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

Background and Purpose

This document provides interim guidance for clinicians and public health professionals in the United States on appropriate testing, specimen collection and processing for patients who may be infected with novel influenza A viruses with the potential to cause severe illness in people. Examples of such viruses include Asian-lineage avian influenza A (H5N2), (H5N8), and (H5N1)¹ viruses, which were detected in wild and domestic birds in North America in December 2014 and January 2015; these viruses may have some or all of their genes from Asian avian influenza viruses, but for simplicity will all be referred to as “newly detected avian influenza A H5” viruses in this guidance document. Other newly detected avian influenza A H5 viruses also may have the potential to cause severe disease in humans. For a list of avian influenza A H5 virus infections identified in birds in the United States, and their locations, please see an update on avian influenza findings maintained by the US Department of Agriculture. CDC will update this guidance as additional information becomes available.

The appearance of newly detected avian influenza A H5 viruses in North America may increase the likelihood of human infection with these viruses in the United States. Because these newly identified avian influenza A H5 viruses are related to avian influenza A viruses associated with severe disease in humans (e.g., highly pathogenic Asian-lineage avian influenza A (H5N1) virus), they should be regarded as having the potential to cause severe disease in humans until shown otherwise. Other CDC guidance provides recommendations for influenza viruses known to be associated with severe disease in humans.

¹ The H5N1 virus isolated in the United States in January 2015 is a new mixed-origin virus (a “reassortant”) that is genetically different from the H5N1 virus found in several other countries (notably in Asia and Africa), which has caused human infections with high mortality. Although it is related to the H5N1 virus that has caused human infections with high mortality, the ability of this new reassortant H5N1 virus to cause severe disease is currently unknown.

Recommendations for Surveillance, Testing, and Investigation

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

1. Consider the possibility of infection with novel influenza A viruses with the potential to cause severe disease in humans in patients with medically-attended influenza-like illness (ILI) and acute respiratory infection (ARI) who have had recent contact¹ (<10 days prior to illness onset) with sick or dead birds in any of the following categories²:
   1. Domestic poultry (e.g., chickens, turkeys, ducks)
   2. Wild aquatic birds (e.g., ducks, geese, swans)
   3. Captive birds of prey (e.g., falcons) that have had contact with wild aquatic birds
2. If infection with a novel influenza A virus with the potential to cause severe disease in humans is possible, respiratory specimens should be collected with appropriate infection control precautions and sent to the state or local health department for immediate testing (see guidance below).
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.

¹ Contact may include: direct contact with birds (e.g., handling, slaughtering, defeathering, butchering, preparation for consumption); or direct contact with surfaces contaminated with feces or bird parts (carcasses, internal organs, etc.); or prolonged exposure to birds in a confined space.

² For questions or concerns about possible human infection in patients with exposures to birds not listed here, please contact CDC. Exposures that occur in geographic regions in the United States where newly detected avian influenza A H5 viruses have been identified are of most concern.

When Specimens Should Be Collected

The duration of shedding of novel influenza A viruses in humans is largely unknown, and there are currently limited data describing prolonged shedding of people infected with these viruses. Therefore, the estimated duration of viral shedding is based upon seasonal influenza virus infection. 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 tested for novel influenza A virus even if obtained after 7 days from illness onset. Note that prolonged shedding of influenza virus 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.

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Infection Control when Collecting Specimens

Standard, contact, and airborne precautions are recommended for patient management; this includes collection of respiratory specimens. Practitioners should employ infection control precautions consistent with precautions recommended for novel influenza A viruses known to cause severe disease in humans. See 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.

Preferred Respiratory Specimens

The following should be collected as soon as possible after illness onset: (i) a nasopharyngeal swab, or (ii) a nasal aspirate or wash, or (iii) two swabs combined into one viral transport media vial (e.g., nasal or nasopharyngeal swab combined with an oropharyngeal swab). If these specimens cannot be collected, a single nasal, or oropharyngeal swab is acceptable. For patients with lower respiratory tract illness, a lower respiratory tract specimen (e.g., an endotracheal aspirate or bronchoalveolar lavage fluid) may be preferred (these specimens have a higher yield for detecting avian influenza A H5N1 and H7N9 viruses and also may facilitate detection of other novel avian influenza A viruses). Specimens should be placed into sterile viral transport media and immediately placed on refrigerant gel-packs or at 4°C (refrigerator) for transport to the laboratory.

If possible, in order to increase the potential for virus detection, multiple respiratory specimens from different sites should be obtained from the same patient on at least two consecutive days.

