Interim Guidance on Specimen Collection and Testing for Patients with Suspected Infection with Novel Influenza A Viruses Associated with Severe Disease or with the Potential to Cause Severe Disease in Humans

Background and Purpose

Novel influenza A viruses are influenza A viruses that have infected people but are antigenically and genetically different from seasonal influenza A viruses that circulate among humans. Novel influenza A viruses are predominantly of avian or swine origin. The clinical spectrum of human infection with novel influenza A viruses varies considerably: from asymptomatic infection to mild illness, including conjunctivitis, fever, and cough; to severe illness, including fulminant pneumonia, acute respiratory distress syndrome (ARDS) and multi-organ failure resulting in death.

This document provides interim guidance for clinicians and public health professionals in the United States on appropriate specimen collection and diagnostic testing for patients who might be infected with novel influenza A viruses associated with severe human disease or with potential to cause severe human disease. Examples include but are not limited to highly pathogenic avian influenza A(H5N1) and A(H5N6) viruses, and avian influenza A(H7N9) virus. A novel influenza A virus is considered to have the potential to cause severe human disease if previous cases of infection with the same hemagglutinin subtype [e.g., HPAI A(H5) or A(H7) viruses] have been associated with severe human disease. More information about specific novel influenza A viruses, including those that have caused severe illness in humans, is available here.

CDC recommends enhanced surveillance efforts by state and local health departments, hospitals, and clinicians among people at increased risk for novel influenza A virus infection. Clinicians should notify their state health department immediately if they decide to test a patient for novel influenza A virus infection so that appropriate testing and follow up of contacts is initiated.

CDC should be notified immediately when any clinical specimens from suspected patients test positive for novel influenza A virus or if the testing results of clinical specimens from suspected cases are inconclusive. Human infection with a novel influenza A virus is a nationally notifiable condition.

CDC will update this guidance as additional information becomes available.

Recommendations for Surveillance, Testing, and Investigation

Clinicians and public health personnel should consider the following for surveillance and testing:

1. Consider the possibility of infection with novel influenza A viruses known to cause severe disease or with the potential to cause severe disease in humans in patients who present with acute respiratory infection (ARI) symptoms or conjunctivitis, and who have had recent direct or close contact (particularly unprotected exposure without use of respiratory protection and eye protection) <10 days prior to illness onset to animals and materials from animals potentially infected or confirmed to be infected with HPAI A(H5N1) virus. A full list of clinical criteria for novel influenza A virus testing is available.
2. If infection with a novel influenza A virus with the potential to cause severe disease in humans is suspected, respiratory specimens should be collected while following recommended infection control precautions. The state health department should be notified as soon as possible, and respiratory specimens should be sent to a public health laboratory such as at the state health department for immediate testing (see guidance below). More information is available on case definitions.
3. If infection with a novel influenza A virus with the potential to cause severe disease in humans is suspected, state health departments are encouraged to initiate a public health investigation with animal health partners and should notify CDC promptly.

For questions or concerns about possible human infection in patients with exposures to birds or other animals not listed here, please contact CDC. Exposures that occur in geographic regions in the United States where HPAI A(H5) viruses have been recently identified are of most concern.

Infection Control when Collecting Specimens

Standard, contact, and airborne precautions, and use of eye protection are recommended for management of patients with suspected or laboratory-confirmed novel influenza A virus infection; this includes collection of respiratory and conjunctival specimens. Practitioners should adhere to infection control precautions recommended for novel influenza A viruses known to cause severe disease in humans. Visit Interim Guidance for Infection Control Within Healthcare Settings When Caring for Confirmed Cases, Probable Cases, and Cases Under Investigation for Infection with Novel Influenza A Viruses Associated with Severe Disease for more information and consult CDC for specific case-by-case infection control recommendations if needed.

When Clinical Specimens Should Be Collected

Specimens should be obtained for novel influenza A virus testing as soon as possible after illness onset, ideally within 7 days of illness onset. However, as some persons who are infected with seasonal influenza viruses are known to shed virus for longer periods (e.g., children and immunocompromised persons), specimens should be collected from symptomatic persons for novel influenza A virus testing even if obtained after 7 days from illness onset. Prolonged shedding of novel influenza A viruses in the lower respiratory tract has been documented for critically ill patients with highly pathogenic avian influenza A(H5N1) virus and avian influenza A(H7N9) virus infections. The duration of shedding of novel influenza A viruses in humans is largely unknown, and there are currently limited data describing shedding of people infected with these viruses.

