The Canadian Pandemic Influenza Plan for the Health Sector

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Annex C
Pandemic Influenza Laboratory Guidelines

Table of Contents

1. Preamble
2. Executive Summary
3. Key PILPN Recommendations
4. Laboratory Testing
   1. Interpandemic Period – Canadian Pandemic Phases 1.0, 1.1, 2.0, 2.1
      1. Methodology
      2. Specimen Type, Collection and Transport
      3. Disinfection Procedures
      4. Biosafety Considerations
      5. Antiviral Susceptibility
      6. Research
   2. Pandemic Alert Period – Canadian Pandemic Phases 3.0, 3.1
      1. Methodology
      2. Specimen Type, Collection and Transport
      3. Disinfection Procedures
      4. Biosafety Considerations
      5. Antiviral Susceptibility
      6. Research
   3. Pandemic Alert Period – Canadian Pandemic Phases 4.0, 4.1, 4.2, 5.0, 5.1, 5.2
      1. Methodology
      2. Specimen Type, Collection and Transport
      3. Disinfection Procedures
      4. Biosafety Considerations
      5. Anti-viral Susceptibility
      6. Research
   4. Pandemic Period – Canadian Pandemic Phases 6.0, 6.1, 6.2
      1. Methodology
      2. Specimen Type, Collection and Transport
      3. Disinfection Procedures
      4. Biosafety Considerations
      5. Antiviral Susceptibility
      6. Research
   5. Postpandemic Period
5. Surveillance and Data Collection
   1. Interpandemic Period – Canadian Pandemic Phases 1.0, 1.1, 2.0, 2.1
   2. Pandemic Alert Period – Canadian Pandemic Phases 3.0, 3.1
   3. Pandemic Alert Period – Canadian Pandemic Phases 4.0, 4.1, 4.2, 5.0, 5.1, 5.2
   4. Pandemic Period – Canadian Pandemic Phases 6.0, 6.1, 6.2
   5. Postpandemic Period
6. Communication Activities
   1. Interpandemic/Pandemic Alert Period – Canadian Pandemic Phases 1.0-5.2
   2. Pandemic Period – Canadian Pandemic Phases 6.0, 6.1, 6.2
   3. Post Pandemic Period
7. Pandemic Preparedness
   1. Planning Assumptions
   2. Business Continuity Considerations
   3. Quality Assurance and Quality Control
8. Appendices
   • Appendix A - Acronyms
   • Appendix B –Minimum requirements for the provision of public health laboratory services during pandemic influenza
   • Appendix C – Province of Québec’s Preparation for an Influenza Pandemic – Rapid Tests: Value and Limitations
   • Appendix D – Nasopharyngeal Swab Procedure
   • Appendix E – Draft Pandemic Influenza Laboratory Advisory
   • Appendix F – Laboratory Pandemic Influenza Communication Flow: Pandemic Alert Period
   • Appendix G –F/P/T Laboratory Services during a Pandemic
   • Appendix H – Members of the Pandemic Influenza Laboratory Preparedness Network

I. Preamble

The Pandemic Influenza Laboratory Preparedness Network (PILPN) of the Canadian Public Health Laboratory Network (CPHLN) has developed this document in accordance with the defined phases of the Canadian Pandemic Influenza Plan for the Health Sector. Laboratory testing, laboratory-based surveillance and data collection, communication issues and pandemic preparedness are addressed from the perspective of the Canadian pandemic phases. This document provides general guidelines to facilitate a consistent approach to laboratory testing for influenza during the Interpandemic, Pandemic Alert and Pandemic Periods and is intended for laboratory professionals.

II. Executive Summary

In the event of pandemic influenza, laboratories will be instrumental in facilitating the delivery of rapid and appropriate public health responses. During a pandemic, laboratory testing will accomplish the following:

• identify the earliest Canadian cases of a novel influenza strain;
• support public health surveillance by monitoring the geographic spread of disease and the impact of interventions;
• facilitate clinical treatment by distinguishing patients infected with the pandemic influenza virus from those with other respiratory diseases until the pandemic strain has been shown to be the predominant virus in a community;
• monitor circulating influenza viruses for antiviral resistance; and
• assess influenza vaccine efficacy.

Annex C – Pandemic Influenza Laboratory Guidelines provides recommendations to Canadian influenza testing facilities on laboratory testing, surveillance and data collection, communication and pandemic preparedness planning. In addition, as the laboratory working group of the Pandemic Influenza Committee (PIC), PILPN has made recommendations as to the minimum requirements necessary for the provision of public health laboratory services during an influenza pandemic (Appendix B)

Laboratory responsibilities will depend upon the pandemic phase. During the Interpandemic and Pandemic Alert Periods, it is recommended that influenza testing laboratories perform testing in support of routine influenza surveillance, laboratory-based detection of novel influenza subtypes and preparedness planning.

Once human-to-human transmission of a novel influenza strain has become established in Canada, the demand for laboratory testing will reach unprecedented levels. Laboratories must be prepared to respond accordingly.

During the Pandemic Period (Phase 6.0–6.2), influenza testing laboratories will support epidemiological efforts to track the spread and trend of the pandemic, and to monitor antiviral resistance. It is assumed that during this period, diagnosis of influenza will be made primarily by clinical assessment.

In order to assist with preparations for a potential pandemic, this document provides a number of planning assumptions. Laboratories are encouraged to review laboratory functions, both influenza and non-influenza related. Human resource issues must also be considered. Most importantly, laboratories should focus on strengthening capacity before the arrival of a pandemic in Canada.

III. Key PILPN Recommendations

1. During seasonal influenza, PILPN recommends the use of rapid detection methods in conjunction with cell culture and nucleic acid testing (NAT) to aid in timely diagnosis, particularly during outbreaks. Serologic testing is of limited usefulness in the diagnosis of acute influenza and therefore is not recommended.

2. PILPN recommends that public health laboratories (PHLs) and local clinical laboratories develop influenza testing strategies for a pandemic. This should include the establishment of protocols to process and identify novel influenza subtypes that may be considered Risk Group 3 pathogens.

3. Laboratory testing algorithms should emphasize the use of NAT complemented by viral culture (in a certified containment level [CL]-3 laboratory) and validated direct fluorescent antibody assay (DFA). Ongoing development of NAT methods for the rapid detection of novel influenza subtypes can be undertaken using conventional or real-time reverse transcriptase polymerase chain reaction (RT-PCR) methods. This may include the utilization of a “universal” detection protocol that would identify any influenza A virus using primers to a conserved region within the genome and subsequent subtyping using primers specific for avian subtypes with pandemic potential, e.g. H5, H7 and human subtypes H1, H2, H3.

4. At present, point-of-care (POC) tests are not recommended for the detection of novel influenza subtypes. POC refers to any rapid test that utilizes rapid antigen detection techniques to identify the influenza virus. Refer to Appendix C for an excerpt on rapid testing taken from Québec’s Preparation for an Influenza Pandemic. The utility of these tests will be reassessed as data on their performance characteristics are obtained.

5. Other novel diagnostic protocols should be explored, such as the development of protocols for the simultaneous detection and subtyping of defined influenza A viruses.

6. In accordance with the response plan outlined by PIC, PILPN recommends that each province and territory (P/T) ensure that at least one laboratory within the P/T has the capability to determine the subtype of the influenza A virus and, if this is not possible, to establish appropriate alternative arrangements. This must include the development of NAT protocols that are capable of identifying a subtype of novel strains of influenza A. The National Microbiology Laboratory (NML) will supply the protocols, primers and reagents necessary to develop and evaluate these assays along with controls required for a quality assurance program.

7. PILPN recommends that all PHLs and other laboratories that routinely test for influenza submit data on influenza testing during the influenza season to the Public Health Agency of Canada (PHAC) on a weekly basis or more frequently if requested by PHAC. These data are reported and disseminated through the Respiratory Virus Detection System and FluWatch.

8. PILPN recommends that NML develop the capacity to produce and evaluate in-house production of alternative sources of reagents, such as monoclonal antibodies for DFA testing, which could be stockpiled and distributed to diagnostic laboratories across the country during a pandemic, when commercial reagents may be in short supply.

