Avian Influenza Virus Outbreaks Research Articles

Highly pathogenic H5N1 avian influenza virus outbreak in cattle: the knowns and unknowns.

Highly pathogenic H5N1 avian influenza virus outbreak in cattle: the knowns and unknowns.

Potential use of poultry farms by wild waterfowl in California's Central Valley varies across space, times of day, and species: implications for influenza transmission risk

Interactions between wildlife and livestock can lead to cross‐species disease transmission, which incurs economic costs and threatens wildlife conservation. Wild waterfowl are natural hosts of avian influenza viruses (AIVs), are often abundant near poultry farms, and have been linked to outbreaks of AIVs in poultry. Interspecific and seasonal variation in waterfowl movement and habitat use means that the risk of disease transmission between wild birds and poultry inevitably varies across species, space, and time. Here, we used GPS telemetry data from 10 waterfowl species in and near California's Central Valley, a region where both wild waterfowl and domestic poultry are abundant, to study selection of poultry farms by waterfowl across diel, seasonal, and annual cycles. We found that waterfowl selected for wetlands, open water, protected areas, and croplands, which meant that they generally avoided habitats that were likely to be used for poultry farming. These selection patterns were linked to species' ecology and diel behavioral patterns, such that avoidance of poultry habitats was stronger for local or partial migrants than for long‐distance migrants, and stronger during daytime than at night. We then combined these habitat selection results with data on poultry farm locations to map risk of waterfowl–poultry contact across the Central Valley. Average selection strength at poultry farms was low, suggesting that current placement of poultry farms is generally effective for limiting risk of contact with wild birds. When we combined these habitat selection results with data on species' abundances and AIV infection prevalence, we found dramatic variation in potential AIV transmission risk among species. These results could be used to prioritize surveillance and biosecurity efforts for regions and times of relatively high risk. More generally, these results highlight that fine‐scale movement data can help identify interspecific, seasonal, and diel patterns in animal behaviors that affect wildlife and poultry health.

HPAIV outbreak triggers short-term colony connectivity in a seabird metapopulation

Disease outbreaks can drastically disturb the environment of surviving animals, but the behavioural, ecological, and epidemiological consequences of disease-driven disturbance are poorly understood. Here, we show that an outbreak of High Pathogenicity Avian Influenza Virus (HPAIV) coincided with unprecedented short-term behavioural changes in Northern gannets (Morus bassanus). Breeding gannets show characteristically strong fidelity to their nest sites and foraging areas (2015–2019; n = 120), but during the 2022 HPAIV outbreak, GPS-tagged gannets instigated long-distance movements beyond well-documented previous ranges and the first ever recorded visits of GPS-tagged adults to other gannet breeding colonies. Our findings suggest that the HPAIV outbreak triggered changes in space use patterns of exposed individuals that amplified the epidemiological connectivity among colonies and may generate super-spreader events that accelerate disease transmission across the metapopulation. Such self-propagating transmission from and towards high density animal aggregations may explain the unexpectedly rapid pan-European spread of HPAIV in the gannet.

Optimizing environmental viral surveillance: bovine serum albumin increases RT-qPCR sensitivity for high pathogenicity avian influenza H5Nx virus detection from dust samples

