MicrobiologyBytes

Although it has not been in the news much recently, the highly pathogenic avian influenza A virus subtype H5N1 has been endemic in some bird species since its emergence in 1996 and its ecology, genetics and antigenic properties continue to evolve. This has allowed diverse virus strains to emerge in endemic areas with altered receptor specificity, including a new H5 sublineage with enhanced binding affinity to the human-type receptor. The pandemic potential of H5N1 viruses is alarming and may be increasing. This article reviews the complex and changing nature of the H5N1 virus that may contribute to the emergence of pandemic strains – with really serious consequences.

The changing nature of avian influenza A virus (H5N1). Tresnd in Microbiology, 5 December 2011

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In this week’s PLoS Medicine, Gabriel Leung from the Government of the Hong Kong SAR and Angus Nicoll from the European Centre for Disease Prevention and Control offer their reflections on the international response to the 2009 H1N1 influenza pandemic, including what went well and what changes need to be made on the part of global and national authorities in anticipation of future flu pandemics.

Summary points:

• Many of the initial responses to the 2009 H1N1 pandemic went well but there are many lessons to learn for future pandemic planning.
• Clear communication of public health messages is crucial, and should not confuse what could happen (and should be prepared for) with what is most likely to happen.
• Decisions regarding pandemic response during the exigencies of a public health emergency must be judged according to the best evidence available at the time.
• Revising pandemic plans – to be more flexible and more detailed – should wait for WHO leadership if national plans are not to diverge. Surveillance beyond influenza should be stepped up, and contingencies drawn up for the emergence or re-emergence of other novel and known pathogens.
• Data collection and sharing are paramount, and include epidemiological and immunological data. Clinical management of severe influenza disease should not be limited to the current antiviral regimen, and include the development of other therapeutics (e.g., novel antivirals and immunotherapy).
• Greater and more timely access to antivirals and influenza vaccines worldwide remains an ongoing challenge.

Reflections on Pandemic (H1N1) 2009 and the International Response. (2010) PLoS Med 7(10): e1000346. doi:10.1371/journal.pmed.1000346

Related:

• The Biology of Influenza
• 10 things you should know about H1N1 (swineflu)

Research published this week in PLoS Medicine presents the most accurate assessment to date of the severity of the swine flu (H1N1) pandemic in the US. Scientists need to measure the severity of swine flu (how often infection with the swine flu virus results in symptoms leading to illness, hospitalization or death) so that appropriate pandemic plans can be put into place. Severity of swine flu has been difficult to measure for two main reasons: first, people with severe influenza are more likely than those with mild cases to seek care, making it difficult to estimate how many total cases have occurred, and second, the sheer number of cases means that recording routine case data can be difficult due to overburdening of public health systems. In this study, researchers from from Milwaukee (where all medically attended cases were recorded, whether hospitalized or not) and New York City (where only hospitalizations, intensive care admission and deaths were recorded, and a telephone survey of flu-like illness was conducted), along with earlier results from studies by the US CDC, used a statistical approach called Bayesian evidence synthesis. This enabled accurate estimations of severity to be made. Their analyses reveal that the autumn-winter pandemic wave of swine flu should have a death toll only slightly higher than, or considerably lower than, that caused by seasonal influenza in an average year, provided swine flu continues to behave as it did during the summer. Seasonal influenza mainly kills elderly adults, but the authors reveal that most deaths from swine flu will occur in non-elderly adults, a shift in age distribution that has been seen in previous pandemics.