9. PILPN recommends that communication between animal and human influenza testing facilities be strengthened.

10. PILPN recommends that each province have an influenza surveillance committee in place. This should include a laboratory representative familiar with issues related to influenza diagnostics and pandemic planning in order to ensure that there is good communication among the provincial laboratory, provincial epidemiologists and health units. The committee will deal primarily with influenza in the event of a pandemic, but it will deal with other surveillance issues at other times as required. The committee should include, at a minimum, a provincial epidemiologist, the provincial laboratory director and the chief medical officer of health or respective designates.

Laboratory Testing

A. Interpandemic Period – Canadian Pandemic Phases 1.0, 1.1, 2.0, 2.1

1. Methodology

Influenza testing facilities will maintain routine laboratory diagnostic services for influenza by performing

• virus isolation in cell culture
• direct antigen detection, i.e. DFA, enzyme immunoassay (EIA), rapid POC testing*
• NAT, such as RT-PCR, or nucleic acid sequence-based amplification

Outside of influenza seasons, severe respiratory illness (SRI) cases will require a comprehensive workup using standard methods for respiratory pathogens.

2. Specimen Type, Collection and Transport

For seasonal influenza, a nasopharyngeal sample (NPS) is recommended as the preferred specimen because it yields the best results in most direct antigen tests as well as in cell cultures. Please see appendix D for the appropriate NPS procedure.

Throat swabs are not recommended because of their poor sensitivity for antigen- and culture-based assays. However, throat swabs and nasopharyngeal (NP) washings may be acceptable or recommended by the manufacturers of specific rapid detection kits.

Nasal swabs may be an acceptable alternative in children, particularly when a NAT is used.

Specimens should be collected as soon as symptoms develop, because viral shedding is maximal at the time of onset of illness and generally decreases to undetectable levels by five days in immunocompetent adults. Viral shedding may last longer in children and immunocompromised patients; hence, collection after five days of illness may still be useful in this setting.

Laboratories will follow appropriate Transportation of Dangerous Goods regulations for the shipment of influenza specimens.

3. Disinfection Procedures

Laboratories will follow appropriate disinfection procedures according to the specimen type.

4. Biosafety Considerations

Laboratories will follow appropriate biosafety guidelines, as prescribed by the Office of Laboratory Security, Centre for Emergency Preparedness and Response (CEPR).

5. Antiviral Susceptibility

Early-season and late-season isolates can be submitted to the NML for resistance testing to amantadine and neuraminidase inhibitors as agreed upon by NML in conjunction with PHLs. NML will undertake investigations related to surveillance for resistance in emerging and currently circulating strains.

6. Research

NML will collaborate in research and development for monitoring influenza vaccine efficacy, immunological response, as well as evolution and determination of cross-reactivity among strains in the population.

NML, in cooperation with the Pandemic Vaccine Working Group of PIC, will develop protocols to test vaccine recipients for immune response. PHLs or designated laboratories in cooperation with NML may also develop and institute similar protocols, depending on available resources and expertise. Assays may include hemagglutination inhibition (HAI) and/or other tests using the antigens that are included in the most current vaccines.

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*Because of the suboptimal positive predictive value during periods of low influenza activity, diagnosis by rapid POC tests must be interpreted with caution and confirmed by DFA, viral culture or NAT. Complete details regarding World Health Organization (WHO) recommendations on the use of rapid testing for influenza, including a review of the currently available kits, can be found at:
http://www.who.int/csr/disease/avian_influenza/guidelines/rapid_testing/en/index.html

B. Pandemic Alert Period – Canadian Pandemic Phases 3.0, 3.1

1. Methodology

During phase 3.0 diagnostic testing services capacity and approach will continue for seasonal influenza as in Phases 1.0, 1.1, 2.0 and 2.1. NML will be responsible for confirming any samples that test positive for novel influenza.

NML will provide the reagents and controls that will be essential in developing assays for newly emerging strains and that will establish quality assurance. NML will be responsible for phenotypic testing and distribution of information to PHLs. The genetic testing (i.e. identification by PCR) will be the responsibility of PHLs.

NML will give priority to reagent preparation for the identification of the new strain in readiness for Phase 3.1. It will distribute NAT and conventional culture protocols as appropriate.

During Phase 3.1, NML is responsible for confirming the results of any samples that test positive for novel influenza. It will also provide information regarding the novel influenza strain to the WHO and coordinate any further testing.

During Phase 3.1, PHLs and other diagnostic laboratories will be on high alert and will focus on the following:

• enhanced laboratory-based surveillance for the emerging new subtype
• implementation or augmentation of NAT protocols
• viral isolation by culture in appropriately equipped laboratories.

There will be an increased demand for testing with emphasis on the identification of the hemagglutinin (HA) type of the viruses identified. RT-PCR will be particularly useful for rapid detection and HA determination. Although viral isolation is encouraged to facilitate detection of the emergence of new subtypes within Canada, the number of laboratories that can perform culture in CL-3 is limited.

The use of commercially available rapid POC tests for the detection of a new subtype is not recommended because of the lack of information on their clinical accuracy. These tests may be able to rapidly identify and differentiate influenza A and B infections, but currently they do not differentiate different HA subtypes of influenza A and cannot differentiate human from avian influenza virus. Any findings from direct antigen or rapid POC tests obtained for patients suspected of being infected with a novel influenza virus must be confirmed by NAT and/or culture. If data become available outlining the efficacy of POC testing, PHLs will communicate these results to laboratories within their jurisdictions.

2. Specimen Type, Collection and Transport

An NPS is the optimal specimen for seasonal human influenza, but this may not be the case for novel influenza viruses, as has been recently reported for H5N1 infecting humans in Eurasia, where throat specimens are preferred. Because the optimal specimen type and timing of collection will not be fully known for novel influenza virus infections, particularly as they continue to evolve, PILPN recommends the collection of different types of respiratory specimens, including NP swabs, NP aspirate, nasal washings, throat swabs and sputa on multiple days. Consideration should be given to testing stool or plasma specimens in patients who have significant gastrointestinal symptoms, as H5N1 has been isolated in the stool and blood of infected patients.

Accurate diagnostic testing is vital, and confirmation of all findings of a novel subtype of influenza is essential. To facilitate this, specimens should ideally be collected in duplicate. If this is not possible or has not been done, the specimen should be divided into two aliquots before testing. One aliquot can then be used for NAT, and the other should be frozen at –70°C for future testing, if required.

Specimens should be shipped at 4^oC to the PHL or designated laboratory as soon as possible. If there is going to be a delay of more than three days, the specimen should be frozen at –70^oC and shipped on dry ice or else maintained at 4^oC until processed. For more information, please see the WHO guidelines Collecting, Preserving and Shipping Specimens for the Diagnosis of Avian Influenza A (H5N1) Virus Infection: Guide for Field Operations

Ongoing training of personnel regarding proper shipment of specimens to reference laboratories and adequate knowledge of procedures and regulations is recommended.

3. Disinfection Procedures

Hypochlorite is considered by the WHO as the best disinfection for use against H5N1 contamination and would likely be applicable to other novel subtypes of influenza A. Hypochlorite is one of the few disinfectants that can safely be used in laboratories where NAT work is undertaken.

Other disinfectants, such as alcohols and quaternary ammonium preparations, can precipitate nucleic acids, which can increase the chance of contamination of subsequent reactions and lead to false-positive results. Chlorine bleach fragments nucleic acids. The WHO suggests two different concentrations to be used under different circumstances:

• Disinfection of specimen spills should be carried out using a 0.5% chlorine bleach.
• Disinfection of surfaces, medical equipment, contaminated waste before disposal and reusable protective clothing before laundering should be carried out using a 0.05% solution as per WHO guidelines. As hypochlorite is caustic, it must be subsequently removed by wiping equipment or surfaces with clean water or 70% ethanol to prevent damage.

For more information on the preparation of and precautions to be taken with chlorine solutions, see the WHO’s Guide for Field Operations (see the end of section 2).

