Quasispecies Theory and the
Behavior of RNA Viruses
Sumeeta Singh, Steve Bowers, Greg Rice, Tom McCarty
BINF 704
02/19/13
Review
Review
Quasispecies Theory and the Behav
Quasispecies Theory and
the Behavior
of RNA Viruses
1
2
Adam S. Lauring , Raul Andino *
Ad am S. Lauring 1, Raul Andino 2*
1 Department of Medicine, University of California, San Francisco, San Francisco, California, United State
1 Department of Medicine, University of California, San Francisco, San Francisco, California, United States of America, 2 Department of Microbiology and Immunology,
University of California, San Francisco, San Francisco, California, United States of America
University of California, San Francisco, San Francisco, California, United States of America
virus sequence may be determined more by its freedom to mutate
virus sequence
into related sequences than by its own replicative capacity.
A largehave
number
important
viruses,
including HIV, hepatitisAbstract:
C virus, and influenza,
RNA of medically
into ofrelated
Second, many
viruses operate
near a threshold
‘‘error s
genomes. These viruses
replicate
with
extremely
high
catastrophe’’
and may behave
combated
by increasing Second,
their replication
including HIV, hepatitis C virus,
and influenza,
RNA
man
mutation rates and exhibit significant genetic diversity.
error
rates.
Third,
increasing
the
fidelity
of
genome
replication
genomes.
with extremely high
This diversity allows a viral
populationThese
to rapidlyviruses
adapt to replicate
catastrophe’’ a
may paradoxically attenuate viruses.
dynamic environmentsmutation
and evolve resistance
to vaccines
rates and
exhibit significant genetic diversity.
error rates. T
and antiviral drugs. For
thediversity
last 30 years,
quasispecies
This
allows
a
viral
population
to
rapidly
adapt
to
Error -Prone Replicat ion and Viral Quasispecies
theory has provided a population-based framework for
may paradoxic
dynamic
environments
and
evolve
resistance to vaccines
understanding RNA viral evolution. A quasispecies is a
M ost viruses encode enzymes responsible for replicating their
cloud of diverse variants
are genetically
and that
antiviral
drugs. linked
For the last
years,
quasispecies
DNA 30
or RNA
genomes.
The intrinsic error rate, or fidelity, of the
through mutation, interact cooperatively on a functional
-Prone
replicase determines
the mutationfor
rate for thatError
virus and
the
theory
has
provided
a
population-based
framework
level, and collectively contribute to the characteristics of
range of genetic
variation upon which
can act.
RNA theory
viral evolution.
A quasispecies
is a natural selection
the population. Many understanding
predictions of quasispecies
M
ost
viruse
Viral
RNA
polymerases
exhibit
characteristically
low
fidelity
with
run counter to traditional
views of
behavior
and
cloud
ofmicrobial
diverse
variants
that
are
genetically
linked
24
measured
mutation
rates
of
roughly
10
mutations
per
DNA or RNA
evolution and have profound implications for our
through
mutation,
interact
cooperatively
on
a
functional
nucleotide
copied,
which
is
orders
of
magnitude
greater
than
understanding of viral disease. Here, we discuss basic
replicase deter
thoseto
of the
nearlycharacteristics
all DNA-based viruses
principles of quasispecies
theory
describe its relelevel,
andand
collectively
contribute
of and organisms [10,15,16].
range
of genet
Given the large population sizes observed in both
experimental
vance for our understanding
of viral fitness, virulence,
the population.
Many and
predictions
of quasispecies theory
antiviral therapeutic strategy.
and natural infections, it is estimated that everyViral
possible
pointpo
RNA
run counter to traditional viewsmutation
of microbial
and are generated with
and manybehavior
double mutations
each
measured
evolution and have profound
implications
our
viral replication
cycle and for
may be
present within the populationmu
at any
time [17].
RNAbasic
viruses exist as swarms
of similarcop
nucleotide
Int roduct ion
understanding of viral disease.
Here,
weBecause
discuss
variants that are continuously regenerated by mutation
of
those ofrelated
nearly
of quasispecies
and
describe
its
releThe rapid evolution of principles
RNA viruses complicates
the manage- theory
sequences, our ability to predict the outcome of an infection or a
ment of chronic infectionsvance
and the control
of
emerging
infectious
Given
the larg
for our understanding oftherapeutic
viral fitness,
virulence,
andof isolated clones
intervention
from studies
is limited.
agents [1–3]. The ongoing global AIDS pandemic and the
into a in
antiviral therapeutic strategy. Even a defined molecular clone will quickly transform
and natural
resurgence of influenza highlight the difficulties associated with
collection of related sequences when introduced into cells. This
mutation and
these genetically labile pathogens [4–6]. RNA viruses have also
Abstract: A large number of medically important viruses,
What is a virus?
•
Viruses are obligate intracellular parasites.
•
Small infectious agents bearing nucleic acid
instructions.
•
Classified based on the form of nucleic acid, DNA
or RNA.
Virus types
•
Plus strand RNA virus
Minus strand RNA virus
Double strand RNA virus
•
Retrovirus
•
Single strand DNA virus
Double strand DNA virus
•
•
•
RNA Virus types
DNA Virus types
“Quasispecies Theory”
•
Mathematical framework describing
evolution of macromolecules (Eigen, 1971)
•
Extends the classic population genetics
ideas of mutation-selection into
quasispecies (Eigen, Schuster, 1977)
•
Eventually borrowed to describe RNA virus
evolution dynamics
Virus Replication Error
•
Plus strand RNA virus
Minus strand RNA virus
Double strand RNA virus
RNA dependentRNA polymerase
Error rate =10-3 to 10-5
•
Retrovirus
Reverse Transcriptase
Error rate =10-4 to 10-5
•
Single strand DNA virus
Double strand DNA virus
DNA polymerase
Error rate =10-7 to 10-9
•
•
•
Requirements and Consequences
•
•
•
•
Polymerase responsible for high error
rates.
Estimated that each single and some double
nucleotide sequence changes occur.
Resulting in a collection of related
sequences around a “master” sequence.
Variation is related to ability to survive
(population genetics) AND probability of
occurring based on sequence neighbors
(quasispecies).
Virus Replication Swarm
Virus Replication Swarm
“Quasispecies Theory”
1971
1977
Quasispecies and RNA viruses
•
Survival of the “flattest”
•
Error Catastrophe
•
Fidelity and Fitness
Survival of the Fittest or Survival of
the Flattest
A flat species is a species which exists in a
genetically diverse group. Not dominated by
one variant.
A quasispecies must be a flat species.
A fit species (in this part of the
presentation) is a species which reproduces
very fast.
Flat Species
Properties of a flat species
A flat species will have high mutation rates.
A flat species is able to mutate without a
major effect on fitness.
Advantages of a flat species
Different mutants in a flat quasispecies can
help each other.
Flat species are better able to adapt.
Dengue-1 Virus
Lives as a quasispecies
 One variant of the virus which is found in
high concentrations cannot survive on its
own.
 It can only survive because other viruses
(infecting the same cell) create the
protein which it lacks.

