DNA vaccine expressing conserved influenza virus proteins protective against H5N1 challenge infection in mice

Influenza vaccination practice, which is based on neutralizing antibodies, requires being able to predict which viral strains will be circulating. If an unexpected strain, as in the 1997 H5N1 Hong Kong outbreak, or even a pandemic emerges, appropriate vaccines may take too long to prepare. Therefore, strategies based on conserved influenza antigens should be explored. We studied DNA vaccination in mice with plasmids expressing conserved nucleoprotein (NP) and matrix (M) from an H1N1 virus. After vaccination, mice were challenged with A/H5N1 viruses of low, intermediate, and high lethality. A/NP+A/M DNA vaccination reduced replication of A/Hong Kong/486/97 (HK/486), a nonlethal H5N1 strain, and protected against lethal challenge with more virulent A/Hong Kong/156/97 (HK/156). After HK/156 exposure, mice survived rechallenge with A/Hong Kong/483/97 (HK/483), although the DNA vaccination alone protected poorly against this highly virulent strain. In the absence of antigenically matched hemagglutinin-based vaccines, DNA vaccination with conserved influenza genes may provide a useful first line of defense against a rapidly spreading pandemic virus.     

Generation and evaluation of a high-growth reassortant H9N2 influenza A virus as a pandemic vaccine candidate

H9N2 subtype avian influenza viruses (AIVs) are widely distributed in avian species and were isolated from humans in Hong Kong and Guangdong province, China in 1999 raising concern of their potential for pandemic spread. We generated a high-growth reassortant virus (G9/PR8) that contains the hemagglutinin (HA) and neuraminidase (NA) genes from the H9N2 avian influenza virus A/chicken/Hong Kong/G9/97 (G9) and six internal genes from A/Puerto Rico/8/34 (PR8) by genetic reassortment, for evaluation as a potential vaccine candidate in humans. Pathogenicity studies showed that the G9/PR8 reassortant was not highly pathogenic for mice or chickens. Two doses of a formalin-inactivated G9/PR8 virus vaccine induced hemagglutination inhibiting antibodies and conferred complete protection against challenge with G9 and the antigenically distinct H9N2 A/Hong Kong/1073/99 (G1-like) virus in a mouse model. These results indicate that the high growth G9/PR8 reassortant has properties that are desirable in a vaccine seed virus and is suitable for evaluation in humans for use in the event of an H9 pandemic.     

Single- and multiple-clade influenza A H5N1 vaccines induce cross protection in ferrets

The rapid evolution, genetic diversity, broad host range, and increasing human infection with avian influenza A (H5N1) viruses highlight the need for an efficacious cross-clade vaccine. Using the ferret model, we compared induction of cross-reactive immunity and protective efficacy of three single-clade H5N1 vaccines and a novel multiple-clade H5N1 vaccine, with and without MF59 adjuvant. Reverse genetics (rg) was used to generate vaccine viruses containing the hemagglutinin (HA) and neuraminidase genes of wild-type H5N1 viruses. Ferrets received two doses of inactivated whole-virus vaccine separated by 3 weeks. Single-clade vaccines (7.5 μg HA per dose) included rg-A/Vietnam/1203/04 (clade 1), rg-A/Hong Kong/213/03 (clade 1), and rg-A/Japanese White Eye/Hong Kong/1038/06 (clade 2.3). The multiple-clade vaccine contained 3.75 μg HA per dose of each single-clade vaccine and of rg-A/Whooper Swan/Mongolia/244/05 (clade 2.2). Two doses of vaccine were required to substantially increase anti-HA and virus neutralizing antibody titers to H5N1 viruses. MF59 adjuvant enhanced induction of clade-specific and cross-clade serum antibody responses, reduced frequency of infection (as determined by upper respiratory tract virus shedding and seroconversion data), and eliminated disease signs. The rg-A/Hong Kong/213/03 vaccine induced the highest antibody titers to homologous and heterologous H5N1 viruses, while rg-A/Japanese White Eye/Hong Kong/1038/06 vaccine induced the lowest. The multiple-clade vaccine was broadly immunogenic against clade 1 and 2 viruses. The rg-A/Vietnam/1203/04 vaccine (the currently stockpiled H5N1 vaccine) most effectively reduced upper respiratory tract virus shedding after challenge with clade 1 and 2 viruses. Importantly, all vaccines protected against lethal challenge with A/Vietnam/1203/04 virus and provided cross-clade protection.     