ABSTRACT The latest outbreaks of high pathogenicity avian influenza viruses (HPAIVs) were the most widespread ever seen in Europe, entailing considerable economic costs and raising public health concerns. Virological surveillance protocols involve reverse transcription quantitative polymerase chain reactions (RT-qPCR) from tracheal and cloacal swabs, which are laborious to perform on a large scale and require special skills. Environmental sampling, and especially dust collection, may represent a relevant alternative, as it is cheap, non-invasive for animals, simpler, and quicker to carry out. The main drawback of dust samples is that they may contain high amounts of organic substances that can inhibit RT-qPCR reactions. Bovine serum albumin (BSA) is a molecule known to facilitate DNA polymerization in the presence of numerous inhibitors, including those from feces, litter, or food. We tested its use on dust samples collected on 107 farms localized in areas affected by epizootics of clade 2.3.4.4b HPAIV H5N1. We used a latent class modeling approach to compare the performance of three detection protocols: (i) BSA addition to the RT-qPCR reaction mix, (ii) dilution of template RNA, and (iii) a control protocol. Our results indicate that BSA addition to the RT-qPCR reaction mix improved the sensitivity of the method, by neutralizing inhibitors’ effect. Indeed, for hemagglutinin (HA) or matrix (M) RNA detection, the sensitivity of the BSA protocol was the highest, followed by that of the control protocol, and that of the dilution protocol. Our results suggest that the use of BSA could be routinely implemented in HPAIV dust monitoring RT-qPCR protocols. IMPORTANCE With the circulation of high pathogenicity avian influenza viruses having intensified considerably in recent years, the European Union is considering the vaccination of farmed birds. A prerequisite for this vaccination is the implementation of drastic surveillance protocols. Environmental sampling is a relevant alternative to animal sampling. However, environmental samples often contain inhibitory compounds in large enough quantities to inhibit RT-qPCR reactions. As bovine serum albumin is a molecule used in many fields to overcome this inhibitory effect, we tested its use on dust samples from poultry farms in areas heavily affected by HPAIV epizootics. Our results show that its use significantly increases the sensitivity of the method.

Control strategies for 2.3.4.4 highly pathogenic avian influenza virus - From vaccines to host disease resistance. Proceeding of The First International Avian Influenza Summit, University of Arkansas- October 16-17, 2023"

Avian influenza virus (AIV) is a highly contagious and lethal disease that can have major impacts on the global poultry industry and food supply. While more frequent in Asia and Europe, highly pathogenic avian influenza virus (HPAIV) outbreaks have traditionally been rare in the U.S. However, in recent years, the U.S. has seen an increase in the incidence of HPAIV outbreaks in wild birds and commercial poultry. In 2014/2015 and in 2021/2022, outbreaks of HPAIV subtype H5NX clade 2.3.4.4 resulted in the death and destruction of over 50 million birds, each costing billions of dollars to the U.S. economy. Here, we present a comprehensive overview of control strategies against the 2.3.4.4 HPAI virus, encompassing both vaccination approaches and host-driven disease resistance mechanisms. The first section outlines the advancements in vaccine development tailored specifically for the 2.3.4.4 HPAI subtype. Through a critical analysis of experimental studies and field trials, we discuss the efficacy, safety, and cross-protection capabilities of various vaccine formulations, including inactivated whole-virus vaccines, vector-based platforms, and novel recombinant technologies. Furthermore, this section explores the challenges associated with vaccine design, including strain diversity, antigenic differences, and logistical hurdles in large-scale vaccination campaigns. The second section delves into host-centric strategies aimed at enhancing natural resistance against 2.3.4.4 HPAI virus infections. Because vaccines are not currently approved for use in the U.S., control strategies for HPAIV are dependent on biosecurity and culling of infected flocks. New strategies for HPAIV control based on gene editing of poultry species could offer solutions for disease control. Two different strategies for improving disease resistance to HPAIV are based on enhancing the innate immunity of the bird and targeting the viral RNA genome inside the cell. By harnessing genetic engineering techniques, including CRISPR/Cas-9 or -13, natural resistance to avian influenza can be achieved in poultry. Taken together, we will examine the current basis of control strategies for the 2.3.4.4 HPAI virus, offering insights into the strengths and limitations of both vaccination and novel gene-editing technologies to enhance host disease resistance to HPAI. By expanding our toolbox, this work contributes to the ongoing efforts to mitigate the impact of 2.3.4.4 HPAI on global poultry industries and public health.

Can Citizen Science Contribute to Avian Influenza Surveillance?