4. Biosafety Considerations

The Office of Laboratory Security of the CEPR has drafted interim biosafety guidelines regarding the handling of clinical specimens associated with novel influenza virus subtypes. These guidelines can be found in appendix E. It is important to note that these are interim guidelines: as information regarding any novel influenza strain becomes available the biosafety guidelines specific to that strain will be disseminated by CEPR in a timely fashion.

5. Antiviral Susceptibility

Isolates can be submitted to the NML for resistance testing to amantadine and neuraminidase inhibitors as agreed upon by NML in conjunction with PHLs. NML will undertake investigations related to surveillance for resistance in emerging and currently circulating strains.

Susceptibility testing of strains will be performed by NML and participating PHLs that have the protocols in place for neuraminidase inhibitors and/or amantadine, depending on the phenotypic characteristics of the pandemic strain. Specimens received by NML will be tested periodically throughout the pandemic as part of surveillance and to monitor the development of antiviral resistance.

In addition to routine surveillance testing, testing for antiviral resistance will be performed on specimens isolated from persistently infected patients (patients who are taking prophylaxis and/or are immunocompromised hosts) in outbreaks. Other testing will be done on specimens as determined by NML in collaboration with PHLs or any submitting diagnostic laboratory.

6. Research

If capacity exists, refer to Interpandemic Period Phase 1.0, 1.1, 2.0, 2.1.

C. Pandemic Alert Period – Canadian Pandemic Phases 4.0, 4.1, 4.2, 5.0, 5.1, 5.2

1. Methodology

During Phases 4.0 and 5.0, PHLs and other diagnostic laboratories will be on high alert and follow the protocol as in Phase 3.0 and 3.1:

• enhanced laboratory-based surveillance for the emerging new subtype
• NAT
• viral isolation by culture.

During Phases 4 and 5, a dramatic increase is expected in the demand for testing, especially in affected areas. NAT will be the primary method for rapid detection. Specimens positive for influenza A from patients with epidemiological features that suggest they are at risk of a novel subtype of influenza will be subtyped using RT-PCR.

Rapid subtyping of positive specimens will be performed by PHLs or designated laboratories.

NML, in consultation with the WHO, will review the primers used in NAT to ensure that they are effective in identifying the novel subtype. Additional supplies of appropriate cell lines may be required. NML, PHLs and designated diagnostic laboratories will share information and reagents for identification of the novel strain and will advise on cell lines, use of rapid test methodologies and the containment level required, etc.

Increased testing by culture in a certified CL-3 laboratory will be required to isolate the novel strain in suspected cases. Isolates or specimens from identified clusters will be forwarded to NML for strain characterization. PHLs and diagnostic laboratories will play a critical role in tracking the potential spread of the novel strain.

Once reference antisera are available, subtyping will be done using HAI and neutralization assays by laboratories with the appropriate containment facilities, as dictated by the containment level requirements of the novel strain. Other laboratories will rely on RT-PCR for rapid subtyping using previously established protocols.

All diagnostic laboratories will be under considerable pressure to provide rapid testing service from the standpoint of infection control and/or isolation as well as treatment and prophylactic options. The current national antiviral strategy is to provide treatment to all those presenting early for medical assessment who are deemed to require treatment, during the pandemic. However, there may be circumstances under which a more targeted approach to antiviral use may be required (e.g. if sporadic cases of novel influenza occur in Canada prior to a pandemic). Diagnostic laboratories will likely play a role, as the use of timely diagnostics may become an integral part of this strategy. PILPN will provide guidance to the Clinical Care Working Group on the utilization of testing, depending upon the available capacity.

2. Specimen Type, Collection and Transport

Refer to Pandemic Alert Phases 3.0, 3.1.

3. Disinfection Procedures

Refer to Pandemic Alert Phases 3.0, 3.1.

4. Biosafety Considerations

Refer to Pandemic Alert Phases 3.0, 3.1.

5. Anti-viral Susceptibility

Refer to Pandemic Alert Phases 3.0, 3.1.

6. Research

If capacity exists, refer to Pandemic Alert Phases 3.0, 3.1.

D. Pandemic Period – Canadian Pandemic Phases 6.0, 6.1, 6.2

1. Methodology

Depending on the extent and duration of the pandemic, the demand for testing could reach an unprecedented level, which may overwhelm the diagnostic abilities of PHLs and other diagnostic laboratories. The laboratories will continue to function as in Phases 4 and 5 with a focus on the following:

• NAT
• reduction of emphasis on viral culture in areas where the pandemic is established
• reduction of testing once the novel subtype is present in the jurisdictional population.

NML will need to prioritize specimen testing in order to prevent overloading its capacity. Specimens routed through PHLs or designated laboratories will have priority.

NML, in collaboration with the WHO, will review the primers used in NAT to ensure that they are effective in identifying the pandemic strain. NML will provide PHLs with information or reagents for identification of the pandemic influenza virus and advice on cell lines.

PHLs will need to redirect resources to give priority to influenza testing. However, when the pandemic has become established, it is assumed that laboratories within the affected regions will scale down testing because clinical case definition may be sufficient for diagnosis in most cases. This will depend on local conditions and available resources. PHLs and designated diagnostic laboratories will focus on tracking the spread and trend of the pandemic and monitoring antiviral resistance, depending on available resources.

PHLs and local laboratories are encouraged to review influenza testing protocols, the availability of reagents and human resource(HR) issues, and to implement the pre-developed strategies to reduce the impact of the pandemic on laboratory testing. Laboratories should review inventory and order necessary supplies and cell lines, more viral transport swabs, influenza antigen tests, antisera for DFA testing and laboratory personnel protective items, etc., as necessary.

Rapid subtyping of isolates will be done by PHLs or designated laboratories with back-up support from NML, which will use culture and NAT-based methods.

2. Specimen Type, Collection and Transport

Refer to Pandemic Alert Phases 3.0, 3.1.

3. Disinfection Procedures

Refer to Pandemic Alert Phases 3.0, 3.1.

4. Biosafety Considerations

Refer to Pandemic Alert Phases 3.0, 3.1.

As the pandemic progresses, PILPN will provide guidelines on testing and updates on antiviral susceptibility of the pandemic strain and other co-circulating strains.

5. Antiviral Susceptibility

Refer to Pandemic Alert Phases 3.0, 3.1.

6. Research

If capacity exists, refer to Pandemic Alert Phases 3.0, 3.1.

E. Postpandemic Period

This will mark a return to interpandemic activities. Any testing issues that arose during the pandemic will be reviewed to determine whether there are changes to the pandemic plan that could be implemented.

V. Surveillance and Data Collection

Laboratory surveillance, in combination with epidemiological data, will be critical during a pandemic to determine the phases that are in effect and thus trigger the necessary responses.

A. Interpandemic Period – Canadian Pandemic Phases 1.0, 1.1, 2.0, 2.1

As part of yearly ongoing surveillance, PHLs or designated laboratories must submit influenza isolates¹ to NML for subtyping and characterization, as noted in Table 1. These isolates must be submitted to NML promptly, along with the results of any subtyping or genotyping performed locally. NML will give priority to processing such specimens. Virus will be amplified in cell culture for subtyping by HAI and/or neutralization assays. For specimens that cannot be amplified by culture, the genotype will be determined after amplification of selected genes by RT-PCR and sequencing. NML will undertake to report the subtype to the submitting laboratory within a few days of receipt.

Table 1: The types of isolates to be submitted by PHLs or designated laboratories to NML for subtyping and characterization as part of the ongoing surveillance program

Phase 2.1 is defined as the period when “an animal influenza virus subtype that poses substantial risk to humans is circulating in animals in Canada”. During this phase laboratories will be testing isolates from cases of suspected animal-to-human transmission or any unusual isolates. Laboratory surveillance data processed during this phase will be used to determine whether the pandemic has moved into phase 3.1.

NML and selected PHLs will share subtyping and susceptibility testing technology as well as the development of rapid test(s) for detection of influenza, and better subtyping and susceptibility testing methods. They will also serve as sites for training other appropriate laboratories in these methods.

