symptoms.
On day 2 of hospitalization the patient’s respiratory rate increased from 22 to 46 and her oxygen saturation dropped while on oxygen administered by nasal cannula. She was transferred to an intensive care unit, where her respiratory status quickly deteriorated, necessitating emergency intubation. A new chest radiograph showed bilateral involvement, and she was begun on vancomycin, aztreonam, and azithromycin. Blood cultures drawn on her admission were negative, and an expectorated sputum sample taken at the same time was not processed due to poor specimen quality. A PCR test performed on a nasopharyngeal swab was positive for a viral agent, revealing the etiology of her infection (Fig. 10.1).
1 1. What is the agent causing her infection? What are the key virulence factors of this agent?
2 2. How does this virus change over time? What made this virus unique in 2009?
3 3. Why was PCR used to diagnose this infection? What do the curves in Fig. 10.1 represent?
4 4. What is the usual outcome of this infection in this patient population? What groups of people are at greater risk of a poor outcome when they are infected with this virus? How did these groups differ in 2009?
5 5. What are the common complications associated with this infection that lead to increased morbidity and mortality? How are these complications diagnosed?
6 6. What antiviral drugs are available to treat this infection, and how do they work? Is there any concern for antiviral resistance?
7 7. Two types of licensed vaccines are available that can prevent this disease. Describe the nature of both of these vaccines and how they are used.
Figure 10.1 Amplification curves of a real-time PCR test.
CASE 10 CASE DISCUSSION
1. Although a number of respiratory viruses could explain this patient’s symptoms, influenza is the most common febrile respiratory illness in adults, particularly during the winter months, when influenza activity normally peaks. In pediatric patients (particularly <1 year old), respiratory syncytial virus should also be considered. The clinical clues of her influenza infection are the abrupt onset of fever and sore throat with nonproductive cough seen at her initial presentation to the ED. Clinically it is difficult to distinguish infections due to influenza A and influenza B, though influenza A tends to be associated with more severe disease, is generally the cause of annual epidemics, and has been responsible for all described pandemics.
Influenza virus has two major envelope proteins that contribute to its pathogenesis: neuraminidase and hemagglutinin. Neuraminidase likely has at least two functions. Its major function seems to be the cleavage of sialic acid from the cell surface and progeny virions, which facilitates the spread of new virions from infected respiratory cells. There is also evidence supporting the role of neuraminidase in viral entry to the cell. One mechanism that has been proposed is that neuraminidase cleaves decoy receptors on mucins, cilia, and cellular glycocalix so that the virus can have greater access to the functional receptors on the cell membrane. Once the virus penetrates to the cell surface, binding to specific sialic acid-rich receptors is mediated by hemagglutinin. Proteolytic cleavage of hemagglutinin by lung serine proteases is required for hemagglutinin activity. After the virus is endocytosed into the cell, hemagglutinin plays a role in the formation of channels through which viral RNA can enter the cytoplasm and initiate the viral replicative cycle.
2. In recent years only two hemagglutinin types (H1 and H3) and two neuraminidase types (N1 and N2) of influenza A virus have been circulating in humans (H1N1 and H3N2). However, due to antigenic variation, there are annual influenza epidemics and, in 2009, a pandemic. Why does this happen? There are two major evolutionary concepts related to influenza virus—antigenic drift and antigenic shift.
A unique property of influenza viruses is that they have single-stranded RNA genomes made of eight segments. Each influenza gene is found on a separate viral RNA segment. Since the mutation rate for RNA is higher than that of DNA (10−3 to 10−5 versus 10−6 to 10−8 per base per generation), point mutations readily accumulate in influenza viruses. Although mutations occur throughout the influenza genome, the accumulation of mutations (and corresponding amino acid changes) in surface antigens, such as hemagglutinin and neuraminidase, have the greatest impact. For influenza A virus, these changes will not necessarily result in the change of the classification of a viral strain (which is based on the subtypes of the H and N antigens), but they may be sufficient to render patients with antibodies to the parent strain susceptible to the new mutant strain. This is the basis for the decision to reevaluate and potentially change the formulation of the influenza vaccine each year to include recent isolates, so that protective antibodies to the most recent isolates will be made in response to the vaccine. Both influenza A and influenza B are constantly changing by antigenic drift.
The more dramatic, and less common, antigenic shift is due to genetic reassortment of genes to form a novel human influenza virus, which typically has different hemagglutinin and/or neuraminidase proteins. Antigenic shift occurs during coinfection of a cell with two different influenza A viruses. Since the packaging of viral RNA segments occurs randomly, a coinfected cell could form a variety of different virions. The result could be a virus with a different classification (e.g., a shift from H1N1 to H5N1) or a virus of the same type but with divergent genomic sequences from nonhuman sources such as pigs or birds. The end result is a new virus that differs dramatically from parent strains.
The influenza A H1N1 pandemic of 2009 was a result of antigenic shift. Although an H1N1 influenza virus had circulated globally for years, a reassortant H1N1 virus was introduced and spread worldwide. The 2009 H1N1 virus was a result of the introduction of Eurasian swine segments (neuraminidase and matrix) into the classical swine influenza strain that previously had only caused swine-to-swine transmission and rare swine-to-human transmission. When an antigenic shift occurs, most of the world’s population has little or no protection against the new virus, resulting in large epidemics or pandemics.
3. There are a variety of ways of diagnosing influenza in the laboratory, including rapid antigen tests, direct fluorescent-antibody assay (DFA), viral culture, and molecular detection. Rapid antigen tests are immunochromatographic assays that have been used for decades and have been favored due to their fast time to result (~15 minutes). However, as diagnostic methods have improved and circulating strains have changed, studies have shown that these tests suffer from lack of sensitivity. Sensitivities down to 10% were reported during the 2009 pandemic. Typical ranges of sensitivity reported are 20 to 90% depending on the strain circulating and the method used as the reference method. A further concern is the positive predictive value of rapid antigen tests when used outside of peak influenza season. Since positive predictive value is dependent on the prevalence of disease, using a test with imperfect specificities (90 to 95%) during times of low prevalence increases the chance that a positive result may actually be false positive rather than true positive. However, the times when laboratory testing for influenza is the most helpful clinically are at the beginning and end of the epidemic season, when the differential diagnosis is much broader. Another rapid method (~2 hours) is DFA testing. DFA uses a pool of monoclonal antibodies to influenza and other common respiratory viruses to directly detect infected cells obtained from the nasopharynx of patients. Although it is more sensitive than rapid antigen tests, DFA also had decreased sensitivity (~47%) for detecting the 2009 H1N1 pandemic strain. DFA sensitivity and specificity are also dependent on the skill of the personnel performing the test. Therefore, if rapid antigen tests or DFA must be used,
