SPECIMEN COLLECTION TO ADENOVIRIDAE Flashcards
[?], from the [?]
Taken at the [?]
RIGHT SPECIMEN; RIGHT SITE
RIGHT TIME
Stored and transported [?]
IMMEDIATELY
To ensure [?]
accurate laboratory results and prevent contamination.
: most crucial procedure
Pre – analytical
Store specimen at [?]
4C
Transportation that will take days at [?]
-70C
SPECIMENS
• Urine
• Throat Swab/Throat Washings
• Blood; Plasma
• Rectal swab
• Biopsy material
• Skin scrapings
• CSF
• Vesicle fluid
• Autopsy tissue
Respiratory Infection
Nasal or throat swabs, postnasal washings
Gastrointestinal Infection
Feces
Vesicular Rash
Vesicular fluid
Hepatitis
Serum, feces
CNS
CSF, throat swab, feces
AIDS
Unclotted blood
VIRAL GROUP
Respiratory Tract
Mouth or Gastrointestinal Tract
Skin
Respiratory Tract
• Adenoviridae
• Herpesviridae
• Poxviridae
• Picornaviridae
• Togaviridae
• Orthomyxovirid ae
Mouth or Gastrointestinal Tract
• Adenoviridae
• Herpesviridae
• Picornaviridae
• Reoviridae
• Flaviviridae
Skin: A. Mild Trauma
• Papovaviridae
• Herpesviridae
• Poxviridae
Skin: B. Injection
• Herpesviridae
• Hepadnaviridae
• Retropiridae
Skin: C. Bites
• Togaviridae
• Rhabdoviridae
• Flaviviridae
THREE APPROACHES
Direct detection of virus
Virus Isolation (Indirect detection)
Serology (Detection of Antibodies)
• Electron Microscopy
Direct detection of virus
• Light Microscopy “Inclusion Bodies”
Direct detection of virus
• Antigen detection tests
Direct detection of virus
• Molecular methods: PCR (Polymerase Chain Reaction) & Nucleic Acid Probes
Direct detection of virus
• Animal inoculation Inoculation of eggs
Virus Isolation (Indirect detection)
• In vitro Cell Culture
Virus Isolation (Indirect detection)
microscopic examination for viral inclusions
CYTOLOGY/HISTOLOGY
less sensitive than culture
CYTOLOGY/HISTOLOGY
helpful for viruses that are difficult to grow
CYTOLOGY/HISTOLOGY
Cytology:
herpes simplex, varicella zoster
Histology:
adenivirus, cytomegalovirus, measles, papilloma virus, parvovirus, rabies (infects the brain)
ELECTRON MICROSCOPY
Advantages
• Economical running costs?
• Detects ?
• Adaptable for the confirmation of ?
• ? required for result; results available the same day (fastest)
• Specific ?
cost
unculturable viruses
cytopathic effect
Short length of time
antiviral chemotherapy
ELECTRON MICROSCOPY
Disadvantages
• Requires ?
• Poor ?
• Large ?
skilled staff
sensitivity
capital outlay
Examples of viruses seen: gastroenteritis
o Calici virus
o Corona virus
o Enteric virus
o Adeno virus
o Norwalk
Examples of viruses seen: encephalitis
o John Cunningham (JC) virus
o Herpes Simplex Virus
o Measles
TYPES OF ELECTRON MICROSCOPY
IMMUNOFLUO RESCENT MICROSCOPY Examples:
Adenovirus, CMV, herpes simplex 1 and 2, influenza virus
antiviral antibodies are used to stain viral antigens in specimen and cultures
IMMUNOFLUO RESCENT MICROSCOPY
Black background
IMMUNOFLUO RESCENT MICROSCOPY
usually solid-phase and membrane Elisa are used
ENZYME IMMUNOASSA Y (EIA)
the basic test consists of antibodies bonded to enzymes, the enzymes remain able to catalyze a reaction yielding a visible discernible end product while attached to antibodies
ENZYME IMMUNOASSA Y (EIA)
ENZYME IMMUNOASSA Y (EIA)
Examples:
enteric adenovirus, rotavirus, RSV
Notes :
• Sample : Stool
• Collecting specimens, have to poke or collect in 3 different sites (e.g. left, right)
• Right amount is placed in a solution → shake → place in a cartridge → results in 10 to 20 mins
ENZYME IMMUNOASSA Y (EIA)
baby hamster kidney cell culture with Bgalactosidase gene that is expressed when cells are infected with the virus
ENZYME LINKED VIRUS INDUCIBLE SYSTEM (ELVIS)
ENZYME LINKED VIRUS INDUCIBLE SYSTEM (ELVIS)
Example:
Herpes simplex 1 and 2
different viruses grow on different cell lines
CONVENTIONAL CELL CULTURE
growth takes about 1 - 28 days
CONVENTIONAL CELL CULTURE
microscopically examined for cytopathic effects (CPE)
CONVENTIONAL CELL CULTURE
Cell rounding, granulation, clumping, vacuolation
cytopathic effects (CPE)
Giant multinucleated cells formation: Herpes simplex virus, Tzanck smear
cytopathic effects (CPE)
Cell fusion, lysis, flux, inclusion bodies
cytopathic effects (CPE)
confirmed with immunofluorescent stains
CONVENTIONAL CELL CULTURE
CONVENTIONAL CELL CULTURE
Examples:
Adenovirus, CMV, enteroviruses, RSV
