FAMILIES OF DNA

VIRUSES
 Viruses in Human Infections and Diseases

DNA or RNA molecules are surrounded by

capsid; obligate parasites that enter a cell,
instruct its genetic and molecular machinery
to produce and release new viruses.
All DNA viruses are double-stranded except
for parvoviruses, which have ssDNA.
All RNA viruses are single-stranded except
for dsRNA reoviruses.

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 Medical Considerations in Viral
Diseases
Viruses are limited to a particular host or cell
type.
Most DNA viruses are budded off the nucleus.
Most RNA viruses multiply in and are released
from the cytoplasm.
Viral infections range from asymptomatic to
mild to life-threatening.
Many viruses are strictly human in origin,
others are zoonoses transmitted by vectors.

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 Course of viral disease: invasion at portal of entry
and primary infection; some viruses replicate locally,
others enter the circulation and infect other tissues
Common manifestations: rashes, fever, muscle
aches, respiratory involvement, swollen lymph
nodes
Body defenses: combined action of interferon,
antibodies and cytotoxic T cells; frequently results in
lifelong immunity

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 Many viral infections have rapid course; lytic
cycle
Some viruses establish long-term persistent
infections that last many years or a lifetime.
2 types of persistent infections:
chronic infections virus is detectable in tissue
samples, multiplying at a slow rate; symptoms mild
or absent
latent infections after a lytic cycle, virus enters
a dormant phase; generally not detectable, no
symptoms; can reactivate and result in recurrent
infections

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 Some persistent viruses are oncogenic.
Several viruses can cross the placenta
causing developmental disturbances and
permanent defects teratogenic.
Diagnosis of viral diseases symptoms,
isolation in cell or animal culture,
serological tests for antibodies; some
tests for antigens

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Survey of DNA Viruses
Animal viruses are categorized according
to nucleic acid, capsid, and presence or
absence of envelope.
7 DNA families, 15 RNA families
DNA viruses causing human disease:
enveloped DNA viruses
nonenveloped DNA viruses
nonenveloped ssDNA viruses

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Insert Table 24.1
DNA viruses

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Enveloped DNA
Viruses

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Parvoviridae
Subfamilies and Genera
Subfamily Parvovirinae
Contains parvoviruses of vertebrates

Genus Parvovirus
Replicate autonomously
Cannot grow in stationary cells
Replicate in cells undergoing mitosis
do not code for enzymes required for
replication, BUT
Rely on the enzymes of the replicating
cell to perform this function

Resistance very stable to

environmental conditions extremes of
pH and heat disinfection DIFFICULT
Parvoviruses

The smallest of the deoxyribonucleic

acid (DNA) viruses.

Only one member of the Parvoviridae,

B19, is known to cause human disease.
Poxviruses: Classification and
Structure
Produce eruptive skin pustules called pocks
or pox, that leave scars
Largest and most complex animal viruses
Have the largest genome of all viruses
dsDNA
Multiply in cytoplasm in factory areas
variola cause of smallpox
vaccinia closely related virus used in
vaccines
monkeypox
cowpox

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Genus Orthopoxvirus

buffalopox virus {buffalo, cattle, human}

camelpox virus {camel} (CMLV)
cowpox virus {rodents, felines, bovines, human}
ectromelia virus {mousepox, reservoir unknown}
monkeypox virus {rodents, primates, human}
rabbitpox virus {colonized rabbit only}
raccoonpoxvirus {North America}
skunkpox virus {North American striped skunk}
taterapox virus {African gerbil}
vaccinia virus {no natural reservoir}
Uasin Gishu disease {Central African horses}
variola virus {human; eradicated from nature}
volepox virus {California pinon mouse and voles}
Insert figure 24.1
poxviruses

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 Properties of Poxviruses
Structure and composition Core
Envelope
Largest viruses
dsDNA, 130-375 kb Lateral
bodies
Brick shaped
dumbbell core (contains nucleic acid)
Lateral bodies (unknown function)

