Avian Influenza (Bird Flu): Implications for Human Disease

Last updated January 7, 2009

Agent
Environmental Survival of Avian Influenza Viruses
Avian Influenza: Brief Background
Laboratory Testing for Avian Influenza in Humans
Summary of Avian Influenza in Humans
The Current Outbreak of H5N1 in Birds and Other Animals
H5N1 in Humans: Epidemiologic Features
H5N1 in Humans: Clinical Features
Treatment and Prophylaxis
Current Status of H5N1 Candidate Vaccines
Current WHO and CDC Travel Recommendations
Use of Seasonal Flu Vaccine in Humans at Risk for H5N1 Infection
Surveillance, Case Evaluation, and Contact Follow-up
Influenza Pandemic Considerations
Infection Control Recommendations
Guidance to Protect Workers From Avian Influenza Viruses
Food Safety Issues
References

Agent

Avian influenza is caused by influenza A viruses.

Family: Orthomyxoviridae

Enveloped virions are 80 to 120 nm in diameter and 200 to 300 nm long and may be filamentous.
They consist of spike-shaped surface proteins, a partially host-derived lipid-rich envelope, and matrix (M) proteins surrounding a helical segmented nucleocapsid (6 to 8 segments).
The family contains five genera, classified by variations in nucleoprotein (NP and M) antigens: influenza A, influenza B, influenza C, thogotovirus, and isavirus.

Genus: Influenzavirus A

Consists of a single species: influenza A virus.
Influenza A viruses are a major cause of influenza in humans.
All past influenza pandemics have been caused by influenza A viruses.
The multipartite genome is encapsidated, with each segment in a separate nucleocapsid. Eight different segments of negative-sense single-stranded RNA are present; this allows for genetic reassortment in single cells infected with more than one virus and may result in multiple strains that are different from the initial ones (see References: Voyles 2002).
The genome consists of 10 genes encoding for different proteins (eight structural proteins and two nonstructural proteins). These include: three transcriptases (PB2, PB1, and PA), two surface glycoproteins (hemagglutinin [HA] and neuraminidase [NA]), two matrix proteins (M1 and M2), one nucleocapsid protein (NP), and two nonstructural proteins (NS1 and NS2).
The virus envelope glycoproteins (HA and NA) are distributed evenly over the virion surface, forming characteristic spike-shaped structures. Antigenic variation in these proteins is used as part of the influenza A virus subtype definition (but not used for influenza B or C viruses).

Influenza A virus subtypes

There are 16 different HA antigens (H1 to H16) and nine different NA antigens (N1 to N9) for influenza A. Until recently, 15 HA types had been recognized, but a new type (H16) was isolated from black-headed gulls caught in Sweden and the Netherlands in 1999 and reported in the literature in 2005 (see References: Fouchier 2005).
All known subtypes of influenza A can be found in birds, and wild aquatic birds are the major reservoir for influenza A viruses (see References: Fouchier 2004). Details about mammalian hosts can be found in the document "Avian Influenza (Bird Flu): Agricultural and Wildlife Considerations" on this site.
Human disease historically has been caused by three subtypes of HA (H1, H2, and H3) and two subtypes of NA (N1 and N2). H1 and H3 are the subtypes that currently cause seasonal influenza in human populations around the globe each year.
More recently, human disease has been recognized to be caused by additional HA subtypes, including H5, H7, and H9. Such cases have predominantly been associated with exposure to infected birds. Person-to-person transmission has occurred in a few isolated situations.

Influenza A nomenclature

Antigenic strain nomenclature is based on: (1) host of origin (if other than human), (2) geographic origin, (3) strain number, (4) year of isolation, and (5) HA and NA type. Examples (for human strains) include: A/Hong Kong/03/68[H3N2], A/swine/Iowa/15/30[H1N1]).
As with other influenza A subtypes, standard nomenclature is used to name avian strains (eg, A/chicken/HK/5/98 [H5N1]).

Environmental Survival of Avian Influenza Viruses

Viruses remain infectious after 24 to 48 hours on nonporous environmental surfaces and less than 12 hours on porous surfaces (see References: Bean 1982). (Note: The importance of fomites in disease transmission has not been determined.)
Influenza A viruses can persist for extended periods in water (see References: WHO: Review of latest available evidence on risks to human health through potential transmission of avian influenza [H5N1] through water and sewage). One study of subtype H3N6 found that virus resuspended in Mississippi River water was detected for up to 32 days at 4°C and was undetectable after 4 days at 22°C (see References: Webster 1978). Another study found that several avian influenza viruses persisted in distilled water for 207 days at 17°C and 102 days at 28°C (see References: Stallknecht 1990).
Recent data from studies of H5N1 in domestic ducks have shown that H5N1 can survive in the environment for 6 days at 37ºC (see References: WHO 2004: Laboratory study of H5N1 viruses in domestic ducks).
Inactivation of the virus occurs under the following conditions (see References: OIE 2002, PHS):

Temperatures of 56°C for 3 hours or 60°C or more for 30 minutes
Acidic conditions
Presence of oxidizing agents such as sodium dodecyl sulfate, lipid solvents, and B-propiolactone
Exposure to disinfectants: formalin, iodine compounds

Avian Influenza: Brief Background

The term "avian influenza" is used to describe influenza A subtypes that primarily affect chickens, turkeys, guinea fowls, migratory waterfowl, and other avian species.
"Avian influenza" is an ecological classification that does not correspond exactly to other classification schemes.
Outbreaks of influenza have been recognized in domestic poultry (chickens and turkeys) for many years. Avian influenza strains in domestic chickens and turkeys are classified according to disease severity, with two recognized forms: highly pathogenic avian influenza (HPAI), also known as fowl plague, and low-pathogenic avian influenza (LPAI). Avian influenza viruses that cause HPAI are highly virulent, and mortality rates in infected flocks often approach 100%. LPAI viruses are generally of lower virulence, but these viruses can serve as progenitors to HPAI viruses. All HPAI strains identified to date have involved H5 and H7 subtypes.
Human infections caused by avian strains have been associated with both HPAI and LPAI strains (H5, H7, and H9) (see References: HHS 2005: Pandemic influenza plan).
Evidence that HPAI strains arise from LPAI strains has led the World Organization for Animal Health (OIE) to classify all H5 or H7 strains as notifiable (see References: Alexander 2003, Capua 2004, OIE 2005).
The 1918 influenza pandemic strain (H1N1) appears to be of avian origin (see References: CDC: Information about pandemic influenza viruses). The pandemic strains of 1957-58 (H2N2) and 1968-69 (H3N2) both involved reassortment events between avian and human influenza strains.
H5 subtypes

H5 subtypes can be found throughout the world and include both LPAI and HPAI strains.
H5N1 is responsible for the current panzootic among domestic poultry and other birds in Asia, the Middle East, Europe, and Africa.
Recent genetic characterization of H5N1 strains involved in the current panzootic has demonstrated two distinct phylogenetic clades (clades 1 and 2) (see References: Webster 2006; WHO Global Influenza Program Surveillance Network; WHO 2008: Antigenic and genetic characteristics of H5N1 viruses and candidate H5N1 vaccine viruses developed for potential use as human vaccines). Six different subclades of clade 2 have been recognized; three of these are primarily responsible for recent human H5N1 cases.

Clade 1 viruses have circulated primarily in Cambodia, Thailand, Hong Kong, and Vietnam.
Clade 2.1 viruses have circulated primarily in Indonesia.
Clade 2.2 viruses have a wide geographic distribution and have spread to over 60 countries in Asia, the Middle East, Europe, and Africa.
Clade 2.3 viruses are genetically diverse and continue to circulate in birds in Asia. Viruses from this group have caused illness in humans in China, Lao People's Democratic Republic (PDR), Myanmar, and Vietnam.

H7 and H9 subtypes

H7 includes HPAI and LPAI strains.
H9 is only known to include LPAI strains. H9N2 viruses had been isolated in multiple avian species throughout Asia, the Middle East, Europe, and Africa.
These subtypes have caused infections in humans on rare occasions (see References: CDC: Avian influenza A viruses; NIAID: Timeline of human pandemics). A recent report, however, suggests that human infections with H9N2 viruses may be more common than previously recognized (see References: Wan 2008). The authors also concluded that H9N2 viruses can evolve extensively and reassort, suggesting that they may be capable of undergoing further adaptation for more efficient transmission among mammals, including humans.

Laboratory Testing for Avian Influenza in Humans

General Considerations

Tests for influenza include: viral culture, polymerase chain reaction (PCR), rapid antigen testing, and immunofluorescence.
Laboratory tests are widely used to identify influenza virus at the genus level (influenza A/B) or at the H-type level (H1, H3, and H5). H subtype-specific tests must be used to identify potential avian strains, including H5N1.
The World Health Organization (WHO) recommends forwarding all H5, H7, and H9-positive isolates to an approved influenza reference laboratory for confirmation and N-typing (see References: WHO: Guidelines for global surveillance of influenza A/H5; WHO: Recommendations and laboratory procedures for detection of avian influenza A [H5N1] virus in specimens from suspected human cases).
The sensitivity and specificity of laboratory tests appears to vary with the involved strain, which has implications for avian influenza and other emerging influenza variants (see References: Weinberg 2005).
Laboratory-based influenza surveillance networks

WHO Global Influenza Surveillance Network (see References)
Centers for Disease Control and Prevention (CDC) National Respiratory and Enteric Virus Surveillance System (NREVSS) (see References)
State or local surveillance health department surveillance networks

Specimen Collection

The following information is taken from a field operations guide for H5N1 influenza that was released by WHO in early November 2006 (see References: WHO 2006: Collecting, preserving, and shipping specimens for the diagnosis of avian influenza A [H5N1] virus infection). Information also was taken from the Department of Health and Human Services (HHS) Pandemic Influenza Plan where noted (see References: HHS 2005: Pandemic influenza plan [Part 2, Supplement 2]).

Specimens to collect from suspect cases

Upper respiratory tract

Posterior-pharyngeal (throat) swabs (provide the highest yield)
Nasal swabs with nasal secretions (from the anterior turbinate areas) or nasopharyngeal aspirates or swabs (these specimens are more appropriate for seasonal influenza, and the yield may be lower for avian influenza)

Lower respiratory tract

A tracheal aspirate or bronchoalveolar lavage specimen (if the patient is intubated)

Blood

Serum (acute and convalescent if possible)

Secondary specimens

Plasma in EDTA (for detection of viral RNA)
Rectal swab (for patients with diarrhea)
Spinal fluid (if meningitis is suspected and a spinal tap is performed for diagnostic purposes)
Pleural tap fluid (referred to in the HHS plan)
Autopsy specimens (referred to in the HHS plan)

When to collect specimens from suspect cases

Ideally, a throat swab should be taken within 3 days after illness onset; if initial specimens are negative, but if a high index of suspicion remains, testing should be repeated as soon as possible. (According to the HHS plan, specimens optimally should be collected within 4 days of illness onset.)
Virus may be detected in tracheal aspirates from onset of lower respiratory symptoms until the second or third week of illness.
An acute phase serum sample should be taken 7 days or less after symptom onset, and a convalescent sample should be taken 3 to 4 weeks following illness onset.
Single serum samples should be collected 14 days or later after symptom onset.
Serum or plasma for detecting viral RNA should be obtained during the first 7 to 9 days after symptom onset.
Ideally specimens should be collected before antiviral therapy, but treatment should not be delayed to take specimens.
Specimens should be collected from deceased patients as soon as possible after death.

Specimen collection and transport

Detailed methods for specimen collection and transport are provided in the WHO field guide (see References: WHO 2006: Collecting, preserving, and shipping specimens for the diagnosis of avian influenza A [H5N1] virus infection).
Infection control precautions should be consistently observed during specimen collection.
Only sterile dacron or rayon swabs with plastic shafts should be used. Calcium alginate or cotton swabs or swabs with wooden sticks should not be used (or used only when appropriate swabs are not available).
Viral transport media (VTM) should be used for nasopharyngeal and oropharyngeal swabs and, according to the HHS plan, specimens should be maintained at refrigerator temperature (4ºC to 8oC) until testing is performed. Freezing at -70ºC is best for maintaining viability during extended storage.
According to the HHS plan, with regard to autopsy specimens, large airways have the highest yield for immunohistochemistry (IHC) tests. Eight blocks or fixed-tissue specimens from each of the following sites should be obtained. Fixed tissue should be transported at room temperature (not frozen); fresh unfixed tissue should be frozen.

Central (hilar) lung with segmental bronchi
Right and left primary bronchi
Trachea (proximal and distal)
Representative pulmonary parenchyma from right and left lung

Biosafety and Biosecurity

Biosafety

Updated safety rules and recommendations for influenza virus have been included in the fifth edition of Biosafety in Microbiological and Biomedical Laboratories (BMBL) (see References: HHS: Biosafety in Microbiological and Biomedical Laboratories [BMBL] 5th Edition). Current recommendations for interpandemic and pandemic alert periods include:

Biosafety level 2 (BSL-2) facilities, practices, and procedures are recommended for diagnostic, research, and production activities utilizing contemporary, circulating human influenza strains (eg, H1/H3/B), LPAI strains (eg, H1-4, H6, H8-16) and equine and swine influenza viruses.

Animal biosafety level 2 (ABSL-2) facilities are appropriate for work with these viruses in animal models.
Use of avian and swine influenza viruses requires a permit from the Animal and Plant Health Inspection Service (APHIS) of the US Department of Agriculture (USDA).
Based on economic ramifications and source of the virus, LPAI H5 and H7 and swine influenza viruses may have additional APHIS permit-driven containment requirements, personnel practices, and/or restrictions.

Noncontemporary, wild-type human influenza (H2N2) strains should be handled with increased caution. Important considerations in working with these strains are the number of years since an antigenically related virus last circulated and the potential for a susceptible population.

BSL-3 and ABSL-3 practices, procedures, and facilities are recommended with rigorous adherence to additional respiratory protection and clothing change protocols.
Negative-pressure, high-efficiency particulate air (HEPA)-filtered respirators, or positive air-purifying respirators (PAPRs) are recommended for use.
Cold-adapted, live attenuated H2N2 vaccine strains may continue to be worked with at BSL-2 facilities.

Any research involving reverse genetics of the 1918 influenza strain should proceed with extreme caution. The risk to laboratory workers is unknown at present, but the pandemic potential is thought to be significant. Until further risk assessment data are available, the following practices and conditions are recommended for manipulation of reconstructed 1918 influenza viruses and laboratory animals infected with the viruses. These practices and procedures are considered minimum standards for work with the fully reconstructed virus.

BSL-3 and ABSL-3 practices, procedures, and facilities
Large laboratory animals, such as nonhuman primates, housed in primary-barrier systems in ABSL-3 facilities
Rigorous adherence to additional respiratory protection and clothing change protocols
Use of negative-pressure, HEPA-filtered respirators or PAPRs
Use of HEPA filtration for treatment of exhaust air
Amendment of personnel practices to include personal showers prior to exiting the laboratory

Manipulating HPAI viruses in biomedical research laboratories requires similar caution, because some strains may pose increased risk to laboratory workers and have significant agricultural and economic implications.

BSL-3 and ABSL-3 practices, procedures, and facilities are recommended, along with clothing change and personal showering protocols.
Loose-housed animals infected with HPAI strains must be contained within agriculture-specific BSL-3 (BSL-3-Ag) facilities.
Negative-pressure, HEPA-filtered respirators or PAPRs are recommended for HPAI viruses with potential to infect humans.