Experimental challenge of chicken vaccinated with commercially available H5 vaccines reveals loss of protection to some highly pathogenic avian influenza H5N1 strains circulating in Hong Kong/China

Highly pathogenic avian influenza (HPAI) H5N1 virus continues to circulate in poultry in Asia and Africa posing a threat to both public and animal health. Vaccination, used as an adjunct to improved bio-security and stamping-out policies, contributed to protecting poultry in Hong Kong from HPAI H5N1 infection in 2004–2008 although the virus was repeatedly detected in dead wild birds. The detection of clade 2.3.4 H5N1 viruses in poultry markets and a farm in Hong Kong in 2008 raised the question whether this virus has changed to evade protection from the H5 vaccines in use. We tested the efficacy of three commercial vaccines (Nobilis, Poulvac and Harbin Re-5 vaccine) in specific pathogen free white leghorn chickens against a challenge with A/chicken/Hong Kong/8825-2/2008 (clade 2.3.4) isolated from vaccinated poultry in Hong Kong and A/chicken/Hong Kong/782/2009 (clade 2.3.2). Harbin Re5 vaccine provided the best, albeit not complete protection against challenge with the clade 2.3.4 virus. All three vaccines provided good protection from death and significantly reduced virus shedding following challenge with the clade 2.3.2 virus. Only Harbin Re-5 was able to completely protect chickens from virus shedding as well as mortality. Sera from vaccinated chickens had lower geometric hemagglutination inhibition titers against A/chicken/Hong Kong/8825-2/08, as compared to two other clade 2.3.4 and one clade 0 virus. Alignment of amino-acid sequences of the haemagglutinin of A/chicken/Hong Kong/8825-2/08 and the other H5 viruses revealed several mutations in positions including 69, 71, 83, 95, 133,140, 162, 183, 189, 194 and 270 (H5 numbering) which may correlate with loss of vaccine protection. Our results indicated that the tested HPAI H5N1 (2.3.4) virus has undergone antigenic changes that allow it to evade immunity from poultry vaccines. This highlights the need for continued surveillance and monitoring of vaccine induced immunity, with experimental vaccine challenge studies being done where indicated.     

Generation of an attenuated H5N1 avian influenza virus vaccine with all eight genes from avian viruses

In the face of disease outbreaks in poultry and the potential pandemic threat to humans caused by the highly pathogenic avian influenza viruses (HPAIVs) of H5N1 subtype, improvement in biosecurity and the use of inactivated vaccines are two main options for the control of this disease. Vaccine candidates of influenza A viruses of H5N1 subtype have been generated in several laboratories by plasmid-based reverse genetics with hemagglutinin (HA) and neuraminidase (NA) genes from the epidemic strains of avian viruses in a background of internal genes from the vaccine donor strain of human strains, A/Puerto Rico/8/34 (PR8). These reassortant viruses containing genes from both avian and human viruses might impose biosafety concerns, also may be do if C4/F AIV would be a live attenuated vaccine or cold-adaptive strain vaccine. In order to generate better and safer vaccine candidate viruses, we genetically constructed attenuated reassortant H5N1 influenza A virus, designated as C4/F AIV, by plasmid-based reverse genetics with all eight genes from the avian strains. The C4/F AIV virus contained HA and NA genes from an epidemic strain A/Chicken/Huadong/04 (H5N1) (C4/H5N1) in a background of internal genes derived from a low pathogenic strain of A/Chicken/F/98(H9N2). The reassortant virus was attenuated by removal of the multibasic amino acid motif in the HA gene by mutation and deletion (from PQRERRRKKR (downward arrow) G to PQIETR (downward arrow) G). The intravenous pathogenicity index (IVPI) of C4/F AIV virus was 0, whereas that of the donor virus C4/H5N1 was 3.0. The virus HA titer of C4/H5N1 in the allantoic fluid from infected embryonated eggs was as high as 1:2048. The inactivated vaccine prepared from the reassortant virus C4/F AIV-induced high HI titer in vaccinated chickens and gave 100% protection when challenged with highly pathogenic avian influenza virus of H5N1 subtype.     

