A comparison of influenza A virus with other viruses that infect the human respiratory tract is shown in Table 1. Influenza virus is considered here.

Table 1. Comparison of Viruses That Infect the Human Respiratory Tract

Pathogenesis and Pathology

 Influenza virus spreads from person to person by airborne droplets or by contact with contaminated hands or surfaces. A few cells of respiratory epithelium are infected if deposited virus particles avoid removal by the cough reflex and escape neutralization by preexisting specific immunoglobulin A (IgA) antibodies or inactivation by nonspecific inhibitors in the mucous secretions. Progeny virions are soon produced and spread to adjacent cells, where the replicative cycle is repeated. Viral NA lowers the viscosity of the mucous film in the respiratory tract, laying bare the cellular surface receptors and promoting the spread of virus-containing fluid to lower portions of the tract. Within a short time, many cells in the respiratory tract are infected and eventually killed.

The incubation period from exposure to virus and the onset of illness varies from 1 day to 4 days, depending on the size of the viral dose and the immune status of the host. Viral shedding starts the day preceding onset of symptoms, peaks within 24 hours, remains elevated for 1–2 days, and then declines over the next 5 days. Infectious virus is very rarely recovered from blood.

Interferon is detectable in respiratory secretions about 1 day after viral shedding begins. Influenza viruses are sensitive to the antiviral effects of interferon, and it is believed that the innate immunity response contributes to host recovery from infection. Specific antibody and cell-mediated responses cannot be detected for another 1–2 weeks.

Influenza infections cause cellular destruction of the superficial mucosa of the respiratory tract but do not affect the basal layer of epithelium. Complete reparation of cellular damage probably takes up to 1 month. Viral damage to the respiratory tract epithelium lowers its resistance to secondary bacterial pathogens, especially staphylococci, streptococci, and Haemophilus influenzae.

Edema and mononuclear infiltrations in response to cytokine release and cell death caused by viral replication probably account for local symptoms. The fever and systemic symptoms associated with influenza reflect the action of cytokines.

Clinical Findings

 Influenza attacks mainly the upper respiratory tract. It poses a serious risk for elderly adults, very young children, and people with underlying medical conditions such as lung, kidney, or heart problems, diabetes, cancer, or immunosuppression .

A. Uncomplicated Influenza

Symptoms of classic influenza usually appear abruptly and include chills, headache, and dry cough followed closely by high fever, generalized muscular aches, malaise, and anorexia. The fever usually lasts 3–5 days, as do the systemic symptoms. Respiratory symptoms typically last another 3–4 days. The cough and weakness may persist for 2–4 weeks after major symptoms subside. Mild or asymptomatic infections may occur. These symptoms may be induced by any strain of influenza A or B. In contrast, influenza C rarely causes the influenza syndrome, causing instead a common cold illness. Coryza and cough may last for several weeks.

Clinical symptoms of influenza in children are similar to those in adults, although children may have higher fever and a higher incidence of gastrointestinal manifestations such as vomiting. Febrile seizures can occur. Influenza A viruses are an important cause of croup, which may be severe, in children younger than 1 year of age. Finally, otitis media may develop.

When influenza appears in epidemic form, clinical findings are consistent enough that the disease can be diagnosed presumptively. Sporadic cases cannot be diagnosed on clinical grounds because disease manifestations cannot be distinguished from those caused by other respiratory tract pathogens. However, those other agents rarely cause severe viral pneumonia, which can be a complication of influenza A virus infection.

B. Pneumonia

Serious complications usually occur only in elderly adults and debilitated individuals, especially those with underlying chronic disease. Pregnancy appears to be a risk factor for lethal pulmonary complications in some epidemics. The lethal impact of an influenza epidemic is reflected in the excess deaths caused by pneumonia and cardiopulmonary diseases.

Pneumonia complicating influenza infections can be viral, secondary bacterial, or a combination of the two. Increased mucous secretion helps carry agents into the lower respiratory tract. Influenza infection enhances susceptibility of patients to bacterial superinfection. This is attributed to loss of ciliary clearance, dysfunction of phagocytic cells, and provision of a rich bacterial growth medium by the alveolar exudate. Bacterial pathogens are most often Staphylococcus aureus, Streptococcus pneumoniae, and H influenzae.