Global change is an important driver of the increase in emerging infectious diseases in recent decades. In parallel, interest in nature has increased, and different citizen science platforms have been developed to record wildlife observations from the general public. Some of these platforms also allow registering the observations of dead or sick birds. Here, we test the utility of live, sick and dead observations of birds recorded on the platform Observation.org for the early detection of highly pathogenic avian influenza virus (HPAIV) outbreaks in the wild in Belgium and The Netherlands. There were no significant differences in the morbidity/mortality rate through Observation.org one to four weeks in advance. However, the results show that the HPAIV outbreaks officially reported by the World Organisation for Animal Health (WOAH) overlapped in time with sudden increases in the records of sick and dead birds in the wild. In addition, in two of the five main HPAIV outbreaks recorded between 2016 and 2021, wild Anseriformes mortality increased one to two months before outbreak declaration. Although we cannot exclude that this increase was related to other causes such as other infectious diseases, we propose that Observation.org is a useful nature platform to complement animal health surveillance in wild birds. We propose possible approaches to improve the utility of the platform for pathogen surveillance in wildlife and discuss the potential for HPAIV outbreak detection systems based on citizen science to complement current surveillance programs of health authorities.

Strong breeding colony fidelity in northern gannets following high pathogenicity avian influenza virus (HPAIV) outbreak

High pathogenicity avian influenza virus (HPAIV) caused the worst seabird mass-mortalities in Europe across 2021–2022. The northern gannet (Morus bassanus) was one of the most affected species, with tens of thousands of casualties in the northeast Atlantic between April–September 2022. Disease outbreaks can modify the movement ecology of animals by diminishing spatial consistency, thereby increasing the potential for disease transmission. To detect potential changes in movement behaviour, we GPS-tracked breeding adults following the initial HPAIV outbreak, at three of the largest northern gannet breeding colonies where major mortality of adults and chicks occurred (Bass Rock, Scotland, UK; Grassholm, Wales, UK; Rouzic, Brittany, France). We also gathered background epidemiological information and northern gannet colony dynamics during the outbreak. Our data indicate that HPAIV killed at least 50 % of northern gannets, and suggest the presence of HPAIV H5N1 antibodies in juveniles. GPS-tracked adult northern gannets remained faithful to their breeding sites despite the HPAIV outbreak and did not prospect other breeding colonies. They performed regular foraging trips at sea, similar to their behaviour before the outbreak. Comparison with GPS-tracking data gathered in 2019, i.e. before the HPAIV outbreak, suggested lower foraging effort in birds which survived HPAIV in 2022, potentially as a consequence of reduced intra- and interspecific food competition. Breeding colony fidelity of surviving adult northern gannets following HPAIV mass-mortalities indicates limited capacity for viral spread during our study. This may contrast with the behaviour of adults during the initial disease outbreak, and with that of younger individuals.

Investigating the Genetic Diversity of H5 Avian Influenza Viruses in the United Kingdom from 2020-2022.

Since 2020, the United Kingdom and Europe have experienced annual epizootics of high-pathogenicity avian influenza virus (HPAIV). The first epizootic, during the autumn/winter of 2020-2021, involved six H5Nx subtypes, although H5N8 HPAIV dominated in the United Kingdom. While genetic assessments of the H5N8 HPAIVs within the United Kingdom demonstrated relative homogeneity, there was a background of other genotypes circulating at a lower degree with different neuraminidase and internal genes. Following a small number of detections of H5N1 in wild birds over the summer of 2021, the autumn/winter of 2021-2022 saw another European H5 HPAIV epizootic that dwarfed the prior epizootic. This second epizootic was dominated almost exclusively by H5N1 HPAIV, although six distinct genotypes were defined. We have used genetic analysis to evaluate the emergence of different genotypes and proposed reassortment events that have been observed. The existing data suggest that the H5N1 viruses circulating in Europe during late 2020 continued to circulate in wild birds throughout 2021, with minimal adaptation, but then went on to reassort with AIVs in the wild bird population. We have undertaken an in-depth genetic assessment of H5 HPAIVs detected in the United Kingdom over two winter seasons and demonstrate the utility of in-depth genetic analyses in defining the diversity of H5 HPAIVs circulating in avian species, the potential for zoonotic risk, and whether incidents of lateral spread can be defined over independent incursions of infections from wild birds. This provides key supporting data for mitigation activities. IMPORTANCE High-pathogenicity avian influenza virus (HPAIV) outbreaks devastate avian species across all sectors, having both economic and ecological impacts through mortalities in poultry and wild birds, respectively. These viruses can also represent a significant zoonotic risk. Since 2020, the United Kingdom has experienced two successive outbreaks of H5 HPAIV. While H5N8 HPAIV was predominant during the 2020-2021 outbreak, other H5 subtypes were also detected. The following year, there was a shift in the subtype dominance to H5N1 HPAIV, but multiple H5N1 genotypes were detected. Through the thorough utilization of whole-genome sequencing, it was possible to track and characterize the genetic evolution of these H5 HPAIVs in United Kingdom poultry and wild birds. This enabled us to assess the risk posed by these viruses at the poultry-wild bird and the avian-human interfaces and to investigate the potential lateral spread between infected premises, a key factor in understanding the threat to the commercial sector.