An inactivated H5N2 vaccine reduces transmission of highly pathogenic H5N1 avian influenza virus among native chickens

Vaccination against H5N1 highly pathogenic avian influenza in endemically affected areas is a potentially attractive option for local prevention and control. In Indonesia the majority of local outbreaks have occurred in back yard flocks with native chickens, and it is therefore of interest to determine whether these birds can be protected against infection by vaccination. To this end two transmission experiments were carried out with H5N1 virus (A/chicken/Legok/2003) in vaccinated and unvaccinated native chickens. The vaccine contained an inactivated heterologous H5N2 strain (A/turkey/England/N28/73 H5N2). Birds were vaccinated at 4 and 7 weeks of age and challenged at 10 weeks of age. During 10 days post-challenge tracheal and cloacal swabs were taken for virus isolation, and serum blood was collected regularly to measure haemaglutinin inhibiting (HI) antibody responses. The results show that transmission of H5N1 virus was rapid and efficient in unvaccinated birds, that infection and transmission were completely prevented in vaccinated birds, and that vaccinated birds that were exposed to unvaccinated inoculated birds were still protected from infection. These findings indicate that vaccination with a heterologous H5N2 vaccine is able to prevent virus transmission in flocks of native chickens.     

Cross-protective immunity in mice induced by live-attenuated or inactivated vaccines against highly pathogenic influenza A (H5N1) viruses

Because of the time required to identify and produce an antigenically well-matched pandemic vaccine, vaccines that offer broader cross-reactive immunity and protection are desirable. We have compared a live attenuated influenza vaccine (LAIV) and inactivated influenza vaccine (IIV) based on a related H5 hemagglutinin (HA) from a nonpathogenic avian influenza virus, A/Duck/Pottsdam/1042-6/86 (H5N2), for the ability to induce cross-reactive immunity and/or cross-protective efficacy against a contemporary highly pathogenic H5N1 viruses. Both LAIV and IIV provided cross-protection from systemic infection, severe disease, and death following lethal challenges with antigenically distinct A/Vietnam/1203/2004 (VN/1203) virus. Substantial levels of serum anti-VN/1203 HA IgG were detected in mice that received either IIV or LAIV, while nasal wash anti-VN/1203 HA IgA was detected in mice that received LAIV. Formulation of IIV with alum adjuvant augmented neutralizing antibody responses and protective efficacy. These results demonstrated that vaccination of mice with H5 IIV or LAIV induced a high degree of cross-protection from illness and death following lethal challenges with a heterologous H5N1 virus.     

A comparison of live and inactivated influenza A (H1N1) virus vaccines. 2. Long-term immunity.

Groups of volunteers were immunized with one of three influenza virus vaccines, and the resistance to challenge infection with attenuated influenza A (H1N1) virus was measured 8 months later. The vaccines were aqueous subunit influenza A/USSR/77 (H1N1) vaccine, aqueous subunit influenza B/Hong Kong/73 vaccine, or attenuated influenza virus A (H1N1) vaccine. The B virus vaccine was included as a control to assess the incidence of natural A virus infection during the study period. A proportion of the B virus vaccinees had pre-existing A (H1N1) virus antibody and were used to study the immunity conferred by natural infection to the live virus challenge. The serum antibody responses were measured at 1 and 8 months after immunization. The results showed that all the vaccines induced serum HI antibody in a proportion of the volunteers; however, after 1 month, higher titres of serum antibody were found in volunteers given inactivated A vaccine than in those given live attenuated A virus vaccine. Eight months post-immunization the titres of serum antibody in volunteers given inactivated vaccine had declined significantly, but there were no changes in the antibody titres of those given live virus vaccine. The incidence of infection by the challenge virus at 8 months post-immunization was directly related to the serum antibody titres 1 month post-immunization; no evidence was obtained to suggest that those given live virus vaccine had a more solid immunity than those given inactivated vaccine.     

