Humoral and cell-mediated immune responses of old horses following recombinant canarypox virus vaccination and subsequent challenge infection

Equine influenza virus is a leading cause of respiratory disease in the horse population; however, the susceptibility of old horses to EIV infection remains unknown. While advanced age in horses (>20 years) is associated with age-related changes in immune function, there are no specific recommendations regarding the vaccination of older horses even though a well-characterized effect of aging is a reduced antibody response to standard vaccination. Therefore, we evaluated the immunological and physiological response of aged horses to a live non-replicating canarypox-vectored EIV vaccine and subsequent challenge infection. Vaccination of the aged horses induced EIV-specific IgGb and HI antibodies. No specific increase in cell-mediated immune (CMI) response was induced by the vaccine as determined by EIV-specific lymphoproliferation and the detection of EIV-specific IFNγ⁺ CD5⁺T cells, IFNγ, IL-2, IL-4 and IL-13 mRNA expression. Non-vaccinated aged horses exhibited clinical signs of the disease (coughing, nasal discharge, dyspnea, depression, anorexia) as well as increased rectal temperature and viral shedding following challenge. Concomitant with the febrile episodes, we also observed increased production of pro-inflammatory cytokine mRNA production in vivo using RT-PCR. Naïve horses were included in this study for vaccine and challenge controls only. As expected, the canarypox-vectored EIV vaccine stimulated significant CMI and humoral immune responses and provided significant protection against clinical signs of disease and reduced virus shedding in naive horses. Here, we show that aged horses remain susceptible to infection with equine influenza virus despite the presence of circulating antibodies and CMI responses to EIV and vaccination with a canarypox-vectored EIV vaccine provides protection from clinical disease.     

Protection from challenge following administration of a canarypox virus-vectored recombinant feline leukemia virus vaccine in cats previously vaccinated with a killed virus vaccine

OBJECTIVE: To compare protection against FeLV challenge obtained following administration of 2 doses of an adjuvanted, chemically inactivated, whole FeLV (FeLV-k) vaccine with protection obtained following administration of 1 dose of an FeLV-k vaccine followed by 1 dose of a canarypox virus-vectored recombinant FeLV (rCP-FeLV) vaccine. DESIGN: Prospective study. ANIMALS: Thirty-two 9-week-old domestic shorthair cats. PROCEDURE: Cats received 2 doses of the FeLV-k vaccine SC, 21 days apart (n = 11); 1 dose of the FeLV-k vaccine SC and, 21 days later, 1 dose of the rCP-FeLV vaccine transdermally (11); or 2 doses of physiologic saline (0.9% NaCl) solution (control; 10). Four weeks after the second vaccine dose, all cats were challenged with FeLV by means of oronasal administration. Blood samples were collected at weekly intervals beginning 21 days after challenge, and serum was tested for FeLV antigen. RESULTS: All 10 control cats became persistently infected (ie, FeLV antigen detected in > or = 3 consecutive serum samples) following FeLV challenge, whereas only 1 of 11 cats that received 2 doses of the FeLV-k vaccine and none of the 11 cats that received 1 dose of the FeLV-k vaccine and 1 dose of the rCP-FeLV vaccine did. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that protection against FeLV challenge obtained following SC administration of a single dose of an FeLV-k vaccine followed, 21 days later, by transdermal administration of a single dose of an rCP-FeLV vaccine was similar to that obtained following SC administration of 2 doses of the FeLV-k vaccine 21 days apart.     

Formulation with CpG ODN enhances antibody responses to an equine influenza virus vaccine