When considering the biocontainment level and attendant practices and procedures for work with other influenza recombinant or reassortant viruses, the local Institutional Biosafety Committee should consider, but not limit consideration, to the following in the conduct of protocol-driven risk assessment.

The gene constellation used
Clear evidence of reduced virus replication in the respiratory tract of appropriate animal models, compared with the level of replication of the wild-type parent virus from which it was derived
Evidence of clonal purity and phenotypic stability
The number of years since a virus that was antigenically related to the donor of the HA and NA genes last circulated
If adequate risk assessment data are not available, a more cautious approach utilizing elevated biocontainment levels and practices is warranted. There may be specific requirements regarding the setting of containment levels in institutions that are subject to NIH guidelines.

Recommendations for testing of clinical specimens from patients suspected to have H5N1 influenza include (see References: CDC: Update on influenza A [H5N1] and SARS):

Culture from patients suspected of having avian influenza, other novel influenza strains, or severe acute respiratory syndrome (SARS) coronavirus should be conducted only under enhanced BSL-3 containment (also see Biosecurity below). This includes controlled access, double-door entry with changing room and shower, use of respirators, decontamination of all waste, and showering out of all personnel. These diagnostic activities must be kept separate from routine influenza diagnostic activities (eg, probable H1 or H3 isolates) to prevent recombination.
Indirect immunofluorescence (IFA) of specimens requires BSL-2 containment and practices. Culture biocontainment recommendations should be implemented when IFA is used for culture identification.
Direct detection methods, including commercial antigen detection assays and reverse transcriptase polymerase chain reaction (RT-PCR), should be conducted under BSL-2 conditions with a class II biological safety cabinet. Serologic methods require BSL-2 containment.
If H5N1 avian influenza virus is presumptively identified by one of the above direct methods, further work should be conducted using the enhanced BSL-3 procedures described for culture.
Any new or re-emergent human influenza strain with suspected pandemic potential should be treated as described for H5N1 avian influenza.
Additional requirements and recommendations apply for laboratory work involving live animals.

Biosecurity

Strains of HPAI and the 1918 influenza virus are Select Agents requiring registration with CDC and/or USDA for possession, use, storage, and/or transfer (see References: HHS: Biosafety in Microbiological and Biomedical Laboratories [BMBL] 5th Edition).
HPAI strains are agricultural Select Agents requiring registration of personnel and facilities with the lead agency for the institution (CDC or USDA-APHIS) (see References: USDA/APHIS: Agricultural Bioterrorism Protection Act of 2002). An APHIS permit is required for working with these agents. Additional containment requirements, personnel practices, and/or restrictions may be added as conditions of the permit.
Both registered and exempt laboratories that identify a Select Agent contained in a specimen presented for diagnosis, verification, or proficiency testing must secure the agent against theft, loss, or release until transfer or destruction. Unregistered laboratories must transfer or destroy select agents within 7 days of identification. Any theft, loss, or release of the agent must be reported to the select agent authority (see References: USDA/APHIS: Questions and answers).

Direct Detection Methods

RT-PCR assays

RT-PCR assays use conserved targets such as the matrix (M) protein for genus-level identification. HA and NA targets are used for specific identification of avian subtypes. PCR generally is not used for strain-level identification, which is based on serologic markers.
The sensitivity of RT-PCR has been reported to be in the range of 90% to 100% when compared with cell culture; however, several researchers have reported significantly higher numbers of total positive specimens with RT-PCR, possibly reflecting its ability to detect nonviable virions (see References: Coiras 2003, Hayden 2002, Herrmann 2001, Pachucki 2004, Wallace 1999).
In February 2006, the Food and Drug Administration (FDA) announced clearance of an Influenza A/H5 (Asian Lineage) Virus Real-Time Reverse Transcription–Polymerase Chain Reaction (RT-PCR) Primer and Probe Set and inactivated virus as a source of positive RNA control for the in vitro detection of highly pathogenic influenza A/H5 virus (Asian lineage) (see References: CDC 2006: New laboratory assay for diagnostic testing of avian influenza A/H5 [Asian lineage]). These reagents and assay protocols have been distributed by the CDC to state and city LRN (Laboratory Response Network) laboratories. Testing with the new assay is limited to LRN-designated laboratories.
In September 2008, the FDA approved the Human Influenza Virus Real-Time RT-PCR Detection and Characterization Panel (rRT-PCR Flu Panel), that can differentiate between seasonal and novel influenza strains within 4 hours (see References: HHS 2008: FDA clears new CDC test to detect human influenza).
Multiplexed real-time RT-PCR assays have been developed for specific detection of virus subtype H5 or H5N1 (See References: Kessler 2004, Ng 2005, Payungporn 2005, Wu 2008). Multiplexed RT-PCR assays also have been developed that can detect and identify 12 HA (H1 through H12) and 9 NA (N1 through N9) subtypes commonly isolated from birds, pigs, and humans (see References: Chang 2008).
Samples positive by RT-PCR for a novel influenza subtype should be forwarded to a public health laboratory (if testing was conducted at a private laboratory) or to the CDC for confirmation (see References: HHS 2005: Pandemic influenza plan).

Immunofluorescence

IFA methods may be used to identify influenza to the species level (influenza A or B) or specific H subtypes (including H5) directly from specimens or cell culture. CDC distributes IFA typing and subtyping reagents to WHO-collaborating laboratories, including many health department laboratories. If HPAI strains are suspected, enhanced BSL-3 containment should be used (see References: WHO: Recommended laboratory tests to identify avian influenza A virus in specimens from humans; FDA: Cautions in using rapid tests for detecting influenza A viruses; HHS 2005: Pandemic influenza plan).
Direct immunofluorescence (DFA) methods are faster and less labor intensive than IFA but are less sensitive and are currently only available for genus-specific detection (see other rapid direct tests).

Molecular microarray tests using flow-through chip technology

A molecular microarray for influenza typing and subtyping using a flow-thru chip platform was initially described in 2004 (see References: Kessler 2004).
Two reports released in August 2006 involved a study of the FluChip-55 diagnostic microarray and showed that the test could be a valuable tool in identifying influenza viruses (see References: Mehlmann 2006, Townsend 2006).
Scientists recently have developed an improved microarray test referred to as the “MChip,” which has several advantages over the FluChip. While the FluChip is based on three influenza genes—HA, NA, and M—the MChip is based on only the M gene segment, which mutates much less rapidly. A recent evaluation demonstrated that the assay exhibited a clinical sensitivity of 97% and clinical specificity of 100% (see References: NIAID 2006).

Rapid tests (see References: Call 2005; CDC: Interim guidance for influenza diagnostic testing during the 2004-05 influenza season; Treanor 2005; WHO: Checklist for influenza pandemic preparedness planning)

The WHO, in its Checklist for Influenza Pandemic Preparedness Planning, recommends against routine use of commercial rapid antigen detection kits and suggests they be used for outbreak investigation only when no other options exist (see References: WHO Writing Committee of WHO Consultation on Human Influenza A/H5 2005).
Rapid tests detect viral antigen (generally nucleoprotein) or enzymatic activity (NA) directly on patient specimens using a variety of platforms. Rapid tests are designed to identify influenza A only, influenza A or B without identifying the type, or influenza A or B with type-specific identification.
Reported sensitivities range from 40% to 80%. Sensitivity is generally greater in children than adults and is greater early in the course of illness.
The predictive value of rapid assays without confirmation by a reference test is strongly correlated with disease prevalence in the community, as is clinical diagnosis without laboratory testing. When the disease prevalence is low, the tests' positive predictive value decreases; therefore, positive results should be confirmed by culture or RT-PCR. When the disease prevalence is high, the negative predictive value of the tests will be lower and clinicians should consider confirming negative tests with culture or RT-PCR. Similarly, the diagnostic predictive value increases when the patient's symptoms are strongly suggestive of influenza.
While the sensitivity and specificity of rapid tests has been evaluated for circulating strains, these measures are largely unknown for detection of emerging strains (including pandemic strains) (see References: FDA: Cautions in using rapid tests for detecting influenza A viruses). For example, only 4 (36%) of 11 culture-positive H5N1 influenza A specimens from patients in Thailand were positive by rapid antigen tests (see References: WHO Writing Committee of WHO Consultation on Human Influenza A/H5 2005).

Serology

Serologic testing can be used for retrospective diagnosis of infection but is rarely useful for patient management and is not widely available (see References: Hayden 2002; Treanor 2005; HHS 2005: Pandemic influenza plan). Acute-phase sera should be collected within 1 week after illness onset, and convalescent sera should be collected 2 to 3 weeks later. Peak antibody response occurs 4 to 7 weeks after infection.
The most common serologic methods are complement fixation (CF), hemagglutination inhibition (HAI), and enzyme immunoassays (EIA). A variety of other methods (such as neutralization, microneutralization, single radial hemolysis, radial immunodiffusion, and Western blot) have been reported (see References: Hayden 2002, Rowe 1999).

HAI and EIAs measure antibody to HA. These tests are more sensitive than CF, but their increased specificity appears to limit their ability to detect new strains.
HAI titers of at least 1:40 or serum neutralizing titers of 1:8 or greater are associated with protection. HAI titers, however, in human avian influenza cases generally have been low or undetectable (see References: HHS 2005: Pandemic influenza plan).
The microneutralization assay can sensitively and specifically detect H5N1 antibody in patients with H5N1 influenza. Since the test uses infectious viruses, HPAI strains should be tested under enhanced BSL-3 containment. As with other tests, paired sera are preferable to single specimens (see References: HHS 2005: Pandemic influenza plan).
CF measures antibody response to nucleoprotein, which is conserved among influenza A strains. This feature could be an advantage for diagnosis of infection with novel pandemic strains.

Since most people are repeatedly exposed to influenza viruses, a fourfold rise in titer between acute and convalescent sera generally is considered necessary for confirmation of influenza infection. While paired sera are optimal, single convalescent specimens may be useful in investigations involving novel viruses, since past exposure to the agent is less likely (see References: HHS 2005: Pandemic influenza plan).

Virus Isolation by Cell Culture

Virus isolation is considered the "gold standard" of influenza testing (see References: Hayden 2002, Treanor 2005).

Specimens for culture optimally should be collected within 3 days of illness onset.
Turnaround time for the standard method is 2 to 14 days.
The time to detection in culture, as measured in one study conducted during two influenza seasons, ranged from 5 days (>90% of positive specimens) to 7 days (100% of positive specimens) (see References: Newton 2002).

Isolates obtained from cell culture are required for strain characterization, which is an integral part of global influenza surveillance and monitoring activities during a pandemic (see References: HHS 2005: Pandemic influenza plan).

Susceptibility Testing

Susceptibility testing generally is conducted at specialized laboratories as part of surveillance or research and is considered an integral component of pandemic influenza response.
Plaque reduction assay (see References: Hayden 1980, McKimm-Breschkin 2003)

The traditional influenza susceptibility testing method for the M2 ion channel inhibitors (amantadine, rimantadine)
Can detect a wide range of resistance phenotypes
Limited utility for neuraminidase inhibitors

Enzyme inhibition assays (see References: McKimm-Breschkin 2003,Wetherall 2003)

Useful for assay of neuraminidase inhibitors
Chemiluminescent or fluorescent substrates

Sequence analysis (see References: McKimm-Breschkin 2003,Wetherall 2003)

Used to detect mutations in genes known or suspected to be responsible for resistance
NA gene sequences from strains isolated prior to introduction of the drugs can be used to evaluate current strain sequences
Mutations in the M2 can be used to detect amantadine resistance (see References: Pachucki 2004)

Researchers have recently reported a PCR assay to efficiently and accurately detect oseltamivir-sensitive and oseltamivir-resistant H5N1 strains (see References: Suwannakarn 2006). The assay is based on the fact that oseltamivir resistance is caused by a single amino acid substitution from histidine (H) to tyrosine (Y) at position 274 of the NA active site.
The Neuraminidase Inhibitor Susceptibility Network (NISN) was established to monitor susceptibility of clinical isolates to zanamivir and oseltamivir. The chemiluminescent neuraminidase enzyme assay was chosen by the NISN as the method of choice for testing neuraminidase inhibitors (see References: Wetherall 2003).

Summary of Avian Influenza in Humans

In the past several years, it has become clear that avian influenza viruses can infect humans.

Human Disease Caused by Avian Influenza Viruses

Situations in which avian influenza virus subtypes have been recognized to be transmitted to humans and cause disease are identified in the following table.

Human Cases of Avian Influenza
Year	Subtype	No. of Cases	Location	Comments
1996	H7N7	1	United Kingdom	The case-patient developed conjunctivitis after cleaning a duck house (see References: CDC: Avian influenza A virus infections of humans).
1997	H5N1	18 (6 deaths)	Hong Kong	Case-patients were linked to an outbreak of H5N1 in poultry. Sustained person-to-person transmission did not occur, and the outbreak stopped when all birds in the Hong Kong commercial poultry industry (about 1.4 million) were slaughtered (see References: Yuen 1998).
1999	H9N2	2 (children ages 4 yr, 13 mo)	Hong Kong	Both case-patients had been hospitalized with influenza-like illness and both recovered uneventfully (see References: Peiris 1999, Uyeki 2002). No additional cases of person-to-person transmission occurred. Further investigation demonstrated that H9N2 strains were circulating in poultry in Hong Kong and China, although the viruses were not highly pathogenic for birds.
2002	H7N2	1	United States (Virginia)	Evidence of infection was found in one person in Virginia following a poultry outbreak (see References: CDC: Avian influenza A virus infections of humans).
2003	H5N1	2 (1 death)	Hong Kong	The 2 case-patients were family members who had recently traveled to China (see References: CDC: Avian influenza infection in humans). A third family member died while in China of an undiagnosed respiratory illness. No direct link between these cases and H5N1infection in poultry was identified.
2003	H7N7	89 (1 death)	The Netherlands	During an outbreak of H7N7 avian influenza in poultry, infection spread to poultry workers and their families in the area (see References: Fouchier 2004, Koopmans 2004, Stegeman 2004). Most patients had conjunctivitis, and several complained of influenza-like illness. The death occurred in a 57-year-old veterinarian. Subsequent serologic testing demonstrated that additional case-patients had asymptomatic infection.
2003	H7N2	1	New York	The source of exposure was not determined (see References: CDC 2004: Influenza activity).
2003	H9N2	1 (child)	Hong Kong	The source of infection remains unknown (see References: CDC 2004: Influenza activity).
2003-2008 (ongoing)	H5N1	More than 390, with a case-fatality rate >60%, according to official WHO numbers	Azerbaijan, Bangladesh, Cambodia, China, Djibouti, Egypt, Indonesia, Iraq, Lao PDR, Myanmar, Nigeria, Pakistan, Thailand, Turkey, Vietnam	Human cases are associated with an ongoing extensive outbreak of avian influenza in poultry (see References: WHO: Cumulative number of confirmed human cases of avian influenza A (H5N1). More information on this situation can be found in the section below.
2004	H7N3	2	Canada (British Columbia)	Two poultry workers became ill during an outbreak of H7N3 avian influenza in poultry (see References: Health Canada 2004). Both had conjunctivitis.
2004	H10N7	2 (infants)	Egypt	One child's father was a poultry merchant (see References: NIAID: Timeline of human flu pandemics).
2006	H7N3	1	United Kingdom	Although a number of exposed persons had symptoms of conjunctivitis or influenza-like illness, only one poultry worker had a laboratory-confirmed infection (see References: Morgan 2009).
2007	H7N2	4	United Kingdom	Case-patients were associated with a poultry outbreak of H7N2 in Wales (see References: CDC: Avian influenza A virus infections of humans). The case-patients had conjunctivitis and influenza-like illness.
2007	H9N2	1 (infant)	Hong Kong	The source of infection is unknown, although the child had visited a bird market with her parents before illness onset (see Mar 28, 2007, CIDRAP News story).
2008	H9N2	1 (infant)	China	The source of infection is unknown (see Dec 29, 2008 CIDRAP News story)

Information on Human Exposure to Avian Influenza Viruses

A serologic survey of 39 duck hunters and 68 wildlife professionals in Iowa conducted in late 2004 and early 2005 found that one duck hunter and two wildlife workers had serologic evidence of past infection to avian influenza virus H11N9. All three had extensive exposure to wild ducks and geese (see References: Gill 2006).

