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In France during winter 2016-2017, 487 outbreaks of clade 2.3.4.4b H5N8 subtype high pathogenicity (HP) avian influenza A virus (AIV) infections were detected in poultry and captive birds. During this epizootic, HPAIV A/decoy duck/France/161105a/2016 (H5N8) was isolated and characterized in an experimental infection transmission model in conventional mule ducks. To investigate options to possibly protect such ducks against this HPAIV, three vaccines were evaluated in controlled conditions. The first experimental vaccine was derived from the hemagglutinin gene of another clade 2.3.4.4b A(H5N8) HPAIV. It was injected at three weeks of age, either alone (Vac1) or after a primer injection at day-old (Vac1 + boost). The second vaccine (Vac2) was a commercial bivalent adjuvanted vaccine containing an expressed hemagglutinin modified from a clade 2.3.2 A(H5N1) HPAIV. Vac2 was administered as a single injection at two weeks of age. The third experimental vaccine (Vac3) also incorporated a homologous 2.3.4.4b H5 HA gene and was administered as a single injection at three weeks of age. Ducks were challenged with HPAIV A/decoy duck/France/161105a/2016 (H5N8) at six weeks of age. Post-challenge virus excretion was monitored in vaccinated and control birds every 2-3 days for two weeks using real-time reverse-transcription polymerase chain reaction and serological analyses (haemagglutination inhibition test against H5N8, H5 ELISA and AIV ELISA) were performed. Vac1 abolished oropharyngeal and cloacal shedding to almost undetectable levels, whereas Vac3 abolished cloacal shedding only (while partially reducing respiratory shedding) and Vac2 only partly reduced the respiratory and intestinal excretion of the challenge virus. These results provided relevant insights in the immunogenicity of recombinant H5 vaccines in mule ducks, a rarely investigated hybrid between Pekin and Muscovy duck species that has played a critical role in the recent H5 HPAI epizootics in France., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

During winter 2020-2021, France and other European countries were severely affected by highly pathogenic avian influenza H5 viruses of the Gs/GD/96 lineage, clade 2.3.4.4b. In total, 519 cases occurred, mainly in domestic waterfowl farms in Southwestern France. Analysis of viral genomic sequences indicated that 3 subtypes of HPAI H5 viruses were detected (H5N1, H5N3, H5N8), but most French viruses belonged to the H5N8 subtype genotype A, as Europe. Phylogenetic analyses of HPAI H5N8 viruses revealed that the French sequences were distributed in 9 genogroups, suggesting 9 independent introductions of H5N8 from wild birds, in addition to the 2 introductions of H5N1 and H5N3., (© 2022 The Authors. Transboundary and Emerging Diseases published by Wiley-VCH GmbH.)

An H3N1 avian influenza virus was detected in a laying hens farm in May 2019 which had experienced 25% mortality in Northern France. The complete sequencing of this virus showed that all segment sequences belonged to the Eurasian lineage and were phylogenetically very close to many of the Belgian H3N1 viruses detected in 2019. The French virus presented two genetic particularities with NA and NS deletions that could be related to virus adaptation from wild to domestic birds and could increase virulence, respectively. Molecular data of H3N1 viruses suggest that these two deletions occurred at two different times., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

