W4: Virology Part 1

Virology

Study of viruses

Obligate Intracellular organisms

They infect cells but are unable to reproduce itself outside host cell

Retroviruses

Most viruses contain only 1 copy of their gene except \_________

Electron microscope

How are viruses visualized?

20-300 nm

Diameter of viruses

Genes, enyzmes, antibiotics

Viruses lack \______ and \______ necessary for energy production and is resistant to \_______

Virion

It is the entire virus particle / infectious unit

Genome / nucleic acid core

It contains the genetic material (DNA or RNA)

Capsid

It is protein in nature and encloses / surrounds the genome

a.) Helical
b.) Icosahedral
c.) Complex

Types of Capsid

Poxviridae

Viruses with a complex capsid symmetry

Nucleocapsid

The genome enclosed in a capsid

Capsomeres

The morphological units on capsid surface / attached to the capsid

Protomers

These are the smallest functional unit equivalent to building units of the capsid

Enveloped

It is composed of lipids and polysaccharides and it is ether-sensitive

Naked / non-enveloped

It is ether-resistant / stable

Protein spikes

It facilitates transfer of nucleic acid from one host cell to another

Prions

These are infectious protein particles causing neurologic diseases in both animals and man

Scrapie

Transmissible spongiform encephalopathy:

Goats / sheeps

Kuru and Creutzfeldt-jakob disease

Transmissible spongiform encephalopathy:

Humans

Bovine Spongiform Encephalopathy (BSE)

It causes Mad-cow disease

a.) Attachment / adsorption
b.) Penetration
c.) Uncoating
d.) Eclipse phase
e.) Morphogenesis / assembly
f.) Release

Steps in viral replication

Tropism

It is the ability of the virus to infect only a number of particular cell types

Attachment / adsorption

It is characterized by interaction between the viral receptor and the host cell receptor

Penetration

The engulfment of the virus through endocytosis

Uncoating

It involves the release of viral contents
into the cell

Eclipse phase

Biosynthesis of viral components

72 hours

Specimens must be collected early in the disease or during the acute phase of the infection within \_________ of illness

2 - 3 weeks

When do we collect as convalescent specimen of the virus?

Directly

When collecting viral specimens, we should sample the infected site \_______

Aspirated secretions

These are best specimens for maximum recovery of the virus but swabs may also be used.

Synthetic swabs

If manner of collection is through swabbing, we need to use \_______

Dacron, nylon fibers

Swabs must be made with \________ or \_________

Calcium alginate

This swab must be avoided as this is toxic to HSV and may inhibit PCR amplification of respiratory virus

7.5% sodium bicarbonate

For urine specimens, what do we use to remove vaginal flora?

3 - 5 mL

Recommended volume for Blood Specimens

Heparin, citrae

If anticoagulated blood is required, the preferred
anticoagulant is \_______ or \_______ for the majority of
viruses

Serum

This is used for serologic testing for viruses

EDTA with polyester material (pearl top tube)

This anticoagulant is used when Nucleic acid testing is requested

EDTA

If anticoagulated blood is required for PCR
procedure, the preferred anticoagulant is \_______

Heparin

This anticoagulant inhibits DNA polymerase and therefore not to be used in nucleic acid testing

Room temperature

Specimens for virus isolation should not be allowed to sit in \________

Refrigerated

f there is a delay in transport, specimens should be \_______

2 - 8°C

Storage temperature for viruses for 72 hours

4°C

Storage temperature for viruses for 5 days

-20°C or -70°C

Storage temperature for viruses for 6 days or longer

Dilute or emulsify in viral transport media (VTM)

What should you do before freezing viral specimens?

1:2 or 1:5

What is the dilution used for viral specimens?

Phenol red

Indicator used for VTM

7.2

pH used for VTM

a.) Buffered Isotonic Solution
b.) Protein
c.) Antibiotics

Content of VTM

Protein

It acts as stabilizing agent for virus and antibiotic in VTM

Antibiotics

It is used to prevent bacterial overgrowth for VTM

Throat swab

Specimen for influenza viruses

Rectal swab

Specimen for Norwalk agent

Urine

Specimen for CMV

Blood and CSF

Specimen for togavirus

Urine

To rule out congenital infection, the preferred
specimen is?

Direct microscopy

It is considered as the least sensitive of all virus detection method

Pox virus

Largest DNA virus

Light microscopy

It is used to visualize Pox viruses

Electron microscopy

It is used to detect virus morphology

Potassium phosphotungstate and Uranyl acetate

Stains for electron microscopy

Phase Contrast Microscope

It is used to detect viral inclusions

Rabies virus

Negri bodies

Fowl Pox virus

Bollinger bodies

Variola and Varicella

Guarneri Paschen Bodies

Yellow Fever Virus

Torres / Councilman bodies

Tzanck smear

It is used to detect VZV and HSV inclusions from a stained smear of cells taken from the base of skin vesicle

Koilocytes

PAP stained smear showing \________ may be diagnostic of HPV infection

Viral culture / Cell culture

Gold standard for virus detection

Viral culture / Cell culture

Most widely used for virus isolation (shell vial)

35 - 37°C

For viral culture, incubation is done at \_______

Primary cell culture

Cell culture that uses normal mature adult cells

Kidney cells (rhesus monkey or rabbit kidney)

Most common primary cell culture

Diploid cell line / Semi-continuous cell lines

Cell culture that uses embryonic or fetal cells

50 times

How many times can Diploid cell line / Semi-continuous cell line be passage?

Continuous cell lines / heteroploid

Cell culture that can be passage in definitely

Continuous cell lines / heteroploid

Cell culture that uses malignant or cancer cells

Human Fetal Diploid

Cell culture for isolation of VZV & HSV

Cervical cancer cells

Source of HeLa cells

Cytopathic Effect (CPE)

It is the change in either nucleus or cytoplasm due to the presence of a virus

Microscope

It is used to examine CPEs

Negative

Quantification:

Uninfected monolayer

Equivocal

Quantification:

Atypical alteration of monolayer involving few cells

1+

Quantification:

1% to 25% of monolayer exhibits cytopathic effects (CPE)

2+

Quantification:

25% to 50% of monolayer exhibits cytopathic effects (CPE)

3+

Quantification:

50% to 75% of monolayer exhibits cytopathic effects (CPE)

4+

Quantification:

76% to 100% of monolayer exhibits cytopathic effects (CPE)

Fluorescein Isothiocyanate

Most common immunofluorescent dye for direct and indirect immunofluorescent

1+

Intensity:

Faint yet unequivocal apple-green fluorescence

2+

Intensity:

Apple-green fluorescence

4+

Intensity:

Brilliant apple-green fluorescence

Hemadsorption

Attachment or adherence of RBCs to the surface
of virus

a.) Respiratory and diarrhea viruses may cause
disease before antibody rise
b.) Extensive antigenic cross reactivity

Limitations of Serology

Parvoviruses

All DNA viruses are double stranded except \______

Poxviruses

All DNA viruses are icosahedral except \________

Poxvirus

DNA virus with a complex capsid symmetry

a.) Papillomaviruses
b.) Adenovirus
c.) Polyomaviruses
d.) Parvovirus

Which DNA viruses does not have an envelope?

Poxviruses

Which DNA virus does not multiply in the nucleus?