Immunity to challenge in volunteers vaccinated with an inactivated current or earlier strain of influenza A(H3N2).

Volunteers were inoculated with vaccine made from the 30c mutant, A/Port Chalmers/73 or B/Hong Kong/8/73. Preliminary experiments showed that the 30 c strain was antigenically quite close to A/HK/8/68. Volunteers given 30c developed haemagglutination inhibiting antibodies against the ''current'' 1973 serotypes (as well as to the vaccine virus) but the titres were less than those after the A/PC/73 vaccine. Volunteers were then challenged with a live attenuated virus, WRL 105, with A/Finland/4/74 antigens, by intranasal inoculation. The rates of infection were 43% after B/Hong Kong/8/73, 20% after 30c and 5% after A/PC/73. This indicated that the 30c gave some protection but that the vaccine prepared from the current strain gave more.     

Immunity to challenge in volunteers vaccinated with an inactivated current or earlier strain of influenza A(H3N2)

Volunteers were inoculated with vaccine made from the 30c mutant, A/Port Chalmers/73 or B/Hong Kong/8/73. Preliminary experiments showed that the 30 c strain was antigenically quite close to A/HK/8/68. Volunteers given 30c developed haemagglutination inhibiting antibodies against the 'current' 1973 serotypes (as well as to the vaccine virus) but the titres were less than those after the A/PC/73 vaccine. Volunteers were then challenged with a live attenuated virus, WRL 105, with A/Finland/4/74 antigens, by intranasal inoculation. The rates of infection were 43% after B/Hong Kong/8/73, 20% after 30c and 5% after A/PC/73. This indicated that the 30c gave some protection but that the vaccine prepared from the current strain gave more.     

Comparative efficacy of North American and antigenically matched reverse genetics derived H5N9 DIVA marker vaccines against highly pathogenic Asian H5N1 avian influenza viruses in chickens

Highly pathogenic (HP) H5N1 avian influenza has become endemic in several countries in Asia and Africa, and vaccination is being widely used as a control tool. However, there is a need for efficacious vaccines preferably utilizing a DIVA (differentiate infected from vaccinated animals) marker strategy to allow for improved surveillance of influenza in vaccinated poultry. Using a reverse genetics approach, we generated Asian rgH5N9 vaccine strain deriving the hemagglutinin gene from A/chicken/Indonesia/7/2003 (H5N1) with modification of the cleavage site to be low pathogenic (LP) and N9 neuraminidase gene from the North American LP A/turkey/Wisconsin/1968 (H5N9) virus. The recombinant rgH5N9, A/turkey/Wisconsin/1968 (H5N9) A/chicken/Hidalgo/232/1994 (H5N2), and wild type HP A/chicken/Indonesia/7/2003 (H5N1) viruses were used to prepare inactivated oil-emulsified whole virus vaccines. Two weeks after vaccination, chickens were challenged with either Asian HP H5N1 viruses, A/chicken/Indonesia/7/2003 (W.H.O. clade 2.1) or A/chicken/Supranburi Thailand/2/2004 (W.H.O. clade 1.0). The H5 HA1 of the North American vaccine strains exhibited 12% amino acid differences including amino acid changes in the major antigenic sites as compared to the Asian HP H5N1 challenge viruses, serologically exhibited substantial antigenic difference, but still provided 100% protection from mortality. However, challenge virus shedding was significantly higher in chickens immunized with antigenically distinct American lineage vaccines as compared to the antigenically matched Asian rgH5N9 and the wild type Asian H5N1 vaccine. The antibody response to the heterologous subtype neuraminidase proteins were discriminated in vaccinated and infected chickens using a rapid fluorescent 2′-(4-methylumbelliferyl)-α-d-N-acetylneuraminic acid sodium salt as substrate for neuraminidase inhibition assay. This study demonstrates the value of using a vaccine containing antigenically matched H5 hemagglutinin for control of HP H5N1 avian influenza in poultry and the potential utility of a heterologous neuraminidase as a DIVA marker.     