Combined viral–bacterial pneumonia is approximately three times more common than primary influenza pneumonia. A molecular basis for a synergistic effect between virus and bacteria may be that some S aureus strains secrete a pro tease able to cleave the influenza HA, thereby allowing pro duction of much higher titers of infectious virus in the lungs.

C. Reye Syndrome

Reye syndrome is an acute encephalopathy of children and adolescents, usually between 2 and 16 years of age. The mortality rate is high (10–40%). The cause of Reye syndrome is unknown, but it is a recognized rare complication of influenza B, influenza A, and herpesvirus varicella-zoster infections. There is a possible relationship between salicylate use and subsequent development of Reye syndrome. The incidence of the syndrome has decreased with the reduced use of salicylates in children with flulike symptoms.

Immunity

 Immunity to influenza is long lived and subtype specific. Whereas antibodies against HA and NA are important in immunity to influenza, antibodies against the other virus encoded proteins are not protective. Resistance to initiation of infection is related to antibody against the HA, but decreased severity of disease and decreased ability to transmit virus to contacts are related to antibody directed against the NA.

Protection correlates with both serum antibodies and secretory IgA antibodies in nasal secretions. The local secretory antibody is probably important in preventing infection. Serum antibodies persist for many months to years; secretory antibodies are of shorter duration (usually only several months). Antibody also modifies the course of illness. A per son with low titers of antibody may be infected but will experience a mild form of disease. Immunity can be incomplete; reinfection with the same virus can occur.

The three types of influenza viruses are antigenically unrelated and therefore induce no cross-protection. When a viral type undergoes antigenic drift, a person with preexisting antibody to the original strain may have only mild infection with the new strain. Subsequent infections or immunizations reinforce the antibody response to the first subtype of influenza experienced years earlier, a phenomenon called “original antigenic sin.”

The primary role of cell-mediated immune responses in influenza is believed to be clearance of an established infection; cytotoxic T cells lyse infected cells. The cytotoxic T lymphocyte response is cross-reactive (able to lyse cells infected with any subtype of virus) and appears to be directed against both internal proteins (NP, M) and the surface glycoproteins.

Laboratory Diagnosis

Clinical characteristics of viral respiratory infections can be produced by many different viruses. Consequently, diagnosis of influenza relies on identification of viral antigens or viral nucleic acid in specimens, isolation of the virus, or demonstration of a specific immunologic response by the patient.

Nasopharyngeal swabs and nasal aspirate or lavage fluid are the best specimens for diagnostic testing and should be obtained within 3 days after the onset of symptoms.

A. Polymerase Chain Reaction

Rapid tests based on detection of influenza RNA in clinical specimens using reverse transcription polymerase chain reaction (RT-PCR) are preferred for diagnosis of influenza. RT-PCR is rapid (<1 day), sensitive, and specific. Multiplex molecular technologies are available that allow for the rapid detection of multiple pathogens in a single test.

B. Isolation and Identification of Virus

 The sample to be tested for virus isolation should be held at 4°C until inoculation into cell culture because freezing and thawing reduce the ability to recover virus. However, if storage time will exceed 5 days, the sample should be frozen at –70°C.

Viral culture procedures take 3–10 days. Classically, embryonated eggs and primary monkey kidney cells have been the isolation methods of choice for influenza viruses, although other cell lines may be used. Inoculated cell cultures are incubated in the absence of serum, which may contain nonspecific viral inhibitory factors, and in the presence of trypsin, which cleaves and activates the HA so that replicating virus will spread throughout the culture.

Cell cultures can be tested for the presence of virus by hemadsorption 3–5 days after inoculation, or the culture fluid can be examined for virus after 5–7 days by hemagglutination or immunofluorescence. If the results are negative, a passage is made into fresh cultures. This passage may be necessary because primary viral isolates are often fastidious and grow slowly.

Viral isolates can be identified by hemagglutination inhibition (HI), a procedure that permits rapid determination of the influenza type and subtype. To do this, reference sera to currently prevalent strains must be used. Hemagglutination by the isolate will be inhibited by specific antiserum to the homologous subtype.

For rapid diagnosis, cell cultures on coverslips in shell vials may be inoculated and stained 1–4 days later with mono clonal antibodies to respiratory agents. Rapid viral cultures can also be tested by RT-PCR to identify a cultured agent.

