Highly Pathogenic Avian Influenza Virus Research Articles

Genetic Characterization and Receptor Binding analysis of a Novel H5N1 HPAI Virus with a H6Nx-Derived PA Gene in Guangdong, China

Genetic Characterization and Receptor Binding analysis of a Novel H5N1 HPAI Virus with a H6Nx-Derived PA Gene in Guangdong, China

High pathogenicity avian influenza virus emergence: Blame it on chickens or on humans raising chickens?

High pathogenicity avian influenza virus emergence: Blame it on chickens or on humans raising chickens?

Dairy cows inoculated with highly pathogenic avian influenza virus H5N1.

Highly pathogenic avian influenza (HPAI) H5N1 hemagglutinin clade 2.3.4.4b was detected in the United States in 2021. These HPAI viruses caused mortality events in poultry, wild birds, and wild mammals. On March 25, 2024, HPAI H5N1 clade 2.3.4.4b was confirmed in a dairy cow in Texas in response to a multi-state investigation into milk production losses.1 Over 200 positive herds were identified in 14 U.S. states. The case description included reduced feed intake and rumen motility in lactating cows, decreased milk production, and thick yellow milk.2,3 The diagnostic investigation revealed viral RNA in milk and mammary tissue with alveolar epithelial degeneration and necrosis and positive immunoreactivity of glandular epithelium. A single transmission event, likely from birds, was followed by limited local transmission and onward horizontal transmission of H5N1 clade 2.3.4.4b genotype B3.13.4 We sought to experimentally reproduce infection with genotype B3.13 in Holstein yearling heifers and lactating cows. Heifers were inoculated by aerosol respiratory route and cows by intramammary route. Clinical disease was mild in heifers, but infection was confirmed by virus detection, lesions, and seroconversion. Clinical disease in lactating cows included decreased rumen motility, changes to milk appearance, and production losses. Infection was confirmed by high levels of viral RNA detected in milk, virus isolation, lesions in mammary tissue, and seroconversion. This study provides the foundation to investigate additional routes of infection, pathogenesis, transmission, and intervention strategies.

An mRNA vaccine candidate encoding H5HA clade 2.3.4.4b protects mice from clade 2.3.2.1a virus infection

Highly pathogenic avian influenza (HPAI) H5 viruses from different clades have been circulating globally, threatening wild/domestic birds and mammals. Given frequent spillovers and high mortality among mammals, coupled with our inability to predict which clade of H5 virus has pandemic potential, cross-clade protective HPAI H5 vaccines are urgently needed. Here, we demonstrate the applicability of a lipid nanoparticle-based mRNA vaccine modality to induce cross-protective immunity against lethal HPAI virus infection.

Continuing evolution of H5N1 highly pathogenic avian influenza viruses of clade 2.3.2.1a G2 genotype in domestic poultry of Bangladesh during 2018–2021

ABSTRACT We characterized 15 H5N1 HPAI viruses from different small- and medium-scale poultry flocks across Bangladesh during 2018–2021 based on their complete genome sequences. The antigenic relatedness of H5N1 HPAI viruses from different timepoints was analysed. During 2020–2021, 42.11% of the flocks tested positive for at least one of the respiratory infections, with 15.79% showing influenza A virus, of which 8.77% tested positive for HPAIV H5N1. Co-infections with two to four pathogens were detected in 15.8% of flocks. Phylogeny and gene constellation analyses based on complete genome sequences of 15 HPAI viruses revealed the continuing circulation of H5 clade 2.3.2.1a genotype G2 viruses. In the HA protein of the study isolates, functionally meaningful mutations caused the loss of an N-linked glycosylation site (T156A), a modified antigenic site A (S141P), and a mutation in the receptor binding pocket (E193R/K). Consequently, antigenic analysis revealed a significant loss of cross-reactivity between viruses from different host species and periods. Most viruses displayed oseltamivir resistance markers at positions V96, I97, S227, and N275 (N1 numbering) of the NA protein. In addition, for the PB2, M1, and NS1 proteins, significant mutations were noticed that have been associated with polymerase activity and increased virulence for mammals in all study isolates. These results highlight the need for intensified genomic surveillance of HPAI circulating in poultry in Bangladesh and for establishing appropriate control measures to decrease the circulation of these viruses in poultry in the country.