In 2021, France faced large avian influenza outbreaks, like in 2016 and 2017. Controlling these outbreaks required the preventive depopulation of a large number of duck farms. A previous study in 2017 showed that the quality of decontamination of trucks and transport crates used for depopulation was often insufficient. A new study was then set up to evaluate cleaning and disinfection (C&D) of trucks and crates used for duck depopulation and whether practices had changed since 2017. Three methods were used to assess decontamination: 1) detection of avian influenza virus (AIV) genome, 2) visual inspection of cleanliness, and 3) microbial counts, considering that 2 and 3 are commonly used in abattoirs. Another objective of the study was to evaluate the correlation between results obtained with the 3 methods. In 5 abattoirs, 8 trucks and their crates were sampled by swabbing to detect AIV genome by rRT-PCR before and after decontamination. Visual cleanliness scores and coliform counts were also determined on crates after C&D. Trucks and crates were decontaminated according to the abattoirs' protocols. Before C&D, 3 quarters of crates (59/79) and 7 of 8 trucks were positive for AIV genome. C&D procedures were reinforced in 2021 compared to 2017; use of detergent solution and warm water were more common. Nevertheless, 28% of the crates were positive for AIV genome after C&D, despite the fact that cleaning scores and microbiological counts were satisfactory for 84% and 91% of the crates, respectively. No correlation was observed between results for AIV genome detection and results from visual control or from coliform counts. Abattoirs are encouraged to use environmental sampling coupled with AIV genome detection to monitor the quality of cleaning and disinfection of trucks and crates during AI outbreaks. Reinforcement of biosecurity measures at abattoirs is still needed to avoid residual contamination of the equipment and cross-contamination during the decontamination process., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

We detected 3 genotypes of highly pathogenic avian influenza A(H5N8) virus in France during winter 2016-17. Genotype A viruses caused dramatic economic losses in the domestic duck farm industry in southwestern France. Our phylogenetic analysis suggests that genotype A viruses formed 5 distinct geographic clusters in southwestern France. In some clusters, local secondary transmission might have been started by a single introduction. The intensity of the viral spread seems to correspond to the density of duck holdings in each production area. To avoid the introduction of disease into an unaffected area, it is crucial that authorities limit the movements of potentially infected birds.

Avian influenza A viruses are a major threat to animal and public health. Since 1997, several highly pathogenic H5N1 avian viruses have been directly transmitted from poultry to humans, caused numerous human deaths, and had considerable economic impact on poultry markets. During 2015-2016, a highly pathogenic avian influenza outbreak occurred in southwestern France. Different subtypes circulated, including the A/chicken/France/150169a/2015 H5N1 highly pathogenic virus, which did not possess the full set of genomic determinants known to promote transmission to humans. In order to evaluate the predicted absence of zoonotic potential, a quick method based on in vitro tests was developed to analyze some genetic and phenotypic host restriction determinants. A receptor-binding assay showed that the virus preferentially recognizes avian cell receptors. Temperature sensitivity revealed a cold-sensitive phenotype of the virus at 33 C as virus replication was reduced in contrast with what is expected for human influenza viruses, according to their primary infection sites. Altogether, our quick evaluation method suggests that the A/chicken/France/150169a/2015 H5N1 highly pathogenic virus has an avian phenotype in vitro, in accordance with in silico predictions based on genomic markers.

Several new highly pathogenic (HP) H5 avian influenza virus (AIV) have been detected in poultry farms from south-western France since November 2015, among which an HP H5N1. The zoonotic potential and origin of these AIVs immediately became matters of concern. One virus of each subtype H5N1 (150169a), H5N2 (150233) and H5N9 (150236) was characterised. All proved highly pathogenic for poultry as demonstrated molecularly by the presence of a polybasic cleavage site in their HA protein - with a sequence (HQRRKR/GLF) previously unknown among avian H5 HPAI viruses - or experimentally by the in vivo demonstration of an intravenous pathogenicity index of 2.9 for the H5N1 HP isolate. Phylogenetic analyses based on the full genomes obtained by NGS confirmed that the eight viral segments of the three isolates were all part of avian Eurasian phylogenetic lineage but differed from the Gs/Gd/1/96-like lineage. The study of the genetic characteristics at specific amino acid positions relevant for modulating the adaptation to and the virulence for mammals showed that presently, these viruses possess most molecular features characteristic of AIV and lack some major characteristics required for efficient respiratory transmission to or between humans. The three isolates are therefore predicted to have no significant pandemic potential., (This article is copyright of The Authors, 2017.)