Another study of 42 veterinarians showed that the veterinarians were significantly more likely to have antibodies to avian influenza subtypes H5, H6, and H7 (indicating past infection with these viruses) compared with a group of 66 healthy nonveterinarian control subjects (see References: Myers 2007). Furthermore, veterinarians who had examined birds had a higher likelihood of having increased antibodies to the three avian subtypes compared with veterinarians who did not have exposure to birds.

A recent study from California involved sampling wild birds and marine mammals for avian influenza viruses from October 2005 through August 2007. The authors then estimated human-wildlife contact based on the prevalence of infection in the bird populations (see References: Siembieda 2008). The investigators defined three levels of contact (casual, recreational, and occupational) and sampled corresponding bird populations. The bird populations included the following: for casual contact, periurban species (eg, sparrows, crows, finches); for recreational contact, hunter-killed waterfowl; and for occupational contact, wild birds and sea mammals admitted to three wildlife hospitals in northern California. The authors found that waterfowl hunters (ie, those with recreational exposure) were eight times more likely to have contact with infected wildlife than those with casual or occupational exposures.

The Current Outbreak of H5N1 in Birds and Other Animals

An outbreak of HPAI caused by a strain of H5N1 avian influenza started in Asia in the fall of 2003 and spread in domestic poultry farms at an historically unprecedented rate. The outbreak tapered off in spring 2004 but in summer re-emerged in several countries in Asia (including Cambodia, China, Lao PDR, Thailand, and Vietnam), where it is ongoing.

In the summer of 2005, H5N1 began expanding its geographic range beyond Asia; this trend has continued into 2008 (see References: WHO 2008: H5N1 avian influenza: timeline of major events). For detailed information on avian influenza, see the document, Avian Influenza (Bird Flu): Agricultural and Wildlife Considerations, on this Web site.

Areas affected by H5N1 avian influenza in poultry or migratory birds as of January 2009 are shown in the following table (see References: FAO 2008).

Countries Affected by H5N1 in Poultry and Wild Birds as of January 2009
East Asia, Southeast Asia	Europe	Siberia, Central Asia, Middle East	Africa
Cambodia China Hong Kong Indonesia Japan Lao PDR Malaysia Myanmar Mongolia South Korea Thailand Vietnam	Albania Austria Bosnia-Herzegovina Bulgaria Croatia Czech Republic Denmark France Germany Greece Hungary Italy Poland Romania Russia (European Russia) Scotland Serbia Slovakia Slovenia Spain Sweden Switzerland United Kingdom	Afghanistan Azerbaijan Bangladesh Cyprus Georgia (former Soviet republic) India Iran Iraq Israel Jordan Kazakhstan Kuwait Pakistan Turkey Ukraine Russia (Siberia) Saudi Arabia West Bank and Gaza Strip	Benin Burkina Faso Cameroon Djibouti Egypt Ghana Ivory Coast Niger Nigeria Sudan Togo

H5N1 in Humans: Epidemiologic Features

Case Occurrence

The WHO has officially recognized more than 390 human cases of H5N1 influenza; cases have been reported from Azerbaijan, Bangladesh, Cambodia, China, Djibouti, Egypt, Indonesia, Iraq, Myanmar, Lao PDR, Nigeria, Pakistan, Thailand, Turkey, and Vietnam (see References: WHO: Cumulative number of confirmed human cases of avian influenza A [H5N1]; WHO: Situation updates).

An epidemiologic report on 340 confirmed H5N1 influenza cases published by the WHO in January 2008 demonstrated that the median age of cases was 18 years and that 90% of infections occurred in persons under 40 years of age (see References: WHO: Writing Committee of the Second World Health Organization Consultation on Clinical Aspects of Human Infection with Avian Influenza A [H5N1] Virus 2008).

The overall case-fatality rate was 61% and was highest among persons 10 to 19 years of age and lowest among persons 50 years of age or older.
Of six infected pregnant women, four died and two had a spontaneous abortion.
Clusters of illness with at least two epidemiologically linked cases have been identified in 10 countries and have accounted for about 25% of all cases. Most clusters have involved two or three people and most have occurred among blood relatives; the largest cluster involved eight people (though the index case did not involve confirmatory tests). Persons involved in case clusters probably acquired infection from common-source exposures to poultry, but person-to-person transmission has occurred on occasion.

Exposure Information

Most cases have involved direct contact with poultry. Types of exposures that have been identified to date include:

Slaughtering, plucking, and preparing diseased birds
Handling fighting cocks or ducks that appear to be well
Playing with or holding diseased or dead poultry
Consumption of raw or undercooked poultry or poultry products (such as duck blood)

Low perceived risk and high population exposures to live chickens appear to be factors that are contributing to the spread of H5N1 from infected birds to humans (see References: Fielding 2005). For example, a survey of households in an area of rural Thailand affected by avian influenza found that 74% of households surveyed owned live poultry (see References: Olsen 2005: Poultry-handling practices during avian influenza outbreak, Thailand).

A case-control study from Vietnam found that the following risk factors were independently associated with H5N1 infection (see References: Dinh 2006):

Preparing sick or dead poultry for consumption in the 7 days before illness onset
Having sick or dead poultry in the household in the 7 days before illness onset
Lack of an indoor water source

Following recognition of a case of avian influenza in a rural village in southern Cambodia in 2005, investigators conducted a retrospective survey of poultry deaths and a seroepidemiologic survey of villagers (see References: Vong 2006). Serologic testing of villagers approximately 2 months after outbreaks in poultry did not demonstrate any recent H5N1 infections, despite close contact with birds likely to have been infected with H5N1. These findings illustrate the following: (1) H5N1 was not easily transmitted from birds to humans and (2) asymptomatic or mildly symptomatic human infections did not occur.

A cross-sectional serologic survey of 322 poultry workers in areas of Thailand where outbreaks of avian influenza had occurred during the previous 6 months did not detect any workers who met the WHO criteria for confirmed infection (see References: Hinjoy 2007). These findings support the perspective that H5N1 avian influenza virus is not easily transmitted from birds to humans. A similar study conducted in Nigeria also found no serologic evidence of H5N1 infection among poultry workers (see References: Ortiz 2007).

The first report of H5N1 disease in humans contracted through exposure to wild birds occurred in the spring of 2006 (see References: Gilsdorf 2006). The discovery was made in a cluster of human cases in Azerbaijan; family members denied any contact with ill domestic poultry, but many wild swans had died in the area and were thought to have played a role. In August 2006, the CDC released a set of guidelines for conducting surveillance on dead birds (see References: CDC: Interim guidance for states conducting avian mortality surveillance for West Nile virus (WNV) and/or highly pathogenic H5N1 avian influenza virus).

In approximately 25% of cases, the source of exposure remains unclear and environment-to-human transmission is considered a possibility (such as through contact with virus-contaminated fomites).

A recent case report suggests that food markets with live birds may be a source of exposure for avian influenza (see References: Wang 2006).
Another report from China involving six cases with no obvious exposure to sick poultry found that all six had visited live poultry markets before illness onset (see References: Yu 2007).

Person-to-Person Transmission

To date, sustained person-to-person transmission has not been recognized, although probable person-to-person spread was identified in Thailand involving transmission from an ill child to her mother and aunt (see References: Ungchusak 2005) and several other familial clusters have been recognized (see References: Olsen 2005: Family clustering of avian influenza A [H5N1]).

In May 2006, WHO reported an H5N1 influenza cluster in Indonesia involving seven cases of person-to-person transmission; one of the cases involved two generations of transmission (see References: WHO: Avian influenza: Situation in Indonesia: Update 14 and see May 24, 2006, CIDRAP News story). An Indonesian official recently put the number of clusters in that country at 10, all involving cases in blood relatives (see Jan 12, 2007, CIDRAP News story).

Inefficient transmission of current H5N1 strains may be related to lack of appropriate avian virus cell receptors in the upper respiratory tracts of humans and the inability of H5N1 strains to recognize human cell receptors (see References: Shinya 2006). A mutation allowing H5N1 avian influenza virus to recognize human cell receptors could enhance person-to-person transmission owing to the potential for greater viral replication in the upper respiratory tract.

H5N1 in Humans: Clinical Features

Clinical Features

H5N1 influenza generally presents as a severe pneumonia that often progresses to acute respiratory distress syndrome (ARDS). The summary table below outlines clinical and laboratory features for H5N1 influenza cases reported to the WHO through mid-December 2007 (see References: WHO: Writing Committee of the Second World Health Organization Consultation on Clinical Aspects of Human Infection with Avian Influenza A [H5N1] Virus 2008).

Clinical and Common Laboratory Features of Influenza A (H5N1) Disease at Hospital Admission
	Vietnam, Thailand, and Cambodia 2004-05 (Clade 1)	Indonesia 2005-06 (Clade 2.1)	China 2005-06 (Clade 2.3)	Egypt 2006-07 (Clade 2.2)	Turkey, Azerbaijan 2006 (Clade 2.2)
Outcome or Measure	No./Total No. (%)	No./Total No. (%)	No./Total No. (%)	No./Total No. (%)	No./Total No. (%)
Fever (>38°C)	41/41 (100)	54/54 (100)	8/8 (100)	34/38 (89)	15/16 (94)
Dyspnea	33/37 (89)	51/54 (94)	4/8 (50)	14/38 (37)	7/16 (44)
Cough	40/41 (98)	50/54 (93)	7/8 (88)	27/38 (71)	12/15 (80)
Pneumonia	41/41 (100)	54/54 (100)	8/8 (100)	23/38 (61)	14/16 (88)
Coryza	9/27 (33)	NR	NR	NR	2/14 (14)
Sore throat	13/41 (32)	NR	NR	26/38 (68)	14/16 (88)
Vomiting	5/31 (16)	6/54 (11)	NR	3/37 (8)	0/7 (0)
Diarrhea	16/32 (50)	6/54 (11)	NR	2/37 (5)	4/14 (29)
Depressed consciousness	NR	NR	NR	3/38 (8)	4/8 (50)
Seizures	NR	9/17 (53)	NR	NR	2/7 (29)
Headache	5/14 (36)	13/17 (76)	NR	19/38 (50)	7/15 (47)
Conjunctivitis	0/22 (0)	NR	NR	14/38 (37)	1/8 (13)
Myalgia	11/37 (30)	7/54 (13)	NR	17/38 (45)	4/15 (27)
Leukopenia	17/22 (77)	41/48 (85)	NR	10/37 (27)	11/15 (73)
Lymphopenia	16/24 (67)	16/29 (55)	NR	4/25 (16)	7/13 (54)
Thrombocytopenia	13/24 (54)	29/45 (64)	NR	8/26 (31)	9/13 (69)
Increased aminotransferase levels	20/28 (71)	NR		15/27 (56)	6/8 (75)
Abbreviations: NR: Not reported. Data were obtained from a recent WHO report (see References: WHO: Writing Committee of the Second World Health Organization Consultation on Clinical Aspects of Human Infection with Avian Influenza A [H5N1] Virus 2008).

The incubation period for most patients with H5N1 influenza is 2 to 5 days; however, the range appears to be as long as 8 or 9 days. A recent report from China that assessed incubation periods for 24 patients with H5N1 avian influenza found that the median incubation period for patients exposed to a wet poultry market was significantly longer than for patients exposed to sick or dead poultry (7 days [range 3.5–9 days] vs. 4.3 days [range 2–9 days) (see References: Huai 2008).

Case-fatality rates by clade are outlined below (see References: WHO: Writing Committee of the Second World Health Organization Consultation on Clinical Aspects of Human Infection with Avian Influenza A [H5N1] Virus 2008).

Clade 1 (Cambodia, Thailand, and Vietnam): 54% (66/123)
Clade 2.1 (Indonesia): 79% (76/96)
Clade 2.2 (Azerbaijan, Djibouti, Egypt, Iraq, Nigeria, and Turkey): 44% (26/59)
Clade 2.3 (China, Lao PDR): 65% (17/26)

A case series reported from Indonesia involving 122 hospitalized patients found that patients for whom treatment was initiated within 2 days after illness onset had a significantly lower mortality than patients for whom treatment was initiated at 5 to 6 days or later than 7 days (see References: Kandun 2008). Similarly, a study of 26 cases identified in China between October 2005 and April 2008 found that a higher proportion of patients who received antiviral drugs survived compared to those who were untreated (see References: Yu 2008).

The high case-fatality rate suggests that the pathogenicity of H5N1 may be similar to (or more severe than) the 1918 H1N1 pandemic strain. Researchers have hypothesized that cytokine storm (ie, overproduction of cytokines) may have played an important role in the pathogenesis of the 1918 pandemic strain. A laboratory-based study involving H5N1 strains taken from ill humans in Asia (during 1997 and 2004) and an ordinary current H1N1 strain (circulating in Asia in 1998) found that all the H5N1 viruses caused human alveolar cells and bronchial epithelial cells to secrete significantly higher levels of various cytokines and chemokines than did the ordinary virus (see References: Chan 2005). Another study demonstrated a strong induction of chemokines and their receptors in macrophages infected by H5N1 and H9N2 avian influenza viruses (see References: Zhou 2006). Finally, a case series reported from Vietnam involving patients with H5N1 influenza showed that high viral load and high chemokine and cytokine levels are central to the pathogenesis of H5N1 influenza (see References: de Jong 2006). These findings support the role of cytokine storm in the pathogenesis of H5N1. A recent report also suggests that avian H5N1 influenza virus leads to substantial cell death in mammalian airway epithelial cells due to the induction of apoptosis (see References: Daidoji 2008).

Some patients have presented with primarily gastrointestinal symptoms. In addition, the case report of a 4-year-old Vietnamese child with H5N1 avian influenza who presented in 2004 with encephalitis demonstrated the following features (see References: De Jong 2005: Fatal avian influenza A (H5N1) in a child presenting with diarrhea followed by coma):

The child presented with a 2-day history of fever, headache, vomiting, and severe diarrhea (approximately 10 episodes per day). The stools were watery without blood or mucus.
Laboratory tests on admission were unremarkable and chest x-ray was normal.
On the third day following initial presentation, the child had a generalized convulsion and became comatose. Respiratory failure developed and he died on the fifth day after initial presentation. Acute encephalitis of unknown origin was reported as the cause of death; no autopsy was performed.
H5N1 influenza A virus was isolated from cerebrospinal fluid, fecal, throat, and serum specimens.
The patient's 9-year-old sister had died 2 weeks earlier from a similar clinical syndrome.