Detection of airborne wild waterbird-derived DNA demonstrates potential for transmission of avian influenza virus via air inlets into poultry houses, the Netherlands, 2021 to 2022.

BackgroundOutbreaks of highly pathogenic avian influenza (HPAI) on poultry farms and in wild birds worldwide persists despite intensified control measures. It causes unprecedented mortality in bird populations and is increasingly affecting mammalian species. Better understanding of HPAI introduction pathways into farms are needed for targeted disease prevention and control. The relevance of airborne transmission has been suggested but research involving air sampling is limited and unequivocal evidence on transmission routes is lacking.AimWe aimed to investigate whether HPAI virus from wild birds can enter poultry houses through air inlets by characterising host materials through eukaryote DNA sequencing.MethodsWe collected particulate matter samples in and around three HPAI-affected poultry farms which were cleared and decontaminated before sampling. Indoor measurements (n = 61) were taken directly in the airflow entering through air inlets, while outdoor air samples (n = 60) were collected around the poultry house. Positive controls were obtained from a bird rehabilitation shelter. We performed metabarcoding on environmental DNA by deep sequencing 18S rRNA gene amplicons.ResultsWe detected waterbird DNA in air inside all three, and outside of two, poultry farms. Sequences annotated at species level included swans and tufted ducks. Waterbird DNA was present in all indoor and outdoor air samples from the bird shelter.ConclusionAirborne matter derived from contaminated wild birds can potentially introduce HPAI virus to poultry houses through air inlets. The eDNA metabarcoding could assess breaches in biosecurity for HPAI virus and other pathogens potentially transmitted through air via detection of their hosts.

Avian influenza overview June-September 2024.

Between 15 June and 20 September 2024, 75 highly pathogenic avian influenza (HPAI) A(H5) and A(H7) virus detections were reported in domestic (16) and wild (59) birds across 11 countries in Europe. Although the overall number of detections in Europe continued to be low compared to previous epidemiological years, an increase in cases along the Atlantic, North Sea and Baltic coasts was notable, particularly an increase in the detection of HPAI viruses in colony-breeding seabirds. Besides EA-2022-BB and other circulating genotypes, these detections also included EA-2023-DT, a new genotype that may transmit more efficiently among gulls. In Germany, HPAI A(H7N5) virus emerged in a poultry establishment near the border with the Netherlands. No new HPAI virus detections in mammals were reported in Europe during this period, but the number of reportedly affected dairy cattle establishments in the United States of America (USA) rose to >230 in 14 states, and HPAI virus was identified in three new mammal species. Between 21 June and 20 September 2024, 19 new human cases with avian influenza virus infection were reported from the USA (six A(H5N1) cases and five A(H5) cases), Cambodia (five A(H5N1) cases, including one fatal), China (one fatal A(H5N6) case and one A(H9N2) case), and Ghana (one A(H9N2) case). Most of the human cases (90%, n = 17/19) had reported exposure to poultry, live poultry markets, or dairy cattle prior to avian influenza virus detection or onset of illness. Human infections with avian influenza viruses remain rare and no evidence of human-to-human transmission has been documented in the reporting period. The risk of infection with currently circulating avian A(H5) influenza viruses of clade 2.3.4.4b in Europe remains low for the general public in the European Union/European Economic Area (EU/EEA). The risk of infection remains low-to-moderate for those occupationally or otherwise exposed to infected animals or contaminated environments.

