to humans. With
extensive logging of tropical rainforest, access to previously remote areas
is now possible, which further sustains the bushmeat trade.
Chimpanzee.
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Male silverback gorilla.
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Sequence analysis
The aim of this exercise is to study the similarities between SIV and HIV-1
sequences. This will allow you to investigate the potential transmission of
these viruses between great apes and humans.
The data provided are 16 nucleotide sequences from the pol gene of the HIV
and SIV viruses from chimpanzee, gorilla and human. There are six human
sequences, two from each of the three HIV subgroups (M, N and O), two
gorilla sequences and eight chimpanzee sequences. The data file is called:
DNA-HIV1andSIV.geneious
The analysis will be performed using a programme called Geneious. This
software can align sequences and build phylogenetic trees. The nucleotide
sequences will first be translated into protein sequences, which will then
be aligned. From the alignment of the protein sequences, you will build a
phylogenetic tree. This will show which sequences are more closely related
to one another.
1. Double click on the document named DNA-HIV1andSIV.
geneious. This will start the Geneious software and load
the file of genetic sequence data into the programme.
Hint: if a box appears over the Geneious start-up screen, saying that
your trial of the ‘Pro’ version has ended, click on ‘Use Geneious Basic’.
2. The 16 DNA sequences will now open in Geneious:
The names of the sequences
are shown here.
Copyright © Anne Fischer and Dean Madden, 2011
The DNA sequences are
in this central window.
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the origins and evolution of hiv
3. You can use the magnifying glass buttons to zoom in on the nucleotide
data:
Zoom buttons
Questions
c. Use the magnifying glass buttons to zoom in on the sequence data. How
can you tell that it is DNA sequence data and not RNA sequence data?
d. What sort of genetic information does HIV (a retrovirus) have?
e. How has the data therefore been processed before it was given to you?
4. Select all 16 sequences at the same time, by clicking on the file name in
the top window so that it is highlighted:
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the origins and evolution of hiv
5. Click on the Translate button to convert the DNA sequence data into
protein sequence data:
Translate button
6. A box will appear, asking you to choose a version of the genetic code to
use. Look at the options available, then choose Standard and click OK.
Note
Although the genetic code is often said
to be ‘universal’ — the same in all
living things — this is not quite true.
There are some minor variations in
different groups of organisms. Hence
this dialogue box, which allows you to
choose which version of the code you
wish to use.
A new file will appear in the top window, containing 16 protein
sequences derived from the original DNA sequences:
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the origins and evolution of hiv
7. Use the magnifying glass button again to zoom in on the sequence data
and check that the sequences are in fact proteins made of amino acids
(the single-letter amino acid codes are used here):
Asp
Glu
Arg
Lys
His
Asn
Gln
Ser
Thr
Tyr
D
Aspartic acid
E
Glutamic acid
RArginine
KLysine
HHistidine
NAsparagine
QGlutamine
SSerine
TThreonine
YTyrosine
Ala AAlanine
Gly GGlycine
Val
VValine
Leu
LLeucine
IleIIsoleucine
Pro
PProline
Phe
FPhenylalanine
Met
MMethionine
Trp
WTryptophan
Cys
CCysteine
Amino acid codes
The three-letter and single letter codes
for the 20 amino acids that are found in
proteins. Geneious uses the single-letter
codes to show the different amino acids.
8. The protein sequences should already be aligned, but before creating
a phylogeny, you will need to ensure that they are. Select the protein
sequences in the top window, then click the Alignment button:
Alignment button
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the origins and evolution of hiv
9. A box will appear, asking you to choose a method of
alignment. Only one method is possible with the basic Geneious
software, so select Geneious Alignment then click OK.
The alignment will take a few minutes to complete (slightly longer on a
slow computer):
Tree
button
10. Select the aligned protein sequences (‘Alignment of 16 sequences’) in
the top window, then click on the Tree button to create a phylogeny.
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the origins and evolution of hiv
A box will appear, offering some options for the tree building. Select
the values shown below and click OK.
Technical note
Select ‘JukesCantor’ and
‘NeighbourJoining’ here.
The Jukes-Cantor distance model
assumes that all amino acid
substitutions (mutations) happen at
the same rate (1 in 20 or 5%). Other
mathematical models assume that
different amino acids mutate at
different rates.
The Neighbor-Joining method is a quick
and popular mathematical model
for calculating genetic distances and
drawing trees. Other methods will
produce slightly different results (and
take longer to do it).
11. A tree will be produced in the lower central window. Re-size the other
windows so that you can study the tree. The software will cluster
similar sequences closer together.
You now have a phylogenetic tree of sequences from the three
subgroups (M, N and O) of the HIV-1 family and their relationship to
SIV sequences from chimpanzees and gorilla.
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the origins and evolution of hiv
Questions
f. Mark, on a paper print out of the tree, using three different colours or
symbols, the branches of the tree that derive from gorilla, chimpanzee
and human viruses.
g. Describe the locations of the HIV sequences in the tree (for example, do
they form any clusters or groups, or are they scattered throughout the
branches of the tree?)
h. Do the HIVs appear to be more closely related to each other, or to some
of the SIV sequences?
i. Do you think that HIV-1 could have originated more than once, and if
so, what was the source on each occasion?
j. Does the geographical distribution of SIV-infected apes overlap with
areas of HIV-1 epidemics? Compare the map on page 2 with the one
below and, if you have access to the internet: www.aidsinafrica.net/
map.php and www.unaids.org/en/
Further reading
Avert, a UK-based AIDS charity, has a website with comprehensive and
authoritative information about HIV/AIDS: www.avert.org
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the origins and evolution of hiv
A phylogenetic tree of sequences from the three subgroups (M, N and
O) of the HIV-1 family and their relationship to SIV sequences from
chimpanzees and gorilla. The codes after the virus names refer to
sampling areas. TAN = Tanzania; CAM = Cameroon; GAB = Gabon.
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