Generation and characterization of a cold-adapted influenza A H9N2 reassortant as a live pandemic influenza virus vaccine candidate

H9N2 subtype influenza A viruses have been identified in avian species worldwide and were isolated from humans in 1999, raising concerns about their pandemic potential and prompting the development of candidate vaccines to protect humans against this subtype of influenza A virus. Reassortant H1N1 and H3N2 human influenza A viruses with the internal genes of the influenza A/Ann Arbor/6/60 (H2N2) (AA) cold-adapted (ca) virus have proven to be attenuated and safe as live virus vaccines in humans. Using classical genetic reassortment, we generated a reassortant virus (G9/AA ca) that contains the hemagglutinin and neuraminidase genes from influenza A/chicken/Hong Kong/G9/97 (H9N2) (G9) and six internal gene segments from the AA ca virus. When administered intranasally, the reassortant virus was immunogenic and protected mice from subsequent challenge with wild-type H9N2 viruses, although it was restricted in replication in the respiratory tract of mice. The G9/AA ca virus bears properties that are desirable in a vaccine for humans and is available for clinical evaluation and use, should the need arise.     

Reverse genetics based H5N2 vaccine provides clinical protection against H5N1, H5N8 and H9N2 avian influenza infection in chickens

The current study aimed to develop broadly protective vaccines for avian influenza. In an earlier study, HA stalk (universal flu vaccine) was found to be broadly protective against different subtypes of influenza virus in mice. Hence, we were interested to know its breadth of protective efficacy either alone or combined with inactivated rgH5N2 (clade 2.3.2.1a) vaccine against challenge viruses of homologous H5N1, heterologous H5N8 (clade 2.3.4.4) and heterosubtypic H9N2 virus in specific pathogen-free chickens. The rgH5N2 vaccine alone or in combination with HA stalk elicited sufficient pre-challenge immunity in the form of haemagglutination inhibiting (HI) antibodies and neutralizing antibodies (MNT) against H5N1, H5N8, and H9N2 in chickens. The rgH5N2 vaccine alone or in combination with HA stalk also attenuated the shedding of H5N1, H5N8 and H9N2 in chickens and protected against the lethal challenge of H5N1 or H5N8. In contrast, all HA stalk immunised chickens died upon H5N1 or H5N8 challenge and H9N2 challenged chickens survived. Our study suggests that the rgH5N2 vaccine can provide clinical protection against H5N1, H5N8 and can attenuate the viral shedding of H9N2 in chickens.     

Preparation of vaccines against H5N1 influenza

In response to the pandemic warning provided by the highly pathogenic H5N1 influenza virus infections in Hong Kong, there were world-wide attempts to develop vaccines. Three strategies were followed and although each was associated with some success, there were also some problems. Pre-clinical vaccine efficacy results are presented from one such strategy, that of using an apathogenic H5N3 avian strain for vaccine production.     

Generation of a reassortant avian influenza virus H5N2 vaccine strain capable of protecting chickens against infection with Egyptian H5N1 and H9N2 viruses

BackgroundAvian influenza H5N1 viruses have been enzootic in Egyptian poultry since 2006. Avian influenza H9N2 viruses which have been circulating in Egyptian poultry since 2011 showed high replication rates in embryonated chicken eggs and mammalian cells.MethodsTo investigate which gene segment was responsible for increasing replication, we constructed reassortant influenza viruses using the low pathogenic H1N1 PR8 virus as backbone and included individual genes from A/chicken/Egypt/S4456B/2011(H9N2) virus. Then, we invested this finding to improve a PR8-derived H5N1 influenza vaccine strain by incorporation of the NA segment of H9N2 virus instead of the NA of H5N1. The growth properties of this virus and several other forms of reassortant H5 viruses were compared. Finally, we tested the efficacy of this reassortant vaccine strain in chickens.ResultsWe observed an increase in replication for a reassortant virus expressing the neuraminidase gene (N2) of H9N2 virus relative to that of either parental viruses or reassortant PR8 viruses expressing other genes. Then, we generated an H5N2 vaccine strain based on the H5 from an Egyptian H5N1 virus and the N2 from an Egyptian H9N2 virus on a PR8 backbone. This strain had better replication rates than an H5N2 reassortant strain on an H9N2 backbone and an H5N1 reassortant on a PR8 backbone. This virus was then used to develop a killed, oil-emulsion vaccine and tested for efficacy against H5N1 and H9N2 viruses in chickens. Results showed that this vaccine was immunogenic and reduced mortality and shedding.DiscussionOur findings suggest that an inactivated PR8-derived H5N2 influenza vaccine is efficacious in poultry against H5N1 and H9N2 viruses and the vaccine seed replicates at a high rate thus improving vaccine production.     

