Assessment of Survival Kinetics for Emergent Highly Pathogenic Clade 2.3.4.4 H5Nx Avian Influenza Viruses.

High pathogenicity avian influenza viruses (HPAIVs) cause high morbidity and mortality in poultry species. HPAIV prevalence means high numbers of infected wild birds could lead to spill over events for farmed poultry. How these pathogens survive in the environment is important for disease maintenance and potential dissemination. We evaluated the temperature-associated survival kinetics for five clade 2.3.4.4 H5Nx HPAIVs (UK field strains between 2014 and 2021) incubated at up to three temperatures for up to ten weeks. The selected temperatures represented northern European winter (4 °C) and summer (20 °C); and a southern European summer temperature (30 °C). For each clade 2.3.4.4 HPAIV, the time in days to reduce the viral infectivity by 90% at temperature T was established (DT), showing that a lower incubation temperature prolonged virus survival (stability), where DT ranged from days to weeks. The fastest loss of viral infectivity was observed at 30 °C. Extrapolation of the graphical DT plots to the x-axis intercept provided the corresponding time to extinction for viral decay. Statistical tests of the difference between the DT values and extinction times of each clade 2.3.4.4 strain at each temperature indicated that the majority displayed different survival kinetics from the other strains at 4 °C and 20 °C.

Pathogenic assessment of avian influenza viruses in migratory birds.

ABSTRACTSeveral subtypes of avian influenza (AI) viruses have caused human infections in recent years; however, there is a severe knowledge gap regarding the capacity of wild bird viruses to infect mammals. To assess the risk of mammalian infection by AI viruses from their natural reservoirs, a panel of isolates from 34 wild birds was examined in animal models. All selected AI virus subtypes were found to predominantly possess Eurasian lineage, although reassortment with North American lineage AI viruses was also noted in some isolates. When used to infect chickens, 20 AI isolates could be recovered from oropharyngeal swabs at 5 days post-infection (dpi) without causing significant morbidity. Similarly, mild to no observable disease was observed in mice infected with these viruses although the majority replicated efficiently in murine lungs. As expected, wild bird AI isolates were found to recognize avian-like receptors, while a few strains also exhibited detectable human-like receptor binding. Selected strains were further tested in ferrets, and 15 out of 20 were found to shed the virus in the upper respiratory tract until 5 dpi. Overall, we demonstrate that a diversity of low-pathogenic AI viruses carried by wild migratory birds have the capacity to infect land-based poultry and mammalian hosts while causing minimal signs of clinical disease. This study reiterates that there is a significant capacity for interspecies transmission of AI viruses harboured by wild aquatic birds. Thus, these viruses pose a significant threat to human health underscoring the need for continued surveillance.

Quick assessment of influenza a virus infectivity with a long-range reverse-transcription quantitative polymerase chain reaction assay.

BACKGROUND: The polymerase chain reaction (PCR) is commonly used to detect viral pathogens because of its high sensitivity and specificity. However, conventional PCR methods cannot determine virus infectivity. Virus infectivity is conventionally examined with methods such as the plaque assay, even though such assays require several days. Long-range reverse-transcription quantitative PCR (RT-qPCR) has previously been suggested for the rapid assessment of RNA virus infectivity where the loss of infectivity is attributable to genomic fragmentation. METHODS: IAV was irradiated with 253.7 nm ultraviolet (UV) rays to induce genomic strand breaks that were confirmed by a full-length RT-PCR assay. The IAV was then subjected to plaque assay, conventional RT-qPCR and long-range RT-qPCR to examine the relationship between infectious titer and copy number. A simple linear regression analysis was performed to examine the correlation between the results of these assays. RESULTS: A long-range RT-qPCR assay was developed and validated for influenza A virus (IAV). Although only a few minutes of UV irradiation was required to completely inactivate IAV, genomic RNA remained detectable by the conventional RT-qPCR and the full-length RT-PCR for NS of viral genome following inactivation. A long-range RT-qPCR assay was then designed using RT-priming at the 3' termini of each genomic segment and subsequent qPCR of the 5' regions. UV-mediated IAV inactivation was successfully analyzed by the long-range RT-qPCR assay especially when targeting PA of the viral genome. This was also supported by the regression analysis that the long-range RT-qPCR is highly correlated with plaque assay (Adjusted R2 = 0.931, P = 0.000066). CONCLUSIONS: This study suggests that IAV infectivity can be predicted without the infectivity assays. The rapid detection of pathogenic IAV has, therefore, been achieved with this sensing technology.

