[Show abstract][Hide abstract] ABSTRACT: H5 low pathogenic avian influenza virus (LPAIV) infection in domestic ducks is a major problem in duck producing countries. Their silent circulation is an ongoing source of potential highly pathogenic or zoonotic emerging strains. To prevent such events, vaccination of domestic ducks might be attempted but remains challenging. Currently licensed vector vaccines derived from H5N1 HPAIV possess clade 0, clade 2.2 or clade 2.3.4 HA sequences: selection of the best HA candidate inducing the largest cross protection is a key issue. For this purpose, DNA immunization of specific pathogen free Muscovy ducks was performed using different synthetic codon optimized (opt) or native HA genes from H5N2 LPAIV and several H5N1 HPAIV clade 2.1, 2.2.1 and 2.3.4. Humoral cross-immunity was assessed 3 weeks after boost by hemagglutination inhibition (HI) and virus neutralization (VN) against three French H5 LPAIV antigens.
Vaccination with LP H5N2 HA induced the highest VN antibody titre against the homologous antigen; however, the corresponding HI titre was lower and comparable to HI titres obtained after immunization with opt HA derived from clades 2.3.4 or 2.1. Compared to the other HPAIV-derived constructs, vaccination with clade 2.3.4 opt HA consistently induced the highest antibody titres in HI and VN, when tested against all three H5 LPAIV antigens and H5N2 LPAIV, respectively: differences in titres against this last strain were statistically significant.
The present study provides a standardized method to assess cross-immunity based on HA immunogenicity alone, and suggests that clade 2.3.4-derived recombinant vaccines might be the optimal candidates for further challenge testing to vaccinate domestic Muscovy ducks against H5 LPAIV.
[Show abstract][Hide abstract] ABSTRACT: Four avian metapneumovirus (AMPV) subgroups (A-D) have been reported previously based on genetic and antigenic differences. However, until now full length sequences of the only known isolates of European subgroup C and subgroup D viruses (duck and turkey origin, respectively) have been unavailable. These full length sequences were determined and compared with other full length AMPV and human metapneumoviruses (HMPV) sequences reported previously, using phylogenetics, comparisons of nucleic and amino acid sequences and study of codon usage bias. Results confirmed that subgroup C viruses were more closely related to HMPV than they were to the other AMPV subgroups in the study. This was consistent with previous findings using partial genome sequences. Closer relationships between AMPV-A, B and D were also evident throughout the majority of results. Three metapneumovirus "clusters" HMPV, AMPV-C and AMPV-A, B and D were further supported by codon bias and phylogenetics. The data presented here together with those of previous studies describing antigenic relationships also between AMPV-A, B and D and between AMPV-C and HMPV may call for a subclassification of metapneumoviruses similar to that used for avian paramyxoviruses, grouping AMPV-A, B and D as type I metapneumoviruses and AMPV-C and HMPV as type II.
PLoS ONE 01/2014; 9(7):e102740. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Avian influenza virus (AIV) subtypes H5 and H7 attracts particular attention because of the risk of their potential pathogenicity in poultry. The haemagglutination inhibition (HI) test is widely used as subtype specific test for serological diagnostics despite the laborious nature of this method. However, enzyme-linked immunosorbent assays (ELISAs) are being explored as an alternative test method.H5 and H7 specific monoclonal antibodies were experimentally raised and used in the development of inhibition ELISAs for detection of serological response specifically directed against AIV subtypes H5 and H7. The ELISAs were evaluated with polyclonal chicken anti-AIV antibodies against AIV subtypes: H1N2, H5N2, H5N7, H7N1, H7N7, H9N9, H10N4 and H16N3.
Both the H5 and H7 ELISA proved to have a high sensitivity and specificity and the ELISAs detected H5 and H7 antibodies earlier during experimental infection than the HI test did. The reproducibility of the ELISA's performed at different times was high with Pearson correlation coefficients of 0.96-0.98.
The ELISAs are a potential alternative to the HI test for screening of large amounts of avian sera, although only experimental sera were tested in this study.
[Show abstract][Hide abstract] ABSTRACT: 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 1.6ducks/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.