B. Pandemic Alert Period – Canadian Pandemic Phases 3.0, 3.1

In addition to subtyping and characterization of isolates as part of routine surveillance activities (Table 1), all influenza A positive specimens obtained from persons with epidemiological risk factors for a novel subtype of influenza A virus must be subtyped and, if they are not, must be forwarded for further testing to NML . PHLs will play a critical role in tracking the potential spread of the novel strain. Any positive influenza specimens obtained from a case with SRI and epidemiological links with a novel influenza virus subtype need to be confirmed by NML and rapidly characterized.

Appendix F outlines the communication flow between provincial and federal agencies in the event of a suspected novel influenza virus.

C. Pandemic Alert Period – Canadian Pandemic Phases 4.0, 4.1, 4.2, 5.0, 5.1, 5.2

Refer to Phases 3.0 and 3.1. Heightened laboratory-based surveillance will continue:

• monitoring genetic drift of the influenza virus;
• monitoring the emergence of new and co-circulating subtypes.

D. Pandemic Period – Canadian Pandemic Phases 6.0, 6.1, 6.2

Enhanced surveillance will continue until the pandemic subtype is established in each laboratory's respective jurisdiction.

E. Postpandemic Period

This will mark a return to interpandemic activities. Any surveillance issues that arose during the pandemic will be reviewed to determine whether there are changes to the pandemic plan that could be implemented.

During this period there will be a sustained increase in the volume of testing. Clinicians will require testing on samples from patients with influenza-like illness in order to identify the viruses present and rule out the continued circulation of the pandemic strain.

During the postpandemic period the laboratory experience should be reviewed at all levels to determine whether modifications to standard operating procedures or diagnostic methods are required to prepare for a potential subsequent wave.

VI. Communication Activities

A. Interpandemic/Pandemic Alert Period – Canadian Pandemic Phases 1.0-5.2

Each PHL will maintain an up-to-date list of laboratories that routinely test for influenza in their jurisdictions. Information from each laboratory, including a contact name, fax and telephone numbers, and e-mail address, should be maintained in a database so that current information regarding novel viral isolates and their diagnostic characteristics can be rapidly disseminated and made accessible to influenza testing facilities.

An up-to-date listing of all influenza testing laboratories will also be maintained by NML and the CPHLN secretariat. The information will also be accessible to all PIPLN and CPHLN members on their respective secure Web sites.

The CPHLN secretariat must set up enhanced communications to link NML , PHLs and other diagnostic laboratories that test for influenza with provincial epidemiologists. This can be done using the Canadian Laboratory Surveillance Network (CLSN) intelligence exchange centre, enabling e-mail, fax, telephone and/or teleconference, and Web-casting communication capabilities for infectious disease outbreak and event management. CPHLN and NML have each established toll-free numbers to enable dedicated contact with their respective leads in the event of an emergency. CPHLN and NML will disseminate these numbers to the appropriate public health authorities at the beginning of an emergency.

Diagnostic expertise from PILPN will be solicited by CPHLN as deemed necessary and appropriate.

NML and PHLs will share sequence information of novel strains as soon as it is available and exchange details of recommended protocols and primers where appropriate.

Information such as subtype, optimal cell lines to use, usefulness of direct antigen testing, antiviral susceptibility, morbidity and mortality from WHO, the Centers for Disease Control and Prevention, NML or laboratories from areas affected by the new subtype will be rapidly disseminated to PHLs by CPHLN secretariat through various means, e.g. CLSN intelligence exchange centres, fax, e-mail, telephone, depending on the circumstances.

Using the laboratory database compiled as part of preparedness activities, PHLs will ensure that other influenza testing laboratories in their province are kept informed. The CPHLN secretariat will coordinate meetings and/or teleconferences of PILPN and PHLs as required.

B. Pandemic Period – Canadian Pandemic Phases 6.0, 6.1, 6.2

Extensive communication within the PILPN will continue, and this will include such means as telephone, fax, e-mail and Canadian Network for Public Health Intelligence (CNPHI) collaboration centres as required.

NML will be responsible for rapid communication of relevant information concerning the evolution of the pandemic to PHLs and other diagnostic laboratories. This will include information concerning the occurrence of small or large clusters in different locations communicated through CPHLN and CLSN intelligence exchange centres, or by fax, e-mail or telephone as appropriate, and the provision of updates on the activity of pandemic strain, cell lines, direct test methods, etc. The PHLs will then be responsible for forwarding information to their respective Chief Medical Officers of Health or the Council of Chief Medical Officers of Health.

PHLs will rapidly communicate through NML their first isolates of the pandemic strain as well as any other local influenza activity (Appendix F).

Communication between PHLs and other influenza testing laboratories within the jurisdiction will continue.

Communications outlining any changes to testing during the pandemic will be released to clients. Alternative strategies available to help reduce the workload of the laboratory may be included.

As the pandemic progresses, NML will keep PHLs informed of influenza activity across the country, changes in susceptibility, and other circulating strains.

PHAC will provide information regarding morbidity and mortality.

Laboratories are encouraged to provide updated educational sessions for staff regarding testing, safety and HR issues, and to prepare communications to physicians regarding reductions in service and indications for limited testing during the pandemic.

C. Post Pandemic Period

This will mark a return to interpandemic activities. Any communication issues that arose during the pandemic will be reviewed to determine whether there are changes to the pandemic plan that could be implemented.

VII. Pandemic Preparedness

A. Planning Assumptions

Planning assumptions are essential in establishing laboratory protocols and procedures, and stockpiling materials and supplies that will be necessary during the pandemic. The assumptions outlined in this annex will help laboratories develop appropriate business continuity plans.

• The pandemic strain will emerge in Canada within three months after it emerges in another part of the world.
• Canada will receive advance warning of the human-to-human spread of disease from other affected geographic areas, and laboratories will use advance notice to finalize details relating to diagnostics, primer sequences, etc.
• Timely and accurate laboratory surveillance and diagnostics in the Pandemic Alert Period will be critical to appropriate clinical and public health management.
• As the pandemic evolves from phase 4 to phase 6, there will be at least a 10 fold increase in the demand for influenza diagnostic testing.
• On the basis of approximately 27,000 influenza tests performed nationally during an eight-week peak of normal influenza activity (Respiratory Virus Detections/Isolations in Canada: http://www.phac-aspc.gc.ca/im/influenza-eng.php), it is expected that 270,000 requests for testing will be processed by sentinel laboratories alone. This may be an underestimate since the number is based only on sentinel laboratories reporting to PHAC’s Respiratory Virus Detection/Isolation Program.
• In phase 6 of the pandemic, laboratory confirmation of clinical influenza diagnoses will likely decrease as the morbidity peaks and a reasonably accurate clinical diagnostic algorithm becomes available.
• In the CPIP it is estimated that over the course of the pandemic, 15%-35% of the population will become clinically ill such that they will be unable to attend work or other activities for at least half a day. Estimated absentee rates of laboratory personnel will correspond to this range.
• It is anticipated that a vaccine will become available four to six months after WHO declares Phase 6.0.
• Antivirals for early treatment of cases will be available throughout Canada. Implementation of this strategy may affect the prioritization of laboratory diagnostics.
• Antiviral resistance will be monitored as part of the antiviral strategy.
• Laboratories will see an increase in testing requests for secondary infections resulting from influenza and an increase in nosocomial testing due to increased numbers of hospital admissions. The extent of these increases is unknown and will depend on the population groups most affected by the pandemic strain.
• There will be shortages of the materials and supplies needed during the pandemic period. Therefore, plans are required to allow for a consistent 16-week supply (i.e. two pandemic waves) of both influenza and non-influenza related materials to address sporadic interruptions of supply chains (e.g. resulting from mail and courier disruptions, border closures, supply limitations).

B. Business Continuity Considerations

As part of pandemic preparation, PILPN recommends that PHLs and other local laboratories assess how the pandemic will affect other clinical laboratory functions and HR issues. Although the true impact is difficult to accurately predict, certain test requests from physicians will increase (e.g. increased respiratory specimens), and others will decrease. Trying to anticipate this in advance may help in developing strategies to maximize workflow and efficiency.