The burden of influenza A and B in Mexico from the year 2010 to 2013: An observational, retrospective, database study, on records from the Directorate General of Epidemiology database.

Despite vaccination programs, influenza still represents a significant disease burden in Mexico. We conducted an observational, retrospective analysis to better understand the epidemiological situation of the influenza virus in Mexico. Analysis of the seasonal patterns of influenza A and B were based on the Directorate General of Epidemiology dataset of influenza-like illness(ILI), and severe acute respiratory infection(SARI) that were recorded between January 2010 and December 2013. Our objectives were 1) to describe influenza A and B activity, by age group, and subtype and, 2) to analyze the number of laboratory-confirmed cases presenting with ILI by influenza type, the regional distribution of influenza, and its clinical features. Three periods of influenza activity were captured: August 2010-January 2011, December 2011-March 2012, and October 2012-March 2013. Cases were reported throughout Mexico, with 50.3% (n = 10,320) of cases found in 18-49 year olds. Over the entire capture period, a total of 76,085 ILI/SARI episodes had swab samples analyzed for influenza, 27% were positive. During the same period, influenza A cases were higher in the 18-49 years old, and influenza B cases in both 5-17 and 18-49 age groups. Peak activity occurred in January 2012 (n = 4,159) and December 2012 (n = 348) for influenza A and B respectively. This analysis confirms that influenza is an important respiratory pathogen for children and adults in Mexico despite vaccination recommendations. School-age children and adolescents were more prone to influenza B infection; while younger adults were susceptible to both influenza A and B viruses. Over the seasons, influenza A and B co-circulated.

Mathematical modeling of postcoinfection with influenza A virus and Streptococcus pneumoniae, with implications for pneumonia and COPD-risk assessment.

BACKGROUND: The interaction between influenza and pneumococcus is important for understanding how coinfection may exacerbate pneumonia. Secondary pneumococcal pneumonia associated with influenza infection is more likely to increase respiratory morbidity and mortality. This study aimed to assess exacerbated inflammatory effects posed by secondary pneumococcal pneumonia, given prior influenza infection. MATERIALS AND METHODS: A well-derived mathematical within-host dynamic model of coinfection with influenza A virus and Streptococcus pneumoniae (SP) integrated with dose-response relationships composed of previously published mouse experimental data and clinical studies was implemented to study potentially exacerbated inflammatory responses in pneumonia based on a probabilistic approach. RESULTS: We found that TNFα is likely to be the most sensitive biomarker reflecting inflammatory response during coinfection among three explored cytokines. We showed that the worst inflammatory effects would occur at day 7 SP coinfection, with risk probability of 50% (likely) to develop severe inflammatory responses. Our model also showed that the day of secondary SP infection had much more impact on the severity of inflammatory responses in pneumonia compared to the effects caused by initial virus titers and bacteria loads. CONCLUSION: People and health care workers should be wary of secondary SP infection on day 7 post-influenza infection for prompt and proper control-measure implementation. Our quantitative risk-assessment framework can provide new insights into improvements in respiratory health especially, predominantly due to chronic obstructive pulmonary disease (COPD).

Preliminary Epidemiologic Assessment of Human Infections With Highly Pathogenic Avian Influenza A(H5N6) Virus, China.

BACKGROUND: Since 2014, 17 human cases of infection with the newly emerged highly pathogenic avian influenza A(H5N6) virus have been identified in China to date. The epidemiologic characteristics of laboratory-confirmed A(H5N6) cases were compared to A(H5N1) and A(H7N9) cases in mainland China. METHODS: Data on laboratory-confirmed H5N6, H5N1, and H7N9 cases identified in mainland China were analyzed to compare epidemiologic characteristics and clinical severity. Severity of confirmed H5N6, H5N1 and H7N9 cases was estimated based on the risk of severe outcomes in hospitalized cases. RESULTS: H5N6 cases were older than H5N1 cases with a higher prevalence of underlying medical conditions but younger than H7N9 cases. Epidemiological time-to-event distributions were similar among cases infected with the 3 viruses. In comparison to a fatality risk of 70% (30/43) for hospitalized H5N1 cases and 41% (319/782) for hospitalized H7N9 cases, 12 (75%) out of the 16 hospitalized H5N6 cases were fatal, and 15 (94%) required mechanical ventilation. CONCLUSION: Similar epidemiologic characteristics and high severity were observed in cases of H5N6 and H5N1 virus infection, whereas severity of H7N9 virus infections appeared lower. Continued surveillance of human infections with avian influenza A viruses remains an essential component of pandemic influenza preparedness.