Previous studies have shown that protection against equine influenza virus (EIV) is partially mediated by virus-specific IgGa and IgGb. In this study we tested whether addition of a CpG ODN formulation to a commercial killed virus vaccine would enhance EIV-specific IgGa and IgGb antibody responses, and improve protection against an experimental EIV challenge. Thirty na?ve horses were assigned to one of three groups and vaccinated as follows: 10 were given vaccine (Encevac TC4, Intervet Inc.) alone, 10 were given vaccine plus 0.25 mg CpG ODN 2007 formulated with 30% Emulsigen (CpG/Em), and 10 controls were given saline. All horses were challenged with live virus 12 weeks after the final vaccination. Antibody responses were tested by single radial hemolysis (SRH) and ELISA, and protection was evaluated by determination of temperature, coughing, and clinical scores. Killed virus vaccine combined with CpG/Em induced significantly greater serologic responses than did the vaccine alone. All antibody isotypes tested increased after the addition of CpG/Em, although no shift in relative antibody isotypes concentrations was detected. Vaccination significantly improved protection against challenge but the differences between the two vaccine groups were not statistically significant. This study is the first demonstration that CpG/Em enhances antigen-specific antibody responses in horses and supports its potential to be used as an adjuvant for vaccines against equine infections.     

An African horse sickness virus serotype 4 recombinant canarypox virus vaccine elicits specific cell-mediated immune responses in horses

A recombinant canarypox virus vectored vaccine co-expressing synthetic genes encoding outer capsid proteins, VP2 and VP5, of African horse sickness virus (AHSV) serotype 4 (ALVAC(?)-AHSV4) has been demonstrated to fully protect horses against homologous challenge with virulent field virus. Guthrie et al. (2009) detected weak and variable titres of neutralizing antibody (ranging from <10 to 40) 8 weeks after vaccination leading us to hypothesize that there could be a participation of cell mediated immunity (CMI) in protection against AHSV4. The present study aimed at characterizing the CMI induced by the experimental ALVAC(?)-AHSV4 vaccine. Six horses received two vaccinations twenty-eight days apart and three horses remained unvaccinated. The detection of VP2/VP5 specific IFN-γ responses was assessed by enzyme linked immune spot (ELISpot) assay and clearly demonstrated that all ALVAC(?)-AHSV4 vaccinated horses developed significant IFN-γ production compared to unvaccinated horses. More detailed immune responses obtained by flow cytometry demonstrated that ALVAC(?)-AHSV4 vaccinations induced immune cells, mainly CD8(+) T cells, able to recognize multiple T-epitopes through all VP2 and only the N-terminus sequence of VP5. Neither VP2 nor VP5 specific IFN-γ responses were detected in unvaccinated horses. Overall, our data demonstrated that an experimental recombinant canarypox based vaccine induced significant CMI specific for both VP2 and VP5 proteins of AHSV4.     

Protection, systemic IFNgamma, and antibody responses induced by an ISCOM-based vaccine against a recent equine influenza virus in its natural host

In the horse, conventional inactivated or subunit vaccines against equine influenza virus (EIV) induce a short-lived antibody-based immunity to infection. Alternative strategies of vaccination have been subsequently developed to mimic the long-term protection induced by natural infection with the virus. One of these approaches is the use of immune-stimulating complex (ISCOM)-based vaccines. ISCOM vaccines induce a strong antibody response and protection against influenza in horses, humans, and a mouse model. Cell-mediated immunity (CMI) has been demonstrated in humans and mice after ISCOM vaccination, but rarely investigated in the horse. The aim of this study was to evaluate EIV-specific immune responses after intra-muscular vaccination with an ISCOM-EIV vaccine (EQUIP F) containing both equine influenza H7N7 (A/eq/Newmarket/77) and H3N8 (A/eq/Borl?nge/91 and A/eq/Kentucky/98) strains. The antibody response was measured by single radial haemolysis (SRH) assay using different H3N8 EIV strains. Stimulation of type-1 immunity was evaluated with a recently developed method that measures EIV-specific IFNgamma synthesis by peripheral blood lymphocytes (PBL). The protective efficacy of this ISCOM-based vaccine against challenge infection with a recent equine influenza (H3N8; A/eq/South Africa/4/03) strain was also evaluated. Vaccinated ponies developed elevated levels of EIV-specific SRH antibody and increased percentage of EIV-specific IFNgamma(+) PBL, whereas these responses were only detected after challenge infection in unvaccinated control ponies. Vaccinates showed minimal signs of disease and did not shed virus when challenged shortly after the second immunisation. In conclusion, evidence of type-1 immunity induced by an ISCOM-based vaccine is described for the first time in horses.     