Strategies may include but are not limited to the following:

Assessing Laboratory Functions

• Develop a list of essential laboratory services.
• Anticipate testing demands that will increase (both influenza diagnostic and non-influenza diagnostic tests) and those that may decrease.
• Consider the appropriate means of streamlining specimen accessioning.
• Develop a prioritization strategy for lowering routine workload and other services to determine which services will be restricted and in what order when there are HR and other resource problems. This will need to include the impact of each test and its volume.
• Consider possible changes in the testing schedule to maximize workflow and efficiency.
• Confirm and provide assurance that enough supplies are available to maintain the ability to diagnose influenza and that these supplies will last at least throughout the first wave of the pandemic.
• Increase inventory of swabs, viral transport media kits and other reagents needed for influenza diagnostics.
• Review inventory and ordering necessary supplies and cell lines (e.g. more viral transport swabs and/or media, NAT reagents and antisera for DFA testing), and personal protective equipment.
• Review testing algorithms for influenza based upon availability of reagents, cell lines and kits, and modify if necessary.
• Review the other non-influenza related tests for which the demand is likely to increase (e.g. blood culture, sputum culture) in addition to influenza-related diagnostics.

PHLs and other viral diagnostic laboratories should also review strategies developed to reduce the impact on clinical laboratories:

• Review the testing prioritization list.
• Review alternative testing strategies and ensure that the supply of reagents is available for testing
• e.g. urine dip sticks for screening urine samples
• e.g. urine dipsticks for general practitioners to encourage in-office testing.
• Prepare any reagents that will be needed for the next several months, e.g. media.
• Begin prioritization exercise.
• Implement specimen deferral, rejection and test minimization strategies.
• Readjust testing schedules as appropriate.
• If capable, act as suppliers or provide seed stocks/protocols for the propagation of cells (e.g. Madin-Darby canine kidney) to other laboratories in need.

Assessing Human Resource Requirements

• Develop an inventory of currently provided services and the HR required to maintain this level of service.
• Develop a list of the HR requirements to maintain essential services.
• Develop a staffing strategy. Ideally staffing issues should be addressed before the pandemic so that potential concerns can be addressed before HR problems develop.
• Encourage laboratory staff to be vaccinated against annual seasonal influenza.

Strengthening Laboratory Capacity

Laboratory capacity must be strengthened to establish optimal levels of laboratory preparedness during a pandemic. At best, according to current operational levels, most laboratories will only be able to double their testing capacity over the anticipated first wave of the pandemic. To meet the projected 10 fold increase in testing over this period, PILPN strongly recommends that influenza testing laboratories increase stockpiled supplies; obtain additional equipment capacity; and increase available trained technical staff:

• Stockpiled supplies must include perishable and non-perishable items that will be in short supply in a pandemic. Although laboratories will strive to use stockpiled perishable supplies before their expiry date, it is unavoidable that a substantial amount of wastage will occur until a pandemic arrives. Careful stock rotation must be implemented to mitigate potential losses.
• Influenza testing laboratories will also require additional nucleic acid extraction and amplification equipment. To ensure that competence in the use of the equipment is maintained, this additional equipment may be used for other routine laboratory testing.
• Laboratory technical staff must be cross-trained to perform NAT for influenza and subsequent subtyping. The increase in trained staff will address additional testing requests and projected absenteeism losses during a pandemic. Trained staff must maintain competence by working on NAT for both seasonal influenza and other viruses.

Participating in Preparedness Exercises

Laboratories should plan to participate in pandemic exercises at the request of the PHAC to test plans and identify areas that need further attention. Assessment of this exercise process will be required by all participants.

Establishing Memoranda of Agreement

Agreements will need to be established both intra- and inter-provincially to outline how PHLs would best redirect testing capacity to help track the spread of the pandemic and to standardize how laboratories will triage critical and non-critical respiratory testing. This will be necessary so that at least some capacity will be available to track the onset and escalation of pandemic activity in Canada.

C. Quality Assurance and Quality Control

Participation in NML proficiency programs for influenza is strongly recommended for all laboratories performing any type of influenza diagnosis. NML will provide proficiency panels to assess the diagnostic sensitivity and specificity of tests available at PHLs and other viral diagnostic laboratories. NML and PHLs will share reagent lots designed to diagnose circulating or emerging influenza subtypes. NML will also provide one influenza proficiency panel per year to any Canadian laboratories that wish to participate in NAT identification of current influenza A strains. This will consist of RNA extracts from key strains of interest for RT-PCR quality control testing.

PILPN recommends participation in other accredited proficiency programs, such as those of the College of American Pathologists.

During the later stages of the Pandemic Alert Period (4.1, 4.2, 5.0, 5.1, 5.2), diagnostic capabilities of laboratories will be strained; however, it will be important to continue quality assurance activities, such as the participation in proficiency panels distributed by NML. NML will be responsible for providing the guidance and materials required.

During the pandemic period, laboratories are encouraged to ensure that the new methods are sensitive and specific through participation in ongoing quality assurance programs. Problems encountered should be reported to NML for investigation and/or sharing of information with PHLs and other diagnostic laboratories.

Appendices

Appendix A
Acronyms

Organizations
Canadian Laboratory Surveillance Network	CLSN
Canadian Network for Public Health Intelligence	CNPHI
Canadian Public Health Laboratory Network	CPHLN
Centers for Disease Control and Prevention (United States)	CDC
Centre for Emergency Preparedness and Response	CEPR
Centre for Immunization and Respiratory Infectious Diseases	CIRID
Federal, provincial and/or territorial	FPT
National Microbiology Laboratory	NML
Pandemic Influenza Committee	PIC
Pandemic Influenza Laboratory Preparedness Network	PILPN
Province and/or territory	P/T
Public Health Agency of Canada	PHAC
Public Health Laboratories	PHLs
World Health Organization	WHO
Diagnostic and Scientific Terms
Containment level	CL
Direct fluorescent antibody assay	DFA
Enzyme immunoassay	EIA
Hemagglutinin	HA
Hemagglutination inhibition	HAI
Human resource(s)	HR
Immunofluorescent assay	IFA
Nasopharyngeal	NP
Nasopharyngeal swab	NPS
Nucleic acid amplification test	NAT
Point of care	POC
Polymerase chain reaction	PCR
Reverse transcriptase polymerase chain reaction	RT-PCR
Ribonucleic acid	RNA
Severe Respiratory Illness	SRI

Appendix B
Minimum requirements for the provision of public health laboratory services during pandemic influenza

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Minimum Requirements

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PILPN recommendation of minimum requirements for the provision of public health laboratory services during pandemic influenza

• Preamble
• Testing Procedures/Capacity
• Specimen Collection
• Business Continuity Plan
• Communication
• PHLs Required to Meet Minimum Requirements

Preamble

The Assumptions about Demands for Testing

• The peak demand for testing services, the duration of that peak period, the availability of testing supplies and reagents, as well as human resources, cannot be accurately predicted.
• The following recommendations are based on PILPN’s best estimates and assumptions.

Testing Procedures/Capacity

• Provincial Public Health Laboratories (PPHLs) or a designated laboratory must be able to diagnose influenza A by nucleic acid amplification tests (NAT). These methods must be broadly reactive to be able to identify novel subtypes of influenza A.
• Each PPHL or designated laboratory must be able to subtype positive influenza A specimens to distinguish seasonal influenza from novel subtypes.
• Each PPHL that has Containment Level 3 (CL3) capabilities must develop standard operational procedures for the culture and identification of novel subtypes of influenza A. PPHL or the designated laboratories that do not have this capability should develop a Memorandum of Understanding with another designated laboratory with CL3 capability or the National Microbiology Laboratory (NML) to provide this service.
• Provincial and other designated laboratories must have the capacity to meet the increased demands for testing for pandemic influenza.
• The NML has the mandate to provide additional, more specialized, testing, including serological, antiviral resistance testing and antigenic characterization. However, PHLs, depending on their resources and expertise, may collaborate with NML to develop these tests in order to provide additional surge capacity for specialized testing.

Specimen Collection

Each PPHL or designate must be able to provide or advise on the appropriate specimen collection devices.