[Show abstract][Hide abstract] ABSTRACT: Newcastle disease, induced by a type 1 Avian Paramyxovirus (APMV-1), is one of the most serious poultry diseases. APMV-1 are divided into two classes based on genetic analysis: class II strains have been recovered from wild or domestic birds and include virulent and avirulent isolates whereas class I strains have been mainly isolated from wild birds and are avirulent. Within class I, a new proposed genotype has recently been reported. The only full genome strain of this group is presently characterised from the point of view of codon usage with reference to class I and class II specificities. Class-specific residues were identified on HN and F proteins that are the two major proteins involved in cell attachment and pathogenicity. Comparison of protein patterns and codon usage for this newly identified APMV-1 strain indicates it is similar to class I viruses but contains a few characteristics close to the viruses of class II. Transmission of viruses from this recently identified divergent group from wild birds to domestic birds could have a major impact on the domestic poultry industry.
[Show abstract][Hide abstract] ABSTRACT: Vaccination protocols were evaluated in one-day old Muscovy ducklings, using an experimental Newcastle disease recombinant vaccine (vNDV-H5) encoding an optimized synthetic haemagglutinin gene from a clade 2.2.1 H5N1 highly pathogenic (HP) avian influenza virus (AIV), either as a single administration or as a boost following a prime inoculation with a fowlpox vectored vaccine (vFP89) encoding a different H5 HP haemagglutinin from an Irish H5N8 strain. These vaccination schemes did not induce detectable levels of serum antibodies in HI test using a clade 2.2.1 H5N1 antigen, and only induced H5 ELISA positive response in less than 10% of vaccinated ducks. However, following challenge against a clade 2.2.1 HPAIV, both protocols afforded full clinical protection at six weeks of age, and full protection against mortality at nine weeks. Only the prime-boost vaccination (vFP89+vNDV-H5) was still fully protecting Muscovy ducks against disease and mortality at 12 weeks of age. Reduction of oropharyngeal shedding levels was also constantly observed from the onset of the follow-up at 2.5 or three days post-infection in vaccinated ducks compared to unvaccinated controls, and was significantly more important for vFP89+vNDV-H5 vaccination than for vNDV-H5 alone. Although the latter vaccine is shown immunogenic in one-day old Muscovy ducks, the present work is original in demonstrating the high efficacy of the successive administration of two different vector vaccines encoding two different H5 in inducing lasting protection (at least similar to the one induced by an inactivated reassortant vaccine, Re-5). In addition, such a prime-boost schedule allows implementation of a DIVA strategy (to differentiate vaccinated from infected ducks) contrary to Re-5, involves easy practice on the field (with injection at the hatchery and mass vaccination later on), and should avoid eventual interference with NDV maternally derived antibodies. Last, the HA insert could be updated according to the epidemiological situation.
[Show abstract][Hide abstract] ABSTRACT: Determining the sequence of non-segmented, negative sense RNA viral genomes is far from routine and often requires the application of several techniques. In this study, an existing method used currently just for determination of the genomic 3' extremity was used to determine both the 3' and 5' sequence extremities of a Newcastle disease virus and an avian metapneumovirus. This was achieved with a single 3' nucleotide tailing reaction of both the genomic RNA and the full length, positive sense, antigenomic RNA, followed by a single reverse transcription reaction targeted to the common poly nucleotide tails, and then individual PCRs specific for each extremity using PCR primers derived from the sequence of the RT primer or from neighbouring virus sequences known previously. For each virus the method was employed separately. Sequences from both viruses were in agreement with those reported previously for other paramyxoviruses, yet one extra base at the 3' and one extra base at the 5' were identified for the avian metapneumovirus In this study. Importantly, the newly determined extremities maintained the complementarity known to exist between the extremities of these viruses. The method was equally successful with both viruses and can be tailored easily to function with other non-segmented, negative sense viruses through minor modification of only the primer sequences.
Journal of virological methods 05/2013; · 2.13 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Newcastle disease (ND) is one of the most lethal diseases of poultry worldwide. It is caused by an avian paramyxovirus 1 that has high genomic diversity. In the framework of an international surveillance program launched in 2007, several thousand samples from domestic and wild birds in Africa were collected and analyzed. ND viruses (NDV) were detected and isolated in apparently healthy fowls and wild birds. However, two thirds of the isolates collected in this study were classified as virulent strains of NDV based on the molecular analysis of the fusion protein and experimental in vivo challenges with two representative isolates. Phylogenetic analysis based on the F and HN genes showed that isolates recovered from poultry in Mali and Ethiopia form new groups, herein proposed as genotypes XIV and sub-genotype VIf with reference to the new nomenclature described by Diel's group. In Madagascar, the circulation of NDV strains of genotype XI, originally reported elsewhere, is also confirmed. Full genome sequencing of five African isolates was generated and an extensive phylogeny reconstruction was carried out based on the nucleotide sequences. The evolutionary distances between groups and the specific amino acid signatures of each cluster allowed us to refine the genotype nomenclature.