It is possible to identify viral antigen directly in exfoliated cells in nasal aspirates using fluorescent antibodies. This test is rapid (taking only a few hours) but is not as sensitive as PCR or viral isolation, does not provide full details about the viral strain, and does not yield an isolate that can be characterized. Rapid influenza antigen detection tests are commercially available that take less than 15 minutes. However, these tests vary in sensitivity and specificity, and a negative result does not rule out influenza infection.

C. Serology

Antibodies to several viral proteins (HA, NA, NP, and matrix) are produced during infection with influenza virus. The immune response against the HA glycoprotein is associated with resistance to infection.

Routine serodiagnostic tests in use are based on HI and enzyme-linked immunosorbent assay. Paired acute and convalescent sera are necessary because normal individuals usually have influenza antibodies. A fourfold or greater increase in titer must occur to indicate influenza infection. Human sera often contain nonspecific mucoprotein inhibitors that must be destroyed before testing by HI.

The HI test reveals the strain of virus responsible for infection only if the correct antigen is available for use. Neutralization tests are the most specific and the best predictor of susceptibility to infection but are more unwieldy and more time consuming to perform than the other tests. The enzyme-linked immunosorbent assay is more sensitive than other assays.

Complications may be encountered in attempting to identify the strain of infecting influenza virus by the patient’s antibody response because anamnestic responses frequently occur.

Epidemiology

 Influenza viruses occur worldwide and cause annual out breaks of variable intensity. It is estimated that annual epidemics of seasonal influenza cause 3–5 million cases of severe illness and 250,000–500,000 deaths worldwide. The eco nomic impact of influenza A outbreaks is significant because of the morbidity associated with infections. Economic costs have been estimated at $10–60 million per million population in industrialized countries, depending on the size of the epidemic.

The three types of influenza vary markedly in their epidemiologic patterns. Influenza C is least significant; it causes mild, sporadic respiratory disease but not epidemic influenza. Influenza B sometimes causes epidemics, but influenza type A can sweep across continents and around the world in massive epidemics called pandemics.

The incidence of influenza peaks during the winter. In the United States, influenza epidemics usually occur from January through April (and from May to August in the Southern Hemisphere). A continuous person-to-person chain of trans mission must exist for maintenance of the agent between epidemics. Some viral activity can be detected in large population centers throughout each year, indicating that the virus remains endemic in the population and causes a few subclinical or minor infections.

A. Antigenic Change

 Periodic outbreaks appear because of antigenic changes in one or both surface glycoproteins of the virus. When the number of susceptible persons in a population reaches a sufficient level, the new strain of virus causes an epidemic. The change may be gradual (hence the term “antigenic drift”) because of point mutations reflected in alterations at major antigenic sites on the glycoprotein or drastic and abrupt (hence the term “antigenic shift”) owing to genetic reassortment during coinfection with an unrelated strain.

All three types of influenza virus exhibit antigenic drift. However, only influenza A undergoes antigenic shift, presumably because types B and C are restricted to humans, but related influenza A viruses circulate in animal and bird populations. These animal strains account for antigenic shift by genetic reassortment of the glycoprotein genes. Influenza A viruses have been recovered from many aquatic birds, especially ducks; from domestic poultry, such as turkeys, chickens, geese, and ducks; from pigs and horses; and even from seals and whales. Serologic surveys indicate a high prevalence of influenza virus infection in domestic cats.

Influenza outbreaks occur in waves, although there is no regular periodicity in the occurrence of epidemics. The experience in any given year will reflect the interplay between extent of antigenic drift of the predominant virus and waning immunity in the population. The period between epidemic waves of influenza A tends to be 2–3 years; the interepidemic period for type B is longer (3–6 years). Every 10–40 years, when a new subtype of influenza A appears, a pan demic results. Seroepidemiology studies suggest the cause of epidemics in 1890 (H2N8) and 1900 (H3N8), with known virologic confirmation of epidemics in 1918 (H1N1), 1957 (H2N2), and 1968 (H3N2). The H1N1 subtype reemerged in 1977, although no epidemic materialized.

In early 2009, a novel swine-origin H1N1 virus appeared and reached pandemic spread by mid-year. It was a quadruple reassortant, containing genes from both North Ameri can and Eurasian swine viruses, as well as from avian and human influenza viruses. The virus was readily transmissible among humans and spread globally, causing more than 18,000 deaths. The severity of illness was comparable to that of seasonal flu. The pandemic virus, designated A(H1N1) pdm09, has become a seasonal influenza virus, continuing to circulate with other seasonal viruses, and appears to have dis placed the previously circulating H1N1 strain virus.