• At present, the ideal specimen for a novel subtype is not known, so these laboratories must recommend collection of nasopharyngeal swabs, throat swabs, nasal swabs and, potentially, plasma and stool from individuals with epidemiological features that put them at risk of a novel subtype of influenza A.
• As the pandemic progresses, information regarding the most appropriate specimen(s) will be provided.

Business Continuity Plan

Each PPHL or designate must develop a business continuity plan for influenza-related diagnostics and other essential non-influenza testing services, to include the following:

1. Determination of the minimum requirements for human resources and materials in order to maintain uninterrupted, essential services during the critical phase of the pandemic. While the initial pandemic wave may last for at least eight weeks (to vary by jurisdiction), laboratories must address limitations in such items as supplies, reagents and human resources. Planning must include provision of supplies and reagents for Phase 4, 5 and 6. Planning should include the Public Health Agency of Canada (PHAC) Mutual Aid Agreement.

2. Development and publication of a prioritized list of laboratory services that will be reduced as needed during a pandemic.

3. Development of stockpiles of reagents and supplies necessary to maintain these essential services and implementation of an inventory management system to maximize utilization of reagents and minimize loss due to reagent expiration. This must take into account the limited shelf life of essential perishable items.

Communication

Minimum Requirements for Timely Communication

• Each PHL or designated laboratory should develop or enhance communication links with its local provincial public health departments and CPHLN. Information regarding novel subtypes will be updated from the Canadian Pandemic Influenza Committee through PILPN and then through CPHLN. (See Figure A: PILPN and Information Flow: Bridging Local and Provincial Public Health Laboratories via CPHLN and the Canadian Pandemic Influenza Committee.)
• Constant and timely updating of laboratory epidemiological information (data, specimen collection and routing, biosafety, etc.) during a pandemic needs to be developed further.
• PPHLs and designated laboratories will take a lead role in developing and guiding specimen triage mechanisms (e.g. an essential clinical information form) (see Figure B).

PHLs Required to Meet Minimum Requirements

*Indicates that the facility has Containment Level 3 certification.

Figure A: PILPN and Information Flow: Bridging Local and Provincial Public Health Laboratories via CPHLN and the Pandemic Influenza Committee

Figure B Severe Respiratory Illness and Pandemic Influenza Laboratory Sheet

Appendix C
Province of Québec’s Preparation for an Influenza Pandemic – Rapid Tests: Value and Limitations²

Excerpt from:

Fauvel, Micheline, Couillard, Michel. October 2006. "Préparation à une pandémie de grippe. Lignes directrices à l’intention des cliniciens et des laboratoires du québec sur l’utilisation des épreuves de laboratoire pour les virus influenza." Institut national de santé publique du Québec.

Sensitivity and Specificity of Two Rapid Tests Approved in the United States

The WHO, the Centers for Disease Control and Prevention (CDC) and the U.S. Food and Drug Administration (FDA) have issued warnings and recommendations on the use of rapid tests for the detection of influenza A according to the suspected presence of the avian flu. Summarized in this section, these documents can be consulted on the following sites.^1-3

• http://www.cdc.gov/flu/professionals/diagnosis/
• http://www.who.int/csr/disease/avian_influenza/
  guidelines/rapid_testing/en/index.html
• http://www.fda.gov/MedicalDevices/Safety/AlertsandNotices/
  TipsandArticlesonDeviceSafety/ucm109385.htm

Several tests for the rapid detection of influenza have been approved in Canada; these can detect the following:

• influenza A and B without viral typing;
• influenza A only;
• influenza A and B with viral typing.

They do not currently distinguish between influenza A subtypes H and N.

The rapid tests have been evaluated with different types of specimens and different populations. They have high sensitivities in pediatric populations, since children excrete more of the virus and for a longer period of time than do adults. The table below presents the data generated with two rapid tests.³ Specificity may also vary according to the age of the population and specimen type.

Sensitivity and Specificity Compiled from Two Rapid Tests*
Specimen	Type of Influenza Virus Detected	Population**	Sensitivity, %†	Specificity, %†
Throat swab	A	Pediatric‡	65-90	81-91
		Adult	24-91	69-94
Throat swab	A and B	Not specified	59-82	81-93
Nasopharyngeal lavage or aspiration	A	Pediatric‡	82-95	98-100
		Adult	53-87	90-100
Nasal lavage	A	Pediatric‡	36-88	92-99
		Adult	9-99	59-100
Nasal lavage or aspiration	A	Not specified	65-84	95-99
Nasal swab	A and B	Not specified	65-87	87-97

*Adapted from FDA. Cautions in Using Rapid Tests for Detecting Influenza A Viruses. November 14, 2005, available at: http://www.fda.gov/cdrh/oivd/tips/rapidflu.html. Kits and manufacturers not identified.
**Data from the U.S., Australia or New Zealand during seasons in which the predominant influenza strains were A H3 and A H1.
†95% confidence intervals.
‡Age not specified but mainly under 10 years old.

Implications

Rapid tests have lower sensitivities than culture or RT-PCR, while their specificities are rather high. Rapid tests can also produce false-positive or false-negative results, at a rate that varies according to the level of influenza activity in the population. In fact, it is the infection’s prevalence that influences the positive or negative predictive values. The table below presents an example of the variation in predictive values when the same kit is used at the start and at the peak of influenza activity for simulated prevalence rates of 1% and 25%.

Predictive Values by Prevalence
	Prevalence of Influenza
	1,000/100,000 (1%)	25,000/100,000 (25%)
Individuals infected	1,000	25,000
Individuals not infected	99,000	75,000
Kit sensitivity	80%	80%
Kit specificity	97%	97%
True positives	800	20,000
False positives	2,970	2,250
True negatives	96,030	72,750
False negatives	200	5,000
PPV	21.2%	89.8%
NPV	99.8%	93.6%

PPV, positive predictive value; NPV, negative predictive value

In the off-season or at the very beginning of an active period, negative predictive values (NPVs) are higher and positive predictive values (PPVs) lower, thereby making false-positive results more likely. For this reason, it is important to confirm the first positive results generated with a rapid antigen detection kit. During the high-activity season, the PPVs will be higher, generating few false positives, whereas the NPVs will be lower.

Limitations of rapid tests

When interpreting results obtained with rapid tests, it is important to consider the laboratory and surveillance data on the circulation of strains and the level of influenza activity. The following points should be considered:

• When activity is low, positive results from a rapid test should be confirmed by culture or by RT-PCR.
• During peak activity periods, when the NPVs are slightly lower, false negatives are more likely.
• At the start of the influenza season, negative results should be interpreted with caution, and confirmation by culture or PCR should be considered, since a negative result does not necessarily rule out an influenza infection.
• Rapid tests do not provide information on viral subtypes and are unable to distinguish influenza A strains infecting humans (H1, H2, H3) from those infecting birds or other animals (H5, H7, H9).

Therefore, it is important that sufficient specimens be collected to enable additional confirmation tests, according to activity periods and when new strains are suspected.

Finally, when commercial kits are evaluated, it is recommended that the published data be analyzed with the following cautions:

• The comparison of sensitivities and specificities of commercial kits published in their manufacturers’ monographs should be interpreted with caution, since their studies are conducted with different groups of patients whose variable specimens are collected at different times after the onset of symptoms.
• Performing a test using samples that were frozen can sometimes result in a higher sensitivity than using fresh samples.
• Inappropriate collection, storage and shipment of specimens can generate false-negative results.
• The tests’ predictive values will depend on the level of influenza activity in the community.

The list of rapid influenza antigen detection kits currently approved in Canada is shown in the table below. To obtain an updated list of approved medical devices used for detection, send an e-mail request to: device_licensing@hc-sc.gc.ca or call 613-957-1909.