In general, asymptomatic or mild infections appear to be uncommon for H5N1, based on available seroepidemiologic data (see References: Vong 2006; WHO: Writing Committee of the Second World Health Organization Consultation on Clinical Aspects of Human Infection with Avian Influenza A [H5N1] Virus 2008). Exceptions are noted below.

Asymptomatic seroconversions were noted among some poultry workers and healthcare workers in the 1997 outbreak in Hong Kong, but that strain was different from later strains (see References: WHO Writing Committee of WHO Consultation on Human Influenza A/H5 2005).
In response to H5N1 outbreaks in poultry in South Korea in 2004, follow-up serologic testing of more than 2,000 poultry workers found nine workers over time with serologic evidence of infection but no history of clinical illness (see Sep 21, 2006, CIDRAP News story). However, the South Korean strain is genetically distinct from Thai and Vietnamese strains and was found to have a low level of pathogenicity in mice (see References: Lee 2005).
Relatively mild illness was noted in three of eight cases detected as part of three case clusters in Indonesia (see References: Kandun 2006). All of the mild cases were in children.

Treatment and Prophylaxis

Available Agents

Two groups of antiviral agents are available for treatment and prophylaxis of influenza: M2 ion-channel inhibitors (the adamantanes [amantadine and rimantadine]) and the neuraminidase inhibitors (NIs) (oseltamivir [Tamiflu] and zanamivir [Relenza]). The WHO considers the NIs to be the major class of antiviral agents for treatment and prophylaxis of H5N1 infection.

Oseltamivir (given orally in capsule form) is approved for treatment and prevention of influenza in adults and children older than 1 year of age (see References: Moscona 2005). Data from uncontrolled clinical trials suggest that use of oseltamivir may improve survival for patients with H5N1 influenza (see References: WHO: Writing Committee of the Second World Health Organization Consultation on Clinical Aspects of Human Infection with Avian Influenza A [H5N1] Virus 2008).
Zanamivir (a powder that is inhaled by mouth) is approved for treatment of influenza in adults and children more than 7 years of age (see References: Moscona 2005). In March 2006, the FDA approved the use of zanamivir for prevention of influenza in adults and children aged 5 and older. The value of zanamivir in treating H5N1 influenza has not been studied to date, although suboptimal delivery to sites of infection in patients with pneumonic or extrapulmonary disease is a concern (see References: WHO: Writing Committee of the Second World Health Organization Consultation on Clinical Aspects of Human Infection with Avian Influenza A [H5N1] Virus 2008).

Other Antiviral Agents Under Consideration

Currently, no parenteral formulations of antiviral agents are available for seriously ill patients. Two NIs, zanamivir and peramivir, have undergone or are undergoing clinical trials for use by intravenous or intramuscular administration, and one long-acting NI (designated CS-8958) is being considered for use by inhalation (see References: Hayden 2008).

Other new agents that are being evaluated are T-705, a polymerase inhibitor, and DAS181, an attachment inhibitor (see References: Hayden 2008). Passive immunization with human H5-specific monoclonal antibodies has been considered as a possible treatment for acute H5N1 influenza virus infection. A recent study found that monoclonal antibodies against conserved regions in the H5 HA may be broadly cross-protective against diverse H5N1variants (see References: Chen 2009).

Antiviral Resistance

Available data on antiviral resistance are outlined below.

Resistance to adamantanes: For influenza strains in general, transmissible amantadine-resistant organisms are shed by about 30% of patients after 2 to 5 days of treatment.

Viral resistance to adamantanes can emerge rapidly, because a single point mutation can confer resistance to both amantadine and rimantadine.
Clade 1 H5N1 viruses and most clade 2 H5N1 viruses from Indonesia are fully resistant to adamantanes; whereas, clade 2 viruses from other lineages in Eurasia and Africa remain susceptible.

Resistance to oseltamivir: Until recently, levels of resistance to oseltamivir have remained relatively low.

During the 2007-08 influenza season, 10.9% of H1N1 viruses tested in the United States were resistant to oseltamivir.
As of mid-December 2008, 98% of 50 H1N1 isolates tested in the United States for the 2008-09 influenza season were resistant to oseltamivir (all were susceptible to zanamivir, amantadine, and rimantadine).

Oseltamivir-resistant H5N1 strains have been isolated from several patients in Vietnam. One was a Vietnamese child who received prophylactic treatment with the drug (see References: Le 2005); another report involved two additional patients, both of whom died of H5N1 influenza (see References: De Jong 2005).
Clade 1 H5N1 viruses appear to be 15 to 30 times more sensitive to oseltamivir than clade 2 H5N1 isolates from Indonesia and Turkey (see References: WHO: Writing Committee of the Second World Health Organization Consultation on Clinical Aspects of Human Infection with Avian Influenza A [H5N1] Virus 2008).
A study of H5N1 influenza isolates from chickens, ducks, geese, and quail from Vietnam and Malaysia (2004), Cambodia (2004 and 2005), and Indonesia (2005) showed that sensitivities to oseltamivir fell into three groups when compared with a reference human H1N1 influenza strain (see References: McKimm-Breschkin 2007).

The clade 1 isolates from 2004 were all more sensitive to oseltamivir than the H1N1 influenza control.
The 2005 Cambodian viruses showed a sixfold to sevenfold decrease in oseltamivir sensitivity in comparison to the 2004 Cambodian isolates.
All of the clade 2 2005 Indonesian viruses demonstrated a 15- to 30-fold decrease in sensitivity specifically to oseltamivir compared with clade 1 viruses.

Genetic markers for reduced susceptibility to oseltamivir were noted in H5N1 isolates from two Egyptian patients who became ill and died in December 2006; both patients had been treated with oseltamivir for 2 days before the isolates were obtained (see References: Influenza Team, European Centre of Disease Prevention and Control 2007).
A recent report suggested that natural variation in sensitivity to oseltamivir may exist among different H5N1 strains, indicating that ongoing surveillance and testing for sensitivity to NIs is essential for tracking trends (see References: Rameix-Welti 2006).

Resistance to zanamivir: No resistance has been detected in previously healthy patients with influenza who have been treated with zanamivir.

One influenza B isolate with reduced sensitivity was obtained from an immunocompromised (post-bone marrow transplant) 18-month-old child after 12 days of treatment (see References: Gubareva 1998).
The study noted above by McKimm-Breschkin et al. showed that all clade 1 and clade 2 viruses from Vietnam, Cambodia, Malaysia, and Indonesia had a similar sensitivity to zanamivir as the reference influenza H1N1 isolate (see References: McKimm-Breschkin 2007).

Current Recommendations for Treatment and Prophylaxis

In May 2006, the WHO released guidelines on use of antiviral agents for H5N1 influenza treatment and prophylaxis (see References: WHO: Rapid Advice Guidelines on pharmacological management of humans infected with avian influenza A [H5N1] virus). Treatment guidelines were updated in August 2007. The updated recommendations on treatment are outlined below (see References: WHO: Clinical management of human infection with avian influenza A [H5N1] virus).

Oseltamivir remains the primary recommended antiviral treatment. Evidence that the H5N1 virus continues to replicate for a prolonged period indicates that treatment with oseltamivir also is warranted when the patient presents to clinical care at a later stage of illness.
Modified regimens of oseltamivir treatment, including twofold higher dosage (ie, 150 mg twice daily for adults), longer duration, and possibly combination therapy with amantadine or rimantadine (in countries where H5N1 viruses are likely to be susceptible to adamantanes) may be considered on a case-by-case basis, especially for patients with pneumonia or progressive disease.

Preclinical studies have shown that combinations of oseltamivir and adamantanes have enhanced antiviral activity and reduced resistance emergence.
Combination therapy should only be considered when the locally circulating H5N1 viruses (clade 2.2 and 2.3) are likely to be susceptible to adamantanes and, whenever possible, with collection of serial respiratory samples for serial virological monitoring.

Corticosteroids should not be used routinely, but may be considered for septic shock with suspected adrenal insufficiency requiring vasopressors. Prolonged or high-dose corticosteroids can result in serious adverse events in H5N1 virus-infected patients, including opportunistic infection.
Antibiotic chemoprophylaxis should not be used. When pneumonia is present, however, antibiotic treatment is appropriate initially for community-acquired pneumonia according to published evidence-based guidelines. When available, the results of microbiologic studies should be used to guide antibiotic usage for suspected bacterial coinfection in patients with H5N1 virus infection.
Monitoring of oxygen saturation (eg, pulse oximetry, arterial blood gases) should be performed whenever possible at presentation and routinely during subsequent care, and supplemental oxygen should be provided to correct hypoxemia.
Therapy for H5N1 virus-associated ARDS should be based upon published evidence-based guidelines for sepsis-associated ARDS, specifically including lung protective mechanical ventilation strategies.

The May 2007 WHO guidelines on use of antiviral agents for H5N1 influenza treatment and prophylaxis outline the following recommendations for chemoprophylaxis (see References: WHO: Rapid Advice Guidelines on pharmacological management of humans infected with avian influenza A [H5N1] virus).

Where NIs are available:

In high-risk exposure groups, including pregnant women, oseltamivir should be administered as chemoprophylaxis, continuing for 7 to 10 days after the last exposure (strong recommendation); zanamivir could be used in the same way (strong recommendation) as an alternative.
In moderate-risk exposure groups, including pregnant women, oseltamivir might be administered as chemoprophylaxis, continuing for 7 to 10 days after the last exposure (weak recommendation); zanamivir might be used in the same way (weak recommendation).
In low-risk exposure groups, oseltamivir or zanamivir should probably not be administered for chemoprophylaxis (weak recommendation). Pregnant women in the low-risk group should not receive oseltamivir or zanamivir for chemoprophylaxis (strong recommendation).
Amantadine or rimantadine should not be administered as chemoprophylaxis (strong recommendation).

Where NIs are not available:

In high- or moderate-risk exposure groups, amantadine or rimantadine might be administered for chemoprophylaxis if local surveillance data show that the virus is known or likely to be susceptible to these drugs (weak recommendation).
In low-risk exposure groups, amantadine and rimantadine should not be administered for chemoprophylaxis (weak recommendation).
In pregnant women, amantadine and rimantadine should not be administered for chemoprophylaxis (strong recommendation).
In the elderly, people with impaired renal function, and individuals receiving neuropsychiatric medication or with neuropsychiatric or seizure disorders, amantadine should not be administered for chemoprophylaxis (strong recommendation).

Stockpiling of Antiviral Agents

If H5N1 avian influenza escalates into a pandemic, stockpiles of antiviral agents may be used for treatment and to curtail spread through prophylaxis targeted to exposed persons. The WHO has stockpiled five million treatment courses of Tamiflu (donated by Roche) for initial pandemic response (see References: Roche: Roche update on Tamiflu for pandemic influenza preparedness). In addition, a number of countries around the globe have developed national antiviral stockpiles (see References: Roche: Roche update on Tamiflu for pandemic influenza preparedness).

One of the cornerstones of the WHO protocol for rapid response and containment is deployment of the Roche international antiviral stockpile to be used initially for targeted antiviral prophylaxis (for known case contacts) and for mass antiviral prophylaxis as needed (either by offering prophylaxis to the affected population within a radius of 5 to 10 km from each detected case or covering at-risk populations in defined administrative areas) (see References: WHO 2006: Pandemic influenza draft protocol for rapid response and containment).
Through a global production network, Roche is currently able to produce in excess of 400 million treatment courses of Tamiflu annually. This network includes eight Roche sites and 19 external manufacturing partners located in nine different countries around the world (see References: Roche: Roche update on Tamiflu for pandemic influenza preparedness).

According to the US federal pandemic influenza plan, HHS intends to ensure a large enough stockpile of antiviral agents to treat approximately 25% of the US population (see References: HHS: Pandemic influenza plan 2005 [Supplement 7]).

The current HHS target is to have 81 million antiviral treatment courses on hand (50 million for distribution to states through the Strategic National Stockpile [SNS] when an influenza pandemic is judged imminent and 31 million in state stockpiles). The total includes six million treatment courses set aside for the early stages of an emerging pandemic and 75 million courses targeted for treatment of ill persons.
The stockpile currently comprises about 80% oseltamivir and 20% zanamivir (see References: HHS 2008: Guidance on antiviral drug use during an influenza pandemic).
In addition, several million courses of rimantadine, purchased in a season of influenza vaccine shortage, are still held in the SNS. Additional stockpiling of M2-inhibitor antiviral drugs (amantadine and rimantadine) has not been recommended, because resistance to these agents among circulating influenza A viruses is frequent and develops rapidly when they are used to treat influenza A virus infections.
Recommendations for use of the federal stockpile and ethical considerations are outlined in a recently released guidance document from the CDC (see References: HHS 2008: Guidance on antiviral drug use during an influenza pandemic).
The Roche supply chain is fully operational in the US, with an annual production capacity of 80 million treatment courses (see References: Roche 2006).
HHS also has suggested in another guidance document that employers should consider stockpiling antiviral agents for their employees (see References: HHS 2008: Considerations for antiviral drug stockpiling by employers in preparation for an influenza pandemic).

Even though antiviral stockpiles are considered to be an important strategy for pandemic preparedness, a number of caveats exist regarding their successful use for treatment or prophylaxis during a pandemic (see References: Democratis 2006).

It is not clear that such agents would be effective against the emergent pandemic strain.
Resistance to widely used antiviral agents could develop quickly during a global pandemic.
Rapid delivery of antiviral agents to newly diagnosed patients or contacts poses substantial logistical challenges, particularly in developing countries.
Even if antiviral agents are shown to be effective, the dose and duration of treatment may depend on the virulence of the pandemic strain. Current antiviral treatment recommendations for influenza are based on studies using circulating H3N2 strains and not on potentially more virulent pandemic strains. For example, since H5N1 strains can be highly virulent, higher doses of antiviral agents given for a longer period of time may be necessary for effective treatment. This was recently demonstrated in a mouse model using an H5N1 strain from Vietnam (see References: Yen 2005).
Even with the increased global capacity for production of oseltamivir, it is not clear whether enough Tamiflu will be available to meet the demand during a pandemic.

Current Status of H5N1 Candidate Vaccines

In April 2004, the WHO made the prototype seed strain for an H5N1 vaccine available to manufacturers (see References: WHO: Avian influenza: situation in Thailand; status of pandemic vaccine development). Over time, the WHO has added additional strains as candidate vaccine viruses. As of February 2008, reassortants with complete regulatory approval included viruses of clade 1, clade 2.1, clade 2.2, and clade 2.3.4. Regulatory approval for several additional viruses is pending, and other viruses have been proposed (see References: WHO 2008: Antigenic and genetic characteristics of H5N1 viruses and candidate H5N1 vaccine viruses developed for potential use as human vaccines). Comparisons of the candidate H5N1 vaccine viruses are ongoing.

Even though progress is being made, a number of barriers exist to actually having effective vaccines against H5N1 influenza available for practical use.

Even with candidate vaccines currently under study, research and development of vaccines that are "market ready" will likely take several more years.
Study of candidate vaccines is hampered by a lack of established correlates of immunity in animals and humans. Developing consistent immunologic end points for clinical trials remains an important challenge (see References: Haque 2007).
Limited global vaccine production capacity exists at this time; currently only about 565 million doses of trivalent influenza vaccine containing 15 mcg HA per strain can be produced each year (see References: WHO 2007: Projected supply of pandemic influenza vaccine sharply increases). This is equivalent to about 1.7 billion doses of monovalent vaccine at the same antigen level. The number of doses could be smaller if an H5N1 vaccine requires higher levels of antigen. Similarly, if two doses are needed, the number of people who could be vaccinated will decrease.