Assessment of Effectiveness of Control Strategies Against Simulated Outbreaks of Highly Pathogenic Avian Influenza in Ontario, Canada.

The North American Animal Disease Spread Model (NAADSM) is a stochastic model framework developed to simulate the spread of highly contagious diseases of livestock and poultry, such as foot-and-mouth disease and highly pathogenic avian influenza (HPAI). The objective of this study was to make recommendations on the most effective HPAI control policy for Canada, specifically, on the effect of different speeds of detection, effectiveness of movement restrictions and stamping-out and ring-culling strategies on the magnitude of an HPAI outbreak. In addition, the effect of introduction of infection in a range of multiple farms simultaneously was also evaluated. A total of 21 060 scenarios, defined as different combinations of parameters for various epidemiological conditions and control measures, were created to simulate the number of poultry flocks that would become infected as a result of an incursion of HPAI. Each scenario was parameterized in NAADSM and replicated 1000 times, generating the median number of flocks infected at the end of the simulated outbreak for each scenario. Negative binomial regression analysis was used to model significant explanatory variables of the median number of flocks infected at the end of each simulated outbreak for each of the 21 060 scenarios. The final model included the following explanatory variables: number and type initially infected flock(s), density of flocks within the county where the initially infected flock(s) was located, probability of transmission through indirect contact, subclinical spread of the infection, speed of detection and a two-way interaction between intensity of bird destruction strategy and movement restriction effectiveness to reduce transmission through direct and indirect contacts. The modelling results suggested that stamping out of the detected infected flocks, without ring culling, in combination with effective movement restrictions on direct and indirect contacts, would be the most appropriate policy for Ontario.

Safe application of regionalization for trade in poultry and poultry products during highly pathogenic avian influenza outbreaks in the USA.

The 2014-2015 H5Nx high pathogenicity avian influenza (HPAI) outbreak affected 211 commercial premises, 21 backyard flocks, 75 individual wild birds and four captive-reared raptors in 21 Western and upper Midwestern states, resulting in death or culling of over 50.4 million poultry in the stamping-out programme that cost the US government $850 million. The outbreak had a negative $3.3 billion impact on the economy. Seventeen trading partners suspended imports of all US-origin poultry and poultry products while 38 trading partners regionalized the United States, and allowed trade in poultry and poultry products to continue from areas of the US not affected by HPAI. Disease response and control activities in addition to the use of comprehensive surveillance and regionalization (zoning) as prescribed by the OIE Terrestrial Animal Health Code are a scientifically valid and effective means to maintain safe trade in poultry and poultry products. This was further realized during the 2016 H7N8 HPAI outbreak in Dubois County, Indiana, with greater acceptance of regionalization and continuity in trade with a more limited cost of $30 million for eradication.

Viral factors in influenza pandemic risk assessment.

The threat of an influenza A virus pandemic stems from continual virus spillovers from reservoir species, a tiny fraction of which spark sustained transmission in humans. To date, no pandemic emergence of a new influenza strain has been preceded by detection of a closely related precursor in an animal or human. Nonetheless, influenza surveillance efforts are expanding, prompting a need for tools to assess the pandemic risk posed by a detected virus. The goal would be to use genetic sequence and/or biological assays of viral traits to identify those non-human influenza viruses with the greatest risk of evolving into pandemic threats, and/or to understand drivers of such evolution, to prioritize pandemic prevention or response measures. We describe such efforts, identify progress and ongoing challenges, and discuss three specific traits of influenza viruses (hemagglutinin receptor binding specificity, hemagglutinin pH of activation, and polymerase complex efficiency) that contribute to pandemic risk.

Risk Assessment of High Pathogenicity Avian Influenza Virus Introduction into Poland via Legal Importation of Live Poultry.

The risk of highly pathogenic avian influenza (HPAI) virus introduction via import of live poultry results from the probability that infected birds are exported from apparently HPAI-free areas during the silent phase of the epidemic, i.e., the period between an incursion of the virus into a susceptible population and a report on the outbreak by an exporting country. In our study we adapted a stochastic model, previously published in 2010 by Sánchez-Vizcaíno et al., with our own modifications in which the probability of HPAI introduction was assessed as the sum of the probabilities of entry of at least one infected bird from each susceptible species exported from each country into each Polish region (county). The mean annual probability of HPAI introduction into Poland via legal trade of live poultry was very low (3.07 × 10(-3), which corresponds to 1 outbreak every 326 yr). The highest risk was associated with the import of turkeys (62%) and chickens (33%). The exporting countries that contributed the most to the overall risk were Italy (31%), the Netherlands (24%), and the Czech Republic (17%). The risk was not evenly distributed across the country and it seemed higher in western, north-central, and eastern Poland while several counties of the north-west, central, or south-east parts of the country were at negligible risk. The applied model provides quantitative evidence that the risk of HPAI introduction through legal trade of poultry does not play a major role and that other paths, such as wild birds migrations or illegal trade, should be considered as the most-likely routes along which the virus can be introduced.