Efficacy of a recombinant equine influenza vaccine against challenge with an American lineage H3N8 influenza virus responsible for the 2003 outbreak in the United Kingdom

Fifteen influenza-naive Welsh mountain ponies were randomly assigned to three groups of five. A single dose of a recombinant ALVAC vaccine was administered intramuscularly to five of the ponies, two doses, administered five weeks apart, were administered to five, and the other five served as unvaccinated, challenge controls. Two weeks after the completion of the vaccination programme, the ponies were all challenged by exposure to an aerosol of influenza virus A/eq/Newmarket/5/03. Their clinical signs were scored daily for 14 days according to a standardised scoring protocol, and nasal swabs were taken daily for 10 days to monitor the excretion of virus. The challenge produced severe clinical signs of influenza (fever, coughing, nasal discharge and dyspnoea) in all five control ponies, but the vaccinated ponies developed only mild disease, consisting of a serous nasal discharge lasting for only one day. The excretion of virus was almost completely suppressed in the vaccinated ponies, but the control ponies shed the virus for up to seven days after the challenge.     

Derivation and characterization of a live attenuated equine influenza vaccine virus

OBJECTIVE: To develop and characterize a cold-adapted live attenuated equine-2 influenza virus effective as an intranasal vaccine. ANIMALS: 8 ponies approximately 18 months of age. PROCEDURES: A wild-type equine-2 virus, A/Equine/Kentucky/1/91 (H3N8), was serially passaged in embryonated chicken eggs at temperatures gradually reduced in a stepwise manner from 34 C to 30 C to 28 C to 26 C. At different passages, infected allantoic fluids were tested for the ability of progeny virus to replicate in Madin-Darby canine kidney (MDCK) cells at 34 C and 39.5 C. Virus clones that replicated at 26 C in eggs and at 34 C in MDCK cells, but not at 39.5 C in MDCK cells, were tested for stability of the cold-adapted, temperature-sensitive (ts), and protein synthesis phenotypes. A stable clone, P821, was evaluated for safety, ability to replicate, and immunogenicity after intranasal administration in ponies. RESULTS: Randomly selected clones from the 49th passage were all ts with plaquing efficiencies of < 10(-6) (ratio of 39.5 C:34 C) and retained this phenotype after 5 serial passages at 34 C in either embryonated eggs or MDCK cells. The clone selected as the vaccine candidate (P821) had the desired degree of attenuation. Administered intranasally to seronegative ponies, the virus caused no adverse reactions or overt signs of clinical disease, replicated in the upper portion of the respiratory tract, and induced a strong serum antibody response. CONCLUSION AND CLINICAL RELEVANCE: A candidate live attenuated influenza vaccine virus was derived by cold-adaptation of a wild-type equine-2 influenza virus, A/Equine/Kentucky/1/91, in embryonated eggs.     

Efficacy and duration of immunity of a combined equine influenza and equine herpesvirus vaccine against challenge with an American-like equine influenza virus (A/equi-2/Kentucky/95)

It has been recommended that modern equine influenza vaccines should contain an A/equi-1 strain and A/equi-2 strains of the American and European-like subtype. We describe here the efficacy of a modern updated inactivated equine influenza-herpesvirus combination vaccine against challenge with a recent American-like isolate of equine influenza (A/equine-2/Kentucky/95 (H3N8). The vaccine contains inactivated Influenza strains A-equine-1/Prague'56, A-equine-2/Newmarket-1/'93 (American lineage) and A-equine-2/ Newmarket-2/93 (Eurasian lineage) and inactivated EHV-1 strain RacH and EHV-4 strain V2252. It is adjuvanted with alhydrogel and an immunostim. Horses were vaccinated at the start of the study and 4 weeks later. Four, six and eight weeks after the first vaccination high anti-influenza antibody titres were found in vaccinated horses, whereas at the start of the study all horses were seronegative. After the challenge, carried out at 8 weeks after the first vaccination, nasal swabs were taken, rectal temperatures were measured and clinical signs were monitored for 14 days. In contrast to unvaccinated control horses, vaccinated animals shed hardly any virus after challenge, and the appearance of clinical signs of influenza such as nasal discharge, coughing and fever were reduced in the vaccinated animals. Based on these observations, it was concluded that the vaccine protected against clinical signs of influenza and, more importantly, against virus excretion induced by an American-like challenge virus strain. In a second experiment the duration of the immunity induced by this vaccine was assessed serologically. Horses were vaccinated at the start of the study and 6 and 32 weeks later. Anti-influenza antibody titres were determined in bloodsamples taken at the first vaccination, and 2, 6, 8, 14, 19, 28, 32, 37, 41, 45 and 58 weeks after the first vaccination. Vaccinated horses had high anti-influenza antibody titres, above the level for clinical protection against influenza, against all strains present in the vaccine until 26 weeks after the third vaccination.     

