Influenza H5/H7 Virus Vaccination in Poultry and Reduction of Zoonotic Infections, Guangdong Province, China, 2017-18.

We compared the detection frequency of avian influenza H7 subtypes at live poultry markets in Guangdong Province, China, before and after the introduction of a bivalent H5/H7 vaccine in poultry. The vaccine was associated with a 92% reduction in H7 positivity rates among poultry and a 98% reduction in human H7N9 cases.

Immunization with live nonpathogenic H5N3 duck influenza virus protects chickens against highly pathogenic H5N1 virus.

Development of an effective, broadly-active and safe vaccine for protection of poultry from H5N1 highly pathogenic avian influenza viruses (HPAIVs) remains an important practical goal. In this study we used a low pathogenic wild aquatic bird virus isolate А/duck/Moscow/4182/2010 (H5N3) (dk/4182) as a live candidate vaccine. We compared this virus with four live 1:7 reassortant anti-H5N1 candidate vaccine viruses with modified hemagglutinin from either A/Vietnam/1203/04 (H5N1) or A/Kurgan/3/05 (H5N1) and the rest of the genes from either H2N2 cold-adapted master strain A/Leningrad/134/17/57 (rVN-Len and rKu-Len) or H6N2 virus A/gull/Moscow/3100/2006 (rVN-gull and rKu-gull). The viruses were tested in parallel for pathogenicity, immunogenicity and protective effectiveness in chickens using aerosol, intranasal and oral routes of immunization. All five viruses showed zero pathogenicity indexes in chickens. Viruses rVN-gull and rKu-gull were immunogenic and protective, but they were insufficiently attenuated and caused significant mortality of 1-day-old chickens. The viruses with cold-adapted backbones (rVN-Len and rKu-Len) were completely nonpathogenic, but they were significantly less immunogenic and provided lower protection against lethal challenge with HPAIV A/Chicken/Kurgan/3/05 (H5N1) as compared with three other vaccine candidates. Unlike other four viruses, dk/4182 was both safe and highly immunogenic in chickens of any age regardless of inoculation route. Single administration of 106 TCID50 of dk/4182 virus via drinking water provided complete protection of 30-days-old chickens from 100 LD50 of the challenge virus. Our results suggest that low pathogenic viruses of wild aquatic birds can be used as safe and effective live poultry vaccines against highly pathogenic avian viruses.

The impact of deposition site on vaccination efficiency of a live bacterial poultry vaccine.

Vaccines are utilized within the poultry industry to minimize disease-associated losses and spray vaccination is a commonly utilized means for the mass application of poultry vaccines. During this process, vaccine-laden particles are deposited upon target areas (e.g., eyes, nares, and oral cavity) resulting in the direct internalization of the vaccine. However, particles are also deposited on nontarget areas such as the exterior of the subject and its surrounding environment. To better determine the fate of particles deposited upon nontarget areas and the impact of deposition site on the efficiency of vaccine application, a live bacterial poultry vaccine (AviPro(®) MG F) was applied via spray using a spray cabinet with a slotted partition allowing for head-only, body-only, and whole-bird spray application. At 11 wk age, Hy-Line(®) W-36 pullets (n = 280) were allocated equally among 7 treatments including: nonvaccinated controls, pullets spray-vaccinated at the manufacturer's recommended dose (1X) in a site-specific manner (head-only, body-only, and whole-bird), pullets spray-vaccinated at 5X the recommended level (body-only), pullets vaccinated by manual eye-drop application (1X), and pullets eye-drop vaccinated at a level approximating that achieved during the spray vaccination process (1/700X). At 6 to 7 wk postvaccination, vaccination efficiency was assessed via serological-based assays [serum plate agglutination (SPA) and ELISA] and the detection of vaccine-derived in vivo populations. Results indicate an additive contribution of the vaccine deposited on the body to the overall vaccination efficiency of this live bacterial live poultry vaccine.

The production of Newcastle disease virus-like particles in Nicotiana benthamiana as potential vaccines.

