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VIROLOGY
 VIROLOGY
GENERAL CHARACTERISTICS OF VIRUSES
Small infectious units (measured in nanometer; about 20 - 300 nm in diameter)

Electron microscopy is required

Parvovirus - smallest animal virus

Poxvirus - largest animal virus

Obligate Intracellular parasite

Do not grow on artificial culture media

Grow on: Tissue culture, Animals, Chick embryo

Contain only one kind of nucleic acid -- DNA or RNA

Not sensitive to antibiotics

 VIRUS STRUCTURE
Viral components -- General:

A nucleic acid genome (RNA or DNA)

A protective protein coat (called the capsid)

May or may not have an envelope made of lipid derived from the host cell
membrane

VIRION NUCLEOCAPSID STRUCTURES

Icosahedral structure

Helical structure

Complex structure (Poxviridae)

MULTIPLICATION CYCLE
Adsorption

Penetration

Uncoating

Synthetic phase

Assembly

Release
 DIAGNOSTIC METHODS
IN VIROLOGY
1. Direct examination

2. Indirect examination (virus isolation)

3. Serology

4. Molecular methods
 Direct Examination
1. Antigen detection - immunofluorescence, ELISA,
etc.

2. Electron microscopy - morphology of virus particles

immune electron microscopy

3. Light microscopy - histological appearance inclusion

bodies

4. Viral genome detection - hybridization with specific

nucleic acid probes polymerase chain reactn (PCR)
 Indirect Examination
Cell culture Cytopathic effect (CPE)

Hemabsorption

Immunofluorescence

Eggs Pocks on CAM

Hemagglutination

Inclusion bodies

Animals Disease or death

Serology
Detection of rising titres of antibody between acute and convalescent
stages of infection, or the detection of IgM is n primary infection.

Classic Techniques New Techniques

1. CFT 1. RIA

2. HAI 2. EIA

3. IF 3. PA

4. NT 4. WB

5. CIE 5. RIBA, Line Immunoassay

 Virus Isolation
Cell Cultures are most widely used for virus isolation

3 Types of Cell Culture

1. Primary cell - Monkey Kidney

2. Semi-continuous cells - Human embryonic kidney and skin fibroblasts

3. Continuous cells - HeLa, Vero, Hep2, LLC-MK2, MDCK

Primary cell culture

the best cell culture systems

they support the widest range of viruses

they are very expensive

often difficult to obtain a reliable supply

Continuous cell line

the most easy to handle

the range of virus supported is often limited

Cell Cultures
Growing virus may produce

1. Cytopathic effect (CPE) - such as the ballooning of

cells or synctia formation, may be specific or non-
specific

2. Hemadsorption - cells acquire the ability to stick to

mammalian red blood cells

Confirmation of the identity of the virus may be carried out

using neutralization, hemadsorption-inhibition or
immunofluorescence tests.
Electron Microscopy
10^6 virus particles per ml required for visualization, 50,000-60,000
magnification normally used. Viruses may be detected in the following
specimens.

Feces Rotavirus, Adenovirus,

Norwalk like viruses

Astrovirus, Calicivirus

Vesicle Fluid HSV, VZV

Skin scrapings Papilomavirus,

Molluscum contagiosum
Serology
Criteria for Diagnosing Primary Infection

4 fold or more increase in titer of IgG or total body antibody between

acute and convalescent sera

Presence of IgM

Seroconversion

A single high titre of IgG ( or total antibody) - very unreliable

Criteria for diagnosing reinfection

4-fold or more increase in titer of IgG or total antibody between

acute and convalescent sera

Absence or slight increase in IgM

POXVIRIDAE
dsDNA genome, BRICK-SHAPED, complex symmetry,
enveloped

LARGEST animal virus

Predilection for epidermal cells (multiply in the cytoplasm --

eosinophilic inclusions -- (Guarnieri bodies)

Vesicular skin lesions in the susceptible host

Variola virus - causes smallpox (declared by the WHO to be

completely eradicated worldwide)
PARVOVIRIDAE
SMALLEST animal virus

ssDNA, icosahedral, naked

Strain B19 - causes

Aplastic crisis:

The infected erythrocytes are destroyed and the patient develops

a profound anemia

Erythema infectiosum ("fifth disease")

"slapped cheeks" appearance of the face

Hydrops fetalis

B19 strain may play a significant role in miscarriages

 HERPES SIMPLEX VIRUS
2 typess: HSV-1 and HSV-2

Transmission:

direct contact with vesicular discharge

sexual transmission

mother-to-infant thru birth canal (perinatal)

Vesicle formation

Primary infection - active replication in skin or mucosal vesicular lesions but is not eliminated

Latent Infection - localized in sensory ganglia:

Trigeminal ganglion - HSV-1

Sacral ganglion - HSV2

Recurrences may occur frequently or not at all

Trigger mech: fever, colds, sunlight, menses, certain diseases

 HERPES SIMPLEX VIRUS
(HSV)
Clinical disease: vesicular lesions

HSV-1 -- perioral lesions not absolute

HSV-2 -- genital lesions

Herpetic gingivostomatitis - most common

Primary

Recurrent (fever blister, herpes labialis, or

cold sores)

Ulcerative pharyngitis

Herpetic whitlow (fingers)

Keratoconjunctivitis

Genital herpes, proctitis

Neonatal herpes
HERPES SIMPLEX VIRUS
Diagnosis:

In most cases - established clinically based typical vesicular lesions

Lab Dx:

Tzanck smear - microscopic exam of scraping from the base of a

vesicle reveal giant multinucleated cells (tzanck cells) with
intranuclear inclusions that push the chromatin to the margins

Culture and isolation

Serology

Direct immunofluorescence - for cases of herpes encephalitis

Test for IgM antibodies

VARICELLA-ZOSTER
VIRUS
Transmission: respiratory
droplets

Vesicular rash

Primary infection: varicella

or chickenpox (scalp, face,
trunk)

Latency: dorsal root

ganlia

Recurrent infection: shingles

or zoster (clustered
vesicular lesions, PAIN
VARICELLA-ZOSTER
VIRUS
Diagnosis:

Clinical

Tzanck test - cytology (Cowdry type A intranuclear inclusions)

does not distinguish those from HSV

Culture from vesicles - not for zoster

Immunofluorescence - more sensitive than culture; choice for

zoster

Serology - may take several weeks for Ab to develop

 EPSTEIN-BARR VIRUS

(EVB)
Infects B cells
Mitogenic and causes polyclonal activation of B cells
inducing IgM production (heterophil antibodies)
Heterophile antibodies - agglutinate sheep and horse rbc
Clinical syndromes
Infectious mononucleosis
Fever
Exudative pharygitis
Lymphadenopathy
Burkitt's lymphoma
Nasopharygeal carcinoma
Viral exanthem (10%)
Other manifestation
Pneumonia with pleural effusion
Aseptic meningitis
EPSTEIN-BARR VIRUS
Diagnosis:

Culture: not routinely done, difficult to grow on usual cell culture

Established by presence of atypical lymphocytes, heterophile antibodies

Serology

Antigens of EBV:

EBV nuclear Ag (EBNA)

Early Ag: 2 types:

EA-R -- restricted to cytoplasm

EA-D -- diffuse in cytoplasm and nucleus

Viral capsid Ag (VCA)

Lymphocyte-determined membrane antigen (LYDMA)

 CYTOMEGALOVIRUS
MOT: transplacental, oral, sexual, and by blood transfusion and tissue
transplant

Pathogenesis: cell-to-cell spread

Latency in wbc

Reactivation following immunosuppression

Clinical syndromes

Congenital and neonatal infection

Adults: asymptomatic or mild mnonucleosis syndrome

Post-transfusion: mononucleosis- like (but heterophile antibody negative)

In immunocompromised patients: chorioretinitis, pneumonia, encephalitis,

colitis, esophagitis (as in AIDS patients)
CYTOMEGALOVIRUS
Diagnosis:

Cytologic exam of urine for congenital CMV (negative in 1/3)

Histologic exam by Paps or HE stain: for cytomegalic cell with dense,

central "owl's eye" basophilic intranuclear inclusion

Ag detection - IF, In-situ hybridization (molecular probe)

Cell culture: definitive; grows only on diploid fobroblasts for congenital

infections, virus should be isolated in urine in the first week for
confirmation

Serology
HUMAN HERPES VIRUS 6

Causes roseola infantum (or exanthem subitum)

HUMAN PAPOVAVIRIDAE
PAPILLOMAVIRUS

tropism for squamous epithelial cells --> warts

Natural transmission: through contact minor abrasions

Several serotypes: ~60 serotypes

Warts (finger, sole, face)

Condylomata acuminata (ano-genital warts)

Cervical cancer

Vulvar

Penile cancer

Squamous cell CA
PAPOVAVIRIDAE
POLYOMAVIRUSES

Epidemiology

Widespread in many parts of the world

Common occurrence during childhood

Route of transmission - not known yet

Clinical associations:

BK virus - 1st isolated from urine of a patient who received a renal & ureter
translant

JC virus - PML Progressive Multifocal Leukoencephalopathy

 ADENOVIRIDAE
dsDNA, icosahedral, naked
Produces latent or persistent infection in tonsils and adenoids
Transmission: respiratory droplets, fecal-oral means
CLINICAL MANIFESTATIONS:
May be isolated from feces of apparently healthy individuals
"Common colds"
Primary Atypical Pneumonia
Acute respiratory disease (ARD)
Conjunctivitis
Pharyngoconjuctival fever
Epidemic keratoconjunctivitis
Gastroenteritis
Acute hemorrhagic cystitis
Pertussis-like syndrome ( whooping cough)
ADENOVIRIDAE
DIAGNOSIS