The Severity of Pandemic H1N1 Influenza in the United States, from April to July 2009: A bayesian Analysis. PLoS Med 6(12): e1000207 doi:10.1371/journal.pmed.1000207
Accurate measures of the severity of pandemic (H1N1) 2009 influenza (pH1N1) are needed to assess the likely impact of an anticipated resurgence in the autumn in the Northern Hemisphere. Severity has been difficult to measure because jurisdictions with large numbers of deaths and other severe outcomes have had too many cases to assess the total number with confidence. Also, detection of severe cases may be more likely, resulting in overestimation of the severity of an average case. We sought to estimate the probabilities that symptomatic infection would lead to hospitalization, ICU admission, and death by combining data from multiple sources. We used complementary data from two US cities: Milwaukee attempted to identify cases of medically attended infection whether or not they required hospitalization, while New York City focused on the identification of hospitalizations, intensive care admission or mechanical ventilation (hereafter, ICU), and deaths. New York data were used to estimate numerators for ICU and death, and two sources of data – medically attended cases in Milwaukee or self-reported influenza-like illness (ILI) in New York – were used to estimate ratios of symptomatic cases to hospitalizations. Combining these data with estimates of the fraction detected for each level of severity, we estimated the proportion of symptomatic patients who died (symptomatic case-fatality ratio, sCFR), required ICU (sCIR), and required hospitalization (sCHR), overall and by age category. Evidence, prior information, and associated uncertainty were analyzed in a Bayesian evidence synthesis framework. Using medically attended cases and estimates of the proportion of symptomatic cases medically attended, we estimated an sCFR of 0.048% (95% credible interval [CI] 0.026%–0.096%), sCIR of 0.239% (0.134%–0.458%), and sCHR of 1.44% (0.83%–2.64%). Using self-reported ILI, we obtained estimates approximately 7–96lower. sCFR and sCIR appear to be highest in persons aged 18 y and older, and lowest in children aged 5–17 y. sCHR appears to be lowest in persons aged 5–17; our data were too sparse to allow us to determine the group in which it was the highest. These estimates suggest that an autumn–winter pandemic wave of pH1N1 with comparable severity per case could lead to a number of deaths in the range from considerably below that associated with seasonal influenza to slightly higher, but with the greatest impact in children aged 0–4 and adults 18–64. These estimates of impact depend on assumptions about total incidence of infection and would be larger if incidence of symptomatic infection were higher or shifted toward adults, if viral virulence increased, or if suboptimal treatment resulted from stress on the health care system; numbers would decrease if the total proportion of the population symptomatically infected were lower than assumed.

Related:

• The good news about influenza
• Spotting pandemic influenza viruses
• How influenza pandemics get started
• The Pathogenicity of Pandemic Influenza Viruses

Influenza viruses contain segmented, negative-strand RNA genomes. Genome segmentation facilitates reassortment between different influenza virus strains infecting the same cell. This phenomenon results in the rapid exchange of RNA segments. In this study, we have developed a method to prevent the free reassortment of influenza A virus RNAs by rewiring their packaging signals. Specific packaging signals for individual influenza virus RNA segments are located in the 5′ and 3′ noncoding regions as well as in the terminal regions of the ORF of an RNA segment. By putting the nonstructural protein (NS)-specific packaging sequences onto the ORF of the hemagglutinin (HA) gene and mutating the packaging regions in the ORF of the HA, we created a chimeric HA segment with the packaging identity of an NS gene. By the same strategy, we made an NS gene with the packaging identity of an HA segment. This rewired virus had the packaging signals for all eight influenza virus RNAs, but it lost the ability to independently reassort its HA or NS gene. A similar approach can be applied to the other influenza A virus segments to diminish their ability to form reassortant viruses.

Rewiring the RNAs of influenza virus to prevent reassortment. PNAS USA September 8 2009 doi:10.1073/pnas.0908897106

For a nice discussion of why this isn’t the answer to influenza pandemics, read What if influenza virus did not reassort? at the Virology Blog.

Related:

• The good news about influenza
• The Biology of Influenza
• How influenza pandemics get started

With the fall in H1N1 “swine flu” influenza cases recently, it has become fashionable for the media to run “What was all the fuss about” stories on the same page as “OMG, it’s going to be bad” stories. The problem with influenza is that it is one of the most unpredictable of all viruses, and while an upsurge in the number of cases can be expected as the winter flu season gets going in the northern hemisphere, the real concern is that this new pandemic virus might “turn nasty” in the second wave, just as in 1918 a much more pathogenic variant of that virus followed the relatively benign first wave of cases.

There are two ways in which this could happen. The first is that the present virus acquires spontaneous mutations which make it more pathogenic. The other possibility is that the virus recombines with a highly pathogenic influenza virus though the process known as reassortment – swapping of genes when two different strains infect the same cell. And there’s a good candidate for that out there – the highly pathogenic H5N1 avian influenza virus. Unlike H1N1, H5N1 has a hard time infecting humans, so it’s unlikely that these viruses would meet. But if they did…

The good news comes from Egypt, where H5N1 is relatively common, and a (worrying) case of H5N1/H1N1 co-infection was recently reported. The Ministry of Health has now discounted the rumour of a co-infection with the two viruses. In addition, a University of Maryland/NIH study suggests that co-infections of H1N1 with seasonal flu viruses do not produce chimeric or reassortant viruses. The H1N1 strain seems to outcompete seasonal viruses, possibly demonstrating this pandemic strain is not under biological pressure and is perhaps more efficiently communicable. Certainly, the past pattern seems to suggest that H1N1 pandemic seem to suppress outbreaks of other strains for some time.