Enveloped
Resistant to inactivation

More than 100 polypeptides

Many target the immune response

Replicate in cytoplasm
Tend to be highly species-specific
Some are being explored as gene therapy vectors
Unique Properties of Poxviruses
The largest, most complex viruses
Have complex, oval to brick-shaped morphology with
internal structure
Have a linear-stranded DNA genome with fused ends.
DNA viruses that replicate in the cytoplasm
Virus encodes and carries all proteins necessary for mRNA
synthesis
It also encodes proteins for functions such as DNA
synthesis, nucleotide scavenging and immune escape
mechanisms
It assembled in inclusion bodies where it acquires its outer
membranes
Importance of studying
poxvirus
The mechanisms of spread of variola virus
within the body represent a model for the
spread of other viral infections
Poxviruses other than variola can cause
human disease
Vaccinia virus has becomr an excellent
vector that can be used for the
development of hybrid vaccines containing
the genes of other infecting agents
 Classification
Family Poxviridae
Subfamilies Genera Members

Chordopoxvirinae Orthopoxvirus Variola, vaccinia, cowpox, monkeypox, camelpox

(vertebrates) Parapoxvirus Sealpox, parapox of deer, pseudocowpox

Canarypox, fowlpox, pigeonpox, turkeypox,
Avipoxvirus
penguinpox
Capripoxvirus Goatpox, sheeppox
Hare fibroma, myoma, rabbit fibroma, squirrel
Leporipoxvirus
fibroma
Suipoxvirus Swinepox

Molluscipoxvirus Molluscum contagium

Yatapoxvirus Tanapox, Yaba monkey tumor

Entomopoxvirinae Capripoxvirus A

(insects) Capripoxvirus B

Capripoxvirus C
Smallpox
First disease to be eliminated by vaccination
Exposure through inhalation or skin contact
Infection associated with fever, malaise,
prostration, and a rash
variola major highly virulent, caused toxemia,
shock, and intravascular coagulation
variola minor less virulent
Routine vaccination ended in U.S. in 1972.
Vaccine reintroduced in 2002 for military and
medical personnel

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 Replication
Attachment, penetration and uncoating
Membrane fusion
Viral core dissociates, releasing
Viral DNA
Viral enzymes
Virally-encoded RNA polymerase (prepackaged) synthesizes
early mRNA
Early proteins
DNA polymerase
Thymidine kinase
Recombinase?
Maturation
Structural proteins are synthesized
Virus synthesizes its own membranes
Some virus exits by budding, but most when the cell dies
 Smallpox
Named smallpox to discriminate it from
largepox (syphilis)
No animal reservoir
Two species
Variola major (20% fatality; Genbank #VVCGAA)
Variola minor (1-2% fatality; Genbank #VMVY16780)
Smallpox has shaped civilization
Earliest evidence: Egyptian mummies
Ramses V (1157 BC)
Introduced to the Americas by European explorers
British army used smallpox as a biological weapon
against the Pontiac Indians
About 40 million native Americans died from
European diseases, including smallpox
Native Americans have limited MHC polymorphisms
Killed 300 million people in the 20th century alone
Total fatalities probably near 1 billion
 Control and Eradication
Vaccine
Edward Jenner observed that milkmaids rarely contracted smallpox
All had recalled earlier cowpox infections, which were nearly
universal in milkmaids
Cowpox only causes a mild infection in humans
Jenner hypothesized that the infectious agent of cowpox protected
against smallpox
He inoculated a nephew by scarification with cowpox crusts
- termed variolation
Todays vaccine is live attenuated vaccinia virus
Vaccinias genome looks similar to cowpox, but it is not identical
In the 1950s, the Soviets proposed a global eradication
program to the United Nations
Some have suggested that the Soviets had a vaccine-resistant
strain of variola, which could be used as a bioweapon
The World Health Organization (WHO) Intensified Smallpox
Eradication Programme program began in 1967
Smallpox was declared eradicated in 1980 (last case in 1977)
Only two nations are supposed to have smallpox viruses today
United States (CDC-Atlanta)
Russia (VECTOR, Moscow)
 Smallpox Pathogenesis
Respiratory transmission
Incubation period 10-14 days
After day 7 or so, humans begin shedding virus
asymptomatically
Principal reason for rapid spread
Outbreaks tended to be in clusters