Cell culture

Immunofluorescence

Neutralization test or restriction endonuclease

mapping of viral DNA

Antigen detection using electron microscopy, ELISA,

RIA, DNA hybridization assay
 PICORNAVIRIDAE
ss-RNA, icosahedral, naked
3 important groups in human:
ENTEROVIRUS - acid-stable
Poliovirus - 3 serotypes
Coxsackie A virus- 26 serotypes
Coxsackie B virus - 6 serotypes
ECHO virus - 33 serotypes
Enterovirus - 4 serotypes
HEPATOVIRUS - acid-stable
Hepatits A
RHINOVIRUS - acid-labile
PICORNAVIRIDAE:
Enteroviruses
POLIOVIRUS

Methods of transmission: Fecal-oral means, respiratory droplets (less often)

Clinical manifestations:

1. Inapparent infection - 90-95% of cases

2. Abortive infection - 4-8%

3. Frank cases - 1-2%

Anterior horn cells of the spinal cord - most frequently affected

Vaccines:

SABIN - live attenuated virus vaccine (oral)

SALK - inactivated virus vaccine - parenteral

PICORNAVIRIDAE:
RHINOVIRUSES

Replicate better at 33C than 37C

Most common cause of "common colds"

>110 serotypes
ORTHOMYXOVIRIDAE
ss-RNA, segmented, helical, enveloped
ORTHOMYXOVIRIDAE
3 Antigenic Groups: based on type of
ribonucleoprotein (RNP)

1. Influenza A - man, animals (birds, pig, etc.) shift,

drift

2. Influenza B - man

3. Influenza C - man
Antigenic Variations
Influenza C Shift, drift Pandemic,
A Epidemic
Man,
animals
Drift
Influenza Epidemic
B
Man
Drift
Sporadic
Influenza
Man
 ORTHOMYXOVIRDAE
M.O.T: airborne

CLINICAL MANIFESTATIONS:

Fever, malaise, headache, gen, aches and pains, sometimes with nasal discharge and
sneezing, non-productive cough, myalgia, anorexia, and shivering

Diagnosis:

Cell culture - no CPE

Hemadsorption technique

Hemagglutination (HA) test - the virus agglutinates rbc

Identification: hemagglutination inhibition test (HAI) - using specific antisera

Fluorescent Ab technique - respiratory secretions

Serology - CF Ab and HAI Ab

PARAMYXOVIRIDAE
ssRNA, helical, nonsegmented, enveloped
PARAMYXOVIRIDAE
MEMBERS:

1. Genus Paramyxovirus No. of Serotypes

Species: Parainfluenza V. 1-4

Mumps V. 1

2. Genus Morbilivirus

Species: Measles V. 1

3. Genus: Pneumovirus

Species: RSV 2
PARAINFLUENZA
VIRUSES
M.O.T: Thru Respiratory Droplets

Adults and older childern - Mild URTI "common colds"

Infants and young children - Severe LRTI

Croup ( laryngotracheobronchitis)

Bronchitis (Bronchiolitis)

Pneumonitis
MUMPS VIRUS
Epidemic parotitis

M.O.T - thru the saliva

CLINICAL MANIFESTATIONS:

Parotitis (involves other salivary glands)

Orchitis

Pancreas, epididymis, prostate, ovary, breast, meninges - rarely

affected

Aseptic meningitis ( rarely meningoencephalitis)

MEASLES (RUBEOLA)
VIRUS
M.O.T. - droplets

Signs and Symptoms:

Fever, cough, coryza, and conjuctivitis (3C's)

KOPLIK spots - like "grains of salt" in the buccal

mucosa

Maculo papular rash

SSPE ( Subacute Sclerosing Panencephalitis)

RESPIRATORY
SYNCYTIAL VIRUS
Similar to paramyxovirus

Recognized as the single most important respiratory pathogen of

infants and young children

MOT - respiratory droplets

CLINICAL FORMS:

1. "Common colds" - adults and older children

2. Croup - 1-3 years of age

3. Bronchiolitis and pneumonitis - <1 yr. most common cause

RUBELLA VIRUS
A Togavirus : ssRNA, icosahedral, enveloped

Only one serotype

Not transmitted by athropods

Transmission: inhalation of contaminated respiratory

droplets, transplacental

1st trimester transmission in pregnancy

 RUBELLA VIRUS
Rubella (German Measles) (3-days rash)

Mild macilopapular rash (face and thorax)

Retroauricular lyphadenopathy ( usually bilateral)