Newcastle disease (ND) is a highly contagious viral respiratory and neurological disease that has a severe impact on poultry production worldwide. In the present study, an expression platform was established for the transient production in N.bethamiana of ND virus-like particles (VLPs) for use as vaccines against ND. The expression of the ND Fusion (F) and/or Hemagglutinin-neuraminidase (HN) proteins of a genotype VII.2 strain formed ND VLPs in planta as visualized under the transmission electron microscope, and HN-containing VLPs agglutinated chicken erythrocytes with hemagglutination (HA) titres of up to 13 log2.The immunogenicity of the partially-purified ND VLPs was confirmed in specific-pathogen-free White leghorn chickens. Birds receiving a single intramuscular immunization with 1024 HA units (10 log2) of the F/HN ND VLPs administered with 20% [v/v] Emulsigen®-P adjuvant, seroconverted after 14 days with F- and HN-specific antibodies at ELISA titres of 5705.17 and HI geometric mean titres (GMTs) of 6.2 log2, respectively. Furthermore, these ND-specific antibodies successfully inhibited viral replication in vitro of two antigenically closely-related ND virus isolates, with virus-neutralization test GMTs of 3.47 and 3.4, respectively. Plant-produced ND VLPs have great potential as antigen-matched vaccines for poultry and other avian species that are highly immunogenic, cost-effective, and facilitate prompt updating to ensure improved protection against emerging ND field viruses.

Cross-Protection Conferred by Combined Vaccine Containing Infectious Bronchitis Virus Attenuated Massachusetts and Recombinant LaSota Virus Expressing Arkansas Spike.

We previously demonstrated that a prime-boost regime with an infectious bronchitis virus (IBV) Massachusetts (Mass)-type vaccine and recombinant LaSota virus (rLS) coexpressing IBV Arkansas (Ark)-type trimeric spike ectodomain (Se) and granulocyte macrophage colony stimulating factor (GMCSF) enhances heterologous protection against virulent Ark challenge. This study evaluates protection against Ark-type challenge conferred by administering the rLS/ArkSe.GMCSF and the attenuated Mass viruses mixed in the same vial as a combined vaccine. Chickens were vaccinated at day of hatch and challenged at 21 days of age with virulent Ark. Protection conferred by vaccination was assessed by respiratory signs, tracheal virus isolation as well as IBV RNA quantitation, and tracheal histomorphometry. Protection conferred by the combined vaccine was compared to protection induced by a commercial attenuated ArkDPI (Delmarva Poultry Industry) vaccine as well as by the attenuated Mass vaccine alone. Vaccination with the combined vaccine (rLS/ArkSe.GMCSF + Mass) as well as Mass alone provided significantly less protection against Ark challenge compared to the control using attenuated live ArkDPI vaccine. Only ArkDPI-vaccinated chickens exhibited "sterilizing immunity," i.e., no virus isolated from ≥10% of chickens after challenge. Chickens vaccinated with the combined vaccine rLS/ArkSe.GMCSF + Mass showed significantly less tracheal damage than birds vaccinated with the attenuated Mass vaccine alone. In addition, the combined vaccine also resulted in less virus isolation from the trachea. We concluded that the combined vaccine containing the recombinant virus and the attenuated Mass enhanced the cross-protective ability of the attenuated Mass vaccine against heterologous challenge.