Allowed the ring immunization containment strategy during the

eradication program
Clinical symptoms
Fever
Malaise
Centrifugally-distributed exanthems
macules
papules
pustules
Clinical
progression
of smallpox
 Smallpox Pathogenesis in an Animal Model

Problem: Smallpox virus was only known to infect humans

Can an animal model be developed to study pathogenesis?
No cases of smallpox since the 1970s
Modern science could learn much from studying the course of
infection and immune response in an animal model
Because of 9/11, CDC and USAMRIID Scientists were granted
permission by the WHO to infect cynomolgus macaques with
smallpox in an attempt to assess pathogenesis using contemporary
laboratory techniques
Jahrling et al. PNAS. 101:15196-15200
Rubins et al. PNAS. 101:15190-15195

Required highest containment: BSL-4

Required >10 pfu iv for infection
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Resulted in fulminant disease fatal in most monkeys

 Fig. 2. Gross lesions associated
with smallpox virus infection of
monkeys. (a) Hemorrhage on the
serosal surface of the distal colon,
monkey I-7, 6 days after
exposure. Note the diffuse
petechial hemorrhages and
hemorrhagic, colic lymph nodes.
(b) Mucosal surface of the distal
colon described in a. Note the
severe congestion, hemorrhage,
and hemorrhagic colic lymph
nodes. (c) Medial surface of the
right arm of monkey I-4, 9 days
after exposure. Smallpox pustules
are predominantly discrete with
occasional coalescence. Pock
lesions developed synchronously
and were more numerous distally,
which is consistent with centrifugal
distribution. (d) Palmar surface of
the left hand of a monkey 11 days
after exposure. Discrete and
coalescing pustules retained
integrity due to heavily keratinized
palmar and digital epidermis. (e)
Upper lip and nostrils from a
cynomolgus monkey 11 days after
exposure. Note synchronous
development of pustules, some of
which have umbilicated. Pustules
that formed on the lightly
keratinized mucosal surfaces of
the nostrils have already
ulcerated, resulting in dried
exudates.
Jahrling et al.
Fig. 4. Immunofluorescence examination of tissues from nonhuman primates infected with variola. (a) Staining for
monocytes/macrophages (green) and viral antigen (red) indicated the presence of infected monocytes/macrophages (gold)
in the lymphoid tissues (lymph node shown) and in circulation. (b) In addition to monocytes/macrophages, virus-infected
endothelial cells (green) were readily observed. (c) Staining for monocytes/macrophages (green) and apoptosis (red)
revealed the presence of numerous tingible body macrophages in lymphoid tissues. The majority of apoptotic cells were not
monocytes/macrophages but instead were lymphocytes. A rare apoptotic monocyte/macrophage (arrow) is shown. (d) A
monocyte/macrophage with a clearly stained nonapoptotic nucleus (blue) is shown engulfing two separate apototic bodies.
Jahrling et al.
 Smallpox as a Biological
Weapon
Variola viruses do not exist in nature
Some nations are suspected of having active
bioweapons programs
North Korea
Syria
Iran
Israel
Russia
Some may have kept smallpox stocks hidden
The world is ill-prepared for smallpox
Vaccination is no longer routine
Thus, a large susceptible population
 Monkeypox
2003 US Outbreak
72 Cases
Wisconsin, Illinois, Indiana
Kansas, Missouri, Ohio

Introduced from Africa to US pet supplier

Gambian giant rat
Prairie dogs
Infection was not fatal to humans
In Africa, disease is fatal in 1-10% of cases
Democratic Republic of Congo - 10%
West Africa - 1-2%

The US strain was from west Africa

Genomic comparisons revealed 5 genes mutant or missing
in West African strains compared to central African strains
WA strains do not have D14L, a protein that disables
complement enzyme activity, while central African do
 Molluscum Contagiosum
In endemic areas, it is primarily an infection of
children.
Transmitted by direct contact and fomites
In US, most commonly an STD
Lesions are small, smooth macules in genital area
and thighs.
AIDS patients suffer an atypical form which attacks
the skin of the face and forms tumorlike growths.
Treatment: freezing, electric cautery, chemical
agents

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Molluscum Contagiosum View Table

Molluscum contagiosum is a common, benign viral infection of

the skin and mucous membranes caused by the molluscum
contagiosum virus. It typically affects children, but can be
transmitted sexually in adults. The mature lesion is an
umbilicated papule.
Molluscum Contagiosum

Lesions begin as small (3 to 6 mm) papules that are

smooth, flesh-colored domes with a central dimple. Inside
the papule is a white, curd-like core that can be easily
expressed.