The WHO anticipates that by 2010 global production capacity for seasonal vaccine will increase to 1 billion doses per year.
WHO officials believe that the amount of antigen needed per dose of H5N1 vaccine may ultimately be about eight times less than trivalent vaccine and that, as a result, global annual production capacity could be as high as 4.5 billion doses of H5N1 vaccine by 2010 (see References: WHO 2007: Projected supply of pandemic influenza vaccine sharply increases). However, these projections may change depending on the actual amount of antigen needed per dose.

Developing an effective vaccine may require having the pandemic strain in hand, which will mean that a vaccine cannot be produced until the onset of the pandemic. Once a virus is identified, it will take at least 19 weeks to develop the appropriate reagents for an inactivated pandemic vaccine (see References: WHO: A description of the process of seasonal and H5N1 influenza vaccine virus selection and development).

Given these caveats, a summary of currently available key information on vaccine studies involving humans is presented in the table below. A number of studies involving animal models also have been published recently; work in this area is ongoing.

Recent Trials in Humans of Prepandemic Influenza Vaccine
Source (see References)	Vaccine Dose^a	Number of Subjects	Key Findings	Comments
Treanor 2006	Subvirion influenza vaccine with 90, 45, 15,or 7.5 mcg of HA or placebo	451 volunteers (age 18 – 64 years)	54% of subjects who received the 90 mcg formulation had neutralization antibody titers ≥1:40 (a subsequent report indicated that 45% responded rather than 54%).	—The vaccine was produced by Sanofi Pastuer and the vaccine was derived from A/Vietnam/1203/2004 (H5N1). —In April 2007, the FDA approved this vaccine and the US federal government has stockpiled at least 13 million doses (see References: FDA 2007).
Bresson 2006	Inactivated split influenza vaccine with 7.5, 15, or 30 mcg of HA with or without aluminium hydroxide adjuvant	300 volunteers (age 18 – 40 years)	67% of subjects who received the adjuvanted 30 mcg formulation had HAI titers ≥1:40.	The vaccine was produced by Sanofi Pastuer and the vaccine was derived from A/Vietnam/1194/2004 (H5N1).
Lin 2006	Inactivated whole-virion influenza vaccine with 1.25, 2.5, 5, or 10 mcg of HA with aluminium hydroxide adjuvant or placebo	120 volunteers (age 18 – 60 years)	78% of subjects who received the 10 mcg formulation had HAI titers ≥1:40.	The vaccine was produced by Sinovac Biotech Co, Ltd (Beijing, China) and the vaccine was derived from A/Vietnam/1194/2004 (H5N1).
GlaxoSmithKline 2008	4 doses, with 3.8 mcg of HA as the lowest dose; adjuvant included (adjuvant not specified)	400 volunteers (age 18 – 60 years)	80% of subjects who received the 3.8 mcg formulation had HAI titers of ≥1:40.	This vaccine was approved by the European Union in May 2008 (see References: GlaxoSmithKline 2008).
Bernstein 2008	Inactivated subvirion vaccine with 45, 30, or 15 mcg per dose; or vaccine with 15 or 7.5 mcg per dose with MF59 adjuvant; or vaccine with 30, 15, or 7.5 mcg per dose with aluminum hydroxide adjuvant	394 volunteers (age 18 – 64 years)	63% of subjects who received the 15 mcg with MF59 formulation had HAI titers ≥1:40.	The vaccine was produced by Chiron Vaccines (now part of Novartis) and the vaccine was derived from A/Vietnam/1203/2004(H5N1).
Erhlich 2008	Whole-virus cell-based vaccine with 3.75, 7.5, 15, or30 mcg of HA with alum adjuvant or 7.5or 15 mcg of HA without adjuvant	275 volunteers (age 18 – 45 years)	76.2% and 70.7% of subjects who received either 7.5 mcg or 15 mcg without adjuvant, respectively, had virus neutralization titers of ≥1:20.	—The vaccine was produced by Baxter and the vaccine was derived from A/Vietnam/1203/2004). —The vaccine also showed evidence of cross-protection against other H5N1 strains.
Nolan 2008	Inactivated, split-virus H5N1 vaccine with 7.5, 15, 30, or 45 mcg HA with or without aluminum ion adjuvant	400 volunteers (age 18 – 64 years)	Between 58% and 59% of participants who received the 30 mcg or 45 mcg adjuvanted formulations had HAI titers ≥1:32 and 73% had MN titers ≥1:20.	—The vaccine was produced by by CSL Limited (Parkville, VIC, Australia) and the vaccine was derived from A/Vietnam/1194/2004 (H5N1). —In June 2008, Australian authorities approved this vaccine (see References: CSL Limited 2008).
Levie 2008	Inactivated, split-virus vaccine of 1.9, 3.8, 7.5, or 15 mcg of HA with oil-in-water emulsion adjuvant or 7.5 mcg without adjuvant	251 volunteers (age 18 – 40 years)	—81% to 89% of subjects who received 3.8, 7.5, or 15 mcg formulations had HI titers ≥1:32. —72% of subjects who received the 1.9 mcg formulation had HAI titers ≥1:32. —34% of subjects who received the 7.5 mcg formulation without adjuvant responded.	The vaccine was produced by Sanofi Pasteur and the vaccine was derived from A/Vietnam/1194/2004 (H5N1).
Abbreviations: HA: hemagglutinin antigen ; HAI: hemagglutination inhibition; MN: virus microneutralization. ^aAll vaccines were administered in a 2-dose series.

Other issues that have been raised regarding antibody protection against H5N1 influenza include:

Researchers have suggested that development and use of an H5N1 vaccine for immunologic priming during the interpandemic period may offset the need for two doses of vaccine once a pandemic begins, even if the strain used in the priming vaccine is somewhat different from an emergent pandemic strain (see References: Ehrlich 2008, Goji 2008, Haque 2007, Monto 2006, Stephenson 2005).
A universal vaccine that would be effective against all types of influenza, including emerging pandemic strains, is being developed by the British company Acambis and is being researched by others as well. Such a vaccine would not have to be reengineered each year. One possible target for a universal vaccine is the relatively conserved M2 homotetramer (see References: Acambis 2005, Haque 2007).
Seasonal influenza vaccine, which includes protection against H1N1 and H3N2, may boost cross-subtype immunity against H5N1 influenza A viruses (see References: Giola 2008). This hypothesis is based on evidence suggesting that the N1 antigen may be a target for both cellular and humoral cross-type immunity.

The WHO is working with vaccine manufacturers to develop an international stockpile of H5N1 vaccine (see References: WHO 2007: WHO and manufacturers move ahead with plans for H5N1 influenza global vaccine stockpile). In June 2007, GlaxoSmithKline pledged to give 50 million doses of H5N1 vaccine to the WHO, and in June 2008, Sanofi Pasteur pledged an additional 60 million doses of vaccine over 3 years (see References: Sanofi Pasteur 2008).

WHO and CDC Travel Recommendations

WHO Recommendations

In November 2005, the WHO updated travel recommendations to be consistent with phase 3 of the WHO six-phase Pandemic Alert (see References: WHO: Advice to international travelers 2005):

Advice to countries

The WHO does not recommend travel restrictions to areas experiencing outbreaks of highly pathogenic H5N1 avian influenza in birds, including countries that have reported associated cases of human infection.
The WHO does not, at present, recommend the routine screening of travelers coming from affected areas. Local authorities may, however, usefully provide information to travelers on risks, risk avoidance, symptoms, and when and where to report should symptoms develop.

Advice to travelers

The WHO advises travelers to avoid contact with high-risk environments in affected countries.

Travelers to areas affected by avian influenza in birds are not considered to be at elevated risk of infection unless direct and unprotected exposure to infected birds (including feathers, feces, and undercooked meat and egg products) occurs.
The WHO continues to recommend that travelers to affected areas avoid contact with live animal markets and poultry farms as well as any free-ranging or caged poultry. Large amounts of the virus are known to be excreted in the droppings of infected birds. Populations in affected countries are advised to avoid contact with dead migratory birds or wild birds showing signs of disease.
Direct contact with infected poultry or with surfaces and objects contaminated by their droppings is considered the main route of human infection. Exposure risk is considered highest during slaughter, defeathering, butchering, and preparation of poultry for cooking. There is no evidence that properly cooked poultry or poultry products can be a source of infection.
Travelers should contact their local health providers or national health authorities for supplementary information.

CDC Recommendations

The CDC also has issued a set of recommendations for travelers to areas affected by H5N1 influenza; these are updated regularly (see References: CDC: Human infection with avian influenza A (H5N1) virus: advice for travelers). In addition to recommendations for travelers, the CDC has published guidelines for personnel who clean, maintain, or remove baggage/packages from commercial and cargo airlines about appropriate precautions related to H5N1 influenza. Recommendations are based on standard infection control practices used in healthcare settings and on available information about the virus that causes H5N1 avian influenza (see References: CDC: Interim guidance for airline cleaning crew, maintenance crew, and baggage/package and cargo handlers for airlines returning from areas affected by avian influenza A [H5N1]).

The CDC recommends the following for travelers:

Before international travel to an area affected by H5N1 avian influenza

Visit the CDC's Travelers' Health Web site (see References) to educate yourself and others who may be traveling with you about any disease risks and CDC health recommendations for international travel in areas you plan to visit.
Be sure you are up-to-date with all your routine vaccinations and see your doctor or healthcare provider, ideally 4 to 6 weeks before travel, to get any additional vaccination medications or information you may need.
Assemble a travel health kit containing basic first aid and medical supplies. Be sure to include a thermometer and alcohol-based hand gel for hand hygiene (see References: CDC: Travelers' health kit).
Identify in-country healthcare resources in advance of your trip.
Check your health insurance plan or get additional insurance that covers medical evacuation in case you become sick. Information about medical evacuation services is provided on the US Department of State website (see References: US Department of State).

During travel to an affected area

Avoid all direct contact with poultry, including touching well-appearing, sick, or dead chickens and ducks. Avoid places such as poultry farms and bird markets where live poultry are raised or kept, and avoid handling surfaces contaminated with poultry feces or secretions.
As with other infectious illnesses, one of the most important preventive practices is careful and frequent hand washing. Cleaning your hands often with soap and water removes potentially infectious material from your skin and helps prevent disease transmission. Waterless alcohol-based hand gels may be used when soap is not available and hands are not visibly soiled.
Influenza viruses are destroyed by heat; therefore, as a precaution, all foods from poultry, including eggs and poultry blood, should be thoroughly cooked.
If you become sick with symptoms such as a fever accompanied by a cough, sore throat, or difficulty breathing, or if you develop any illness that requires prompt medical attention, a US consular officer can assist you in locating medical services and informing your family or friends. The book Health Information for International Travel provides information about what to do if you become ill while abroad (see References: CDC: Seeking health care abroad). You should defer further travel until you are free of symptoms, unless your travel is health-related. Inform your healthcare provider of any possible exposures to avian influenza.

After your return

Monitor your health for 10 days.
If you become ill with a fever plus cough, sore throat, or trouble breathing during this 10-day period, consult a healthcare provider. Important: Before you visit a healthcare setting, tell the provider the following: (1) your symptoms, (2) where you traveled, and (3) if you have had direct contact with poultry or close contact with a severely ill person.
Do not travel while ill, unless you are seeking medical care. Limiting contact with others as much as possible can help prevent the spread of an infectious illness.

Use of Seasonal Flu Vaccine in Humans at Risk for H5N1 Infections

On January 30, 2004, the WHO released guidelines for the use of seasonal influenza vaccine among persons at risk for H5N1 influenza (see References: WHO: Guidelines for the use of seasonal influenza vaccine in humans at risk of H5N1 infection). The WHO recommends targeted use of seasonal influenza vaccine to reduce the potential for humans to be infected with H5N1 at the same time that they are harboring a human influenza strain. This will decrease the opportunity for genetic reassortment of the avian H5N1 strain with genes from a human (H1 or H3) strain and thereby reduce the likelihood that a novel pandemic strain will emerge from the current situation in Asia.

Groups recommended for vaccination include:

All persons expected to be in contact with poultry or poultry farms suspected or known to be affected with avian influenza (H5N1), especially:

Cullers involved in destruction of poultry
People living and working on poultry farms where H5N1 has been reported or is suspected or where culling takes place

Healthcare workers involved in the daily care of confirmed human cases of influenza H5N1
Healthcare workers in emergency care facilities in areas where there is confirmed occurrence of influenza H5N1 in birds (provided that sufficient supplies of vaccine are available)

Surveillance, Case Evaluation, and Contact Follow-up

Case Definitions

In August 2006, the WHO released case definitions for H5N1 influenza (see References: WHO 2006: Case definitions for human infections with influenza A [H5N1] virus). The case definitions apply to the current phase of pandemic alter (phase 3) and may change as new information about the disease or its epidemiology becomes available. The current case definitions are:

Person under investigation: A person whom public health authorities are investigating for possible H5N1 infection.
Suspected H5N1 case: A patient who has unexplained acute lower respiratory illness with a fever greater than 38°C (100.4°F) and cough, shortness of breath, breathing difficulty, and one or more of the following exposures 7 days before symptom onset:

Close contact (within 1 meter, eg, caring for, speaking with, or touching) with a person who is a suspected, probable, or confirmed H5N1 case
Exposure to (eg, handling, slaughtering, defeathering, butchering, or preparing for consumption) poultry or wild birds, their remains, or their feces where H5N1 infections in animals or humans have been suspected or confirmed in the last month
Consumption of raw or undercooked poultry where H5N1 infections in animals or humans have been suspected or confirmed in the last month
Close contact with a confirmed H5N1-infected animal other than poultry or wild birds (eg, cat or pig)
Handling human or animal samples suspected of containing the H5N1 virus in a laboratory or other setting

Probable H5N1 case (notify WHO):

Definition 1: A person who meets the criteria for a suspected case and has either (1) evidence of acute pneumonia on a chest radiograph plus respiratory failure (hypoxemia, severe tachypnea) or (2) laboratory confirmation of influenza A but insufficient laboratory evidence for H5N1.
Definition 2: A person dying of an unexplained respiratory illness who is epidemiologically linked by time, place, and exposure to a probable or confirmed H5N1 case.

Confirmed H5N1 case (notify WHO): A patient who meets the criteria for a suspected or probable case and has had one of the following test results from a national, regional, or international influenza laboratory whose H5N1 test results are accepted by the WHO:

Isolation of an H5N1 virus
Positive H5 PCR results from tests using two different PCR targets (eg, primers specific for influenza A and H5)
A fourfold or greater rise in neutralization antibody titer for H5N1 based on testing of an acute serum specimen (collected 7 days or less after symptom onset) and a convalescent serum specimen; the convalescent neutralizing antibody titer must be 1:80 or higher
A microneutralization antibody titer for H5N1 of 1:80 or greater in a single serum specimen collected at day 14 or later after symptom onset and a positive result using a different serologic assay, such as a horse red blood cell HAI titer of 1:160 or more or an H5-specific Western blot positive result

Case Evaluation

Recommendations from the CDC for evaluation of suspected cases were published in February 2004 (see References: CDC: Outbreaks of avian influenza A (H5N1) in Asia and interim recommendations for evaluation and reporting of suspected cases, United States, 2004).