A West Nile virus (WNV) recombinant canarypox virus vaccine elicits WNV-specific neutralizing antibodies and cell-mediated immune responses in the horse

Successful vaccination against West Nile virus (WNV) requires induction of both neutralizing antibodies and cell-mediated immune responses. In this study, we have assessed the ability of a recombinant ALVAC-WNV vaccine (RECOMBITEK WNV) to elicit neutralizing antibodies and virus-specific cell-mediated immune responses in horses. In addition, we examined whether prior exposure to ALVAC-WNV vaccine would inhibit B and cell-mediated immune responses against the transgene product upon subsequent booster immunizations with the same vaccine. The results demonstrated that the recombinant ALVAC-WNV vaccine induced neutralizing antibodies and prM/E insert-specific IFN-gamma(+) producing cells against WNV in vaccinated horses. Prior exposure to ALVAC-WNV vaccine did not impair the ability of horses to respond to two subsequent booster injections with the same vaccine, although anti-vector-specific antibody and cell-mediated immune responses were induced in vaccinated horses. This report describes, for the first time, the induction of antigen-specific cell-mediated responses following vaccination with an ALVAC virus recombinant vaccine encoding WNV antigens. Moreover, we showed that both WNV-specific IFN-gamma producing cells and anti-WNV neutralizing antibody responses, are not inhibited by subsequent vaccinations with the same vector vaccine.     

Comparison of two modern vaccines and previous influenza infection against challenge with an equine influenza virus from the Australian 2007 outbreak

During 2007, large outbreaks of equine influenza (EI) caused by Florida sublineage Clade 1 viruses affected horse populations in Japan and Australia. The likely protection that would be provided by two modern vaccines commercially available in the European Union (an ISCOM-based and a canarypox-based vaccine) at the time of the outbreaks was determined. Vaccinated ponies were challenged with a representative outbreak isolate (A/eq/Sydney/2888-8/07) and levels of protection were compared. A group of ponies infected 18 months previously with a phylogenetically-related isolate from 2003 (A/eq/South Africa/4/03) was also challenged with the 2007 outbreak virus. After experimental infection with A/eq/Sydney/2888-8/07, unvaccinated control ponies all showed clinical signs of infection together with virus shedding. Protection achieved by both vaccination or long-term immunity induced by previous exposure to equine influenza virus (EIV) was characterised by minor signs of disease and reduced virus shedding when compared with unvaccinated control ponies. The three different methods of virus titration in embryonated hens’ eggs, EIV NP-ELISA and quantitative RT-PCR were used to monitor EIV shedding and results were compared. Though the majority of previously infected ponies had low antibody levels at the time of challenge, they demonstrated good clinical protection and limited virus shedding. In summary, we demonstrate that vaccination with current EIV vaccines would partially protect against infection with A/eq/Sydney/2888-8/07-like strains and would help to limit the spread of disease in our vaccinated horse population.     