Preferred Clinical Specimens

The following respiratory specimens should be collected as soon as possible after illness onset: (i) a nasopharyngeal swab, and (ii) a nasal swab combined with an oropharyngeal swab (e.g., two swabs collected separately and combined into one viral transport media vial). The nasopharyngeal swab and the combined nasal-throat swabs should be tested separately. If these specimens cannot be collected, a single nasal swab, or oropharyngeal swab is acceptable. If the person has conjunctivitis (with or without respiratory symptoms), both a conjunctival swab and nasopharyngeal swab should be collected. Patients with severe respiratory disease with suspected novel influenza A virus infection also should have lower respiratory tract specimens (e.g., an endotracheal aspirate or bronchoalveolar lavage (BAL) fluid from intubated patients or induced sputum) collected, if possible, because these specimens have a higher yield for detecting HPAI A(H5N1) and A(H7N9) viruses and may facilitate detection of other novel influenza A viruses. For severely ill persons, multiple respiratory tract specimens from different sites should be obtained to increase the potential for novel influenza A virus detection. Specimens should be placed into sterile viral transport media or universal transport media and immediately placed on refrigerant gel-packs or at 4°C (refrigerator) for transport to the laboratory. BAL specimens in phosphate buffered saline are acceptable.

To increase the potential for novel influenza A virus detection, multiple respiratory specimens from different sites should be obtained from the same patient on at least two consecutive days for hospitalized patients.

Swabs

Swab specimens should be collected using swabs with a synthetic tip (e.g., polyester or Dacron®) and an aluminum or plastic shaft. Swabs with cotton tips and wooden shafts are not recommended. Specimens collected with swabs made of calcium alginate are not acceptable. The swab specimen collection vials should contain 1-3ml of sterile viral transport medium or universal transport medium (e.g., containing protein stabilizer, antibiotics to discourage bacterial and fungal growth, and buffer solution).

Transporting and Storing Clinical Specimens

Human respiratory specimens to be tested within 72 hours post-collection should be transported refrigerated at 2-8°C. Alternatively, clinical specimens can be frozen at ≤-70°C and transported promptly. Avoid freezing and thawing specimens.

Shipping Clinical Specimens to State Public Health Laboratories

Clinical specimens sent to state public health laboratories should be shipped in the appropriate packaging and according to instructions by the laboratory. Store frozen at ≤-70°C and ship on dry ice. Avoid freezing and thawing specimens because viability of some viruses from specimens that were frozen and then thawed is greatly diminished. All specimens should be labeled clearly and include information requested by the local or state public health laboratory.

Testing techniques

Diagnostic Testing

The performance of current Food and Drug Administration (FDA) cleared diagnostic tests for detecting influenza viruses in respiratory specimens has been demonstrated for seasonal human influenza A and B viruses as described by the manufacturer's package insert. Performance has not been demonstrated with most novel influenza A viruses. FDA-cleared influenza diagnostic tests do not specifically identify infection with novel influenza A viruses or distinguish between infection with seasonal influenza A or novel influenza A viruses. Testing of symptomatic human cases of suspected novel influenza A virus infection should be referred to the nearest public health laboratory.

Existing, commercially available FDA-cleared molecular assays [e.g., Real-Time RT-PCR (rRT-PCR)] may fail to detect some novel influenza A viruses or may detect with results that only indicate "influenza A positive", but with subtype not identified. For these assays a novel influenza A virus may give an influenza A "unsubtypeable" result. Clinicians and laboratorians using molecular assays that can detect all currently circulating influenza A virus subtypes in people (i.e., "seasonal influenza" A virus subtypes) who identify an unsubtypeable result should contact CDC and their state or local public health laboratory as soon as possible for additional testing (see below).

Rapid influenza diagnostic tests (RIDTs) and immunofluorescence assays are antigen detection tests that only identify whether an influenza A virus is detected and have unknown sensitivity and specificity to detect human infection with novel influenza A viruses in respiratory specimens. Some studies suggest that antigen detection tests have low sensitivity to detect novel influenza A viruses. Therefore, negative results from either type of test do not exclude novel influenza A virus infection, especially in patients with signs and symptoms suggestive of influenza. A negative influenza A test result could be a false negative and should not be used as a final diagnostic test for influenza, including novel influenza A virus infection. These tests may yield a positive influenza A result for a specimen containing novel influenza A virus but cannot identify the subtype and cannot distinguish novel influenza A virus from seasonal influenza A virus infection. Therefore, influenza A testing by rRT-PCR is recommended at public health laboratories and CDC for any patient with suspected novel influenza A virus infection.