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 viral transport medium (e.g., containing protein stabilizer, antibiotics to discourage bacterial and fungal growth, and buffer solution).

Storing Clinical Specimens

Respiratory specimens should be kept at 4°C for no longer than 3 days. Specimens can alternatively be frozen at ≤-70°C. Avoid freezing and thawing specimens if at all possible.

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. If clinical specimens will be examined within 72 hours after collection, keep the specimen at 4°C (2-8°C) and ship on refrigerant gel-packs, otherwise store frozen at ≤-70°C and ship on dry ice. Avoid freezing and thawing specimens. Viability of some pathogens from specimens that were frozen and then thawed is greatly diminished. All specimens should be labeled clearly and include information requested by your state public health laboratory.

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Diagnostic Testing

The performance of current Food and Drug Administration (FDA) cleared diagnostic tests for influenza has been demonstrated for seasonal human influenza viruses as described by the manufacturer package insert. Performance has not been demonstrated with most novel influenza A viruses. Although some diagnostic assays may detect the presence of some novel influenza A viruses, a negative result should not be used to rule out influenza when testing possible human cases. Testing of symptomatic human cases for novel influenza A virus infections should be referred to the nearest public health laboratory.

Existing, commercially available FDA-cleared molecular assays (e.g., rRT-PCR) may fail to detect novel influenza A viruses or may detect with results that indicate “influenza A positive”, but with subtype undetected. For these assays a novel influenza A virus may give an influenza A “unsubtypable” result. Clinicians and laboratorians using molecular assays that are capable of detecting all currently circulating influenza A subtypes (i.e., “seasonal influenza” subtypes) who identify an unsubtypable result should contact CDC and their state or local public health laboratory for additional testing (see below).

Rapid influenza diagnostic tests (RIDTs) and immunofluorescence assays are antigen detection tests that also have unknown sensitivity and specificity to detect human infection with novel influenza A virus in clinical specimens. Some studies suggest that antigen detection tests have low sensitivity to detect H5N1 viruses. Therefore, negative results from either type of test do not exclude novel influenza virus infection, especially in patients with signs and symptoms suggestive of influenza. A negative 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 give a positive influenza A result for a specimen containing novel influenza A virus, but cannot identify the subtype and cannot distinguish a novel influenza A virus from a seasonal influenza A virus. Therefore, testing by rRT-PCR is recommended at state health laboratories for any patient with suspected novel influenza A virus infection.

Clinicians should always consider diagnostic testing for other pathogens that can cause acute febrile respiratory illness since novel influenza A virus infections of humans are very rare, even in exposed persons.

Testing at State Health Departments

Clinicians should notify their state health department immediately when they wish to test a patient for suspected infections with novel influenza A viruses. Specimens to be tested for novel influenza A viruses should be sent first to the state or local 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., H7N9 or H5N1 virus, other avian H5 viruses, or variant influenza viruses² such as H3N2v), and clinical specimens should be shipped to CDC for confirmatory testing.

CDC Flu rRT-PCR Dx Panel testing algorithms should be used as described in the package insert to rule out seasonal influenza virus infection. Public Health officials should contact CDC immediately if they obtain unsubtypable results when testing an influenza specimen.

Specimens that are unsubtypable or that are presumptive positive for novel influenza A 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 Human Influenza Real-Time rRT-PCR Flu Diagnostic 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: H1, H3, pdmInfA and pdmH1. Detailed guidance for testing can be found in the influenza surveillance diagnostic testing algorithm disseminated recently by Association of Public Health Laboratories [27 KB, 1 page].
• Where patients may be infected with influenza A/H5 viruses (see Recommendations for Testing for H5N2 and H5N8 Virus Infections above), test also with H5 primer/probe set. Specimens that are positive for 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 viruses that normally circulate in pigs are termed “variant viruses” when found in humans.

Antiviral Treatment

Until more information is available, antiviral treatment should be given to 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. Empiric treatment with a neuraminidase inhibitor antiviral drug (oral oseltamivir, inhaled zanamivir, or IV peramivir) should be administered immediately according to current guidelines. For discussion of dosing and duration of therapy, see CDC’s interim guidance on the use of antiviral agents for treatment of human infections with avian influenza A (H7N9) viruses.

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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 (all other times).

Ship specimens to CDC at the following address:

Virology, Surveillance and Diagnosis Branch
ATTN: Steve Lindstrom
Influenza Division, NCIRD
Centers for Disease Control and Prevention
1600 Clifton Road NE MS G-16
Atlanta, GA 30333

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