Name of the Kit	Manufacturer	Approval No.
BD DIRECTIGEN EZ FLU A+B	Becton Dickinson and Company	66969
DIRECTIGEN FLU A TEST KIT	Becton Dickinson and Company	10819
DIRECTIGEN FLU A + B	Becton Dickinson and Company	23834
BINAX NOW INFLUENZA A & B	Binax Inc.	71036
FLU OIA TEST KIT	Thermo Biostar Inc.	23519
ACTIM INFLUENZA A & B TEST	Medix Biochemica OY AB	66656
QUICK S INFLU A/B TEST	Innovatek Medical Inc.	66848
IMMUNOCARD STAT FLU A & B	Meridian Bioscience Inc.	65947
QUICKVUE INFLUENZA A+B TEST	Quidel Corporation	20158
NOW FLU A NOW FLU B TEST KIT	Binax Inc.	61340
XPECT FLU A/B	Remel	63374
CLEARVIEW FLU A/B TEST	Wampole Laboratories Inc.	66506
INFLU A RESPI STRIP	Coris Bioconcept	63448
INFLU A&B TEST KITS	Coris Bioconcept	69985

Note: Data from June 2006. Source: Medical Devices Bureau, Health Canada.

References

1. World Health Organization. WHO recommendations on the use of rapid testing for influenza diagnosis. July 2005. Available at: http://www.who.int/csr/disease/avian_influenza/guidelines/rapid_testing/en/index.html.

2. US Food and Drug Administration. Cautions in using rapid tests for detecting influenza A viruses. November 14, 2005. Available at: http://www.fda.gov/MedicalDevices/Safety/AlertsandNotices/TipsandArticlesonDeviceSafety/ucm109385.htm.

3. US Food and Drug Administration. Guidance for industry and FDA staff. In vitro diagnostic devices to detect influenza A viruses: labeling and regulatory path. April 2006. Available at: www.fda.gov/OHRMS/DOCKETS/98fr/06d-0121-gdl0001.pdf.

4. FDA. Cautions in using rapid tests for detecting influenza A viruses. November 14, 2005. Available at: http://www.fda.gov/MedicalDevices/Safety/AlertsandNotices/TipsandArticlesonDeviceSafety/ucm109385.htm.

Appendix D
Nasopharyngeal Swab Procedure

1. Use the swab supplied with the viral transport media.

2. Explain the procedure to the patient.

3. When collecting the specimens, wear eye protection, gloves, and a mask. Change gloves and wash your hands between each patient.

4. If the patient has a lot of mucus in the nose, this can interfere with the collection of cells. Either ask the patient to use a tissue to gently clean out visible nasal mucus or clean the nostril yourself with a cotton swab (e.g. Q-Tip).

5. How to estimate the distance to the nasopharynx: prior to insertion, measure the distance from the corner of the nose to the front of the ear and insert the shaft approximately 2/3 of this length.

6. Seat the patient comfortably. Tilt the patient’s head back slightly to straighten the passage from the front of the nose to the nasopharynx to make insertion of the swab easier.


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7. Insert the swab provided along the medial part of the septum, along the floor of the nose, until it reaches the posterior nares; gentle rotation of the swab may be helpful. (If resistance is encountered, try the other nostril; the patient may have a deviated septum.)

   

   *Image obtained from http://www.nlm.nih.gov/medlineplus/ency/imagepages/9687.htm

8. Allow the swab to sit in place for 5–10 seconds.

9. Rotate the swab several times to dislodge the columnar epithelial cells. Note: Insertion of the swab usually induces a cough.

10. Withdraw the swab and place it in the collection tube.

11. Refrigerate immediately.

12. Remove gloves.

13. Wash hands.

14. Attach completed requisition.

15. Transport to the laboratory.

Appendix E
Draft Pandemic Influenza Laboratory Advisory

Office of Laboratory Security, Centre for Emergency Preparedness and Response, Public Health Agency of Canada

As part of the emergency preparedness plan, the Office of Laboratory Security is providing the following draft biosafety advisory regarding the laboratory handling of clinical specimens associated with new influenza virus subtypes that may pose a pandemic threat. Please note that the recommendations below are subject to change as specific information on the pandemic strain becomes available, and changes will be disseminated by CEPR in a timely fashion.

Precautions for laboratories receiving and processing human clinical specimens and tissue samples from suspected human cases of pandemic influenza:

• Specimens may be processed for packaging and distribution to diagnostic laboratories for further testing in a Containment Level 2 laboratory1 using the additional operational practices as outlined below.
• Diagnostic testing (excluding viral culture) to rule out pandemic influenza virus strains may be performed in a Containment Level 2 laboratory1 using the additional operational practices as outlined below.
• Viral culture should be performed in a certified CL3 laboratory.

Additional operational practices:

• Laboratory workers should wear protective clothing (e.g. protective solid-front gowns, gloves and NB95 respiratory protection) in accordance with the risk of exposure when handling specimens.
• Manipulations are to be carried out in a certified biological safety cabinet.
• Centrifugation of clinical specimens and tissue samples should be carried out using sealed centrifuge cups or rotors, both of which are unloaded in a biological safety cabinet.

Precautions for laboratories handling human clinical specimens from confirmed cases of pandemic influenza for isolation and further manipulation of the agent:

• Specimens may be processed for packaging and distribution to laboratories for further testing in a Containment Level 2 laboratory¹ using the additional operational practices as outlined above.
• Manipulation of the agent should be in a Containment Level 3 laboratory¹ using Containment Level 3 operational practices.¹
• PCR testing of extracted, non-infectious genetic material may be performed in a Containment Level 2 laboratory.¹

For information on biosafety precautions for activities involving pandemic influenza virus strains in vivo and handling of animal specimens, contact the Biohazard Containment and Safety Division at the Canadian Food Inspection Agency, (613) 221-7088.

Transportation of clinical specimens:

• Packaging, shipping and transport of specimens must, as a minimum, comply with the requirements of the Transportation of Dangerous Goods Regulations, Transport Canada² and the Dangerous Goods Regulations, International Air Transport Association.³

Further biosafety and transportation information may be obtained from the Office of Laboratory Security, Centre for Emergency Preparedness and Response, Public Health Agency of Canada at (613) 957-1779, fax (613) 941-0596 or Web site http://www.phac-aspc.gc.ca/ols-bsl/.

References

1. Health Canada. Laboratory biosafety guidelines, 3rd ed. Available at: http://www.phac-aspc.gc.ca/ols-bsl/lbg-ldmbl/index-eng.php

2. Transport Canada. Transport of Dangerous Goods Regulations. Available at: http://www.tc.gc.ca/tdg/menu.htm

3. International Air Transport Association. Dangerous Goods Regulations. Available at: http://www.iataonline.com

Appendix F
Laboratory Pandemic Influenza Communication Flow: Pandemic Alert Period

Appendix G
F/P/T Laboratory Services during a Pandemic

This appendix is a working document and may change as new information becomes available.

INTERPANDEMIC PERIOD

Federal

• Establish the minimum laboratory proficiency testing standards required for pandemic preparedness and ensure that federal laboratories (or designates) meet these standards.
• Conduct surveillance to detect, subtype and characterize circulating seasonal influenza viruses, to provide an early warning of novel influenza strains that could pose a pandemic risk.
• Provide reference laboratory services to provincial/territorial laboratories.
• NML will represent Canada internationally as the WHO National Influenza Reference Laboratory and provide a liaison function with the CDC and WHO.
• Ensure that the information and reagents requisite for the diagnosis of novel human and animal influenza strains posing a risk to humans are shared in a timely fashion between the Canadian Food Inspection Agency and the Public Health Agency of Canada, and that relevant information and reagents are transferred to any provincial/territorial PHLs that may require them.
• NML will provide a leadership role in the communication of laboratory test results confirming the diagnosis of pandemic influenza and will establish case definitions for the laboratory diagnosis of pandemic influenza.
• Disseminate information and reagents necessary for diagnosis of novel influenza virus strains.
• Maintain a national reference collection of influenza virus isolates and antisera.
• Provide regular, annual or semi-annual influenza proficiency panels, recommend diagnostic protocols, and provide training workshops.
• Provide surge capacity for additional emergency laboratory testing in any jurisdiction where it is needed.
• NML will maintain accreditation at international standard level in quality assurance for key tests.
• Monitor for antiviral resistance.
• Perform technology transfer to Canadian PHLs.
• Participate in vaccine evaluation studies.
• Facilitate, coordinate and participate in ongoing collaborative diagnostics research.
• Maintain an up-to-date business continuity plan for service provision during a pandemic.
• Provide ongoing biosafety guidance on containment requirements.