Use of DNA and recombinant canarypox viral (ALVAC) vectors for equine herpes virus vaccination

In this study, experimental canarypox virus (ALVAC) and plasmid DNA recombinant vaccines expressing the gB, gC and gD glycoproteins of EHV-1 were assessed for their ability to protect conventional ponies against a respiratory challenge with EHV-1. In addition, potential means of enhancing serological responses in horses to ALVAC and DNA vaccination were explored. These included co-administration of the antigen with conventional adjuvants, complexation with DMRIE-DOPE and co-expression of the antigen along with equine GM-CSF. Groups of EHV primed ponies were vaccinated twice intra-muscularly with one dose of the appropriate test vaccine at an interval of 5 weeks. Two to 3 weeks after the second vaccination, ponies were infected intra-nasally with the virulent Ab4 strain of EHV-1 after which they were observed clinically and sampled for virological investigations. The results demonstrated that DNA and ALVAC vaccination markedly reduced virus excretion after challenge in terms of duration and magnitude, but failed to protect against cell-associated viremia. Noteworthy was the almost complete absence of virus excretion in the group of ponies vaccinated with ALVAC-EHV in the presence of Carbopol adjuvant or DNA plasmid formulated with aluminium phosphate. The administration of the DNA vaccine in the presence of GM-CSF and formulated in DMRIE-DOPE and of the ALVAC vaccine in the presence of Carbopol adjuvant significantly improved virus neutralising antibody responses to EHV-1. These findings indicate that DNA and ALVAC vaccination is a promising approach for the immunological control of EHV-1 infection, but that more research is needed to identify the immunodominant protective antigens of EHV-1 and their interaction with the equine immune system.     

Antibody responses induced by Japanese whole inactivated vaccines against equine influenza virus (H3N8) belonging to Florida sublineage clade2

In 2010, the World Organisation for Animal Health recommended the inclusion of a Florida sublineage clade2 strain of equine influenza virus (H3N8), which is represented by A/equine/Richmond/1/07 (Richmond07), in equine influenza vaccines. Here, we evaluate the antigenic differences between Japanese vaccine strains and Richmond07 by performing hemagglutination inhibition (HI) assays. Ferret antiserum raised to A/equine/La Plata/93 (La Plata93), which is a Japanese vaccine strain, reacted with Richmond07 at a similar titer to La Plata93. Moreover, two hundred racehorses exhibited similar geometric mean HI antibody titers against La Plata93 and Richmond07 (73.1 and 80.8, respectively). Therefore, we can expect the antibody induced by the current Japanese vaccines to provide some protection against Richmond07-like viruses.     

Efficacy of a canarypox-vectored recombinant vaccine expressing the hemagglutinin gene of equine influenza H3N8 virus in the protection of ponies from viral challenge

OBJECTIVE: To determine onset and duration of immunity provided by a 2- or 3-dose series of a new canarypox-vectored recombinant vaccine for equine influenza virus (rCP-EIV vaccine) expressing the hemagglutinin genes of influenza H3N8 virus strains A/eq/Kentucky/94 and A/eq/Newmarket/2/93 in ponies. ANIMALS: Forty-nine 1- to 3-year-old male Welsh Mountain Ponies that were seronegative for equine influenza virus. PROCEDURES: Vaccinated and control ponies were challenged with aerosolized influenza virus A/eq/Sussex/89 (H3N8), representative of the Eurasian lineage of circulating influenza viruses. In trial 1, control ponies and ponies that received rCP-EIV vaccine were challenged 2 weeks after completion of the 2-dose primary vaccination program. In trial 2, ponies were challenged 5 months after 2 doses of rCP-EIV vaccine or 1 year after the first boosting dose of rCP-EIV vaccine, administered 5 months after completion of the primary vaccination program. After challenge, ponies were observed daily for clinical signs of influenza and nasal swab specimens were taken to monitor virus excretion. RESULTS: The challenge reliably produced severe clinical signs consistent with influenza infection in the control ponies, and virus was shed for up to 7 days. The vaccination protocol provided clinical and virologic protection to vaccinates at 2 weeks and 5 months after completion of the primary vaccination program and at 12 months after the first booster. CONCLUSION AND CLINICAL RELEVANCE: The rCP-EIV vaccine provided protection of ponies to viral challenge. Of particular importance was the protection at 5 months after the second dose, indicating that this vaccine closes an immunity gap between the second and third vaccination.     

Efficacy of a cold-adapted, intranasal, equine influenza vaccine: challenge trials.