Mild fever, joint pain

Congenital Rubell

Fetal defects

Blueberry muffin baby

Cataracts, cardiac lesions, microcephaly and deafness

Risk and severity

Infection during the first trimester - greatest risk for the fetus
RUBELLA VIRUS
Diagnosis:

INFANTS:

TORCH titers - usually obtained because of difficulty of differentiating congenital infections from
one another

High or rising CF of HAI titers

Elevated specific IgM (Infection)

Viral culture from respiratory secretions or urine

PREGNANT

Serologic test: positive serum for rubella virus antibodies - previous infection

Specific anti-rubella IgM - recent infection

Negative serologic test:

Test again n 28 days (18 days incubation time plus 10 days to develop antibody)

If positive on second test - recent infection

Vaccine: live attenuated virus

 RABIES
(RHABDOVIRIDAE)
ssRNA, helical, envelop, BULLET-SHAPED

Transmission:

Animal bite: majority

Via scratch: occassionally

Inhalation of contamainated aerosolized animal materials: rarely

Sylvatic Rabies - rabies in wild animals

Urban Rabies - rabies in domestic animals

Clinical Syndrome

Incubation period: 14-32 months

The location of the bite influences the length of the incubation period

Hydrophobia s the most characteristic symptom

Coma and death

Death occurs in more than 90% of cases

RABIES
(RHABDOVIRDAE)
Diagnosis

Immunofluorescence - demostration of NEGRI

BODIES in the brain of the dog
 DENGUE VIRUS
FLAVIVIRIDAE (arboviruses)

4 SUBTYPES

Vector: Aedes aegypti, Aedes albopictus

"Saddleback fever"' "breeakbone fever"

Disease manifest in the form of

Classic Dengue fever (mild)

Acute Hemorrhagic Fever. Reinfection with another serotype

Dengue Shock Syndrome (most severe)

LABORATORY DIAGNOSIS:

Thrombocytopenia and hemoconcentration

Virus isolation

Serology - 4 fold rise in antibody titer, NS1 Ag

TREATMENT: Supportive

PREVENTION: Eradication of vector

RETROVIRODAE
ssRNA, helical or icosahedral (depending on
subfamily), enveloped

3 concepts unique to retroviridae

Carry a reverse transcriptase

Can cause cancer in cell they infect

Cytotoxic to certain cell

 HIV
Modes of Transmission

Sexual activity - most common; present in seminal fluid and vaginal and
cervical secretions

Blood product transfusion

IV drug use with needle sharing

Transplacentally 30%, during delivery and perinatally

Stick with a needle contaminated with infected blood (0.3%)

Tropic for cells bearng the CD4 molecule

T helper cells

Macrophages

Glial cells
 OPPORTUNISTIC INFECTIONS IN
AIDS
VIRAL
Disseminated CMV (lungs, retina, brain)
HSV (lungs, GIT, CNS, skin)
JC papovavirus (brain- PML)
EBV ( hairy leukoplakia)
BACTERIAL
Mycobacterial (e.g. Mycobacterium avium-intracellulare, M. tuberculosis) disseminated,
extrapulmonary
Salmonella (recurrent, disseminated) septicemia
Pyogenic bacteria (Haemophilus, Streptococcus, Pneumococcus)
FUNGAL
Pneumocystis jerovici (pneumonia)
Candida albicans (esophagus, lung infection)
Cryptococcus neformans CNS
Histoplasmosis (disseminated, extrapulmonary)
Coccidiodes (disseminated and extrapulmonary)
 HIV
MARKERS OF INFECTIONS

(+) of virus itself

Viral antigens

HIV Abs - 6 weeks to develop

LABORATORY DIAGNSIS

Test use HIV Ags derived from disruption of

whole virus cultured in human derived cell
line

Methods

ELISA - screening test

WESTERN BLOT - confirmatory

gp120, gp41, p24

PCR
REOVIRIDAE
1 in infantile diarrhea

dsDNA, 10 segments, icosahedral, naked

Human pathogen

Rotavirus - wagon wheel like

4 serotypes

Transmission: fecal oral

Most common non bacterial cause of diarrhea in infants

May also cause diarrhea in adults

Diagnosis - usually does not require the laboratory

Stool: ELISA or RIA

UNCONVENTIONAL
AGENTS PRIONS
GENERAL CHARACTERISTICS

contain protein only ( no nucleic acid)

are small, proteinaceous particles

Humans and other animals encode a protein PrPc ( cellular prion

protein) which is in extended form

A modified prion (PrPsc)

Globular - resistant to proteases

Responsible for the disease

PRION- CAUSED
DISEASES
SCRAPIE ( sheep or goat)

KURU

CREUTZFELDT-JAKOB DISEASE

GERSTMANN-STRAUSSLER SCHEINKER
SYNDROME