Avian Influenza – Real-time PCR

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This page provides molecular testing information for avian influenza at Public Health Ontario (PHO). For information regarding other testing options, refer to the following PHO webpages:

Testing Indications

PHO’s laboratory is currently responding to cases for highly pathogenic avian influenza (H5N1) in birds across Ontario. The investigation number ONT-2022-00001 should be included on all requisitions for local exposures related to infected birds. Testing for symptomatic individuals will include COVID-19, avian influenza and multiplex respiratory virus PCR. 

Specimen Collection and Handling

Specimen Requirements

Test Requested Required Requisition(s) Specimen Type Minimum Volume Collection Kit

Avian Influenza

Nasopharyngeal swab, or throat swab

1 swab in the collection kit media

Virus Respiratory Collection Kit (Order number 390082)

Avian Influenza

Bronchial Alveolar Lavages (BAL), or pleural fluid

2.0 ml

Sterile container

Virus-Enteric culture: Order number # 390087

Avian Influenza

Respiratory tract tissue

1.0 g

Sterile container

Virus-Enteric culture: Order # 390087

Avian Influenza

Stool (Do not submit until approved by PHOL microbiologist)

2.0 grams

Sterile container

Virus Enteric Collection Kit (sterile container, Order number 390087)

Avian Influenza

Blood/Serum (Do not submit until approved by PHOL microbiologist)

5 ml whole blood or 1.0 ml serum

Serum separator tubes (SST)

Submission and Collection Notes


Contact the local health unit if there is a suspected avian Influenza infection.


Prior to specimen submission, contact PHO’s Laboratory Customer Service Centre at 416-235-6556 or 1-877-604-4567 for further instructions and approval.


Complete all fields of the PHO’s General Test Requisition Form, including

  1. Tests requested (Avian Influenza, COVID-19, and Multiplex Respiratory Virus PCR/MRVP) and reason for test
  2. Patient setting, specimen types and sites
  3. Outbreak investigation number ONT-2022-00001
  4. Travel history and clinical information (see special instructions section below)

Label the specimen container(s) with the patient’s first and last name, date of collection, and one other unique identifier such as the patient’s date of birth or Health Card Number. Failure to provide this information may result in rejection or testing delay.


The following specimens should be collected on all patients being tested for avian influenza:

  1. respiratory tract samples (e.g., nasopharyngeal (NP) swab plus bronchoalveolar lavage (BAL) when possible); lung tissue if obtained (e.g. biopsy, post-mortem)
  2. a viral throat swab in viral transport media should also be collected on all hospitalized patients.

Sample types other than nasopharyngeal swabs, nasopharyngeal aspirates, bronchial washes (if available) or throat swabs require approval by a PHO microbiologist. Testing may be arranged on weekends if approved by a PHO microbiologist. Contact the After-hours Emergency Duty Officer at 416-605-3113 for after-hours consultation.


Stool and blood are not routinely tested for avian influenza but may be tested if avian influenza is detected in respiratory specimens. Submission requires approval of a PHO microbiologist.

Note: serology testing is not recommended for influenza diagnosis, including avian influenza, and is not available for clinical testing in Canada.

Storage and Transport

Place specimen in biohazard bag and seal. Specimens should be stored at 2-8°C following collection. Package and ship primary clinical samples to the local PHO laboratory in accordance with the Transportation of Dangerous Goods Regulations.

Mark the outer packaging containing avian Influenza specimens: ‘Attn: Molecular Diagnostics’.

Special Instructions

Complete all fields of the PHO General Test Requisition Form.

In Section 3 – Test(s) Requested order avian influenza, COVID-19 and multiplex respiratory virus PCR (MRVP) (See example below)
Travel history and clinical information must be provided. Indicate the following: travel to farms or areas experiencing avian influenza; symptoms including pneumonia; high risk status for respiratory viral infection complications; patient setting; admission status at time of collection (if known).

Avian Flu Test Requested

Requisitions and Kit Ordering

Test Frequency and Turnaround Time (TAT)

Influenza A testing is performed Monday to Friday.

The TAT for influenza typing is 1-2 days from specimen receipt at testing site.

Preliminary subtyping will be completed within 2-3 days from specimen receipt at testing site.

Positive samples will be forwarded to the National Microbiology Laboratory (NML) for confirmation.

Test Methods

Influenza A and B is tested with CDC in-house Real Time PCR assay.

Influenza A positive specimens will be subtyped with a separate real time RT-PCR.


Results are reported to the physician, authorized health care provider (General O. Reg 45/22, s.18), or submitter as indicated on the requisition.

Specimens that are positive for avian Influenzaare reported to the Medical Officer of Health as per Health Protection and Promotion Act.


Results are reported to the ordering physician or health care provider as indicated on the requisition.

Specimens that are positive for avian Influenza are reported to the Medical Officer of Health as per Health Protection and Promotion Act.

Mis à jour le 23 mai 2023