Phosphatidylcholine and cholesterol-loaded-cyclodextrins can replace egg yolk in the cryopreservation of equine spermatozoa

In the horse breeding industry, cryopreservation of spermatozoa has become an integral aspect of modern-day protocols. To avoid sperm damage during the freezing process, freezing media containing egg yolk are used. Recently, urgency to find an alternative to egg yolk has been increased due to the prevalence of Avian Influenza Virus (AIV). In the UK, the World Organization for Animal Health reports that between July and December 2022, an AIV outbreak led to the death of 4.62 million birds (56.73 million globally), with substantial consequences for agriculture, environment and egg availability (WAHIS. 2023; https://wahis.woah.org/#/dashboard/qd-dashboard: Accessed: 12 January 2023). Recent research has also called into question the biosafety of egg yolk, with RNA from highly pathogenic and low pathogenic AIV found on and in eggs (Stephens.et.al. Avian Diseases. 2020; 64:143-148). The cryoprotectant potential of phosphatidylcholine, the main phospholipid in egg yolk, has been investigated in diluents used for cryopreservation of human spermatozoa with some success (Sicchieri.et.al. Translational Andrology and Urology. 2021; 10:397-407). The chemically defined origin and increased biosafety make phosphatidylcholine a desirable alternative to egg yolk for use in cryopreservation. Ejaculates from four horses (n=4) were frozen using four different freezing extenders: a reference medium containing egg yolk and 3% dimethylformamide (control) and media without egg yolk supplemented with 1.5mg/ml2-hydroxypropyl-β-cyclodextrin cholesterol complex, 3% dimethylformamide and either 2% (P1), 4% (P2) or 6% phosphatidylcholine (P3). Samples were diluted and centrifuged on a density gradient medium (1000xg’ for 20 minutes). The cell band was aspirated and resuspended (200 × 10⁶ cells/ml) with allotted treatment extender. Samples were packed into 0.25ml straws, cooled to 5˚C for 25 minutes, then frozen using a programmable freezing unit (IceCube14s, Minitube, Germany). Temperature of the straws was reduced from 4 ˚C to -143˚C in 9 minutes, after which they were plunged directly into liquid nitrogen (-196˚C). Viability was assessed using NucleoCounter® NC-100™. Total motility analyses were performed with computer assisted semen analysis (Hamilton Thorne, USA). No difference in post-thaw viability was found between any of the media tested (P>0.05). No difference in total motility was found between the control (36.75±1.93σM), P1 (29.25±3.82σM) or P2 media (19.5±6.35σM) (P>0.05). However, post-thaw total motility of sperm cryopreserved in P3 media (16±6.01σM) was lower than the control (P<0.05). Media supplemented with 2-4% phosphatidylcholine and 1.5mg/ml cholesterol was effective in limiting loss of total motility and membrane integrity of equine spermatozoa, establishing the need for further breeding trials into the use of phospholipids for egg yolk lipid replacement to improve biosafety. The authors wish to recognize the support of Tullis Matson and Stallion AI Services Ltd in this study.