THREE BASIC TYPES OF CELL CULTURES
made by dispersing cell (usually with trypsin) from freshly removed host tissues
PRIMARY CELL LINE
Unable to grow for more than a few passages in culture
PRIMARY CELL LINE
PRIMARY CELL LINE Example :
Primary Monkey Kidney
that have undergone a change that allows their limited culture (range from 50 up to 100 passages/generations) but that retain their normal chromosome pattern
DIPLOID CELL LINE
DIPLOID CELL LINE Example :
Human diploid fibroblast and Human diploid vaccine for Lassa virus/Rabies
cultures capable of more prolonged growth (“immortal”) that have been derived from diploid cell lines or from malignant tissues
CONTINUOUS CELL LINE
INTRACYTOPLASMIC INCLUSION BODIES
INTRANUCLEAR INCLUSION BODIES
BOTH INTRACYTOPLASMIC AND INTRANUCLEAR
NEGRI BODIES
Rabies virus
MOLLUSCUM BODIES
Molluscum contagiosum virus
GUARNIERI BODIES
Vaccinia virus
OWL’S EYE INCLUSION BODIES
Cytomegalovirus
COWDRY TYPE A BODIES
HSV, VZV
COWDRY TYPE B BODIES
Polio virus
TORRES BODIES
Yellow Fever Virus
WARTHIN-FINKELDEY BODIES
Measles virus
• Grow via conventional cell culture (Cytomegalovirus and Varicella zoster virus)
SHELL VIAL CULTURE
• Advantage: Rapid, within 1-2 days
SHELL VIAL CULTURE
• Direct test, latex coated with antibodies, rapid and inexpensive
LATEX AGGLUTINATION
• Example: Rotavirus
LATEX AGGLUTINATION
• Used in IS
• Agglutination = (+)
LATEX AGGLUTINATION
• Limited commercial availability
POLYMERASE CHAIN REACTION (PCR)
• Enterovirus in CSF
POLYMERASE CHAIN REACTION (PCR)
• Most common: used during the pandemic (confirmatory test for + result)
POLYMERASE CHAIN REACTION (PCR)
• Specific IgG assays are good indications of prior infections
SEROLOGICAL TEST
• IgM assays are good indicators of recent or recent infections
SEROLOGICAL TEST
• Allow retrospective diagnosis if no acute clinical samples are available
SEROLOGICAL TEST
• Rapid
SEROLOGICAL TEST
• May be automated
SEROLOGICAL TEST
• Allows diagnosis if uncultured viruses are involved
SEROLOGICAL TEST
• Can make use of noninvasive clinical samples such as urine saliva
SEROLOGICAL TEST
• CF test is usually insensitive
SEROLOGICAL TEST
• Cross-reactivity of antibodies is a source of false positive results v Insensitive for congenital infections
SEROLOGICAL TEST
• Not appropriate for immunocompromised patients
SEROLOGICAL TEST
SEROLOGICAL TEST Example : in Isolating the virus
Antigen detection test by Immunofluorescence
Complement fixation
Hemagglutination inhibition
Enzyme immunoassay
DETECTED IN SERUM ONLY
SEROLOGICAL TEST
Useful in evaluating immune status or diagnosing viral infections where culture is difficult or impossible
SEROLOGICAL TEST
SEROLOGICAL TEST Example virus :
Arbo virus
Cytomegalovirus
Epstein- Barr virus
Hepatitis
Herpes simplex
Measles
Rubella
Varicella zoster virus
Adenoviridae
a. Human Adenovirus
• Hepadnaviridae
a. Hepatitis B Virus
• Herpes viridae
a. Herpes Simplex Virus 1
b. Herpes Simplex Virus 2
c. Varicella-Zoster Virus
d. Cytomegalovirus
e. Epstein-Barr Virus
f. Human Herpes Virus 6,7 and 8
• Papovaviridae
a. Human Papilloma Virus
b. JC and BK Polyoma Virus
• Parvoviridae
a. Parvovirus B-19
• Poxviridae
a. Variola
b. Vaccina
c. Orf
d. Molluscum contagiosum
• Double-stranded (except)
Parvoviridae
• Icosahedral (except)
Poxviridae
• Replication in the nucleus (except)
Poxviridae
• Naked:
Papoviridae, Parvoviridae, Adenoviridae
• Enveloped:
Poxviridae, Herpesviridae, Hepadnaviridae
subspecie
Human Adenovirus
over 80 species with about 40 are pathogenic to man
Human Adenovirus
70-90 nm
Human Adenovirus
Isolated from humans and animals
Human Adenovirus
produces cytopathic changes
ADENOVIRIDAE
rounding, enlargement and aggregation
ADENOVIRIDAE
Diseases
• Acute Febrile Pharyngitis: common in young children
• Pharyngeal-conjunctival Fever
• Eye infections
• Pneumonia and gastroenteritis
- Common in newborn, immunosuppressed and military men
Eye infections
No ab produced
ADENOVIRIDAE
Cytotoxic T-cells kill infected cell early
ADENOVIRIDAE
Common to children below 6 y/o
ADENOVIRIDAE
Predominant in cold climate
flu-like symptoms; not fatal
ADENOVIRIDAE
ADENOVIRIDAE SAMPLE COLLECTION
throat swab, eye samples, stool
ADENOVIRIDAE DIAGNOSIS
cell culture, serologic test and control
ADENOVIRIDAE CONTROL
vaccine; Anti-flu vaccine