Surveillance for influenza outbreaks is necessary to identify the early appearance of new strains, with the aim of pre paring vaccines against them before an epidemic occurs. That surveillance may extend into animal populations, especially birds, pigs, and horses. Isolation of a virus with an altered HA in the late spring during a mini-epidemic signals a possible epidemic the following winter. This warning sign, termed a “herald wave,” has been observed to precede influenza A and B epidemics.

B. Avian Influenza

 Sequence analyses of influenza A viruses isolated from many hosts in different regions of the world support the theory that all mammalian influenza viruses derive from the avian influenza reservoir. Of the 18 HA subtypes found in birds, only a few have been transferred to mammals (H1, H2, H3, H5, H7 and H9 in humans; H1, H2 and H3 in swine; and H3 and H7 in horses). The same pattern holds for NA; 11 NA subtypes are known for birds, only 2 of which are found in humans (N1, N2).

Avian influenza ranges from inapparent infections to highly lethal infections in chickens and turkeys. Most influenza infections in ducks are avirulent. Influenza viruses of ducks multiply in cells lining the intestinal tract and are shed in high concentrations in fecal material into water, where they remain viable for days or weeks, especially at low temperatures. It is likely that avian influenza is a waterborne infection, moving from wild to domestic birds and pigs.

To date, all human pandemic strains have been reassortants between animal and human influenza viruses. Evidence supports the model that pigs serve as mixing vessels for reassortants because their cells contain receptors recognized by both human and avian viruses (Figure 1). The pandemic strain of 2009 was a novel reassortant that contained swine origin viral genes as well as those from avian and human influenza viruses. School-age children are the predominant vectors of influenza transmission. Crowding in schools favors the aerosol transmission of virus, and children take the virus home to their families.

In 1997, in Hong Kong, the first documented infection of humans by avian influenza A virus (H5N1) occurred. The source was domestic poultry. By 2006, the geographic pres ence of this highly pathogenic H5N1 avian influenza virus in both wild and domestic birds had expanded to include many countries in Asia, Africa, Europe, and the Middle East. Out breaks were the largest and most severe on record. Of about 425 laboratory-confirmed human cases by May 2009, more than half were fatal. So far, isolates from human cases have contained all RNA gene segments from avian viruses, indicating that, in those infections, the avian virus had jumped directly from bird to human. All evidence to date indicates that close contact with diseased birds has been the source of human H5N1 infection. The concern is that, given enough opportunities, the highly pathogenic H5N1 avian influenza virus will acquire the ability to spread efficiently and be sustained among humans, either by reassortment or by adaptive mutation. This would result in a devastating influenza pandemic. Other avian influenza strains have been found in human infections following close bird exposure, including H7N9 H9N2 viruses.

Fig1. Pigs may act as an intermediate host for the generation of human–avian reassortant influenza viruses with pandemic potential. (Reprinted by permission from Macmillan Publishers Ltd: Claas ECJ, Osterhaus ADME: New clues to the emergence of flu pandemics. Nat Med 1998;4:1122. Copyright © 1998).

C. Reconstruction of 1918 Influenza Virus

PCR technology has yielded gene fragments of influenza virus from archival lung tissue specimens from victims of the 1918 Spanish flu epidemic. The complete coding sequences of all eight viral RNA segments have been determined, and the sequences document that it was an H1N1 influenza A virus. It appears that the 1918 virus was not a reassortant but was derived entirely from an avian source that adapted to humans. Using reverse genetics, an infectious virus containing all the gene segments from the 1918 pandemic virus was constructed. In contrast to ordinary influenza viruses, the 1918 virus was highly pathogenic, including being able to kill mice rapidly. The 1918 HA and polymerase genes appeared to be responsible for the high virulence.

 Prevention and Treatment by Drugs

Amantadine hydrochloride and an analog, rimantadine, classed as adamantane drugs, are M2 ion channel inhibitors for systemic use in the treatment and prophylaxis of influenza A. The NA inhibitors zanamivir and oseltamivir (approved in 1999), and peramivir (approved in 2014) are useful for treatment of both influenza A and influenza B. To be maximally effective, the drugs must be administered very early in the disease. Resistant viruses emerge more frequently during therapy with M2 inhibitors than with NA inhibitors and more frequently in children than adults. Resistance to oseltamivir has been associated with the H275Y mutation in NA gene. During 2013–2014, all circulating influenza viruses were resistant to M2 inhibitors, but most were sensitive to NA inhibitors. Depending on the susceptibility of the predominantly circulating strains, sub typing may be useful to determine optimal therapy.