Viroid Experiment
Theory: More genetically diverse (flatter)
species are better able to adapt to
mutations.
 Viroid 1 - CSVd – very fit
 Viroid 2- CChVMd – more flat, but less fit
 Procedure:
Infect plants with each viroid
Subject the plants to two environment
either normal, or UVC light

Results
UVC light will cause mutations
Results:
Under normal conditions the fitter viroid
did better, then the flatter viroid.
Under the UVC light the two were about
equal
Under the UVC light the flatter became
more diverse.
The fitter did not become more diverse.
Fitter or Flatter, which is better able
to survive?
The fit species will grow faster in an ideal
environment.
 The flat species will be able to adapt
more quickly.

Error Threshold
RNA Viruses have a high mutation rate
The point at which accumulated mutations reduce fitness:
- Too much mutation can lead to loss of vital information
- Too little mutation can lead to host defenses overcoming the virus
Error Threshold: position in informational space where a phase
transition occurs such that the genomic sequence information can no
longer be perpetuated.
The greatest fitness is when mutation rates
approach the error threshold
Model of Error Catastrophe
http://www.pnas.org/content/98/12/6895.full
Error Catastrophe
Extinction of a virus as a result of excessive RNA
mutations – lethal mutagenesis
Decrease viral fitness by increasing the rate at which new
mutations appear.
There is an intrinsic limit to the maximum variability of
viral genetic information before it loses meaning. If an
RNA virus quasispecies goes beyond that mutation limit,
the population will no longer be viable.
Increasing Mutation Rate

Ionizing radiation (eg X-rays) – cause
mutations by damaging DNA.

Base Analogs – chemicals that replace one
of the usual nucleotides in the DNA.
These mutagens cause copying errors.
APOBEC3G
Humans have the ability to induce lethal
mutagenesis
 Protein found inside cells that has a very
specific antiviral role
 Cytidine deaminase enzyme
 Unfortunately, HIV has the ability to bind
to APOBEC3G proteins and cause their
degradation

APOBEC3G
It de-aminates the cytosine base, thus mutating it to a uracil base
APOBEC3G
Fidelity and Fitness: Mutation
Rate
Evolutionary Theory: viral error
rates
• RNA Virus: low fidelity generates
diverse population of variants
• Homogenous population vs
dynamic environment
•
29
Fidelity and Fitness: Mutation Rate
Ribavirin and Lethal
Mutagenesis
• Hypothesis: mutant
with low mutation
rate less sensitive to
LM and resistant to
ribavirin
• Poliovirus
experimental groups:
G64S polymerase
mutation
•
30
Fidelity and Fitness: Mutation
Rate/Pathogenicity
3D-G64S Virus Is Less Pathogenic
than Wild-Type Virus in Mice
Competition between 3D-G64S and
Wild-Type Virus in Mice
31
Fidelity and Fitness: Virulence
•
•
•
G64S species attenuated in transgenic mouse model for
poliovirus infection
Virulence determined by diversity of coinfecting
population
Quasispecies diversity, rather than the selection of
individual variants, correlates with enhanced virulence
32
Fidelity and Fitness: Attenuation
•
•
•
Vaccine Design
Vignuzzi(2008): G64 engineered
mutants stimulated high titers of
neutralizing antibodies in mice
Fidelity modulation as therapeutic
strategy
33
Future Perspectives
•
•
•
•
•
How do models apply to infected hosts?
What is best measure of viral fitness in
dynamic population?
How does population diversity influence
pathogenesis (subpopulation cooperation)?
Improved assays for characterizing viral
populations
Modern techniques: Deep sequencing,
Molecular barcoding
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