According to current CDC recommendations, testing for H5N1 of patients hospitalized in the United States is indicated for patients who have both of the following conditions:

Radiographically confirmed pneumonia, ARDS, or other severe respiratory illness for which an alternative diagnosis has not been established
A history of travel within 10 days of symptom onset to a country with documented H5N1 avian influenza infection in poultry or humans

Testing for influenza A (H5N1) also should be considered for patients with all of the following:

Documented temperature of over 100.4ºF (38ºC)
Cough, sore throat, or shortness of breath
History of contact with poultry or domestic birds (eg, visited a poultry farm, a household raising poultry, or a bird market) or a known or suspected patient with influenza A [H5N1] in an H5N1-affected country within 10 days of symptom onset)

The CDC recommends the following for laboratory testing of clinical specimens from patients with suspected H5N1 influenza A:

Virus isolation studies on respiratory specimens should not be performed unless all BSL-3 laboratory conditions are met.
Clinical specimens can be tested by PCR assays by using standard BSL-2 work practices in a class II biological safety cabinet.
Commercially available antigen-detection tests can be used under BSL-2 levels to test for influenza.
Specimens from suspected cases should be sent to the CDC if they test positive for influenza A either by PCR or antigen-detection testing, or if PCR assays for influenza are not locally available.

Follow-up of Case Contacts

In November 2008, the CDC released a guidance document for follow-up of close contacts of persons with suspected, probable, or confirmed H5N1 infection (see References: CDC 2008: Interim guidance for follow-up of contacts of persons with suspected infection with highly pathogenic avian influenza A (H5N1) virus in the United States). Key points from that guidance are outlined below.

Types of Contacts

Close contacts are defined by the CDC as "persons who were within about 6 feet of a suspected, probable, or confirmed H5N1 case while the case was symptomatic." Potential close contacts include the following identifiable persons:

Household and family contacts
Healthcare personnel
Laboratory workers
Other persons who were known to be within about 6 feet of the suspected case

Follow-up Period

According to the CDC, all identified close contacts should be monitored daily for 7 days after the last known exposure to an ill person suspected to be infected with H5N1. The following should be assessed each day during this period:

Measured temperature
Presence of any illness symptoms

Any close contacts who have a temperature of 38°C or higher or any illness symptoms should be referred for prompt medical evaluation and possible testing for H5N1. Monitoring of close contacts may be discontinued if H5N1 infection is ruled out for the suspect case (ie, when laboratory testing by RT-PCR of appropriately collected respiratory specimens at a state health department laboratory or at the CDC has excluded infection with H5N1 virus) or upon absence of any illness symptoms among contacts during the 7-day follow-up.

Antiviral Chemoprophylaxis for Close Contacts

Daily antiviral chemoprophylaxis with an NI medication (oseltamivir or zanamivir) should be provided to close contacts for 7 days after the last known exposure. Oseltamivir is the recommended antiviral medication for chemoprophylaxis of H5N1; zanamivir is considered an alternative. Chemoprophylaxis should be provided to close contacts in the following order of priority:

Highest-risk exposure groups
Household or close family member contacts
Moderate-risk exposure groups
Healthcare personnel in close contact with a suspected or confirmed H5N1 patient
Laboratory workers who had unprotected exposure to H5N1 virus–containing samples
Social contacts
Low-risk exposure groups
Healthcare personnel not in close contact with a suspected or confirmed H5N1 case or who used appropriate personal protective equipment (PPE) during exposure to a suspected or confirmed case

When to Discontinue Antiviral Chemoprophylaxis of Close Contacts

Chemoprophylaxis of close contacts may be discontinued if H5N1 infection is ruled out for the suspect case or upon absence of any illness symptoms among contacts during the 7-day follow-up.

Influenza Pandemic Considerations

Influenza pandemics occurring during the 20th century apparently all arose from the Eurasian avian lineage of viruses.

Influenza pandemic strains can emerge from the avian lineage either through the process of genetic reassortment between human and animal strains (referred to as "antigenic shift") or through gradual adaptation to humans.
Current research indicates that the pandemic strains of 1957 and 1968 occurred through genetic reassortment. The 1918 pandemic strain, however, apparently did not originate through a reassortment event; rather, it is likely that an avian strain initially infected humans and then adapted gradually to the human population over time to become a pandemic strain (see References: Taubenberger 2005).
At this point, there is no evidence of genetic reassortment between avian H5N1 viruses and humans; however, if H5N1 continues to circulate widely among poultry, the potential for emergence of a reassorted pandemic strain remains a concern. For example, H5N1 viruses have been found in pigs in southern China and in Indonesia (see Oct 10, 2006, CIDRAP News story and see References: Cyranoski 2004, Cyranoski 2005), and human H3N2 influenza viruses are endemic in pigs in China. Thus, the conditions exist for exchange of genetic material between the different viruses in the pig host (see References: Li 2004; WHO: Avian influenza: update: implications of H5N1 infections in pigs in China).
The potential for gradual adaptation of H5N1 to humans over time with evolution into a pandemic strain is also a possibility. As more humans become infected with the virus, this issue becomes a greater concern. Influenza viruses that are adapted to humans have HAs (ie, virus surface proteins) that bind specifically to the long alpha 2-6 glycan receptors of epithelial cells in the human upper respiratory tract (see References: Chandrasekaran A 2008). Current strains of H5N1 do not have binding affinity for these receptors.
A study published in July 2006 attempted to determine whether a synthetic influenza virus made by combining an H5N1 avian flu virus and a human flu virus would be more contagious than the natural H5N1 virus. Findings showed that it was not more easily spread in a ferret model (see References: Maines 2006). However, the H5N1 virus used was a strain collected in 1997.

According to the WHO, at this time the pandemic alert level for H5N1 influenza is at phase 3: A new viral subtype is causing disease in humans but is not yet spreading efficiently and sustainably (see References: WHO: Current WHO phase of pandemic alert).

Of the avian influenza subtypes, H5N1 is of greatest concern for the following reasons (see References: WHO: Avian influenza: assessing the pandemic threat):

The subtype mutates rapidly.
It causes severe disease in humans, with a high case-fatality rate.
The virus has spread rapidly throughout poultry flocks in Asia, increasing the likelihood of infecting humans or pigs, where genetic reassortment with human strains could occur, leading to a new pandemic strain.
The virus is continuing to spread to other areas of the world, including Central Asia, Europe, the Middle East, and Africa.
Recent genetic sequencing performed on viral isolates from Turkey demonstrates that the strains contain two mutations that may make the virus better adapted to humans (see References: Butler 2006). These mutations could enhance transmission from birds to humans and between humans.

More information can be found in the Pandemic Influenza section of this Web site.

Infection Control Recommendations

Infection Control Guidelines for H5N1 Avian Influenza

In May 2004, the CDC and WHO issued infection control guidelines for prevention of transmission of H5N1 influenza in healthcare settings; the WHO revised its guidelines in April 2006 (see References: CDC: Interim recommendations for infection control in health-care facilities caring for patients with known or suspected avian influenza; WHO 2006: Avian influenza, including influenza A (H5N1), in humans: WHO interim infection control guideline for health care facilities). Summaries from the CDC and WHO of recommended isolation precautions are outlined in the table below. Both agencies recommend that Airborne Precautions be implemented, if possible. A recent review of the potential for airborne transmission of influenza suggests that use of Airborne Precautions may be appropriate for protecting healthcare workers who care for patients with influenza (see References: Tellier 2006).

Isolation Precautions for Patients With H5N1 Avian Influenza

CDC Recommendations

Standard Precautions
Pay careful attention to hand hygiene before and after all patient contact or contact with items potentially contaminated with respiratory secretions.

Contact Precautions
—Use gloves and gown during all patient contact.
—Use dedicated equipment such as stethoscopes, disposable blood pressure cuffs, and disposable thermometers.

Eye protection (ie, goggles or face shields)
Wear when within 3 ft of patient.

Airborne Precautions
—Place patient in an AIR. Such rooms should have monitored negative air pressure in relation to corridor, with 6 to 12 ACH, and should exhaust air directly outside or have recirculated air filtered by a HEPA filter. If an AIR is unavailable, contact the healthcare facility engineer to assist or use portable HEPA filters to augment ACH.
—Use a fit-tested respirator, at least as protective as a NIOSH-approved N95 filtering facepiece (ie, disposable) respirator, when entering room.

WHO Recommendations

Standard Precautions
Droplet Precautions
Contact Precautions
Airborne Precautions (including use of high-efficiency masks and negative-pressure rooms when available)

Abbreviations: ACH, air changes per hour; AIR, airborne isolation room; HEPA, high-efficiency particulate air; NIOSH, National Institute of Occupational Safety and Health.

A recent study demonstrated that hand hygiene using soap and water or an alcohol-based hand rub is highly effective in reducing influenza A virus on human hands; therefore, good hand hygiene is an important component of influenza A infection control (see References: Grayson 2008).

In June 2006, researchers in Thailand reported isolating H5N1 virus from the blood of a 5-year-old patient with avian influenza (see References: Chutinimitkul 2006). This finding supports the importance of using Standard Precautions (which include wearing gloves when handling blood or body fluids from any patient).

Guidance to Protect Workers From Avian Influenza Viruses

In November 2006, the US Department of Labor released a revised Occupational Safety and Health Administration (OSHA) guidance to help employers protect their workers from job-related exposure to H5N1 avian influenza (see References: OSHA 2006). The document provides guidance for the following sets of workers:

Poultry employees
Animal handlers other than poultry employees
Laboratory employees
Healthcare workers who treat patients with known or suspected avian influenza
Food handlers
Airport personnel exposed to passengers suspected of being infected with avian influenza
Travelers on temporary work assignment abroad
US employees stationed abroad
Other employee groups that may be at risk

In February 2008, the National Institute of Occupational Safety and Health (NIOSH) released an alert on steps for protecting poultry workers against avian influenza (see References: CDC/NIOSH 2008).

Also in February 2008, the WHO updated its guidance on protection of workers and other individuals at risk of exposure (see References: WHO: Protection of individuals with high poultry contact in areas affected by avian influenza H5N1: Consolidation of pre-existing guidance). High-contact activities include handling, collecting, transporting, culling, and disposal of birds, and cleaning/disinfection of contaminated areas. According to the WHO, all individuals involved in high-risk activities should perform the following:

Be registered with the local animal health authority (or by the public health authority in collaboration with the animal health authority).
Wear appropriate PPE, including protective clothing, heavy gloves and boots, goggles, and masks, and receive adequate training on putting on, taking off, and hygienic disposal/disinfection of PPE.
Maintain diligence in personal hygiene, including frequent hand washing.
Receive adequate instruction on disinfection/disposal of potentially contaminated personal clothing and other personal articles.
Be monitored twice daily for fever (>38°C) and influenza-like illness for 7 days after the last day of contact with poultry/contaminated environments. Any person experiencing fever or influenza-like illness should immediately report to health authorities for diagnostic testing and appropriate treatment.

A recent report suggests that strict compliance with PPE (including protective coveralls, protective footwear, disposable gloves, face-fitted mask or other mask, and protective goggles) can reduce hazardous exposure to humans and illness during poultry outbreaks (see References: Morgan 2009).

Food Safety Issues

Several human cases of H5N1 apparently have resulted from consumption of improperly cooked or raw poultry products (see References: WHO Writing Committee of the Second World Health Organization Consultation on Clinical Aspects of Human Infection with Avian Influenza A [H5N1] Virus 2008).

In September 2007, German officials reported that they had found H5N1 virus in 18 frozen ducks from a batch sample at a poultry company slaughterhouse, but no cases were traced to consumption of the contaminated ducks (see Sep 10, 2007, CIDRAP News story).

Concerns also have been raised about the potential for contamination of water sources with H5N1 virus. A recent study found that free chlorine concentrations typically used in drinking water treatment are sufficient to inactivate the virus by >3 orders of magnitude (see References: Rice 2007).

In November 2005, the WHO issued a statement on food safety issues (see References: WHO/INFOSAN 2004). This statement includes the following information:

The H5N1 avian influenza virus is not transmitted to humans through properly cooked food. The virus is sensitive to heat, and normal temperatures used for cooking (so that food reaches 70ºC in all parts) will kill the virus.
To date, no evidence indicates that any person has become infected with the H5N1 virus following the consumption of properly cooked poultry or poultry products, even when the food item contained the virus prior to cooking. However, several cases have involved consumption of raw poultry ingredients, such as uncooked duck blood.
Poultry and poultry products from areas free of the disease can be prepared and consumed as usual, with no fear of acquiring H5N1 infection.
Most strains of avian influenza virus are found only in the respiratory and gastrointestinal tracts of infected birds, not in meat. However, available studies indicate that highly pathogenic viruses, including the H5N1 virus, spread to virtually all parts of an infected bird, including meat. For this reason, proper handling of poultry and poultry products during food preparation and proper cooking are extremely important in areas experiencing outbreaks of H5N1 avian influenza in poultry.
Consumers in areas with outbreaks need to be aware of the risks of cross-contamination between raw poultry and other foods that will not be cooked prior to their consumption. Juices from raw poultry or poultry products should never be allowed during food preparation to touch or mix with items eaten raw. When handling raw poultry or raw poultry products, persons involved in food preparation should wash their hands thoroughly and clean and disinfect surfaces in contact with the poultry products. Soap and hot water are sufficient for this purpose.
In countries with outbreaks, thorough cooking is imperative. Consumers need to be sure that all parts of the poultry are fully cooked (no "pink" parts) and that eggs, too, are properly cooked (no "runny" yolks).
The H5N1 virus can survive for at least 1 month at low temperatures. For this reason, common food preservation measures, such as freezing and refrigeration, will not substantially reduce the concentration of virus in contaminated meat or kill the virus. In countries with outbreaks, poultry stored under refrigeration or frozen should be handled and prepared with the same precautions as fresh products.
In countries with outbreaks, eggs may contain virus both on the outside (shell) and inside (white and yolk). Eggs from areas with outbreaks should not be consumed raw or partially cooked. Raw eggs should not be used in foods that will not be treated by heat high enough to kill the virus (70ºC).
To date, a large number of human infections with the H5N1 virus have been linked to the home slaughter and subsequent handling of diseased or dead birds prior to cooking. These practices represent the highest risk of human infection and are the most important to avoid. Proper handling and cooking of poultry and poultry products can further lower the risk of human infections.

In November 2008, the USDA released a draft risk assessment tool for contracting HPAI from eating poultry products, shell eggs, and egg products (see References: USDA 2008). The tool is intended to be used by risk managers for decision-making when determining possible interventions following detection of HPAI in a poultry flock in the United States. The risk assessment also can be used to target risk communication messages, identify and prioritize research needs, and provide a framework for coordinating prevention and control efforts with stakeholders.