Assessment of the Potential Distance of Dispersal of High Pathogenicity Avian Influenza Virus by Wild Mallards.

This work presents the results of studies aimed at assessing the median and maximum distances covered by wild mallards (Anas platyrhynchos; n = 38), hypothetically infected with the high pathogenicity avian influenza virus (HPAIV) during spring migrations, using GPS-GSM tracking and published data on the susceptibility to HPAIV infection and duration of shedding. The model was based on the assumptions that the birds shed virus in the absence of clinical signs during infectious periods (IP) that were assumed to last 1 day (IP1), 4 days (IP4), and 8 days (IP8) and that each day of migration is a hypothetical day of the onset of IP. Using the haversine formula over a sliding timeframe corresponding to each IP, distances were estimated for each duck that undertook migration and then the maximum distance (Dmax) was selected. Ten mallards undertook spring migrations but, due to the loss of signal in the GPS-GSM devices, only three ducks were observed during autumn migrations. The following ranges of Dmax values were calculated for spring migrations: 124-382 km for IP1 (median 210 km), 208-632 km for IP4 (median 342 km), and 213-687 km for IP8 (median 370 km). The present study provides information that can be used as a data source to perform risk assessment related to the contribution of wild mallards in the dispersal of HPAIV over considerable distances.

Assessment of transmission, pathogenesis and adaptation of H2 subtype influenza viruses in ferrets.

After their disappearance from the human population in 1968, influenza H2 viruses have continued to circulate in the natural avian reservoir. The isolation of this virus subtype from multiple bird species as well as swine highlights the need to better understand the potential of these viruses to spread and cause disease in humans. Here we analyzed the virulence, transmissibility and receptor-binding preference of two avian influenza H2 viruses (H2N2 and H2N3) and compared them to a swine H2N3 (A/swine/Missouri/2124514/2006 [swMO]), and a human H2N2 (A/England/10/1967 [Eng/67]) virus using the ferret model as a mammalian host. Both avian H2 viruses possessed the capacity to spread efficiently between cohoused ferrets, and the swine (swMO) and human (Eng/67) viruses transmitted to naïve ferrets by respiratory droplets. Further characterization of the swMO hemagglutinin (HA) by x-ray crystallography and glycan microarray array identified receptor-specific adaptive mutations. As influenza virus quasispecies dynamics during transmission have not been well characterized, we sequenced nasal washes collected during transmission studies to better understand experimental adaptation of H2 HA. The avian H2 viruses isolated from ferret nasal washes contained mutations in the HA1, including a Gln226Leu substitution, which is a mutation associated with α2,6 sialic acid (human-like) binding preference. These results suggest that the molecular structure of HA in viruses of the H2 subtype continue to have the potential to adapt to a mammalian host and become transmissible, after acquiring additional genetic markers.

Economic analysis of HPAI control in the Netherlands I: epidemiological modelling to support economic analysis.

Economic analysis of control strategies for contagious diseases is a necessity in the development of contingency plans. Economic impacts arising from epidemics such as highly pathogenic avian influenza (HPAI) consist of direct costs (DC), direct consequential costs (DCC), indirect consequential costs (ICC) and aftermath costs (AC). Epidemiological models to support economic analysis need to provide adequate outputs for these critical economic parameters. Of particular importance for DCC, ICC and AC is the spatial production structure of a region. Spatial simulation models are therefore particularly suited for economic analysis; however, they often require a large number of parameters. The aims of this study are (i) to provide an economic rationale of epidemiological modelling in general, (ii) to provide a transparent description of the parameterization of a spatially based epidemiological model for the analysis of HPAI control in the Netherlands and (iii) to discuss the validity and usefulness of this model for subsequent economic analysis. In the model, HPAI virus transmission occurs via local spread and animal movements. Control mechanisms include surveillance and tracing, movement restrictions and depopulation. Sensitivity analysis of key parameters indicated that the epidemiological outputs with the largest influence on the economic impacts (i.e. epidemic duration and number of farms in the movement restriction zone) were more robust than less influential indicators (i.e. number of infected farms). Economically relevant outputs for strategy comparison were most sensitive to the relative role of the different transmission parameters. The default simulation and results of the sensitivity analysis were consistent with the general outcomes of known HPAI models. Comparison was, however, limited due to the absence of some economically relevant outputs. It was concluded that the model creates economically relevant, adequate and credible output for subsequent use in economic analysis. A detailed economic analysis is presented in a subsequent article.