Long lasting immunity in chickens induced by a single shot of influenza vaccine prepared from inactivated non-pathogenic H5N1 virus particles against challenge with a highly pathogenic avian influenza virus

An influenza vaccine was prepared from inactivated whole particles of the non-pathogenic strain A/duck/Hokkaido/Vac-1/04 (H5N1) virus using an oil adjuvant containing anhydromannitol-octadecenoate-ether (AMOE). The vaccine was injected intramuscularly into five 4-week-old chickens, and 138 weeks after vaccination, they were challenged intranasally with 100 times 50% chicken lethal dose of the highly pathogenic avian influenza (HPAI) virus A/chicken/Yamaguchi/7/04 (H5N1). All 5 chickens survived without exhibiting clinical signs of influenza, although 2 days post-challenge, 3 vaccinated chickens shed limited titres of viruses in laryngopharyngeal swabs.     

Further evidence of antigenic drift and protective efficacy afforded by a recombinant HVT-H5 vaccine against challenge with two antigenically divergent Egyptian clade 2.2.1 HPAI H5N1 strains

In this study, we have compared the protection afforded by a recombinant turkey herpesvirus vaccine expressing the H5 gene from a clade 2.2 H5N1 strain (rHVT-H5) and a Mexican-origin H5N2 inactivated vaccine, alone or in combination, against two antigenically divergent H5N1 Egyptian strains isolated in 2007 and 2008. Our results confirm the existence of a major antigenic drift among the Egyptian H5N1 strains such that, although protection against the “classical” 2007 HPAI H5N1 Egyptian strain could be obtained with both types of vaccines, only vaccination with the rHVT-H5 vaccine protected against challenge with the “variant” 2008 HPAI H5N1 Egyptian strain.     

Prevention and control of influenza. Recommendations of the Advisory Committee on Immunization Practices (ACIP).

This report updates the 2002 recommendations by the Advisory Committee on Immunization Practices (ACIP) on the use of influenza vaccine and antiviral agents (CDC. Prevention and Control of Influenza: Recommendations of the Advisory Committee on Immunization Practices [ACIP]. MMWR 2002;51 [No. RR-3]:1-31). The 2003 recommendations include new or updated information regarding 1) the timing of influenza vaccination by age and risk group; 2) influenza vaccine for children aged 6-23 months; 3) the 2003-2004 trivalent inactivated vaccine virus strains: A/Moscow/10/99 (H3N2)-like, A/New Caledonia/20/99 (H1N1)-like, and B/Hong Kong/330/2001-like antigens (for the A/Moscow/10/99 [H3N2]-like antigen, manufacturers will use the antigenically equivalent A/Panama/2007/99 [H3N2] virus, and for the B/Hong Kong/330/2001-like antigen, manufacturers will use either B/Hong Kong/330/2001 or the antigenically equivalent B/Hong Kong/1434/2002); 4) availability of certain influenza vaccine doses with reduced thimerosal content, including single 0.25 mL-dose syringes; and 5) manufacturers of influenza vaccine for the U.S. market. Although the optimal time to vaccinate against influenza is October and November, vaccination in December and later continues to be strongly recommended A link to this report and other information regarding influenza can be accessed at http://www.cdc.gov/ncidod/diseases/flu/fluvirus.htm.     