Exploring Culinary Films as a Form of Health Literacy: A Case Study of Aruna dan Lidahnya in Indonesia

The lack of Indonesian culinary-themed films that discuss public health literacy is reflected in the limited number of titles available. The film, Aruna dan Lidahnya, is a topic of discussion as it raises issues related to culinary tourism, religion, public policy, and public health literacy, specifically the avian influenza virus outbreak, which has objective and subjective dimensions. This study aims to discover how culinary films are constructed in health literacy and why the film is constructed as a form of health literacy. This qualitative study uses a case study approach to investigate how and why culinary films are constructed as a form of health literacy, specifically focusing on the film Aruna dan Lidahnya. The researchers used primary and secondary data from the film from literature studies and observations and applied data analysis techniques to interpret the data. The film Aruna dan Lidahnya promotes the potential of local cuisine as a unique attraction to grow the economy of tourist destinations. It also emphasizes the importance of health communication in raising public awareness about the avian influenza virus and following health protocols. The film highlights the need for cultural and local wisdom-based tourism development for a more ethical and aesthetic tourist attraction. Aruna dan Lidahnya is a film that highlights the potential of local cuisine in developing tourist destinations in four cities affected by the avian influenza virus. Developing tourist destinations based on local cuisine can improve social welfare and the area's economy and be a unique attraction for cultural tourism. The film also emphasizes the importance of health communication, following health protocols, and spreading awareness about the avian influenza virus in the context of local culinary opportunities.

CRISPR, Quorum Sensing, and Arbitrium as strategic control measures for avian influenza virus. Proceeding of The First International Avian Influenza Summit, University of Arkansas- October 16-17, 2023”

Avian Influenza Virus (AIV) poses a significant threat to both avian populations and human health due to its potential for zoonotic transmission. The emergence of highly pathogenic strains necessitates the development of innovative control strategies. This abstract presents a comprehensive overview of the potential of CRISPR-based technologies, Quorum Sensing (QS) systems, and the application of the synthetic inducer molecule Arbitrium as strategic control measures for mitigating AIV outbreaks. CRISPR-based technologies have revolutionized genetic engineering and hold immense promise in combating AIV. By leveraging the adaptive immune system of prokaryotes, CRISPR offers precise and targeted gene editing capabilities. This enables the creation of AIV-resistant poultry strains by introducing specific antiviral sequences, thereby reducing viral transmission within avian populations. Quorum Sensing, a cell-to-cell communication mechanism utilized by bacteria, can be harnessed to modulate AIV infectivity. By engineering QS systems to detect AIV-specific molecular signatures, it becomes possible to trigger population-wide responses in avian hosts. This includes activating immune responses and initiating antiviral signaling cascades, effectively conferring enhanced resistance to AIV infection. The incorporation of Arbitrium, a synthetic signaling molecule, represents a novel approach to controlling AIV. By engineering avian hosts to recognize Arbitrium, it is possible to orchestrate precise temporal and spatial responses to viral challenges. This programmable control system empowers the host organism to mount robust antiviral defenses upon exposure to AIV, providing an additional layer of protection. Furthermore, a synergistic approach integrating CRISPR, QS, and Arbitrium technologies holds the potential to create a comprehensive and adaptable defense system against AIV. This multi-pronged strategy addresses viral challenges at multiple levels, from genetic modification to population-wide immune responses, thereby bolstering the overall resilience of avian populations to AIV outbreaks. In conclusion, integrating CRISPR-based technologies, Quorum Sensing, and Arbitrium signaling represents a promising avenue for developing strategic control measures against Avian Influenza Virus. The synergistic application of these technologies offers precise and targeted genetic modifications and enables dynamic, population-level responses to AIV challenges. This innovative approach holds significant potential for revolutionizing AIV control strategies and safeguarding both avian and human health.

Airborne Avian Influenza Virus in Ambient Air in the Winter Habitats of Migratory Birds.