Survivability of low pathogenic avian influenza virus in aqueous poultry manure fertilizer

Avian influenza has been a growing problem in the United States for decades. Low pathogenic avian influenza (LPAI) virus causes low morbidity, while highly pathogenic avian influenza (HPAI) has a devastating effect on the poultry industry causing high morbidity and mortality worldwide. Manure from large and small poultry operations is spread on crop fields as a fertilizer. During an outbreak of avian influenza in a farm it is critical to adequately process the manure to kill the virus before its use in agriculture. Aqueous extraction of poultry manure at high temperature (55-65°C) is an attractive method for treatment of poultry manure, which extracts the nutrients and kills harmful microbes from the manure. The end product is a nutrient rich liquid that can be safely applied to fields or used in hydroponic operations, with nutrients that are more bioavailable to the plant compared to dry litter. However, we do not know whether aqueous high temperature extraction processes completely kill avian influenza virus present in the manure. Our aim in this study was to determine the competence of the aqueous extraction of poultry manure on avian influenza virus inactivation. We optimized the procedure using a LPAI virus isolate in a biosafety level 2 cabinet. Our data indicated that LPAI virus including the hemagglutinin protein of the virus were completely destroyed when the poultry manure was processed by aqueous extraction at temperatures ≥ 55°C for one hour. Future research will identify the conditions required for inactivating HPAI virus using this optimized procedure by performing the study in a BSL3 facility.

The Resurgent Threat of Highly Pathogenic Avian Influenza HPAI A (H5N1) Virus: A Zoonotic Infectious Disease and Public Health Concern

The highly pathogenic avian influenza (HPAI) A (H5N1) virus of clade 2.3.4.4b was identified among waterfowl (wild birds and geese) and domestic poultry across Southeast Asia, Europe, and Africa in 2021. By 2022, the HPAI virus was detected in North America, signifying further global dissemination. There is no human immunity to clade 2.3.4.4b A (H5N1). With the spread of the A (H5N1) virus throughout the United States and increasing viral infection in humans, resurgence of the A (H5N1) virus is of great public health concern that it could lead to the next infectious zoonotic pandemic after Covid-19. This paper describes the characteristics of HPAI A (H5N1) virus of clade 2.3.4.4b and recommendations for clinical treatment and prevention.

Molecular Characterization of a Clade 2.3.4.4b H5N1 High Pathogenicity Avian Influenza Virus from a 2022 Outbreak in Layer Chickens in the Philippines.

H5 subtype high-pathogenicity avian influenza (HPAI) viruses continue to devastate the poultry industry and threaten food security and public health. The first outbreak of H5 HPAI in the Philippines was reported in 2017. Since then, H5 HPAI outbreaks have been reported in 2020, 2022, and 2023. Here, we report the first publicly available complete whole-genome sequence of an H5N1 high-pathogenicity avian influenza virus from a case in Central Luzon. Samples were collected from a flock of layer chickens exhibiting signs of lethargy, droopy wings, and ecchymotic hemorrhages in trachea with excessive mucus exudates. A high mortality rate of 96-100% was observed within the week. Days prior to the high mortality event, migratory birds were observed around the chicken farm. Lungs, spleen, cloacal swabs, and oropharyngeal-tracheal swabs were taken from two chickens from this flock. These samples were positive in quantitative RT-PCR assays for influenza matrix and H5 hemagglutinin (HA) genes. To further characterize the virus, the same samples were subjected to whole-virus-genome amplification and sequencing using the Oxford Nanopore method with mean coverages of 19,190 and 2984, respectively. A phylogenetic analysis of the HA genes revealed that the H5N1 HPAI virus from Central Luzon belongs to the Goose/Guangdong lineage clade 2.3.4.4b viruses. Other segments also have high sequence identity and the same genetic lineages as other clade 2.3.4.4b viruses from Asia. Collectively, these data indicate that wild migratory birds are the likely source of H5N1 viruses from the 2022 outbreaks in the Philippines. Thus, biosecurity practices and surveillance for HPAI viruses in both domestic and wild birds should be increased to prevent and mitigate HPAI outbreaks.

The global H5N1 influenza panzootic in mammals.