PLoS ONE 01/2013; 8(10):e76413. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We report the first complete genome sequence of a strain that presents some pathogenicity and that belongs to a recently characterized genotype of avian paramyxovirus type 1 (APMV-1). This virus, isolated from the common teal, presents the most divergent genome within class I of APMV-1.
[Show abstract][Hide abstract] ABSTRACT: We report here the complete genome of a new avian paramyxovirus (APMV-11) isolated from common snipes. Sequence data from this virus showed that it has the largest genome of APMV and unusual P gene mRNA editing.
Journal of Virology 07/2012; 86(14):7710. · 5.08 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Since 2006, the members of the molecular epidemiological working group of the European "EPIZONE" network of excellence have been generating sequence data on avian influenza and avian paramyxoviruses from both European and African sources in an attempt to more fully understand the circulation and impact of these viruses. This review presents a timely update on the epidemiological situation of these viruses based on sequence data generated during the lifetime of this project in addition to data produced by other groups during the same period. Based on this information and putting it all into a European context, recommendations for continued surveillance of these important viruses within Europe are presented.
[Show abstract][Hide abstract] ABSTRACT: Wild aquatic birds are the main reservoir for avian influenza viruses (AIV) and play a major epidemiological role in the persistence and spread of the infection (7)....
Journal of clinical microbiology 04/2011; 49(6):2376-7. · 4.16 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Recurrent outbreaks of H5N1 HPAIV occurred in several Central European countries in 2007. In-depth phylogenetic analyses which included full-length genomic sequences of the viruses involved were performed to elucidate possible origins of incursions and transmission pathways. Tree reconstructions as well as host-shift and ancestral area inferences were conducted in a maximum likelihood framework. All viruses belonged to a separate subgroup (termed "EMA-3") within clade 2.2, and, thus, were distinct from two lineages of HPAIV H5N1 viruses (termed "EMA-1" and "EMA-2") present in the same geographic area in 2006. Analysis of concatenated coding regions of all eight genome segments significantly improved resolution and robustness of the reconstructed phylogenies as compared to single gene analyses. At the same time, the methodological limits to establish retrospectively transmission networks in a comparatively small geographic region and spanning a short period of time became evident when only few corroborating field-epidemiological data are available. Ambiguities remained concerning the origin of the EMA-3 viruses from a region covering Southeast Germany and the Czech Republic as well as routes of spread to other European countries. AIV monitoring programmes in place for wild birds and poultry in these countries did not reveal presence of these viruses in either population. Host switches between domestic poultry and wild bird populations occurred several times. Analysis of outbreaks in Northeast Germany and nearby Northern Poland in December 2007 demonstrated that geographic and even temporal vicinity of outbreaks does not necessarily indicate a common source of incursion.