EU annual serosurveillance programs show that domestic duck flocks have the highest seroprevalence of H5 antibodies, demonstrating the circulation of notifiable avian influenza virus (AIV) according to OIE, likely low pathogenic (LP). Therefore, transmission characteristics of LPAIV within these flocks can help to understand virus circulation and possible risk of propagation. This study aimed at estimating transmission parameters of four H5 LPAIV (three field strains from French poultry and decoy ducks, and one clonal reverse-genetics strain derived from one of the former), using a SIR model to analyze data from experimental infections in SPF Muscovy ducks. The design was set up to accommodate rearing on wood shavings with a low density of 1.6 ducks/m(2): 10 inoculated ducks were housed together with 15 contact-exposed ducks. Infection was monitored by RNA detection on oropharyngeal and cloacal swabs using real-time RT-PCR with a cutoff corresponding to 2-7 EID50. Depending on the strain, the basic reproduction number (R0) varied from 5.5 to 42.7, confirming LPAIV could easily be transmitted to susceptible Muscovy ducks. The lowest R0 estimate was obtained for a H5N3 field strain, due to lower values of transmission rate and duration of infectious period, whereas reverse-genetics derived H5N1 strain had the highest R0. Frequency and intensity of clinical signs were also variable between strains, but apparently not associated with longer infectious periods. Further comparisons of quantitative transmission parameters may help to identify relevant viral genetic markers for early detection of potentially more virulent strains during surveillance of LPAIV., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Following the emergence of highly pathogenic avian influenza (AI), active surveillance of infections due to the H5 and H7 subtypes in poultry has increased and been made compulsory in Europe since 2002, by means of annual serological surveys using the haemagglutination inhibition (HI) test. Domestic anseriforms, particularly ducks and geese, are more frequently infected by H5 low pathogenic AI virus, often subclinically, and represent a threat for other terrestrial poultry. 1783 sera, mainly from ducks, have been used to evaluate and compare a commercial ELISA kit detecting H5 antibodies with the currently recommended HI test. Different approaches to calculating specificity and sensitivity have been used, including the original Bayesian method. Results were similar when data were analyzed at the individual and batch levels, and when using different methods of calculation. However, results showed that H5 ELISA had both a higher sensitivity and a lower specificity than the HI test. Given that sensitivity is the most important factor for a screening test, H5 ELISA could therefore be recommended for AI surveillance, followed in cases of positivity by molecular tests aimed at detecting the virus gene., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Prevalence of avian influenza infection in free-range mule ducks (a cross between Muscovy [Cairina moschata domesticus] and Pekin ducks [Anas platyrhychos domesticus]) is a matter of concern and deserves particular attention. Thus, cloacal swabs were collected blindly from 30 targeted mule flocks at 4, 8, and 12 wk of age between October 2004 and January 2005. They were stored until selection. On the basis of a positive H5 antibody detection at 12 wk of age with the use of four H5 antigens, the samples from eight flocks were selectively analyzed. Positive samples were first screened with a matrix gene-based real-time reverse transcriptase-polymerase chain reaction assay before virus isolation. Eight avian influenza subtypes (H5N1, H5N2, H5N3, H6N2, H6N8, and H11N9) and three avian paramyxovirus type 1 viruses were isolated. All 11 are characterized as low pathogenicity (LP) and avirulent, respectively, by in vivo tests and, when relevant, nucleotide sequencing of the hemagglutinin (or fusion [F]) protein cleavage site. Regarding H5 isolates, all of their eight genes belong to the avian lineage and some particular genetic traits were determined. H5 genes were fully sequenced and phylogenetically analyzed; they all belong to the Eurasian lineage, lack additional glycosylation sites, and do not cluster, suggesting separate introductions from the wild reservoir. None were grouped with the Asian isolates. The N1 gene (H5N1 isolate) was very close genetically to an Italian LP-H7N1 gene. Antigenic relationships between these H5 isolates and others were assessed comparatively by crossed hemagglutination inhibition tests. All these data are very useful to control the evolution of H5 viruses at the genetic and antigenic level to better understand the source of new outbreaks (new introductions from wild birds or the result of spread among poultry) and to assess the immunity afforded by available vaccines. These data are useful also to update antigens for avian influenza survey and to choose the most suitable vaccine in the case of preventive vaccination of ducks.