Prevention and Control by Vaccines

 Inactivated viral vaccines are the primary means of prevention of influenza in the United States. However, certain characteristics of influenza viruses make prevention and control of the disease by immunization especially difficult. Existing vaccines are continually being rendered obsolete as the viruses undergo antigenic drift and shift. Surveillance programs by government agencies and the World Health Organization constantly monitor subtypes of influenza circulating around the world to promptly detect the appearance and spread of new strains.

A major advance would be the ability to design a vaccine that stimulates production of a broadly neutralizing antibody response effective against many influenza subtypes.

Several other problems are worthy of mention. Although protection can reach from 70% to 100% in healthy adults, frequency of protection is lower (30–60%) among the elderly and among young children. Inactivated viral vaccines usually do not generate good local IgA or cell-mediated immune responses. The immune response is influenced by whether the person is “primed” by having had prior antigenic experience with an influenza A virus of the same subtype.

A. Preparation of Inactivated

Viral Vaccines Inactivated influenza A and B virus vaccines are licensed for parenteral use in humans. Federal bodies and the World Health Organization make recommendations each year about which strains should be included in the vaccine. The vaccine is usually a cocktail containing one or two type A viruses and one type B virus of the strains isolated in the previous winter’s outbreaks.

Selected seed strains are grown in embryonated eggs, the substrate used for vaccine production. Sometimes the natural isolates grow too poorly in eggs to permit vaccine production, in which case a reassortant virus is made in the laboratory. The reassortant virus, which carries the genes for the surface antigens of the desired vaccine with the replication genes from an egg-adapted laboratory virus, is then used for vaccine production. A cell-based vaccine using animal cell cultures became available in 2012, which overcomes some limitations of egg-based production.

Virus is harvested, purified, concentrated by zonal centrifugation, and inactivated with formalin or β-propiolactone. The quantity of HA is standardized in each vaccine dose (~15 μg of antigen), but the quantity of NA is not standardized because it is more labile under purification and storage conditions. Each dose of vaccine contains the equivalent of about 10 billion virus particles.

Vaccines are either whole virus (WV), subvirion (SV), or surface antigen preparations. The WV vaccine contains intact, inactivated virus; the SV vaccine contains purified virus disrupted with detergents; and the surface antigen vaccines contain purified HA and NA glycoproteins. All are efficacious.

B. Live-Virus Vaccines

 A live-virus vaccine must be attenuated so as not to induce the disease it is designed to prevent. In view of the constantly changing face of influenza viruses in nature and the extensive laboratory efforts required to attenuate a virulent virus, the only feasible strategy is to devise a way to transfer defined attenuating genes from an attenuated master donor virus to each new epidemic or pandemic isolate.

A cold-adapted donor virus, able to grow at 25°C but not at 37°C—the temperature of the lower respiratory tract—should replicate in the nasopharynx, which has a cooler temperature (33°C). A live attenuated, cold-adapted, temperature-sensitive, trivalent influenza virus vaccine administered by nasal spray was licensed in the United States in 2003. It was the first live virus influenza vaccine approved in the United States, as well as the first nasally administered vaccine in the United States.

C. Use of Influenza Vaccines

 The only contraindication to vaccination is a history of allergy to egg protein, although the cell-based vaccine over comes this limitation. When the vaccine strains are grown in eggs, some egg protein antigens are present in the vaccine.

Annual influenza vaccination is recommended for all children ages 6 months to 18 years and for high-risk groups. These include individuals at increased risk of complications associated with influenza infection (those with either chronic heart or lung disease, including children with asthma, or metabolic or renal disorders; residents of nursing homes; persons infected with the human immunodeficiency virus; and those 65 years of age and older) and persons who might transmit influenza to high-risk groups (medical personnel, employees in chronic care facilities, household members). The live-virus intranasal vaccine is not currently recommended for individuals in the high-risk groups.

 Prevention by Hand Hygiene

Although transmission of influenza virus occurs primarily by aerosol spread, hand transmission also is potentially important. Studies have shown that handwashing with soap and water or the use of alcohol-based hand rubs is highly effective at reducing the amount of virus on human hands.