Infection, genetics and evolution: journal of molecular epidemiology and evolutionary genetics in infectious diseases 10/2010; 10(7):1075-84. · 3.22 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Highly pathogenic (HP) H5N1 avian influenza (AI) is enzootic in several countries of Asia and Africa and constitutes a major threat, at the world level, for both animal and public health. Ducks play an important role in the epidemiology of AI, including HP H5N1 AI. Although vaccination can be a useful tool to control AI, duck vaccination has not proved very efficient in the field, indicating a need to develop new vaccines and a challenge model to evaluate the protection for duck species. Although Muscovy duck is the duck species most often reared in France, the primary duck-producing country in Europe, and is also produced in Asia, it is rarely studied. Our team recently demonstrated a good cross-reactivity with hemagglutinin from clade 2.2 and inferred that this could be a good vaccine candidate for ducks. Two challenges using two French H5N1 HP strains, 1) A/mute swan/France/06299/06 (Swan/06299), clade 2.2.1, and 2) A/mute swan/France/070203/07 (Swan/070203), clade 2.2 (but different from subclade 2.2.1), were performed (each) on 20 Muscovy ducks (including five contacts) inoculated by oculo-nasal route (6 log10 median egg infectious doses per duck). Clinical signs were recorded daily, and cloacal and oropharyngeal swabs were collected throughout the assay. Autopsies were done on all dead ducks, and organs were taken for analyses. Virus was measured by quantitative reverse transcriptase-PCR based on the M gene AI virus. Ducks presented severe nervous signs in both challenges. Swan/070203 strain led to 80% morbidity (12/15 sick ducks) and 73% mortality (11/15 ducks) at 13.5 days postinfection (dpi), whereas Swan/06299 strain produced 100% mortality at 6.5 dpi. Viral RNA load was significantly lower via the cloacal route than via the oropharyngeal route in both trials, presenting a peak in the first challenge at 3.5 dpi and being more stable in the second challenge. The brain was the organ containing the highest viral RNA load in both challenges. Viral RNA load in a given organ was similar or statistically significantly higher in ducks challenged with Swan/06299 strain. Thus, the Swan/06299 strain was more virulent and could be used as a putative challenge model. Moreover, challenged ducks and contacts contained the same amounts of viral RNA load, demonstrating the rapid and efficient transmission of H5N1 HP in Muscovy ducks in our experimental conditions.
[Show abstract][Hide abstract] ABSTRACT: Highly pathogenic (HP) avian influenza A viruses (AIVs) subtype H5N1 (subclade 2.2) were detected in wild birds during outbreaks in France during winter 2006 and summer 2007 in la Dombes wetlands (eastern France) and in Moselle wetlands (northeastern France), respectively. Blood samples from apparently healthy wild birds were collected in 2006 and 2007 from the end of the outbreak to several weeks after the influenza A outbreak inside and outside the contaminated areas, and in 2008 outside the contaminated areas. The samples were tested for the presence and/or quantitation of serum antibodies to influenza A subtypes H5 and N1 using hemagglutination inhibition tests (HITs), a commercial N1-specific enzyme-linked immunosorbent assay kit, and virus neutralization assay. In the HIT, low pathogenicity (LP) and HP H5 AIVs (belonging to H5N1, H5N2, and H5N3 subtypes) were used as antigens. One hundred mute swans were bled in the la Dombes outbreak area in 2006. During 2007, 46 mallards, 69 common pochards, and 59 mute swans were sampled in the Moselle outbreak area. For comparison, blood samples were also collected in 2007 from 60 mute swans from the Marne department where no HP H5N1 influenza A cases have been reported, and in 2008 from 111 sacred ibises in western France where no HP H5N1 influenza A infections in wild birds have been reported either. Mute swans (irrespective of their origin and time of sampling) and sacred ibises (from an area with no known outbreaks) had the highest prevalence of positive sera in the H5 HIT (49-69% and 64%, respectively). The prevalence of anti-H5 antibodies in mallards and common pochards was lower (28% and 27%, respectively). Positive H5- and N1-antibody responses were also significantly associated in swans (irrespective of their origin and time of sampling) and in sacred ibises. However, in swans from the area without outbreaks, the HIT titer against an H5N1 LPAIV was significantly higher than against an H5N1 2.2.1 HPAIV, whereas no difference could be shown for swans from the outbreak areas sampled in 2006 and 2007. These results suggest that ibises and swans from areas without declared outbreaks had acquired humoral immunity after AIV infections with subtypes H5 and N1 but independently from HP H5N1 infection. However, for swans living in outbreak areas, it cannot be excluded that this immunity might result from either a subclinical or a nonlethal infection by HP H5N1.
[Show abstract][Hide abstract] ABSTRACT: In Madagascar, Newcastle disease (ND) has become enzootic after the first documented epizootics in 1946, with recurrent annual outbreaks causing mortality up to 40%. Four ND viruses recently isolated in Madagascar were genotypically and pathotypically characterised. By phylogenetic inference based on the F and HN genes, and also full-genome sequence analyses, the NDV Malagasy isolates form a cluster distant enough to constitute a new genotype hereby proposed as genotype XI. This new genotype is presumably deriving from an ancestor close to genotype IV introduced in the island probably more than 50 years ago. Our data show also that all the previously described neutralising epitopes are conserved between Malagasy and vaccine strains. However, the potential implication in vaccination failures of specific amino acid substitutions predominantly found on surface-exposed epitopes of F and HN proteins is discussed.