References

Acambis. Acambis enters flu vaccine arena with launch of flu vaccine development programme. Aug 4, 2005 [Full text]

Alexander DJ. Should we change the definition of avian influenza for eradication purposes? Avian Dis 2003;47(3 Suppl):976-81 [Abstract]

Bean B, Moore BM, Sterner B, et al. Survival of influenza viruses on environmental surfaces. J Infect Dis 1982 Jul;146(1):47-51 [Abstract]

Bernstein DI, Edwards KM, Dekker CL, et al. Effects of adjuvants on the safety and immunogenicity of an avian influenza H5N1 vaccine in adults. J Infect Dis 2008 Mar 1;197(suppl 2):667-75 [Abstract]

Bresson J-L, Perronne C, Launay O, et al. Safety and immunogenicity of an inactivated split-virion influenza A/Vietnam/1194/2004 (H5N1) vaccine: phase 1 randomised trial. Lancet 2006 May 20;367(9523):1657-64 [Abstract]

Butler D. Alarms rising over bird flu mutations. Nature 2006 Jan 19;439:248-9 [Full text]

Call SA, Vollenweider MA, Hornung CA, et al. Does this patient have influenza? JAMA 2005 Feb 23;293(8):987-97 [Abstract]

Capua I, Alexander DJ. Avian influenza: recent developments. Avian Pathol. 2004 Aug;33(4):393-404 [Abstract]

CDC. Avian influenza A viruses [Web page]

CDC. Avian influenza A virus infections of humans. [Full text]

CDC. Human infection with avian influenza A (H5N1) virus: advice for travelers. Released Sep 23, 2005; updated frequently [Full text]

CDC. Influenza activity: United States and worldwide, 2003-04 season, and composition for the 2004-05 influenza vaccine. MMWR 2004 Jul 2;53(25):547-52 [Full text]

CDC. Information about pandemic influenza viruses [Web page]

CDC. Interim guidance for airline cleaning crew, maintenance crew, and baggage/package and cargo handlers for airlines returning from areas affected by avian influenza A (H5N1). Released Dec 9, 2005; updated regularly [Full text]

CDC. Interim guidance for follow-up of contacts of persons with suspected infection with highly pathogenic avian influenza A (H5N1) virus in the United States. Nov 7, 2008 [Full text]

CDC. Interim guidance for influenza diagnostic testing during the 2004-05 influenza season. Nov 2004 [Full text]

CDC. Interim guidance for states conducting avian mortality surveillance for West Nile virus (WNV) and/or highly pathogenic H5N1 avian influenza virus. Aug 23, 2006 [Full text]

CDC. Interim recommendations for infection control in health-care facilities caring for patients with known or suspected avian influenza [Full text]

CDC. National Respiratory and Enteric Virus Surveillance System (NREVSS) [Web page]

CDC. New laboratory assay for diagnostic testing of avian influenza A0H5 (Asian lineage). MMWR 2006 Feb 3;55(Early release):1 [Full text]

CDC. Outbreaks of avian influenza A (H5N1) in Asia and interim recommendations for evaluation and reporting of suspected cases: United States, 2004. MMWR 2004 Feb 13;53(5):97-100 [Full text]

CDC. Travelers' health [Web page]

CDC. Seeking health care abroad. In: Health information for international travel: the 'yellow book' [Full text]

CDC. Travelers' health kit. In: Health information for international travel: the 'yellow book' [Full text]

CDC. Update on influenza A(H5N1) and SARS: Interim recommendations for enhanced U.S. surveillance, testing, and infection controls [Full text]

CDC/NIOSH. Protecting poultry workers from avian influenza (bird flu). February 2008 [Full text]

Chan M CW, Cheung CY, Chui WH, et al. Proinflammatory cytokine responses induced by influenza A (H5N1) viruses in primary human alveolar and bronchial epithelial cells. Respir Res 2005 Nov 11;6(1) [Abstract]

Chan PK. Outbreak of avian influenza A (H5N1) virus infection in Hong Kong in 1997. Clin Infect Dis 2002 May 1;34(suppl 2):S58-S64 [Full text]

Chang HK, Park JH, Song MS, et al. Development of multiplex RT-PCR assays for rapid detection and subtyping of influenza type A viruses from clinical specimens. J Microbiol Biotechnol 2008 Jun;18(6):1164-9

Chen Y, Qin K, Wu WL, et al. Broad cross-protection against H5N1 avian influenza virus infection by means of monoclonal antibodies that map to conserved viral epitopes. J Infect Dis 2009 Jan 1;199(1):49-58 [Abstract]

Cheung C-L, Rayner JM, Smith GJD, et al. Distribution of amantadine-resistant H5N1 avian influenza variants in Asia. J Infect Dis 2006 Jun 15;193(12):1626-9 [Full text]

Chotpitayasunondh T, Ungchusak K, Hanshaoworakul W, et al. Human disease from influenza A (H5N1), Thailand, 2004. Emerg Infect Dis 2005 Feb;11(2) [Full text]

Chutinimitkul S, Bhattarakosol P, Srisuratanon S, et al. H5N1 influenza A virus and infected human plasma. (Letter) Emerg Infect Dis 2006 Jun;12(6):1041-2 [Full text]

Coiras MT, Perez-Brena P, Garcia ML, et al. Simultaneous detection of influenza A, B, and C viruses, respiratory syncytial virus, and adenoviruses in clinical samples by multiplex reverse transcription nested-PCR assay. J Med Virol 2003 Jan;69(1):132-44 [Abstract]

CSL Limited. CSL receives TGA approval to register Panvax, avian flu vaccine. Jun 17, 2008, press release [Full text]

Daidoji T, Koma T, Du A, et al. H5N1 avian influenza virus induces apoptotic cell death in mammalian airway epithelial cells. J Virol 2008 Nov;82(22):11294-307 [Abstract]

De Jong MD, Simmons CP, Thanh TT, et al. Fatal outcome of human influenza A (H5N1) is associated with high viral load and hypercytokinemia. (Letter) Nat Med 2006 (published online Sep 10) [Abstract]

De Jong MD, Thanh TT, Khanh TH, et al. Oseltamivir resistance during treatment of influenza A (H5N1) infection. N Engl J Med 2005 Dec 22;353:2667-72 [Full text]

De Jong MD, Van Cam B, Tu Qui P, et al. Fatal avian influenza A (H5N1) in a child presenting with diarrhea followed by coma. N Engl J Med 2005;352:686-91 [Abstract]

Democratis J, Pareek M, Stephenson I. Use of neuraminidase inhibitors to combat pandemic influenza. J Antimicrob Chemother 2006 (published online Sep 6) [Abstract]

Dinh PN, Long HT, Tien NTK, et al. Risk factors for human infection with avian influenza A H5N1, Vietnam, 2004. Emerg Infect Dis 2006 Dec [cited Oct 27, 2006] [Full text]

Ehrlich HJ, Muller M, Oh HML, et al. A clinical trial of a whole-virus H5N1 vaccine derived from cell culture. N Engl J Med 2008 Jun 12;358(24):2573-84 [Full text]

FAO. H5N1 HPAI Global Overview: May 2008. FAO AIDEnews 2008 Jun 16, Issue No. 54 [Full text]

FDA. FDA approves first U.S. vaccine for humans against the avian influenza virus H5N1. Apr 17, 2007, press release [Full text]

FDA. Cautions in using rapid tests for detecting influenza A viruses [Web site]

Fielding R, Lam WTW, Ho EYY, et al. Avian influenza risk perception, Hong Kong. Emerg Infect Dis 2005 May;11(5):677-82 [Full text]

Fouchier RAM, Schneeberger PM, Rozendaal FW, et al. Avian influenza A virus (H7N7) associated with human conjunctivitis and a fatal case of acute respiratory distress syndrome. Proc Natl Acad Sci 2004 Feb 3;101(5):1356-61 [Abstract]

Fouchier RAM, Munster V, Wallensten A, et al. Characterization of a novel influenza A virus hemagglutinin subtype (H16) obtained from black-headed gulls J Virol 2005 Mar;79(5):2814-22 [Abstract]

GlaxoSmithKline. First pre-pandemic vaccine approved to help protect against pandemic influenza. May 19, 2008 [Full text]

GlaxoSmithKline. GSK reports significant advance in H5N1 pandemic flu vaccine programme. July 26, 2006, press release [Full text]

Gill JS, Webby R, Gilchrist MJR, et al. Avian influenza among waterfowl hunters and wildlife professionals. Emerg Infect Dis 2006 Aug;12(8):1284-6 [Full text]

Gilsdorf A, Boxall N, Gasimov B, et al. Two clusters of human infection with influenza A/H5N1 virus in the Republic of Azerbaijan, February-March 2006. Eurosurveillance Monthly 2006 May;11(5) [Full text]

Gioia C, Castilletti C, Tempestilli M, et al. Cross-subtype immunity against avian influenza in persons recently vaccinated for influenza. Emerg Infect Dis 2008 Jan;14(1):121-8 [Full text]

Goji NA, Nolan C, Hill H, et al. Immune responses of healthy subjects to a single dose of intramuscular inactivated influenza A/Vietnam/1203/2004 (H5N1) vaccine after priming with an antigenic variant. J Infect Dis 2008 Sep 1;198(5):635-41 [Abstract]

Grayson ML, Melvani S, Druce J, et al. Efficacy of soap and water and alcohol-based hand-rub preparations against live H1N1 influenza virus on the hands of human volunteers. Clin Infect Dis 2008 (published online Dec 30) [Abstract]

Gubareva LV, Matrosovich MN, Brenner MK, et al. Evidence of zanamivir resistance in an immunocompromised child infected with influenza B virus. J Infect Dis 1998 Nov;178(5):1257-62 [Abstract]

Haque A, Hober D, Kasper LH. Confronting potential influenza A (H5N1) pandemic with better vaccines. Emerg Infect Dis 2007 Oct;13(10):1512-8 [Full text]

Hayden F. Developing new antiviral agents for influenza treatment: what does the future hold? Clin Infect Dis 2009 Jan 1;48(suppl 1):S3-13 [Abstract]

Hayden FG, Cote KM, Douglas RG Jr. Plaque inhibition assay for drug susceptibility testing of influenza viruses. Antimicrob Agents Chemother 1980 May;17(5):865-70 [Full text]

Hayden FG, Palese P. Influenza virus. In: Richman DD, Whitley RJ, Hayden FG, eds. Clinical virology. Ed. 2. Washington, DC: ASM Press, 2002

Health Canada. Avian influenza: British Columbia (update). Infectious Diseases News Briefs. Apr 8, 2004 [Full text]

Herrmann B, Larsson C, Zweygberg BW. Simultaneous detection and typing of influenza viruses A and B by a nested reverse transcription-PCR: comparison to virus isolation and antigen detection by immunofluorescence and optical immunoassay (FLU OIA). J Clin Microbiol 2001 Jan;39(1):134-8 [Full text]

HHS. Biosafety in Microbiological and Biomedical Laboratories (BMBL) 5th Edition. Feb 2007 [Web Page]

HHS. Considerations for antiviral drug stockpiling by employers in preparation for an influenza pandemic. Released Jun 2008 [Full text]

HHS. FDA clears new CDC test to detect human influenza. Sep 30, 2008, press release [Full text]

HHS. Guidance on antiviral drug use during an influenza pandemic. Dec 16, 2008 [Full text]

HHS. HHS awards contracts totaling more than $1 billion to develop cell-based influenza vaccine. May 4, 2006 [Press release]

HHS. Pandemic influenza plan. Released Nov 2, 2005 [Links to overview and full text]

HHS. Q&A: What age groups are most likely to be affected during an influenza pandemic? [Web page]

Hinjoy S, Puthavathana P, Laosiritaworn H, et al. Low frequency of infection with avian influenza virus (H5N1) among poultry farmers, Thailand 2004. Emerg Infect Dis 2008 Mar;14(3):499-501 [Full text]

HSC (Homeland Security Council). National strategy for pandemic influenza: implementation plan. May 2006 [Full text]

Huai Y, Xiang N, Zhou L, et al. Incubation period for human cases of avian influenza A (H5N1) infection, China. (Letter) Emerg Infect Dis 2008 Nov;14(11)1819-20 [Full text]

Influenza Team, European Centre of Disease Prevention and Control. Avian influenza update: recent outbreaks of H5N1 in poultry worldwide. Euro Surveill 2007 Jan 25;12(1):E070125.2 [Full text]

Kandun IN, Wibisono H, Sedyaningsih ER, et al. Three Indonesian clusters of H5N1 virus infection in 2005. N Engl J Med 2006;355(21):2186-94 [Full text]

Kandun IN, Tresnaningsih E, Purba WH, et al. Factors associated with case fatality of human H5N1 virus infections in Indonesia: a case series. Lancet 2008 (published online Aug 15) [Abstract]

Kessler N, Ferraris O, Palmer K, Marsh W, Steel A. Use of the DNA flow-thru chip, a three-dimensional biochip, for typing and subtyping of influenza viruses. J Clin Microbiol. 2004 May;42(5):2173-85 [Full text]

Koopmans M, Wilbrink B, Conyn M, et al. Transmission of H7N7 avian influenza A virus to human beings during a large outbreak in commercial poultry farms in the Netherlands. Lancet 2004 Feb 21;363(9409):587-93 [Abstract]

Le QM, Kiso M, Someya K, et al. Avian flu: isolation of drug-resistant H5N1 virus. Nature 2005 Oct 20;437(7062):1108 [Abstract]

Lee CW, Suarez DL, Tumpey TM, et al. Characterization of highly pathogenic H5N1 avian influenza A viruses isolated from South Korea. J Virol 2005 Mar;79(6):3692-702 [Abstract]

Levie K, Leroux-Roels I, Hoppenbrouwers K, et al. An adjuvanted, low-dose, pandemic influenza A (H5N1) vaccine candidate is safe, immunogenic, and induces cross-reactive immune responses in healthy adults. J Infect Dis 2008 Sep 1;198(5):642-9 [Abstract]

Li KS, Guan Y, Wang J, et al. Genesis of a highly pathogenic and potentially pandemic H5N1 influenza virus in eastern Asia. Nature 2004 Jul 8;430:209-13 [Abstract]

Lin J, Zhang J, Dong X, et al. Safety and immunogenicity of an inactivated adjuvanted whole-virion influenza A (H5N1) vaccine: a phase 1 randomised controlled trial. Lancet 2006 Sep 15;368(9540):991-7 [Abstract]

Lipatov AS, Kwon YK, Sarmento LV, et al. Domestic pigs have low susceptibility to H5N1 highly pathogenic avian influenza viruses. PLoS Pathog 2008 Jul 11;4(7):e1000102 [Full text]

Maines TR, Chen LM, Matsuoka Y, et al. Lack of transmission of H5N1 avian-human reassortant influenza viruses in a ferret model. Proc Natl Acad Sci 2006 Aug 8;103(32):1221-6 [Abstract]

McKimm-Breschkin J, Trivedi T, et al. Neuraminidase sequence analysis and susceptibilities of influenza virus clinical isolates to zanamivir and oseltamivir. Antimicrob Agents Chemother 2003 Jul;47(7):2264-72 [Full text]

McKimm-Breschkin JL, Selleck PW, Usman TB, et al. Reduced sensitivity of influenza A (H5N1) to oseltamivir. Emerg Infect Dis 2007 Sep;13(9):1354-7 [Full text]

Mehlmann M, Dawson ED, Townsend MB, et al. Robust sequence selection method used to develop the FluChip diagnostic microarray for influenza virus. J Clin Microbiol 2006 Aug;44(8):2857-62 [Abstract]

Myers KP, Setterquist SF, Capuano AW, et al. Infection due to 3 avian influenza subtypes in United States veterinarians. Clin Infect Dis 2007 Jul 1;45(1):4-9 [Abstract]