Influenza A viruses (IAV) have caused more documented global pandemics in human history than any other pathogen1,2. High pathogenicity avian influenza (HPAI) viruses belonging to the H5N1 subtype are a leading pandemic risk. Two decades after H5N1 "bird flu" became established in poultry in Southeast Asia, its descendants have resurged3, setting off an H5N1 panzootic in wild birds that is fueled by (a) rapid intercontinental spread, reaching South America and Antarctica for the first time4,5; (b) fast evolution via genomic reassortment6; and (c) frequent spillover into terrestrial7,8 and marine mammals9. The virus has sustained mammal-to-mammal transmission in multiple settings, including European fur farms10,11, South American marine mammals12-15, and US dairy cattle16-19, raising questions about whether humans are next. Historically, swine are considered optimal intermediary hosts that help avian influenza viruses (AIV) adapt to mammals before jumping to humans20. However, the altered ecology of H5N1 has opened the door to new evolutionary pathways. Could dairy cattle, farmed mink, or South American sea lions serve as new mammalian gateways to humans? Here we explore the molecular and ecological factors driving H5N1's sudden expansion in host range and assess the likelihood of different zoonotic pathways leading to an H5N1 pandemic.

Review of the Highly Pathogenic Avian Influenza in Argentina in 2023: Chronicle of Its Emergence and Control in Poultry.

Highly pathogenic avian influenza (HPAI) is a highly contagious viral disease that represents a significant threat to poultry production worldwide. Variants of the HPAI virus (HPAIV) H5A/Goose/GuangDong/1/96 (H5 Gs/GD/96) lineage have caused five intercontinental epizootic waves, with the most recent, clade 2.3.4.4b, reaching Argentina in February 2023. Initially detected in wild birds, the virus quickly spread to backyard and commercial poultry farms, leading to economic losses, including the loss of influenza-free status (IFS). By March/April 2023 the epidemic had peaked and vaccination was seriously considered. However, the success of strict stamping-out measures dissuaded the National Animal Health Authority (SENASA) from authorizing any vaccine. Suspected cases sharply declined by May, and the last detection in commercial poultry was reported in June. The effective control and potential eradication of HPAIV in Argentina were due to SENASA's early detection and rapid response, supported by private companies, veterinarians, and other stakeholders. Stamping-out measures have been effective for virus elimination and reduced farm-to-farm transmission; however, as the virus of this clade may remain present in wild birds, the risk of reintroduction into poultry production is high. Therefore, maintaining continuous active surveillance will be crucial for promptly detecting any new HPAIV incursion and taking appropriate action to contain virus dissemination.

Genomic characterization of highly pathogenic H5 avian influenza viruses from Alaska during 2022 provides evidence for genotype-specific trends of spatiotemporal and interspecies dissemination

: The ongoing panzootic of highly pathogenic H5 clade 2.3.4.4b avian influenza (HPAI) spread to North America in late 2021, with detections of HPAI viruses in Alaska beginning in April 2022. HPAI viruses have since spread across the state, affecting many species of wild birds as well as domestic poultry and wild mammals. To better understand the dissemination of HPAI viruses spatiotemporally and among hosts in Alaska and adjacent regions, we compared the genomes of 177 confirmed HPAI viruses detected in Alaska during April – December 2022. Results suggest multiple viral introductions into Alaska between November 2021 and August or September 2022, as well as dissemination to areas within and outside of the state. Viral genotypes differed in their spatiotemporal spread, likely influenced by timing of introductions relative to population immunity. We found evidence for dissemination of HPAI viruses between wild bird species, wild birds and domestic poultry, as well as wild birds and wild mammals. Continued monitoring for and genomic characterization of HPAI viruses in Alaska can improve our understanding of the evolution and dispersal of these economically costly and ecologically relevant pathogens.

Kleptoparasitism in seabirds—A potential pathway for global avian influenza virus spread

AbstractWild birds have experienced unprecedented, near‐global mass mortalities since 2021, driven by outbreaks of high‐pathogenicity avian influenza virus (HPAIV) H5N1 lineage 2.3.4.4b. Managing this panzootic requires identification of transmission pathways. We investigated potential HPAIV transmission via kleptoparasitism (food theft) by examining the distribution, behaviors, and movements of two globally widespread and commonly kleptoparasitic seabird families: Fregatidae (frigatebirds) and Stercorariidae (skuas). These kleptoparasites force other seabirds (targets) to regurgitate food, which the kleptoparasite then ingests, potentially facilitating direct transfer of viral particles from target to kleptoparasite. Scavenging and predation probably contribute further to viral spread. Although frigatebirds use kleptoparasitism on a year‐round basis, skuas more commonly do so outside of the breeding season. Both frequently forage, disperse, or migrate across oceans and hemispheres. Dense aggregations of kleptoparasitic and target seabirds at breeding and/or roosting sites may facilitate the spread of HPAIV. In addition, the migration of these species could also facilitate broadscale geographic spread of HPAIV. Surveillance of kleptoparasites for HPAIVs could aid in early detection and may be important for seabird conservation.