Provincial/Territorial

• Designate provincial reference laboratories responsible for influenza laboratory testing in a pandemic.
• Ensure that all laboratories involved in national influenza surveillance and responsible for influenza testing in a pandemic participate fully in the national influenza proficiency program administered by NML.
• Ensure that equitable access to influenza testing by First Nations and Inuit populations is maintained through existing P/T laboratory service plans or covered by federal laboratories.
• Implement minimum criteria for PHL diagnostic capabilities as recommended by the CPIP Laboratory Task Group.
• Ensure that provincial or territorial laboratories (or designates) meet the minimum standards established for pandemic preparedness.
• Ensure that any laboratory test results indicating a positive diagnosis of pandemic influenza are confirmed by the NML.
• Ensure that any communications announcing the laboratory diagnosis of pandemic influenza within Canada are coordinated through the NML.
• Maintain routine comprehensive laboratory diagnostic services for influenza testing.
• Participate in the national influenza surveillance program by submitting representative strains to NML.
• Maintain a business continuity plan for laboratory services, including both front-line clinical and surveillance testing, biosafety facilities, stockpiling, and table-top exercises and training.

Public Health Network Council

• Communicate Council decisions that affect laboratories in a timely fashion to laboratory stakeholders

PANDEMIC PERIOD

Federal

Continue interpandemic activities and include the following:

• Monitor the performance characteristics of existing tests.
• Provide laboratory services for the assessment of pandemic vaccine effectiveness.
• Rapidly communicate relevant information concerning the evolution of the pandemic to provincial and territorial clients, as well as internationally to WHO.

Provincial/Territorial

Continue interpandemic activities and include the following:

• Review of pandemic stockpiling as per pandemic plan.
• Ensure that the minimum standards for quality assurance are maintained.
• Submit strains from index cases to NML for confirmation and antigenic characterization.

Public Health Network Council

Communicate Council decisions that affect laboratories in a timely fashion to laboratory stakeholders

Appendix H
Members of the Pandemic Influenza Laboratory Preparedness Network

The Pandemic Influenza Laboratory Preparedness Network (PILPN) operates under the auspices of the Canadian Public Health Laboratory Network, and it is responsible for the preparation of this laboratory annex. PILPN members are given below.

Berry, Jody
Public Health Agency of Canada,
NML
1015 Arlington St.
Winnipeg, MB  R3E 3R2

Booth, Tim
Public Health Agency of Canada
NML
Director of Viral Diseases
1015 Arlington St.
Winnipeg, MB  R3E 3R2

Couillard, Michel
Laboratoire de santé publique du Québec, Coordonnateur Scientifique
20045, chemin Sainte-Marie
Sainte-Anne-de-Bellevue, QC
H9X 3R5

Drews, Steven
Clinical Microbiologist
Head, Molecular Diagnostics
Ontario Public Health Laboratories
Ministry of Health and Long-Term Care
Public Health Laboratories Branch
81 Resources Road
Toronto, ON  M9P 3T1

Fearon, Margaret
Canadian Blood Services, Executive Medical Director, Medical Microbiology
67 College St.
Toronto, ON M5G 2M1

Fonseca, Kevin
Provincial Laboratory for Public Health, Clinical Virologist
3030 Hospital Drive NW
Calgary, AB  T2W 4W4

Fox, Julie
Provincial Laboratory Alberta, Associate Professor, University of Calgary, Microbiologist and Program Leader
3030 Hospital Drive NW
Calgary, AB  T2N 4W4

Jean-Nicolas Gagnon
Jean-Nicolas Gagnon
A/Head, Importation and Biosafety Programs
Office of Laboratory Security
Center for Emergency Preparedness and Response
Public Health Agency of Canada
100 Colonnade Road
Ottawa, ON  K1A 0K9

Hatchette, Todd
QEII Health Sciences Centre, Director, Immunology/Virology,
Rm 315 MacKenzie Bld,
Victoria General Site
QE II Health Sciences Centre
1278 Tower Rd.
Halifax, NS  B3H 2Y9

Horsman, Greg
Saskatchewan Health, Medical Director
3211 Albert Street
Regina, SK  S4V 5W6

Li, Yan
Public Health Agency of Canada, Chief, Respiratory Viruses
1015 Arlington St.
Winnipeg, MB  R3E 3R2

Majury, Anna
Ontario Public Health Laboratories, Microbiologist/Chair Laboratory Working Group--OHPIP
181 Barrie Street, POB 240
Kingston, ON  K7V 4V8

Petric, Martin
BCCDC, Clinical Virologist
655 W 12th Ave.
Vancouver, BC  V5Z 4R4

Ratnam, Sam
Newfoundland Public Health Laboratory, Director
100 Forest Road - P.O. Box 8800
St. John’s, NL  A1B 3Z9

Tam, Theresa
Director - Immunization and Respiratory Infections Division
Public Health Agency of Canada
2nd Floor, Room 2363, Building 6
100 Eglantine Driveway, Tunney's Pasture
Ottawa, Ontario  K1A 0K9

Van Caeseele, Paul
Cadham Provincial Laboratory, Laboratory Director
Box 8450, 750 William Avenue
Winnipeg, MB  R3C 3Y1

Wong, Tom
PHAC
Director of Community Acquired Infections Division
Rm 3444, LCDC Bldg,
Tunney's Pasture
P/L 0603B
Ottawa, ON

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Antiviral Liaison

Huston, Patricia
Public Health Agency of Canada, Chief
Emerging Infectious Diseases
LCDC Bldg, A.L. # 0602C
Tunney's Pasture
Ottawa, ON  K1A 0K9

Tamblyn, Susan
Public Health Consultant
85 Neal Ave,
Stratford, ON
Canada N5A 5A7

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Clinical Care Liaison

Huston, Patricia
Public Health Agency of Canada, Chief
Emerging Infectious Diseases
LCDC Bldg, A.L. # 0602C
Tunney's Pasture
Ottawa, ON  K1A 0K9

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Disease Surveillance Liaison

Macey, Jeannette
Public Health Agency of Canada, Acting Head of Disease Surveillance
Building 6, Tunney's Pasture, Ottawa, Ontario
Ottawa, ON  K1H 0L2

Watkins, Kerri
Public Health Agency of Canada, A/Senior Epidemiologist
Building #6, Tunney's Pasture
A.L. #0602C
Ottawa, ON  K1A 0K9

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Facilitator

Ebsworth, Anthony
Provincial Laboratory Alberta, Corporate Quality
3030 Hospital Drive N.W.
Calgary, AB  T2N 4W4

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Hospital Liaison

Chernesky, Max
McMaster University,
50 Charlton Avenue East, Room L424
Hamilton, ON  L8N 4A6

Smieja, Marek
McMaster University, Microbiologist & Infectious Diseases Physician
L424 St.Joseph's Hospital
50 Charlton Ave. E.
Hamilton, ON  L8N 4A6

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Public Health Measure Liaison

Sciberras, Jill
Senior Epidemiologist
Public Health Agency of Canada
3rd Floor, Building 6
100 promenade Eglantine Driveway, Tunney's Pasture
Ottawa, Ontario  K1A 0K9

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Canadian Public Health Laboratory Network Secretariat

DeWinter, Leanne
Standards Development Officer [Former]
National Microbiology Laboratory

Guercio, Steven
Operational Liaison Officer
National Microbiology Laboratory
1015 Arlington St.
Winnipeg, MB R3E 3R2

Jorowski, Catherine
Information Services Officer
National Microbiology Laboratory
1015 Arlington Street
Winnipeg, MB  R3E 3R2

Kuschak, Theodore
Manager
National Microbiology Laboratory
1015 Arlington Street
Winnipeg, MB  R3E 3R2

Vegh-Yarema, Niki
Standards Development Officer
National Microbiology Laboratory
1015 Arlington Street
Winnipeg, MB  R3E 3R2

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Vaccine Liaison

Skowronski, Danuta
BC Centre for Disease Control
655 West 12th Ave.
Vancouver BC V5Z 4R4

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