Lesions can occur anywhere on the skin and mucous

membranes, but are usually grouped in one or two areas.
Occasionally, they may be widely disseminated. Typically
fewer than 20 lesions are present, but some individuals
may have hundreds.

The head, eyelids, trunk, and genitalia are most

commonly affected, the genitalia being the predominant
site in adults.

The lesions are characteristically asymptomatic, but a few

patients may complain of itching or may develop an
eczematous reaction around the lesions.
Differential Dx
View Table
Molluscum Contagiosum

Variola (Smallpox) Molluscum Contagiosum

Lesions are widespread Lesions usually grouped
Associated with severe in one or two areas
clinical symptoms Not associated with
Lesions progress from clinical symptoms
macules to vesicles to Lesions are umbilicated
papules to crusts papules only
(progression occurs all at
the same time in
smallpox but at different
times in chickenpox)
 Other Poxviruses
Many mammalian groups host some poxvirus
cowpox, rabbitpox, mousepox, elephantpox.
Humans are susceptible to monkeypox and
cowpox.
Monkeypox in humans skin pocks, fever,
swollen lymph nodes
Cowpox in humans rare, usually confined to
hands; other cutaneous sites can be involved

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 HUMAN INFECTION
Cowpox
acquired by humans by milking cows
ulcerative lesions ("milkers nodules") on the hands
protects against smallpox
rodents are the main reservoir of
spreads secondarily to cows and domestic cats.
 Molluscum HUMAN
contagiosum
minor infectious
INFECTION
warty papule of the
skin with a central
umbilication,
transferred by
direct contact
sometimes as a
veneral
disease.
 Monkey pox HUMAN
INFECTION
acquired from monkeys or
wild squirrels
occasionally spreads from
human to human in
unvaccinated communities
Antigenically cross-reacts
with other poxviruses
Sick monkeys have not
been identified
apparently healthy
animals have antibodies.
Distribution
of Human Sierra
Ivory
coast
Nigeria

Monkeypox Leone
Liberia

(1970 -
1982)
Tropical rain forest Zaire

Location of human
Monkeypox cases
HUMAN INFECTION
Pseudocowpox
occurs worldwide
disease primarily of cattle.
In humans it causes non-ulcerating "milker's nodes".
ORF
worldwide occupational disease associated with
handling sheep and goats afflicted with "scabby
mouth".
In humans it manifests as a single painless, papulo-
vesicular lesion on the hand, forearm or face.
Non human infections

Ectromelia
mouse pox
Fowl pox
EXPRESSION VECTORS:
homologous
recombination
in infected
cells
introduces
foreign DNA
coupled to VV
promoter (e.g.
tk) into virus
genome.
Use to make
vaccines by
introducing
gene for key
antigen(s)
 Vaccinia Expression
Vector
Plasmid
DNA Foreign Recombinant plasmid
gene cloning vector
Vaccinia
DNA
Introduced by transfection
Nucleus
Gene from Recomb.
vector of vaccinia
Gene from sequences
rescuing
Replication
virus
Maturation
Vaccination w/
Infection of cell Release of
recombinant live virus capable
Infectious vaccinia virus of producing
vaccinia virus containing foreign foreign
Foreign
gene
rescuing virus gene immunizing antigen
EXPRESSION VECTORS:

Problems for human vaccines:

% of the human population already vaccinated
lifelong protection may result in poor
response to recombinant vaccines
Dangerous in immunocompromised hosts.
solution use Avipoxvirus vectors, e.g. fowlpox or
canarypox
'suicide vectors' undergo abortive replication in
mammalian cells:
can express high levels of foreign proteins
no danger of pathogenesis
no natural immunity in humans