Use of “ARRIAH-AviFluVac” vaccine in turkeys, geese and ducks

“ARRIAH-AviFluVac” vaccine against H5 avian influenza was demonstrated to be effective for ducks, geese and turkeys both in the laboratory and production environment. When administered to ducks at 0.5; 1.0 and 1.5 cm3, the vaccine provided 100%-effective protection of birds against the disease and death after challenge with the relevant high pathogenicity avian influenza virus of subtype H5N1, clade 2.3.4.4b. Singular 0.5–1.5 cm3 inoculation induced formation of antibodies, which were detected in the hemagglutination inhibition test at the titres that ranged from 4.3 to 6.1 log2. The vaccine facilitated 9–26-fold decrease in the virulent virus shedding by the ducks. Protection of turkeys vaccinated at the dose of 1.0 cm3 was maintained at the level of 87.5% after challenge with high pathogenicity avian influenza virus of subtype H5N1, clade 2.3.4.4b. The vaccine induced formation of antibodies at the titres of 4.9 and 5.5 log2 in turkeys after singular and double vaccination at the dose of 1.0 cm3, respectively. It was demonstrated, that after double administration of 1.0 cm3 of “ARRIAH-AviFluVac” vaccine, the post-vaccinal avian influenza antibody level exceeded 5.0 log2 in 75.9–90.0% of the geese population. The most appropriate way of the vaccine use in turkeys, ducks and geese involves at least its double administration at the double commercial dose. Higher species resistance of ducks to the challenge with avian influenza virus of subtype H5, clade 2.3.4.4bas compared to turkeys was also demonstrated.

Contrasting dynamics of two incursions of low-pathogenicity avian influenza virus into Australia.

The current panzootic of high pathogenicity avian influenza virus H5N1 demonstrates how viral incursions can have major ramifications for wildlife and domestic animals. Herein, we describe the recent incursion into Australia of two low pathogenicity avian influenza virus subtypes, H4 and H10, that exhibited contrasting evolutionary dynamics. Viruses detected from national surveillance and disease investigations between 2020 and 2022 revealed 27 genomes, 24 of which have at least one segment more closely related to Eurasian or North American avian influenza lineages than those already circulating in Australia. Phylogenetic analysis revealed that H4 viruses circulating in shorebirds represent a recent incursion from Asia that is distinct from those circulating concurrently in Australian waterfowl. Analysis of the internal segments further demonstrates exclusive, persistent circulation in shorebirds. This contrasts with H10, where a novel lineage has emerged in wild waterfowl, poultry, and captive birds across Australia and has likely replaced previously circulating H10 lineages through competitive exclusion. Elucidating different dynamics for avian influenza incursions supports effective disease risk identification and communication that better informs disease preparedness and response.

Susceptibility of Synanthropic Rodents (Mus musculus, Rattus norvegicus and Rattus rattus) to H5N1 Subtype High Pathogenicity Avian Influenza Viruses.

Synanthropic wild rodents associated with agricultural operations may represent a risk path for transmission of high pathogenicity avian influenza viruses (HPAIVs) from wild birds to poultry birds. However, their susceptibility to HPAIVs remains unclear. In the present study, house mice (Mus musculus), brown rats (Rattus norvegicus), and black rats (Rattus rattus) were experimentally exposed to H5N1 subtype HPAIVs to evaluate their vulnerability to infection. After intranasal inoculation with HA clade 2.2 and 2.3.2.1 H5N1 subtype HPAIVs, wild rodents did not show any clinical signs and survived for 10- and 12-day observation periods. Viruses were isolated from oral swabs for several days after inoculation, while little or no virus was detected in their feces or rectal swabs. In euthanized animals at 3 days post-inoculation, HPAIVs were primarily detected in respiratory tract tissues such as the nasal turbinates, trachea, and lungs. Serum HI antibodies were detected in HA clade 2.2 HPAIV-inoculated rodents. These results strongly suggest that synanthropic wild rodents are susceptible to infection of avian-origin H5N1 subtype HPAIVs and contribute to the virus ecosystem as replication-competent hosts. Detection of infectious viruses in oral swabs indicates that wild rodents exposed to HPAIVs could contaminate food, water, and the environment in poultry houses and play roles in the introduction and spread of HPAIVs in farms.