Monto AS. Vaccines and antiviral drugs in pandemic preparedness. Emerg Infect Dis 2006 Jan;12(1):55-60 [Full text]

Morgan O, Kuhne M, Nair P, et al. Personal protective equipment and risk for avian influenza (H7N3). Emerg Infect Dis 2009 Jan;15(1):59-62 [Full text]

Moscona A. Neuraminidase inhibitors for influenza. N Engl J Med 2005 Sep 29;353:13:1363-73 [Extract]

Newton DW, Mellen CF, Baxter BD, et al. Practical and sensitive screening strategy for detection of influenza virus. J Clin Microbiol 2002 Nov;40(11):4353-6 [Full text]

NIAID. Inexpensive test detects H5N1 infections quickly and accurately. Nov 13, 2006, press release [Full text]

NIAID. Timeline of human flu pandemics [Web page]

Ng EK, Cheng PK, Ng AY, et al. Influenza A H5N1 detection. Emerg Infect Dis 2005 Aug;11(8):1303-5 [Full text]

Nolan TM, Richmond PC, Skelio MV, et al. Phase I and II randomised trials of the safety and immunogenicity of a prototype adjuvanted inactivated split-virus influenza A (H5N1) vaccine in healthy adults. Vaccine 2008 Aug 5;26(33):4160-7 [Abstract]

OIE. Avian influenza. Chap 2.7.12. In: Manual of diagnostic tests and vaccines for terrestrial animals. May 2005 [Full text]

OIE. Highly pathogenic avian influenza. Technical disease card database. Apr 22, 2002 [Web page]

OIE. Highly pathogenic avian influenza. International Health Code. Chap 2.7.12. 2004 [Full text]

OIE. Update on highly pathogenic avian influenza in animals (type H5 and H7) [Web page]

Olsen SJ. Ungchusak K, Sovann L, et al. Family clustering of avian influenza A (H5N1). (Letter) Emerg Infect Dis 2005 Nov;11(11):1799-1801 [Full text]

Olsen SJ, Laosiritaworn Y, Pattanasin S, et al. Poultry-handling practices during avian influenza outbreak, Thailand. Emerg Infect Dis 2005 Oct;11(10):1601-3 [Full text]

Ortiz JR, Katz MA, Mahmoud MN, et al. Lack of evidence of avian-to-human transmission of avian influenza A (H5N1) virus among poultry workers, Kano, Nigeria, 2006. J Infect Dis 2007 Dec 1;196(11):1685-91 [Abstract]

OSHA. OSHA guidance update on protecting employees from avian flu (avian influenza) viruses. November 2006 [Full text]

Pachucki CT, Khurshid MA, Nawrocki J. Utility of reverse transcriptase PCR for rapid diagnosis of influenza a virus infection and detection of amantadine-resistant influenza a virus isolates. J Clin Microbiol 2004 Jun;42(6):2796-8 [Full text]

Payungporn S, Chutinimitkul S, Chaisingh A, et al. Single step multiplex real-time RT-PCR for H5N1 influenza A virus detection. J Virol Methods 2005 [Published online Sep 22] [Abstract]

Peiris M, Yuen KY, Leung CW, et al. Human infection with influenza H9N2. Lancet 1999;354:916-7 [Abstract]

PHS (Poultry Health Services). Fowl plague, avian influenza highly pathogenic. PHS Avian Influenza Forum [Web page]

Rameix-Welti MA, Agou F, Buchy P, et al. Natural variation can significantly alter the sensitivity of influenza A (H5N1) viruses to oseltamivir. Antimicrob Agents Chemother 2006 Nov;50(11):3809-15 [Abstract]

Rice EW, Adcock NJ, Sivaganesan M. Chlorine inactivation of highly pathogenic avian influenza virus (H5N1). Emerg Infect Dis 2007 Oct;13(10):1568-70 [Full text]

Roche. Roche update on Tamiflu for pandemic influenza preparedness. Apr 26, 2007 [Press release]

Roche. U.S.-based supply chain for Tamiflu fully operational. Sep 14, 2006 [Press release]

Rowe T, Abernathy RA, Hu-Primmer J, et al. Detection of antibody to avian influenza A (H5N1) virus in human serum by using a combination of serologic assays. J Clin Microbiol 1999 Apr;37(4):937-43 [Full text]

Sanofi Pasteur. Sanofi Pasteur to donate 60 million doses of H5N1 vaccine to WHO over 3 years for its influenza vaccine global stockpile June 16, 2008, press release [Full text]

Shinya K, Ebina M, Yamada S, et al. Avian flu: influenza virus receptors in the human airway. Nature 2006 Mar 23;440:435-6 [Abstract]

Siembieda J, Johnson CK, Boyce W, et al. Risk for avian influenza virus exposure at human–wildlife interface. Emerg Infect Dis 2008 Jul:14(7):1151-3 [Full text]

Stallknecht DE. Shane SM, Kearney MT, et al. Persistence of avian influenza viruses in water. Avian Dis 1990 Apr-Jun;34(2):406-11 [Abstract]

Stegeman A, Bouma A, Elbers ARW, et al. Avian influenza A virus (H7N7) epidemic in The Netherlands in 2003: course of the epidemic and effectiveness of control measures. J Infect Dis 2004 Dec 15;190:2088-95 [Abstract]

Stephenson I, Bugarini R, Nicholson KG, et al. Cross-reactivity to highly pathogenic avian influenza H5N1 viruses after vaccination with nonadjuvanted and MF59-adjuvanted influenza A/duck/Singapore/97 (H5N3) vaccine: a potential priming strategy. J Infect Dis 2005 Apr 15;191(8):1210-5 [Full text]

Suwannakarn K, Chutinimitkul S, Payungporn S, et al. Assay to detect H5N1 oseltamivir resistance. Emerg Infect Dis 2006 Dec;12(12):1995-6 [Full text]

Taubenberger JK, Reid AH, Lourens RM, et al. Characterization of the 1918 influenza virus polymerase genes. (Letter) Nature 2005;437(7060):889-93 [Full text]

Tellier R. Review of Aerosol transmission of influenza A virus. Emerg Infect Dis 2006; 12(11) [Full text]

Townsend MB, Dawson ED, Mehlmann M, et al. Experimental evaluation of the FluChip diagnostic microarray for influenza virus surveillance. J Clin Micro 2006 Aug;44(8):2863-71 [Abstract]

Tran TH, Nguyen TL, Nguyen TD, et al. Avian influenza A (H5N1) in 10 patients in Vietnam. N Engl J Med 2004 Mar 18;350(12):1179-88 [Abstract]

Treanor JJ, Campbell JD, Zangwill KM , et al. Safety and immunogenicity of an inactivated subvirion influenza (H5N1) vaccine. N Engl J Med 2006;354:1343-51 [Full text]

Treanor JJ. Influenza virus. In: Mandell GL, Douglas RG, Bennett JE, eds. Mandell, Douglas, and Bennett's principles and practice of infectious diseases. Ed 6. Philadelphia: Elsevier, 2005

Ungchusak K, Auewarakul P, Dowell SF, et al. Probable person-to-person transmission of avian influenza (H5N1). N Engl J Med 2005 Jan 27;352(4): 323-4 [Full text]

USDA/APHIS. Agricultural Bioterrorism Protection Act of 2002: Possession, use, and transfer of biological agents and toxins. Final Rule 7 CFR Part 331 and 9 CFR Part 121. Fed Reg 2005 Mar 18;70(52):13242-92 [Full text]

USDA/APHIS. Questions and answers on the Agricultural Bioterrorism Protection Act of 2002: Possession, use, and transfer of select agents and toxins. APHIS factsheet. May 2005 [Full text]

USDA. Draft interagency risk assessment for the public health impact of highly pathogenic avian influenza virus (HPAIV) in poultry, shell eggs, and egg products. Nov 2008 [Full text]

Uyeki TM, Chong YH, Katz JM, et al. Lack of evidence for human-to-human transmission of avian influenza A (H9N2) viruses in Hong Kong, China, 1999. Emerg Infect Dis 2002 Feb;8(2):154-9 [Full text]

US Department of State. Medical information for Americans traveling abroad [Web page]

Vong S, Coghlan B, Mardy S, et al. Low frequency of poultry-to-human H5N1 transmission, southern Cambodia, 2005. Emerg Infect Dis 2006 Oct;12(10) [Full text]

Voyles BA. Orthomyxoviruses. In: The biology of viruses. Ed 2. New York, NY: McGraw-Hill, 2002:147

Wallace LA, McAulay KA, Douglas JD, et al. Influenza diagnosis: from dark isolation into the molecular light. West of Scotland Respiratory Virus Study Group. J Infect 1999 Nov;39(3):221-6 [Abstract]

Wan H, Sorrell EM, Song H, et al. Replication and transmission of H9N2 influenza viruses in ferrets: evaluation of pandemic potential. PLoS One 2008 Aug;3(8):1-13 [Full text]

Wang M, Di B, Zhou D-H, et al. Food markets with live birds a source of avian influenza. Emerg Infect Dis 2006;12(11) [Full text]

Webster RG, Peiris M, Chen H, et al. H5N1 outbreaks and enzootic influenza. Emerg Infect Dis 2006 Jan;12(1):3-8 [Full text]

Webster RG, Yakhno M, Hinshaw VS, et al. Intestinal influenza: replication and characterization of influenza viruses in ducks. Virology 1978;84(2):268-78

Weinberg A. Evaluation of three influenza a and B rapid antigen detection kitsupdate. Clin Diagn Lab Immunol 2005 Aug;12(8):1010 [Listing with link]

Wetherall NT, Trivedi T, Zeller J, et al. Evaluation of neuraminidase enzyme assays using different substrates to measure susceptibility of influenza virus clinical isolates to neuraminidase inhibitors: report of the neuraminidase inhibitor susceptibility network. J Clin Microbiol 2003 Feb;41(2):742-50 [Full text]

WHO. A description of the process of seasonal and H5N1 influenza vaccine virus selection and development. Nov 19, 2007 [Full text]

WHO. Advice to international travelers. Nov 2005 [Full text]

WHO. Antigenic and genetic characteristics of H5N1 viruses and candidate H5N1 vaccine viruses developed for potential use as human vaccines. Feb 2008 [Full text]

WHO. Antiviral drugs: their role during a pandemic. Nov 2005 [Full text]

WHO. Avian influenza: assessing the pandemic threat [Full text]

WHO. Avian influenza, including influenza A (H5N1), in humans: WHO interim infection control guideline for health care facilities [Full text]

WHO. Avian influenza: situation in Indonesia: update 14 [Full text]

WHO. Avian influenza: situation in Thailand; status of pandemic vaccine development. Oct 4, 2004 [Web page]

WHO. Avian influenza [Home page]

WHO. Avian influenza surveillance [Full text]

WHO. Avian influenza: update: implications of H5N1 infections in pigs in China. Aug 25, 2004 [Web page]

WHO. Case definitions for human infections with influenza A(H5N1) virus [Full text]

WHO. Checklist for influenza pandemic preparedness planning. 2005 [Full text]

WHO. Collecting, preserving, and shipping specimens for the diagnosis of avian influenza A (H5N1) virus infection. Guide for field operations [Full Text]

WHO. Cumulative number of confirmed human cases of avian influenza A(H5N1) [Web page—open most recent report available]

WHO. Current WHO phase of pandemic alert [Web page]

WHO. Development of a vaccine effective against avian influenza H5N1 infection in humans: update 4. Jan 20, 2004 [Web page]

WHO. Guidelines for global surveillance of influenza A/H5 [Web page]

WHO. Guidelines for the use of seasonal influenza vaccine in humans at risk of H5N1 infection. Jan 30, 2004 [Web page]

WHO. H5N1 avian influenza: timeline of major events. Mar 25, 2008 [Full text]

WHO. Laboratory study of H5N1 viruses in domestic ducks: main findings. Oct 29, 2004 [Full text]

WHO. Manual on animal influenza diagnosis and surveillance [Full text]

WHO. Pandemic influenza draft protocol for rapid response and containment. May 30, 2006 [Full text]

WHO. Projected supply of pandemic influenza vaccine sharply increases. Oct 23, 2007 [Full text]

WHO. Protection of individuals with high poultry contact in areas affected by avian influenza H5N1: consolidation of pre-existing guidance. Feb 2008 [Full text]

WHO. Rapid Advice Guidelines on pharmacological management of humans infected with avian influenza A (H5N1) virus [Full text]

WHO. Recommendations and laboratory procedures for detection of avian influenza A (H5N1) virus in specimens from suspected human cases. Revised Aug 2007 [Full text]

WHO. Review of latest available evidence on risks to human health through potential transmission of avian influenza (H5N1) through water and sewage. Mar 24, 2006 [Full text]

WHO. Situation updates: avian flu [Web page]

WHO. Update: WHO-confirmed human cases of avian influenza A(H5N1) infection, 25 November 2003 - 24 November 2006. Wkly Epidemiol Rec 2007 Feb 9;82(6):41-8 [Full text]

WHO. WHO and manufacturers move ahead with plans for H5N1 influenza global vaccine stockpile. Jun 13, 2007 [Full text]

WHO. WHO global influenza preparedness plan: the role of WHO and recommendations for national measures before and during pandemics. Apr 2005 [Full text]

WHO Global Influenza Program Surveillance Network. Evolution of H5N1 avian influenza viruses in Asia. Emerg Infect Dis 2005 October 11(10):1515-21[Full text]

WHO/INFOSAN. Highly pathogenic H5N1 avian influenza outbreaks in poultry and in humans: food safety implications. Nov 4, 2005 [Full text]

WHO Writing Committee of WHO Consultation on Human Influenza A/H5. Avian influenza A (H5N1) infection in humans. N Engl J Med 2005 Sep 29;353(13):1374-85 [Full text]

WHO Writing Committee of the Second World Health Organization Consultation on Clinical Aspects of Human Infection with Avian Influenza A (H5N1) Virus. Update on avian influenza A (H5N1) virus infection in humans. N Engl J Med 2008 Jan 17;358(3):261-73 [Full text]

Wu C, Cheng X, He J, et al. A multiplex real-time RT-PCR for detection and identification of influenza virus types A and B and subtypes H5 and N1. J Virol Methods 2008 Mar;148(1-2):81-8 [Abstract]

Yen HL, Monto AS, Webster RG, et al. Virulence may determine the necessary duration and dosage of oseltamivir treatment for highly pathogenic A/Vietnam/1203/04 influenza virus in mice. J Infect Dis 2005 Aug 15;192(4):665-72 [Abstract]

Yu H, Feng Z, Zhang X, et al. Human influenza A (H5N1) cases, urban areas of People's Republic of China, 2005-2006. Emerg Infect Dis 2007 Jul;13(7):1061-4 [Full text]

Yu H, Gao Z, Feng Z, et al. Clinical characteristics of 26 human cases of highly pathogenic avian influenza A (H5N1) virus infection in China. PLoS ONE 2008 Aug 21;3(8):e2985 [Full text]

Yuen KY, Chan PKS, Peiris M, et al. Clinical features and rapid viral diagnosis of human disease associated with avian influenza A H5N1 virus. Lancet 1998;351(9101):467-71 [Abstract]

Zhou J, Law HKW, Cheung CY, et al. Differential expression of chemokines and their receptors in adult and neonatal macrophages infected with human or avian influenza viruses. J Infect Dis 2006 Jul 1;194(1):61-70 [Abstract]


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