Clinicians should always consider diagnostic testing for other respiratory pathogens that can cause acute febrile respiratory illness depending on the local epidemiology of circulating respiratory viruses (e.g., SARS-CoV-2) since novel influenza A virus infections of humans are very rare, even in exposed persons.

Testing at State Health Departments

Clinicians should notify their local and state health department immediately when they wish to test a patient for infection with novel influenza A viruses. Specimens to be tested for novel influenza A viruses should be sent first to the local or state public health laboratory.

Testing can be performed by public health laboratories on a portion of the specimen, while a portion of the sample should be reserved in case there is a need to ship it to CDC. CDC should be notified immediately in the event that any clinical specimens from suspected cases test positive for any novel influenza A virus [e.g., LPAI or HPAI A(H7N9), HPAI A(H5N1), HPAI A(H5N6) or other avian A(H5) viruses, or variant influenza A viruses² such as A(H3v)], and clinical specimens should be shipped to CDC for confirmatory testing.

The CDC Human Influenza Real-Time RT-PCR Flu Diagnostic Panel (CDC Flu rRT -PCR Dx Panel) testing algorithms should be used as described in the package insert to rule out seasonal influenza A or B virus infection. Public Health officials should contact CDC immediately if they obtain unsubtypeable results when testing an influenza A virus positive specimen.

Specimens that are unsubtypeable or that test presumptive positive for novel influenza A virus at the state public health laboratory should be sent to CDC, Influenza Division, Virology Surveillance and Diagnosis Branch Laboratory for confirmatory testing. Laboratories should not attempt to isolate novel influenza A viruses using viral culture.

The following protocol may be used when testing for novel influenza A viruses with the potential to cause severe disease in humans:

• All state public health laboratories should use the CDC Flu rRT-PCR Dx Panel to screen specimens for InfA, InfB, and RP.
• State public health laboratories should test all InfA-positive specimens with the CDC Influenza A Subtyping kit using all primer/probe sets: H3, pdmInfA and pdmH1. Detailed guidance for testing can be found in the influenza surveillance diagnostic testing algorithm disseminated by the Association of Public Health Laboratories.
• For specimens collected from patients with suspected infection with HPAI A(H5) viruses, testing should be performed using the CDC H5 primer/probe set. Specimens that are positive or inconclusive for A(H5) virus by rRT-PCR at the state health department should be sent to CDC Influenza Division for additional testing as soon as possible.

² Influenza A viruses that normally circulate in pigs are termed "variant viruses" when identified in infected humans.

Antiviral Treatment

Empiric antiviral treatment should be started as soon as possible for all patients with possible infection with novel influenza A viruses with the potential to cause severe disease in humans. Antiviral treatment should not be withheld or delayed pending collection of specimens or laboratory testing. More information on dosing and duration of antiviral therapy is available at CDC's interim guidance on the use of antiviral medication for treatment of human infections with novel influenza A viruses associated with severe human disease. For management of patients with laboratory-confirmed novel influenza A virus infection who are hospitalized with severe pneumonia, the CDC Influenza Division should be consulted.

Sending specimens

Shipping Clinical Specimens to CDC

Specimens to be tested for novel influenza A virus that are shipped from state public health laboratories to CDC should include all information required for seasonal influenza surveillance isolate or specimen submission.

Before sending specimens, state and local health departments should contact the CDC Influenza Division Epidemiology and Prevention Branch at (404) 639-3747 Monday – Friday, 8:30 AM – 5:00 PM (or the on-call epidemiologist at (770) 488-7100 at all other times) or flusupport@cdc.gov.

Specimen Submission Form

Prior to shipping specimens, please reach out to flusupport@cdc.gov for shipping guidelines. Ship specimens to CDC at the following address:

Send diagnostic samples to:
Marie K. Kirby, Ph.D.
Centers for Disease Control and Prevention
Influenza Division, H23-6 (Unit 198)
c/o STAT
1600 Clifton Rd, NE
Atlanta, GA 30329

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