Outbreaks of avian influenza virus (AIV) have raised public concerns recently. Airborne AIV has been evaluated in live poultry markets and case farms; however, no study has discussed airborne AIV in ambient air in the winter habitats of migratory birds. Therefore, this study aimed to evaluate airborne AIV, specifically H5, H7, and H9, in a critical winter habitat of migratory birds and assess the factors influencing airborne AIV transmission in ambient air to provide novel insights into the epidemiology of avian influenza. A total of 357 ambient air samples were collected in the Aogu Wetland, Taiwan, Republic of China, between October 2017 and December 2019 and analyzed using quantitative real-time polymerase chain reaction. The effects of environmental factors including air pollutants, meteorological factors, and the species of the observed migratory birds on the concentration of airborne AIV were also analyzed. To our knowledge, this is the first study to investigate the relationship between airborne AIV in ambient air and the influence factors in the winter habitats of migratory birds, demonstrating the benefits of environmental sampling for infectious disease epidemiology. The positive rate of airborne H7 (12%) was higher than that of H5 (8%) and H9 (10%). The daily mean temperature and daily maximum temperature had a significant negative correlation with influenza A, H7, and H9. Cold air masses and bird migration were significantly associated with airborne H9 and H7, respectively. In addition, we observed a significant correlation between AIV and the number of pintails, common teals, Indian spot-billed ducks, northern shovelers, Eurasian wigeons, tufted ducks, pied avocets, black-faced spoonbills, and great cormorants. In conclusion, we demonstrated the potential for alternative surveillance approaches (monitoring bird species) as an indicator for influenza-related risks and identified cold air masses and the presence of specific bird species as potential drivers of the presence and/or the airborne concentration of AIV.

Habitat loss exacerbates pathogen spread: An Agent-based model of avian influenza infection in migratory waterfowl

Habitat availability determines the distribution of migratory waterfowl along their flyway, which further influences the transmission and spatial spread of avian influenza viruses (AIVs). The extensive habitat loss in the East Asian-Australasian Flyway (EAAF) may have potentially altered the virus spread and transmission, but those consequences are rarely studied. We constructed 6 fall migration networks that differed in their level of habitat loss, wherein an increase in habitat loss resulted in smaller networks with fewer sites and links. We integrated an agent-based model and a susceptible-infected-recovered model to simulate waterfowl migration and AIV transmission. We found that extensive habitat loss in the EAAF can 1) relocate the outbreaks northwards, responding to the distribution changes of wintering waterfowl geese, 2) increase the outbreak risk in remaining sites due to larger goose congregations, and 3) facilitate AIV transmission in the migratory population. In addition, our modeling output was in line with the predictions from the concept of “migratory escape”, i.e., the migration allows the geese to “escape” from the location where infection risk is high, affecting the pattern of infection prevalence in the waterfowl population. Our modeling shed light on the potential consequences of habitat loss in spreading and transmitting AIV at the flyway scale and suggested the driving mechanisms behind these effects, indicating the importance of conservation in changing spatial and temporal patterns of AIV outbreaks.

Rapid, in-field deployable, avian influenza virus haemagglutinin characterisation tool using MinION technology

Outbreaks of avian influenza virus (AIV) from wild waterfowl into the poultry industry is of upmost significance and is an ongoing and constant threat to the industry. Accurate surveillance of AIV in wild waterfowl is critical in understanding viral diversity in the natural reservoir. Current surveillance methods for AIV involve collection of samples and transportation to a laboratory for molecular diagnostics. Processing of samples using this approach takes more than three days and may limit testing locations to those with practical access to laboratories. In potential outbreak situations, response times are critical, and delays have implications in terms of the spread of the virus that leads to increased economic cost. This study used nanopore sequencing technology for in-field sequencing and subtype characterisation of AIV strains collected from wild bird faeces and poultry. A custom in-field virus screening and sequencing protocol, including a targeted offline bioinformatic pipeline, was developed to accurately subtype AIV. Due to the lack of optimal diagnostic MinION packages for Australian AIV strains the bioinformatic pipeline was specifically targeted to confidently subtype local strains. The method presented eliminates the transportation of samples, dependence on internet access and delivers critical diagnostic information in a timely manner.

Emergence of clade 2.3.4.4b novel reassortant H5N1 high pathogenicity avian influenza virus in South Korea during late 2021.

High pathogenicity H5N1 avian influenza viruses pose a threat to both animal and human health worldwide. In late 2020, outbreaks of H5 high pathogenicity avian influenza viruses belonging to clade 2.3.4.4b emerged in Europe, following on from outbreaks in East Asia in earlier years. However, very recent studies show that clade 2.3.4.4b H5N1, rather than 2.3.4.4b H5N8, has become predominant in wild birds and has infected poultry in several countries. In this study, we describe isolation of a novel H5N1 virus from a captured mandarin duck in South Korea, and another H5N1 virus from a quail farm. We performed genetic analysis of these two viruses to identify their origin and to determine their relationship with the clade 2.3.4.4b H5N1 viruses currently circulating in Europe. Based on our results, it is presumed that the novel H5N1 virus isolated in Korea originated from an unknown reassortant between clade 2.3.4.4b H5N8 viruses circulating from 2020 and other Eurasian viruses, with additional reassortment of genes and point mutations that discriminate them from the recently reported H5N1 virus in Europe.