Detection and spread of high pathogenicity avian influenza virus H5N1 in the Antarctic Region

Until recent events, the Antarctic was the only major geographical region in which high pathogenicity avian influenza virus (HPAIV) had never previously been detected. Here we report on the detection of clade 2.3.4.4b H5N1 HPAIV in the Antarctic and sub-Antarctic regions of South Georgia and the Falkland Islands, respectively. We initially detected H5N1 HPAIV in samples collected from brown skuas at Bird Island, South Georgia on 8th October 2023. Since this detection, mortalities were observed in several avian and mammalian species at multiple sites across South Georgia. Subsequent testing confirmed H5N1 HPAIV across several sampling locations in multiple avian species and two seal species. Simultaneously, we also confirmed H5N1 HPAIV in southern fulmar and black-browed albatross in the Falkland Islands. Genetic assessment of the virus indicates spread from South America, likely through movement of migratory birds. Critically, genetic assessment of sequences from mammalian species demonstrates no increased risk to human populations above that observed in other instances of mammalian infections globally. Here we describe the detection, species impact and genetic composition of the virus and propose both introductory routes and potential long-term impact on avian and mammalian species across the Antarctic region. We also speculate on the threat to specific populations following recent reports in the area.

Phylodynamics of high pathogenicity avian influenza virus in Bangladesh identifying domestic ducks as the amplifying host reservoir

ABSTRACT High pathogenicity avian influenza (HPAI) virus H5N1 first emerged in Bangladesh in 2007. Despite the use of vaccines in chickens since 2012 to control HPAI, HPAI H5Nx viruses have continued to infect poultry, and wild birds, resulting in notable mass mortalities in house crows (Corvus splendens). The first HPAI H5Nx viruses in Bangladesh belonged to clade 2.2.2, followed by clade 2.3.4.2 and 2.3.2.1 viruses in 2011. After the implementation of chicken vaccination in 2012, these viruses were mostly replaced by clade 2.3.2.1a viruses and more recently clade 2.3.4.4b and h viruses. In this study, we reconstruct the phylogenetic history of HPAI H5Nx viruses in Bangladesh to evaluate the role of major host species in the maintenance and evolution of HPAI H5Nx virus in Bangladesh and reveal the role of heavily impacted crows in virus epidemiology. Epizootic waves caused by HPAI H5N1 and H5N6 viruses amongst house crows occurred annually in winter. Bayesian phylodynamic analysis of clade 2.3.2.1a revealed frequent bidirectional viral transitions between domestic ducks, chickens, and house crows that was markedly skewed towards ducks; domestic ducks might be the source, or reservoir, of HPAI H5Nx in Bangladesh, as the number of viral transitions from ducks to chickens and house crows was by far more numerous than the other transitions. Our results suggest viral circulation in domestic birds despite vaccination, with crow epizootics acting as a sentinel. The vaccination strategy needs to be updated to use more effective vaccinations, assess vaccine efficacy, and extension of vaccination to domestic ducks, the key reservoir.

The Alarming Situation of Highly Pathogenic Avian Influenza Viruses in 2019-2023.

Avian influenza viruses (AIVs) have the potential to cause severe illness in wild birds, domestic poultry, and humans. The ongoing circulation of highly pathogenic avian influenza viruses (HPAIVs) has presented significant challenges to global poultry industry and public health in recent years. This study aimed to elucidate the circulation of HPAIVs during 2019 to 2023. Specifically, we assess the alarming global spread and continuous evolution of HPAIVs. Moreover, we discuss their transmission and prevention strategies to provide valuable references for future prevention and control measures against AIVs.