病毒性呼吸道檢體採集 之安全措施 衛生署 疾病管制局 中區傳染病防治醫療網 王任賢 指揮官

Learning Objectives Know the basic components of specimen collection kits for use with suspect avian influenza cases Know what specimens to collect, how to collect them, store them, and transport them to the laboratory Understand laboratory safety precautions

Session Overview Specimen collection Specimen storage, handling, shipping Safety precautions in the laboratory Working with reference laboratories Managing laboratory data

The Specimen Collection Kit

Specimen Collection Kit Personal protective equipment Collection vials with VTM Polyester fiber- tipped applicators Tongue depressors Items for blood collection Secondary container/ cooler Ice packs Suspect case forms A pen or marker for labeling samples Labels

Personal Protective Equipment Gloves Mask Gown Eye protection

Polyester Fiber-Tipped Applicator Should be drayon, rayon, or polyester fiber swabs Do not use calcium alginated or cotton swabs nor ones with wooden sticks; they inhibit PCR

How to choose VTM Can be made in a lab or purchased Different types of VTM: –For collection of animal specimens –For viral isolation –For molecular testing (Do not use phosphate-based media If VTM is not available, 100% ethanol can be used for molecular testing

Viral Transport Media

Storing VTM Sterile collection vials containing 2-3 ml of VTM Vials can be stored in a freezer at -20 ºC until use Vials can be stored for short periods of time at ºC

Managing Media Stock Keep records of when the VTM was made Do not use vials if the liquid becomes cloudy

Phlebotomy Supplies Tourniquet Disposable needles Vacuum tubes with EDTA Plastic needle holder Alcohol and iodine swabs Gauze Band-aids Biohazard sharps container

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 Specimens Image obtained from Target region for seasonal influenza Target region for H5N1 detection

Clinical Specimen Collection Responsibility Usually a hospital staff function May be a rapid responder function if hospital staff are unavailable Designate at least one member of the team to be trained to collect specimens

When to Collect CDC recommends laboratory testing for: Suspected cases –Symptoms consistent with influenza –Epidemiologic link to avian influenza A (H5N1)

Updated Interim Guidance for Laboratory Testing of Persons with Suspected Infection with Avian Influenza A (H5N1) Virus in the United States Testing for avian influenza A (H5N1) virus infection is recommended for a patient who has an illness that: requires hospitalization or is fatal; AND has or had a documented temperature of ≥100.4° F; AND has radiographically confirmed pneumonia, acute respiratory distress syndrome (ARDS), or other severe respiratory illness for which an alternate diagnosis has not been established; AND has at least one of the following potential exposures within 10 days of symptom onset:

A) History of travel to a country with influenza H5N1 documented in poultry, wild birds, and/or humans, AND had at least one of the following potential exposures during travel: direct contact with (e.g., touching) sick or dead domestic poultry; direct contact with surfaces contaminated with poultry feces; consumption of raw or incompletely cooked poultry or poultry products; direct contact with sick or dead wild birds suspected or confirmed to have influenza H5N1; close contact (approach within 1 meter [approx. 3 feet]) of a person who was hospitalized or died due to a severe unexplained respiratory illness;

B) Close contact (approach within 1 meter [approx. 3 feet]) of an ill patient who was confirmed or suspected to have H5N1; or C) Worked with live influenza H5N1 virus in a laboratory.

Case by Case Considerations! Mild or atypical disease (hospitalized or ambulatory) with one of the exposures listed above Severe or fatal respiratory disease whose epidemiological information is uncertain, unavailable, or otherwise suspicious but does not meet the criteria above

What to Collect Preferred specimens Oropharyngeal swabs Lower respiratory tract specimens –Bronchoalveolar lavage or tracheal aspirates Other specimens Nasopharyngeal swabs Nasal swabs and aspirates Acute and convalescent sera Sputum specimens Collect samples on several different days

When to Collect Respiratory Specimens As soon as possible after symptoms begin Ideally before antiviral medications are administered Collect multiple specimens on multiple days

Serological Samples Paired serum samples are most useful Acute sample Within 7 days after symptom onset Convalescent sample 2 to 4 weeks after acute sample

Personal Protective Equipment Masks (N-95 or N/P/R-100) Gloves Protective eyewear (goggles) Hair covers Boot or shoe covers Protective clothing (gown or apron)

Collecting Specimens

Suspect Case Form Patient name Unique identification number Patient symptoms and date of onset Specimens and collection date Whether or not patient is hospitalized Patient contact information Patient demographic information

Specimen Tracking System Maintain a database to track: Identification number Subject information Type of specimen Specimen collection date Specimen collection location Date of laboratory receipt of specimen Date tested Diagnostic test results

Nasopharyngeal Swab 1.Insert dry swab into nostril and back to nasopharynx 2.Leave in place for a few seconds 3.Slowly remove swab while slightly rotating it

Nasopharyngeal Swab continued 4.Use a different swab for the other nostril 5.Put tip of swab into vial containing VTM, breaking applicator’s stick

Oropharyngeal Swab 1.Ask the subject to open his or her mouth 2.Depress the tongue 3.Swab the posterior pharynx 4.Avoid the tonsils

Nasopharyngeal Aspirate Collection Process 1.Attach mucus trap to vacuum source 2.Place catheter into nostril parallel to palate 3.Apply vacuum 4.Slowly remove catheter while slightly rotating it 5.Repeat with other nostril using the same catheter 6.After collection, flush catheter with 3 ml VTM and return VTM to a plastic vial

Blood Collection for Serology Only by trained persons Collect red-top tubes Freeze sera separated from cells at - 20 º C

How to Label Samples Use pre-printed barcode labels: –On the specimen container –On the field data collection form –On the log book Label each specimen with: –Subject’s name –Subject’s unique identification number

Specimen Storage, Handling, and Transportation

How to Store Respiratory 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 dry ice or in refrigerator Avoid freeze-thaw cycles –Better to keep on ice for a week than to have repeat freeze and thaw

Handling Infectious Materials in the Field Always wear personal protective equipment Be careful with sharp objects Treat all clinical samples as potentially infected with avian influenza

Packing Specimens for Transportation Use three packaging layers First layer should be water tight Use absorbent material in all layer No more than 500 mL should be in the specimen container

Transporting Specimens from Field to Lab

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 Packing Specimens for Transportation

Transporting Specimens Initial testing at State Public Health Laboratory Call CDC Hotline before sending specimens for reference testing Send overnight or using appropriate means based on recommendations from the State Department of health or CDC Include inventory sheet and CDC case ID number Coordinate with the laboratory

Laboratory Biosafety Laboratory testing of suspected novel influenza viruses should be conducted in biosafety level 3 (BSL-3) laboratories –Infectious agents that may be transmitted via the airborne route –PPE for laboratory personnel may include respirators

Summary When handling infectious materials in the laboratory or in the field, take safety precautions. Proper specimen storage, handling, and shipping are vital to successful laboratory tests. Maintain adequately stocked specimen collection kits and store them properly when they are not in use.

Summary Oropharyngeal swabs and lower respiratory specimens are the best specimens to collect for avian influenza A (H5N1). Collect multiple specimens (respiratory and blood) on multiple days. Keep track of information on the specimens collected in a database or logbook. 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. Results should be confirmed by the CDC. When you report on the progress of an outbreak investigation, share investigation results with local, regional, and federal health officials.