Multiple dose vaccination with heterologous H5N2 vaccine: immune response and protection against variant clade 2.2.1 highly pathogenic avian influenza H5N1 in broiler breeder chickens

Circulation of an antigenically variant lineage of highly pathogenic avian influenza (HPAI) H5N1 virus in chicken breeder flocks in Egypt is a continuing problem. The protective efficacy of multiple repeated vaccinations using the currently available H5N2 vaccines is unclear. Here, broiler breeder chickens were vaccinated at weeks 6, 12 and 18 with an inactivated H5N2 commercial vaccine. HI-titer against an Egyptian H5N1 field isolate of classic clade 2.2.1 (EGYcls/H5N1) were significantly lower after the first immunization but increased after booster vaccinations. In contrast, no HI titers were induced against an antigenically distinct field virus of the variant lineage of clade 2.2.1 (EGYvar/H5N1). Upon challenge at week 50 mild, if any, clinical signs were observed in the group infected with EGYcls/H5N1 although one of eight (12.5%) birds died. Mortality reached 6/8 (75%) in the EGYvar/H5N1 challenge group. Virus excretion in all vaccinated groups was reduced in amplitude, but in vaccinated surviving birds, time of virus excretion was extended to up to ten days. Strikingly, challenged vaccinated birds kept laying eggs almost throughout the observation period. Virus was detected on the outer egg-shell of 17 of 40 eggs. The majority of the infected eggs were derived from the EGYcls/H5N1 challenged animals; here the virus was detected also in the yolk and albumin. Repeated vaccination using a commercial H5N2-based vaccine broadened the antigen profile of induced antibodies but did not provide adequate protection against heterologous virus variant. In addition, the observation of AIV contaminated eggs from infected flocks highlights the risk of silent virus spread by vaccinated animals and point to eggs as a possible vector.     

Comparison of immunogenicity between candidate influenza A(H3N2) virus vaccine strains in Japan: A randomized controlled trial using a monovalent vaccine of A/Saitama/103/2014 (CEXP-002) and A/Hong Kong/4801/2014 (X-263)

BackgroundFor the 2017–18 influenza season, A/Saitama/103/2014 (CEXP-002) (Saitama strain) was antigenically more similar to prior circulating strains than A/Hong Kong/4801/2014 (X-263) (Hong Kong strain) in a ferret model and was selected as the A(H3N2) vaccine virus strain in Japan. However, the Saitama strain grew poorly, and the Japanese government switched to the Hong Kong strain, raising public concerns of poor effectiveness. To enhance understanding of the correlation between antigenicity in experimental models and immunogenicity, as a surrogate measure of vaccine effectiveness, in the human population, we compared the immunogenicity of specially-prepared single dose monovalent influenza A(H3N2) vaccines containing the Saitama or the Hong Kong strain.MethodsA randomized controlled trial of 100 healthy adults aged 20–64 years (n = 50/group) was conducted. Virus neutralization assay was performed on sera from days 0 (pre-vaccination) and 21 (post-vaccination). Geometric mean titer (GMT), mean fold rise (MFR), seroconversion proportion (SCP), and seroprotection proportion (SPP) were calculated for vaccine strains and a representative circulating A(H3N2) virus strain (A/Osaka/188/2017).ResultsFor the Hong Kong strain, post-vaccination GMT was significantly higher in the Hong Kong vaccine recipients (1:546 vs 1:260, p < 0.01), but MFR, SCP, and SPP were similar for both vaccine groups. For the Saitama strain, post-vaccination GMT (1:116 vs 1:61, p = 0.01) and SPP (86% vs 68%, p = 0.03) were significantly higher in the Hong Kong vaccine recipients, but MFR and SCP were similar for both vaccine groups. Against A/Osaka/188/2017, post-vaccination GMT and MFR were similar in both vaccine groups, but SCP (32% vs 4%, p < 0.01) and SPP (28% vs. 6%, p < 0.01) were significantly higher in the Hong Kong vaccine recipients.ConclusionThe Hong Kong vaccine induced better or equivalent immunogenicity in comparison to the Saitama vaccine. Our trial showed that antigenic similarity in experimental models does not necessarily correlate with immunogenicity in the human population.Clinical trial registration: UMIN000029293.