A randomised, controlled, double-blind, influenza virus, aerosol challenge of horses was undertaken to determine the efficacy of a cold-adapted, temperature sensitive, modified-live virus, intranasal, equine influenza vaccine. Ninety 11-month-old influenza-na?ve foals were assigned randomly to 3 groups (20 vaccinates and 10 controls per group) and challenged 5 weeks, 6 and 12 months after a single vaccination. Challenges were performed on Day 0 in a plastic-lined chamber. Between Days 1 and 10, animals were examined daily for evidence of clinical signs of influenza. Nasal swabs for virus isolation were obtained on Day 1 and Days 1 to 8 and blood samples for serology were collected on Days 1, 7 and 14. There was no adverse response to vaccination in any animal. Following challenge at 5 weeks and 6 months, vaccinates had significantly lower clinical scores (P = 0.0001 and 0.005, respectively), experienced smaller increases in rectal temperature (P = 0.0008 and 0.0007, respectively) and shed less virus (P<0.0001 and P = 0.03, respectively) over fewer days (P<0.0001 and P = 0.002, respectively) than did the controls. After the 12 month challenge, rectal temperatures (P = 0.006) as well as the duration (P = 0.03) and concentration of virus shed (P = 0.04) were significantly reduced among vaccinated animals. The results of this study showed that 6 months after a single dose of vaccine the duration and severity of clinical signs were markedly reduced amongst vaccinated animals exposed to a severe live-virus challenge. Appropriate use of this vaccine should lead to a marked reduction in the frequency, severity and duration of outbreaks of equine influenza in North America.     

A new modified live equine influenza virus vaccine: phenotypic stability, restricted spread and efficacy against heterologous virus challenge.

Flu Avert IN vaccine is a new, live attenuated virus vaccine for equine influenza. We tested this vaccine in vivo to ascertain 1) its safety and stability when subjected to serial horse to horse passage, 2) whether it spread spontaneously from horse to horse and 3) its ability to protect against heterologous equine influenza challenge viruses of epidemiological relevance. For the stability study, the vaccine was administered to 5 ponies. Nasal swabs were collected and pooled fluids administered directly to 4 successive groups of na?ve ponies by intranasal inoculation. Viruses isolated from the last group retained the vaccine's full attenuation phenotype, with no reversion to the wild-type virus phenotype or production of clinical influenza disease. The vaccine virus spread spontaneously to only 1 of 13 nonvaccinated horses/ponies when these were comingled with 39 vaccinates in the same field. For the heterologous protection study, a challenge model system was utilised in which vaccinated or na?ve control horses and ponies were exposed to the challenge virus by inhalation of virus-containing aerosols. Challenge viruses included influenza A/equine-2/Kentucky/98, a recent representative of the 'American' lineage of equine-2 influenza viruses; and A/equine-2/Saskatoon/90, representative of the 'Eurasian' lineage. Clinical signs among challenged animals were recorded daily using a standardised scoring protocol. With both challenge viruses, control animals reliably contracted clinical signs of influenza, whereas vaccinated animals were reliably protected from clinical disease. These results demonstrate that Flu Avert IN vaccine is safe and phenotypically stable, has low spontaneous transmissibility and is effective in protecting horses against challenge viruses representative of those in circulation worldwide.     

Antibody and T cell responses induced in chickens immunized with avian influenza virus N1 and NP DNA vaccine with chicken IL-15 and IL-18