Universal influenza vaccine: the holy grail?

Influenza vaccines have been available since the 1950s and have seen increasingly wide use as public health authorities expanded recommendations. Recent events including shortages and avian influenza outbreaks have renewed interest in influenza vaccines, particularly improved vaccines.

PATH Influenza Vaccine Project: accelerating the development of new influenza vaccines for low-resource countries.

The 2009 influenza A/H1N1 pandemic demonstrated that a pandemic influenza virus has the potential to spread more rapidly in today's highly interconnected world than in the past. While pandemic morbidity and mortality are likely to be greatest in low-resource countries, manufacturing capacity and access to influenza vaccines predominantly exist in countries with greater resources and infrastructure. Even with recently expanded manufacturing capacity, the number of doses available within a 6-month timeframe would be inadequate to fully immunize the global population if the decision to implement a global vaccination program were made today. Improved, affordable vaccines are needed to limit the consequences of a global influenza outbreak and protect low-resource populations. PATH's Influenza Vaccine Project is supporting a range of activities in collaboration with private- and public-sector partners to advance the development of promising influenza vaccines that can be accessible and affordable for people in low-resource countries.

Cost analysis of various low pathogenic avian influenza surveillance systems in the Dutch egg layer sector.

BACKGROUND: As low pathogenic avian influenza viruses can mutate into high pathogenic viruses the Dutch poultry sector implemented a surveillance system for low pathogenic avian influenza (LPAI) based on blood samples. It has been suggested that egg yolk samples could be sampled instead of blood samples to survey egg layer farms. To support future decision making about AI surveillance economic criteria are important. Therefore a cost analysis is performed on systems that use either blood or eggs as sampled material. METHODOLOGY/PRINCIPAL FINDINGS: The effectiveness of surveillance using egg or blood samples was evaluated using scenario tree models. Then an economic model was developed that calculates the total costs for eight surveillance systems that have equal effectiveness. The model considers costs for sampling, sample preparation, sample transport, testing, communication of test results and for the confirmation test on false positive results. The surveillance systems varied in sampled material (eggs or blood), sampling location (farm or packing station) and location of sample preparation (laboratory or packing station). It is shown that a hypothetical system in which eggs are sampled at the packing station and samples prepared in a laboratory had the lowest total costs (i.e. € 273,393) a year. Compared to this a hypothetical system in which eggs are sampled at the farm and samples prepared at a laboratory, and the currently implemented system in which blood is sampled at the farm and samples prepared at a laboratory have 6% and 39% higher costs respectively. CONCLUSIONS/SIGNIFICANCE: This study shows that surveillance for avian influenza on egg yolk samples can be done at lower costs than surveillance based on blood samples. The model can be used in future comparison of surveillance systems for different pathogens and hazards.

A qualitative approach to measure the effectiveness of active avian influenza virus surveillance with respect to its cost: a case study from Switzerland.

The aim of the project was to apply cost-effectiveness analysis to the economic appraisal of avian influenza virus (AIV) surveillance, using the implemented surveillance programme in Switzerland as a case study. First a qualitative risk assessment approach was used to assess the expected impact of surveillance on the transmission and spread of AIV. The effectiveness of surveillance was expressed as the difference in defined probabilities between a scenario with surveillance and a scenario without surveillance. The following probabilities were modelled (i) transmission of highly pathogenic AIV (HPAIV) from wild birds to poultry, (ii) mutation from low pathogenic AIV (LPAIV) into HPAIV in poultry, and (iii) transmission of HPAIV to other poultry holdings given a primary outbreak. The cost-effectiveness ratio was defined conventionally as the difference in surveillance costs (ΔC) divided by the change in probability (ΔP), the technical objective, on the presumption that surveillance diminishes the respective probabilities. However, results indicated that surveillance in both wild birds and poultry was not expected to change the probabilities of primary and secondary AIV outbreaks in Switzerland. The overall surveillance costs incurred were estimated at 31,000 €/year, which, to be a rational investment of resources, must still reflect the value policy makers attribute to other benefits from having surveillance (e.g. peace of mind). The advantage of the approach adopted is that it is practical, transparent, and thus able to clarify for policy makers the key variables to be taken into account when evaluating the economic efficiency of resources invested in surveillance, prevention and intervention to exclude AIV.