Chicken meat taste preferences, perceived risk of human infection with avian influenza virus, and self-reported chicken meat consumption in China

Chinese consumers often prefer to buy live chickens for cooking at home rather than chilled or frozen chicken meat because of the superior flavor. Even though the purchase of live chickens for consumption was restricted by the Chinese government after the avian influenza (AI) virus outbreak in 2013, people continue to buy live chickens. Thus, it is important to understand why people make this decision. The goal of this study was to investigate the relationships among chicken meat taste preferences, perceived risk of human infection with AI virus, and chicken meat consumption behaviors in China. To address this, a survey of 1108 individuals was carried out in Jiangsu Province. This province was selected because its per capita consumption of poultry products is higher than the national average, and it is at the highest risk for avian influenza A H7N9 and H5N1 virus epidemics out of all provinces in China. Descriptive statistics, multivariate linear regression, and hierarchical regression modeling were used to analyze the data. The results showed that Chinese consumers still buy live chickens at a high rate due to taste preferences, and have a very low perceived risk of infection with AI virus. Multivariate linear and hierarchical regression analyses showed that total monthly household income was significantly and positively associated with the perceived importance of chicken taste; the number of people in the household and the perceived importance of chicken taste were significantly and positively associated with the purchase of live chickens for consumption; age was significantly and negatively associated with the purchase of chilled and frozen chicken for consumption; and education and expertize in livestock-related fields were significantly and positively associated with the purchase of live chickens for consumption. Expertize in livestock-related fields was significantly and positively associated with perceived risk of human infection with AI virus, and perceived risk was significantly and positively associated with the purchase of chilled chicken for consumption. However, the perceived risk of human infection with AI virus had no effect on the relationship between the perceived importance of chicken taste and the purchase of live chickens for consumption. The findings from this study suggest that, over time and with increasing public education, the purchase of live poultry for consumption will gradually decline in China. However, to accelerate this process, measures should be taken to reduce consumer perception of taste differences between chilled chicken meat and meat from freshly killed chickens. In addition, educating consumers about livestock to enhance their risk perception could also be another effective means of promoting the purchase of chilled and frozen chicken meat.

Host Correlates of Avian Influenza Virus Infection in Wild Waterfowl of the Sacramento Valley, California.

Avian influenza viruses (AIVs) are distributed globally in members of the family Anatidae (waterfowl), and significant disease may occur when these viruses infect commercial poultry or humans. Early detection of AIV through surveillance of wild waterfowl is one measure to prevent future disease outbreaks. Surveillance efforts that are designed to account for host and environmental determinants of susceptibility to infection are likely to be most effective. However, these determinants have not been clearly delineated and may vary with location. Because some regions are at greater risk for AIV outbreaks, the factors that contribute to AIV infection of waterfowl in these areas are of interest. We investigated the prevalence of AIVs in hunter-killed waterfowl at wintering sites in California's Central Valley. Overall, AIV prevalence was 10.5% and, after controlling for age and sex, was greatest in northern shovelers (Spatula clypeata) and lowest in wood ducks (Aix sponsa). Overall, AIV prevalence was higher in females than in males, but this trend was driven by one sampling year and one waterfowl species (green-winged teal, Anas crecca). AIV prevalence in waterfowl was lower in samples collected from brackish wetlands compared with those collected from freshwater wetlands, suggesting that wetland type or other environmental factors contribute to AIV prevalence. This study adds to our understanding of the ecology of AIV infection in waterfowl and may assist in developing more efficient, targeted surveillance efforts for the detection of potentially harmful viruses circulating in North American waterfowl.