A phase I clinical trial conducted by scientists from the University of Leicester tested 100 healthy volunteers with an H1N1 vaccine to see how their immune system responded. Trial leader Dr Iain Stephenson found 80 per cent of the volunteers showed a “strong, potentially protective” response after one dose, with more than 90 per cent showing the same response after two doses. The results suggest that one vaccine dose may be sufficient to protect against A(H1N1) swine flu, rather than two. Larger trials are now under way around the world involving up to more than 6,000 adults and children.

Reasons to be cheeful.

Related:

• H5N1 and 1918 pandemic influenza virus infection and the lungs
• Spotting pandemic influenza viruses
• How influenza pandemics get started
• The Pathogenicity of Pandemic Influenza Viruses
• The Great Flu – a free online game

This summer on MicrobiologyBytes we’ll be revisiting a few old favourites – some of the most popular posts on this site. Today’s post is:

The results of a new study have provided fresh insights into influenza pandemics by raising the possibility that all three pandemic influenza strains of the 20th century may have been generated through a series of multiple reassortment events and emerged over a period of years before pandemic recognition. The results indicate that each of these strains was produced by reassortment between the previously circulating human virus and at least one virus of animal origin. The novel gene segments for the H2N2/1957 and H3N2/1968 pandemics seem to have originated from avian hosts, but the zoonotic sources of the introduced viral gene segments for the 1918 pandemic remain ambiguous. However, evidence suggests that, over a number of years, avian gene virus segments have entered mammalian populations where the viruses may have undergone reassortment with the prevailing human virus. Given the frequent interspecies transmission of influenza viruses between swine and humans, it is most likely that such reassortment events occurred in pigs before pandemic emergence.

This work suggests that in the 1918 and 1957 pandemics, novel NA and internal genes may have been introduced into the prevailing human virus strains before the acquisition of the novel pandemic HA. Frequent detection of seasonal human influenza strains in pigs indicates that pandemic precursor viruses probably have circulated in either swine or human populations. The precursors to the H2N2 and H3N2 pandemics have not been detected, probably because they originated in Asia where little or no surveillance was conducted at that time.

If future pandemics arise in this manner, this interval may provide the best opportunity for health authorities to intervene to mitigate the effects of a pandemic or even to abort its emergence. However, the findings argue the need for highthroughput characterization of all 8 gene segments of human virus isolates, even those that have unremarkable HA antigens, particularly of human viruses isolated in hotspots for zoonotic infections with avian influenza viruses. At present, global influenza surveillance in humans focuses attention primarily on hemagglutinin. Although this focus will continue to be required for strain selection for seasonal influenza vaccines, our findings argue that this surveillance will not suffice for early warning of an incipient pandemic.

Dating the emergence of pandemic influenza viruses. PNAS USA July 13, 2009. doi: 10.1073/pnas.0904991106
Pandemic influenza viruses cause significant mortality in humans. In the 20th century, 3 influenza viruses caused major pandemics: the 1918 H1N1 virus, the 1957 H2N2 virus, and the 1968 H3N2 virus. These pandemics were initiated by the introduction and successful adaptation of a novel hemagglutinin subtype to humans from an animal source, resulting in antigenic shift. Despite global concern regarding a new pandemic influenza, the emergence pathway of pandemic strains remains unknown. Here we estimated the evolutionary history and inferred date of introduction to humans of each of the genes for all 20th century pandemic influenza strains. Our results indicate that genetic components of the 1918 H1N1 pandemic virus circulated in mammalian hosts, i.e., swine and humans, as early as 1911 and was not likely to be a recently introduced avian virus. Phylogenetic relationships suggest that the A/Brevig Mission/1/1918 virus (BM/1918) was generated by reassortment between mammalian viruses and a previously circulating human strain, either in swine or, possibly, in humans. Furthermore, seasonal and classic swine H1N1 viruses were not derived directly from BM/1918, but their precursors co-circulated during the pandemic. Mean estimates of the time of most recent common ancestor also suggest that the H2N2 and H3N2 pandemic strains may have been generated through reassortment events in unknown mammalian hosts and involved multiple avian viruses preceding pandemic recognition. The possible generation of pandemic strains through a series of reassortment events in mammals over a period of years before pandemic recognition suggests that appropriate surveillance strategies for detection of precursor viruses may abort future pandemics.

Related:

• How influenza pandemics get started
• The Pathogenicity of Pandemic Influenza Viruses

Laurie Garrett: What can we learn from the 1918 flu?