PLoS ONE 01/2010; 5(11):e13987. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: H5 low-pathogenic avian influenza virus (LPAIV) has the potential to become highly pathogenic and to cause serious problems in animal and public health. AIV surveillance and characterization in both wild and domestic species is therefore necessary. In order to acquire molecular information and to identify possible reassortments in French viruses, we analysed the entire genome of five H5N3, three H5N2 and two H5N1 LPAIV, isolated in France between 2002 and 2008 mostly from captive ducks (free-range commercial poultry or decoy ducks). Some of the genome sequences showed atypical characteristics, such as an insertion of 1 aa in the PB1 protein of one H5N3, a highly truncated PB1-F2 protein (11 aa in length instead of 90 aa) in one H5N2, and an insertion of 8 aa in the NS1 protein of H5N1. These two last molecular characteristics have not been described previously. Phylogenetic analysis demonstrated that all genes of French LPAIV, except the closely related matrix protein genes, clustered within the Eurasian avian influenzavirus lineage and fell into at least two phylogenetic subgroups. In addition, the French H5 LPAIV were segregated into eight genotypes, suggesting that many reassortment events have occurred in H5 LPAIV in Europe. However, it is not known whether the reassortment events have occurred in wild waterfowl and/or in captive birds in direct or indirect contact with wild birds.
Journal of General Virology 12/2009; 91(Pt 4):960-70. · 3.13 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Influenza A viruses have been isolated from a wide range of animal species, aquatic birds being the reservoir for their genetic diversity. Avian influenza viruses can be transmitted to humans, directly or indirectly through an intermediate host like pig. This study aimed to define in vitro conditions that could prove useful to evaluate the potential of influenza viruses to adapt to a different host. Growth of H1N1, H1N2 and H3N2 influenza viruses belonging to different lineages isolated from birds or pigs prior to 2005 was tested on MDCK or NPTr cell lines in the presence or absence of exogenous trypsin. Virus multiplication was compared at 33, 37 and 40 degrees C, the infection site temperatures in human, swine and avian hosts, respectively. Temperature sensitivity of PB2-, NP- and M-RNA replication was also tested by quantitative real-time PCR. Multiplication of avian viruses was cold-sensitive, whatever cell type. By contrast, temperature sensitivity of swine viruses was found to depend on the virus and the host cell: for an H1N1 swine isolate from 1982, multiplication was cold-sensitive on NPTr cells and undetectable at 40 degrees C. From genetic analyses, it appears that temperature sensitivity could involve other residues than PB2 residue 627 and could affect other steps of the replication cycle than replication.
[Show abstract][Hide abstract] ABSTRACT: In late 2000, Italy was the first country of the European Union (EU) to implement an emergency vaccination programme against notifiable avian influenza. Vaccination with a conventional vaccine containing a seed strain with a different neuraminidase subtype from that of the field virus was used to complement biosecurity and restriction measures as part of an overall eradication strategy. This vaccination technique, in line with the Differentiating Infected from Vaccinated Animals system (DIVA), was applied several times until March 2008. This strategy enabled the identification of field exposed flocks and ultimately the eradication of low pathogenic H7N1, H7N3 and H5N2 infections. Italy was also the first country to implement a bivalent H5/H7 prophylactic vaccination programme of defined poultry populations, which was discontinued in December 2006. Following the incursion of highly pathogenic H5N1 into Europe, in 2005 and 2006, two other EU Member States, namely France and the Netherlands, implemented preventive vaccination programmes in 2006 but they targeted selected poultry populations different from those targeted in Italy and were implemented for short periods of time. Data generated during six years of experience with vaccination against avian influenza in Italy indicate that it is a useful tool to limit secondary spread and possibly prevent the introduction of low pathogenic avian influenza viruses in a susceptible population. The experience of France and the Netherlands provides data on vaccination of ducks and hobby poultry respectively and monitoring programmes associated with vaccination and difficulties related to their application. The advantages and disadvantages of vaccination need to be considered in the decision-making process, including the financial aspects of vaccination.
Revue scientifique et technique (International Office of Epizootics) 05/2009; 28(1):245-59. · 0.69 Impact Factor