Characterization and use in neutralization assays of avian influenza codon-optimized H5 and H7 retroviral pseudotypes

Influenza is a relevant problem for public and animal health, with a significant economic impact. In recent years, outbreaks of avian influenza virus have resulted in devastating losses in the poultry industry worldwide, and although its transmission to humans is very rare, there is always a potential risk for an even more severe outbreak. Currently, vaccination is considered the most effective tool for the control and prevention of influenza infections in both humans and animals. The maintenance of animal welfare and the successful implementation of animal health programs depend on the timely administration of vaccines, which must comply with quality specifications indicated by health authorities; for example, the capability to ensure a minimum antibody titer. The production of viral antigens used in these tests can pose a biosafety risk, and some viral strains can be difficult to grow. Therefore, new biotechnological alternatives are required to overcome these disadvantages. In this study, we produced pseudotypes carrying H5 and H7 hemagglutinins from lowly and highly pathogenic avian influenza viruses. These pseudotypes were used in neutralization assays to detect neutralizing antibodies in avian sera, which were confirmed positive by inhibition of the hemagglutination test. Our results showed that the pseudotype neutralization assay is a viable alternative for the detection of neutralizing antibodies, by demonstrating subtype specificity and requiring reduced biosafety requirements. Therefore, it represents a versatile platform that can facilitate technology transfer protocols between laboratories, and an immediate application in serological tools for quality control of veterinary vaccines against avian influenza.

Two-stage algorithms for visually exploring spatio-temporal clustering of avian influenza virus outbreaks in poultry farms

The development of visual tools for the timely identification of spatio-temporal clusters will assist in implementing control measures to prevent further damage. From January 2015 to June 2020, a total number of 1463 avian influenza outbreak farms were detected in Taiwan and further confirmed to be affected by highly pathogenic avian influenza subtype H5Nx. In this study, we adopted two common concepts of spatio-temporal clustering methods, the Knox test and scan statistics, with visual tools to explore the dynamic changes of clustering patterns. Since most (68.6%) of the outbreak farms were detected in 2015, only the data from 2015 was used in this study. The first two-stage algorithm performs the Knox test, which established a threshold of 7 days and identified 11 major clusters in the six counties of southwestern Taiwan, followed by the standard deviational ellipse (SDE) method implemented on each cluster to reveal the transmission direction. The second algorithm applies scan likelihood ratio statistics followed by AGC index to visualize the dynamic changes of the local aggregation pattern of disease clusters at the regional level. Compared to the one-stage aggregation approach, Knox-based and AGC mapping were more sensitive in small-scale spatio-temporal clustering.

Genetic Characterization and Pathogenesis of Three Novel Reassortant H5N2 Viruses in South Korea, 2018.

The outbreaks of H5N2 avian influenza viruses have occasionally caused the death of thousands of birds in poultry farms. Surveillance during the 2018 winter season in South Korea revealed three H5N2 isolates in feces samples collected from wild birds (KNU18-28: A/Wild duck/South Korea/KNU18-28/2018, KNU18-86: A/Bean Goose/South Korea/KNU18-86/2018, and KNU18-93: A/Wild duck/South Korea/KNU18-93/2018). Phylogenetic tree analysis revealed that these viruses arose from reassortment events among various virus subtypes circulating in South Korea and other countries in the East Asia–Australasian Flyway. The NS gene of the KNU18-28 and KNU18-86 isolates was closely related to that of China’s H10N3 strain, whereas the KNU18-93 strain originated from the H12N2 strain in Japan, showing two different reassortment events and different from a low pathogenic H5N3 (KNU18-91) virus which was isolated at the same day and same place with KNU18-86 and KNU18-93. These H5N2 isolates were characterized as low pathogenic avian influenza viruses. However, many amino acid changes in eight gene segments were identified to enhance polymerase activity and increase adaptation and virulence in mice and mammals. Experiments reveal that viral replication in MDCK cells was quite high after 12 hpi, showing the ability to replicate in mouse lungs. The hematoxylin and eosin-stained (H&E) lung sections indicated different degrees of pathogenicity of the three H5N2 isolates in mice compared with that of the control H1N1 strain. The continuing circulation of these H5N2 viruses may represent a potential threat to mammals and humans. Our findings highlight the need for intensive surveillance of avian influenza virus circulation in South Korea to prevent the risks posed by these reassortment viruses to animal and public health.