Larry Brilliant: Help stop the next pandemic

H5, H7, and H9 avian influenza subtypes top the World Health Organization’s list of strains with the greatest pandemic potential. A transition from avian-like 2,3-linked sialic acid (SA2,3) receptors to human-like 2,6-linked sialic acid (SA2,6) receptors appears to be a crucial step for avian influenza viruses to replicate efficiently and transmit in humans. An increasing number of contemporary avian H9N2 viruses contain leucine (L) at position 226 in the hemagglutinin (HA) receptor-binding site (RBS), supporting the preferential binding to SA2,6 receptors and the ability to replicate efficiently in human respiratory epithelial cells and in the ferret model, an in vivo model which closely resembles human airway epithelium and clinical infection. Since the mid-1990’s, H9N2 influenza viruses have become endemic in poultry throughout Eurasia and have occasionally transmitted to humans and pigs. In addition to possessing human virus-like receptor specificity, avian H9N2 viruses induce typical human flu-like illness, which can easily go unreported, and therefore have the opportunity to circulate, undergo reassortment, and increase in transmissibility.

Seroepidemiological studies in Asia suggest that the incidence of human H9N2 infections could be more prevalent than what has been reported and possible human-to-human transmission cannot be completely excluded. These direct infections with avian H9N2 confirm that interspecies transmission of H9N2 from avian species to mammalian hosts occurs and it is not uncommon. Reassortment between the current human epidemic strain and an avian virus of a different subtype is postulated to generate the next pandemic strain. Given the receptor specificity of avian H9N2 viruses and their repeated introduction into humans, as recent as December 2008, the opportunity for their reassortment and/or adaptation for human-to-human transmission is ever present. However the question remains what is missing for the H9N2 virus to transmit from human-to-human and possibly lead to the next pandemic.

A new study describes respiratory droplet transmission of an avian–human H9N2 influenza virus in ferrets and pinpoints the minimal changes necessary for respiratory droplet transmission in this model. After only 10 passages of nasal washes researchers were able to establish infection and sustain respiratory droplet transmission that was reproducible in multiple studies. This adaptation required only 5 amino acid changes in the entire influenza virus genome, implying that little is needed for currently circulating avian H9N2 viruses to transmit human-to-human following reassortment with a human strain. Studies to identify the minimal changes necessary indicated three changes in the surface HA and NA as the key point mutations essential for respiratory droplet transmission. The scientists also identified and located a change that dramatically alters the antigenicity of the virus, bringing to light the inherent limitations in the selection of vaccine seed stocks for avian H9N2 viruses and the possible inefficiency regarding the seed stock selection of other avian influenza strains. Whether these changes can affect transmission phenotypes of additional avian H9N2 strains and possibly other influenza subtypes, most notably H5 and H7, remains to be determined.

Minimal molecular constraints for respiratory droplet transmission of an avian–human H9N2 influenza A virus. PNAS USA April 20, 2009
Pandemic influenza requires interspecies transmission of an influenza virus with a novel hemagglutinin (HA) subtytpe that can adapt to its new host through either reassortment or point mutations and transmit by aerosolized respiratory droplets. Two previous pandemics of 1957 and 1968 resulted from the reassortment of low pathogenic avian viruses and human subtypes of that period; however, conditions leading to a pandemic virus are still poorly understood. Given the endemic situation of avian H9N2 influenza with human-like receptor specificity in Eurasia and its occasional transmission to humans and pigs, we wanted to determine whether an avian–human H9N2 reassortant could gain respiratory transmission in a mammalian animal model, the ferret. Here we show that following adaptation in the ferret, a reassortant virus carrying the surface proteins of an avian H9N2 in a human H3N2 backbone can transmit efficiently via respiratory droplets, creating a clinical infection similar to human influenza infections. Minimal changes at the protein level were found in this virus capable of respiratory droplet transmission. A reassortant virus expressing only the HA and neuraminidase (NA) of the ferret-adapted virus was able to account for the transmissibility, suggesting that currently circulating avian H9N2 viruses require little adaptation in mammals following acquisition of all human virus internal genes through reassortment. Hemagglutinin inhibition (HI) analysis showed changes in the antigenic profile of the virus, which carries profound implications for vaccine seed stock preparation against avian H9N2 influenza. This report illustrates that aerosolized respiratory transmission is not exclusive to current human H1, H2, and H3 influenza subtypes.

Related:

• The Biology of Influenza
• Influenza pandemic – when?
• The Pathogenicity of Pandemic Influenza Viruses
• H5N1 and 1918 pandemic influenza virus infection and the lungs