We had examined the immunogenicity of a series of plasmid DNAs which include neuraminidase (NA) and nucleoprotein (NP) genes from avian influenza virus (AIV). The interleukin-15 (IL-15) and interleukin-18 (IL-18) as genetic adjuvants were used for immunization in combination with the N1 and NP AIV genes. In the first trial, 8 groups of chickens were established with 10 specific-pathogen-free (SPF) chickens per group while, in the second trial 7 SPF chickens per group were used. The overall N1 enzyme-linked immunosorbent assay (ELISA) titer in chickens immunized with the pDis/N1 + pDis/IL-15 was higher compared to the chickens immunized with the pDis/N1 and this suggesting that chicken IL-15 could play a role in enhancing the humoral immune response. Besides that, the chickens that were immunized at 14-day-old (Trial 2) showed a higher N1 antibody titer compared to the chickens that were immunized at 1-day-old (Trial 1). Despite the delayed in NP antibody responses, the chickens co-administrated with IL-15 were able to induce earlier and higher antibody response compared to the pDis/NP and pDis/NP + pDis/IL-18 inoculated groups. The pDis/N1 + pDis/IL-15 inoculated chickens also induced higher CD8+ T cells increase than the pDis/N1 group in both trials (P < 0.05). The flow cytometry results from both trials demonstrated that the pDis/N1 + pDis/IL-18 groups were able to induce CD4+ T cells higher than the pDis/N1 group (P < 0.05). Meanwhile, pDis/N1 + pDis/IL-18 group was able to induce CD8+ T cells higher than the pDis/N1 group (P < 0.05) in Trial 2 only. In the present study, pDis/NP was not significant (P > 0.05) in inducing CD4+ and CD8+ T cells when co-administered with the pDis/IL-18 in both trials in comparison to the pDis/NP. Our data suggest that the pDis/N1 + pDis/IL-15 combination has the potential to be used as a DNA vaccine against AIV in chickens.     

Analysis of the quality of protection induced by a porcine influenza A vaccine to challenge with an H3N2 virus

Antigenic drift of swine influenza A (H3N2) viruses away from the human A/Port Chalmers/1/73 (H3N2) strain, used in current commercial swine influenza vaccines, has been demonstrated in The Netherlands and Belgium. Therefore, replacement of this human strain by a more recent swine H3N2 isolate has to be considered. In this study, the efficacy of a current commercial swine influenza vaccine to protect pigs against a recent Dutch field strain (A/Sw/Oedenrode/96) was assessed. To evaluate the level of protection induced by the vaccine it was compared with the optimal protection induced by a previous homologous infection. Development of fever, virus excretion, and viral transmission to unchallenged group mates were determined to evaluate protection. The vaccine appeared efficacious in the experiment because it was able to prevent fever and virus transmission to the unchallenged group mates. Nevertheless, the protection conferred by the vaccine was sub-optimal because vaccinated pigs excreted influenza virus for a short period of time after challenge, whereas naturally immune pigs appeared completely protected. The immune response was monitored, to investigate why the vaccine conferred a sub-optimal protection. The haemagglutination inhibiting and virus neutralising antibody responses in sera, the nucleoprotein-specific IgM, IgG, and IgA antibody responses in sera and nasal secretions and the influenza-specific lymphoproliferation responses in the blood were studied. Vaccinated pigs developed the same or higher serum haemagglutination inhibiting, virus neutralising, and nucleoprotein-specific IgG antibody titres as infected pigs but lower nasal IgA titres and lymphoproliferation responses. The lower mucosal and cell-mediated immune responses may explain why protection after vaccination was sub-optimal.     

Duration of circulating antibody and immunity following infection with equine influenza virus

The duration of immunity as measured by virological, serological and clinical responses following infection with influenza A/equine/Newmarket/79 (H3N8) was assessed in repeated challenge experiments in which ponies were infected by exposure to aerosols of infectious virus. Previous infection stimulated complete clinical protection which persisted for at least 32 weeks as demonstrated by the absence of febrile responses and coughing in two groups of ponies infected 16 weeks or 32 weeks after the first infection. Partial clinical protection persisted for over a year as demonstrated by the absence of coughing and a reduction in the number of febrile responses in a group of ponies infected 62 weeks after their first infection. These results contrasted with those observed in immunologically naive control ponies which developed pyrexia, dyspnoea and nasal discharge and coughing. The kinetics of virus specific antibody production in primary and secondary infections with equine influenza were studied by the single radial haemolysis test and a radioisotopic antiglobulin binding assay which measured virus specific IgGab antibody isotype. Antibody to the haemagglutinin, as measured by the single radial haemolysis test, declined rapidly after primary infection whereas the IgGab responses to whole virus antigens persisted for longer. The single radial haemolysis test was therefore particularly useful for the detection of antibody responses in multiple infections or exposures to influenza antigens. The radioisotopic antiglobulin binding assay was more sensitive for identifying infections which had occurred more than six months previously, as evidenced by anamnestic IgGab responses in ponies with low levels of antibody before rechallenge.