Transcript Slide 1

Laboratory Diagnostics,
Specimen Collection,
and Biosafety Issues
Learning Objectives
• Prepare and maintain collection kits
• Collect and transport specimens safely and
correctly
• Manage laboratory specimens safely
• Interpret laboratory data
The Specimen
Collection Kit
Specimen Collection Kit
• Collection vials with
VTM
• Polyester fiber-tipped
applicators
• Sterile saline
(0,85% NaCl)
• Sputum or mucus trap
• Tongue depressors
• Specimen collection
cups or Petri dishes
• Transfer pipettes
• Secondary container
• Ice packs
• Items for blood
collection
• Personal protective
equipment
• Field collection forms
• A pen or marker for
labeling samples
How to Manage Kits
• Store specimen collection kits in a dry,
cool place
• Store specimen collection kit where it
will be accessible after hours and on
weekends
How to Safely And Correctly
Collect Samples
Clinical Specimen Sources
Be prepared to collect specimens
before you leave for the field
• Suspected cases
– Symptoms consistent with influenza
• Contacts
– Including people living or working with
suspected cases
What to Collect
Preferred specimens
• Oropharyngeal swabs
• Lower respiratory tract specimens
– Bronchoalveolar lavage or tracheal aspirates
Other specimens
• Nasal swab
• Nasopharyngeal swabs
Collect the sample on several different days
When to Collect
Respiratory Specimens
• As soon as possible after symptoms begin
• Before antiviral medications are
administered
• Even if symptoms began more than one
week ago
• Collect multiple specimens on multiple days
Personal Protective
Equipment
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Masks (N-95 or N/P/R-100)
Gloves
Protective eye ware (goggles)
Hair covers
Boot or shoe covers
Protective clothing (gown or apron)
Field Data Collection Form
• Patient name
• Unique identification number
• Patient demographic information
• Patient’s health status
Specimen Tracking System
Maintain a database to track:
• Identification number
• Subject information
• Specimen collection date
• Specimen collection location
• Diagnostic test results
Specimen Storage, Handling,
and Transportation
How to Store Specimens
For specimens in VTM:
• Transport to laboratory as soon as possible
• Store specimens at 4 °C before and during
transportation within 48 hours
• Store specimens at -70 °C beyond 48 hours
• Do not store in standard freezer – keep on ice or in
refrigerator
• Avoid freeze-thaw cycles
– Better to keep on ice for a week than to have repeat freeze
and thaw
How to Store Specimens
For sera:
• Store specimen at 4 °C
• For both VTM specimens and sera,
avoid repeated freeze-thaw cycles
Packing Specimens for
Transportation
• Keep specimens at 4 ºC
– Fill a cooler with ice packs or coolant
packs
– Double-bag specimens if you use dry ice
• Include an itemized list of specimens
with identification numbers and
laboratory instructions
Transporting Specimens
• Initial testing at State Public Health Laboratory
• Call CDC Hotline before sending specimens for
reference testing
770-488-7100
• Send Priority Overnight
• Include inventory sheet and CDC case ID number
• Coordinate with the laboratory
Managing and Analyzing
Laboratory Data
Data Management Rules
• Double check data entry accuracy
• Include unique identification numbers
• Keep subject names confidential
• Track testing dates and results
• Back up the database
Computer Software
• Epi-info (CDC)
– Free, at http://www.cdc.gov/epiinfo/
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Microsoft Excel and Access
Oracle
MySQL
Filemaker Pro
Data Analysis Measures
Prevalence
The number of cases divided by the size of the
population
Sensitivity
The number of cases identified by the test divided by
the number of true positives which were identified by
the gold standard technique
Specificity
The number of non-cases identified by the test
divided by the number of true negatives that were
identified by the gold standard technique
How to Present Results
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Report:
Time and place of the outbreak
Prevalence of infection
Clinical information about cases
Epidemic curve
Share results with local health officials
and CDC
Laboratory Tests
Laboratory Diagnosis for
Avian Influenza
Tests for respiratory samples:
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PCR-based techniques
Virus isolation
Immunofluorescence
Rapid antigen detection
Blood used for:
• Measurement of specific antibodies (sera)
• Viral isolation (whole blood)
• PCR-based techniques (sera)
Laboratory Tests
Virus Isolation
– Gold standard for detection of avian influenza
– Technically difficult and requires BSL-3 laboratory
PCR Based Techniques
• Sensitivity depends on the particular test, the
influenza strain, and the type of specimen
used
– Sensitivity:
– Specificity:
80 - 100%
90 - 100%
Serological Tests
Paired serum samples are most useful
Acute sample
Within 7 days after symptom onset
Convalescent sample
More than 12 days after symptom onset
Interpreting Serology Tests
Acute
Sample
low
low
high
Convalescent
Sample
Interpretation
low
No evidence of
Infection
high
Seroconversion
(evidence of infection)
high
Inconclusive; past
exposure to avian
influenza possible
Summary
• Maintain adequately stocked specimen
collection kits and store them properly
when they are not in use.
• Oropharyngeal swabs and lower
respiratory specimens are the best
specimens to collect.
• Collect multiple specimens (respiratory
and blood) on multiple days.
Summary
• Proper specimen storage, handling, and shipping is
vital to a successful laboratory test.
• Keep track of information on the specimens
collected in a database or logbook.
• When handling infectious materials in the laboratory
or in the field, take safety precautions.
• Properly dispose of any infectious material.
Summary
• Most states will be able to perform some
basic influenza identification tests on a
clinical specimen, but more specific testing
may need to be done at a higher level
national or regional laboratory.
• When you report on the progress of an
outbreak investigation, share investigation
results with local health officials and CDC.
Laboratory Practice
Exercise
Glossary
Centrifuge
A machine that uses high-speed rotation to separate
materials with different densities.
Culture
The growing of microorganisms in a nutrient-rich
medium.
Nucleic acid
A component genetic material such as DNA or RNA
found in all cells in humans, animals, bacteria, and
viruses. Every species and organism has a unique
pattern.
Glossary
Pipette
A glass or plastic tube used to measure or transfer
small amounts of liquid.
Saline
A liquid solution made of salt and water.
Viral Transport Medium (VTM)
The preservative liquid in which specimens are
stored until they are tested.
References and Resources
• Recommended laboratory tests to identify avian
influenza A virus in specimens from humans. World
Health Organization, June, 2005.
http://www.who.int/csr/disease/avian_influenza/guide
lines/avian_labtests2.pdf
• WHO guidelines for the collection of human
specimens for laboratory diagnosis of avian
influenza infection, 12 January 2005.
http://www.who.int/csr/disease/avian_influenza/guide
lines/humanspecimens/en/index.html