Nota de investigación- Protección cruzada conferida por una vacuna combinada que contiene el virus atenuado de la bronquitis infecciosa serotipo Massachusetts y un virus de Newcastle LaSota recombinante que expresa la proteína de la espícula del serotipo Arkansas. Previamente se demostró que un esquema de refuerzo con una vacuna tipo Massachusetts (Mass) del virus de la bronquitis infecciosa (IBV) y con un virus de Newcastle LaSota recombinante (rLS) que co-expresa el ectodominio de pico trimérico del serotipo Arkansas (Ark) del virus de la bronquitis infecciosa (Se) y el factor de estimulación de colonias de granulocitos y macrófagos (GMCSF) (virus rLS/ArkSe.GMCSF) mejora la protección heteróloga contra el desafío con serotipo Arkansas virulento. Este estudio evaluó la protección contra el desafío con el serotipo Arkansas conferida por la administración del virus recombinante rLS/ArkSe.GMCSF y el virus Massachussets atenuado mezclados en el mismo vial como una vacuna combinada. Los pollos se vacunaron el día de la eclosión y se desafiaron a los 21 días de edad con el serotipo Arkansas virulento. La protección conferida por la vacunación se evaluó mediante signos respiratorios, el aislamiento del virus traqueal, cuantificación del ARN del virus de bronquitis y por histomorfometría traqueal. La protección conferida por la vacuna combinada se comparó con la protección inducida por una vacuna comercial atenuada ArkDPI (Delmarva Poultry Industry), y con la vacuna atenuada Massachussets por si sola. La vacunación con la vacuna combinada (rLS/ArkSe.GMCSF + Mass), así como con Massachussets sola, proporcionó una protección significativamente menor contra la exposición con Arkansas en comparación con el control que utilizó la vacuna ArkDPI viva atenuada. Solo los pollos vacunados con ArkDPI exhibieron "inmunidad esterilizante", es decir, que no se aisló ningún virus en ≥10 % de los pollos después del desafío. Los pollos vacunados con la vacuna combinada rLS/ArkSe.GMCSF + Mass mostraron significativamente menos daño traqueal que las aves vacunadas con la vacuna Massachusetts atenuada sola. Además, la vacuna combinada también resultó en un menor aislamiento del virus de la tráquea. Se concluye que la vacuna combinada que contiene el virus recombinante y el serotipo Massachussets atenuado mejoró la capacidad de protección cruzada de la vacuna de Massachusetts atenuada contra el desafío heterólogo.

Efficacy of inactivated and RNA particle vaccines against a North American Clade 2.3.4.4b H5 highly pathogenic avian influenza virus in chickens.

Highly pathogenic avian influenza virus (HPAIV) has caused widespread outbreaks in poultry in the Americas. Because of the duration and extent of these outbreaks, vaccine use may be an additional tool to limit virus spread. Three vaccines were evaluated for efficacy in chickens against a current North American clade 2.3.4.4b H5 HPAIV isolate, A/turkey/Indiana/3703-003/2022 H5N1. The vaccines included: 1) a commercial inactivated reverse genetics (rg) generated H5N1 product with a clade 2.3.4.4c H5 hemagglutinin (HA) (rgH5N1); 2) a commercial alphavirus RNA particle (RP) vaccine with the TK/IN/22 HA; and 3) an in-house inactivated rg produced vaccine with the TK/IN/22 HA and a North American lineage N9 neuraminidase (NA) (SEP-22-N9). Both inactivated vaccines were produced with HA genes that were modified to be low pathogenic and with the remaining genes from the PR8 influenza strain. All vaccines provided 100% protection against mortality and morbidity and all vaccines reduced virus shed by the oropharyngeal and cloacal routes significantly compared to sham vaccinates. However, differences were observed among the vaccines in quantities of virus shed at two- and four-days post challenge (DPC). To determine if infected birds could be identified after vaccination to aid surveillance programs, serum was collected from the RP and SEP-22-N9 vaccine groups at 7, 10, and 14 DPC to detect antibody to the NA and nucleoprotein (NP) of the challenge virus by enzyme linked lectin assay (ELLA) and ELISA. As early as 7DPC ELLA detected antibody in sera from 100% of the chickens in the RP vaccinated group and 70% of the chickens in the SEP-22-N9 vaccinated group. Antibody to the NP was detected by commercial ELISA in more than 50% of the birds in the RP vaccinated group at each time point.

A Computational Reverse Vaccinology Approach for the Design and Development of Multi-Epitopic Vaccine Against Avian Pathogen Mycoplasma gallisepticum.

Avian mycoplasma is a bacterial disease causing chronic respiratory disease (CRD) in poultry industries with high economic losses. The eradication of this disease still remains as a challenge. A multi-epitope prophylactic vaccine aiming the antigenic proteins of Mycoplasma gallisepticum can be a capable candidate to eradicate this infection. The present study is focused to design a multi-epitope vaccine candidate consisting of cytotoxic T-cell (CTL), helper T-cell (HTL), and B-cell epitopes of antigenic proteins, using immunoinformatics strategies. The multi-epitopic vaccine was designed, and its tertiary model was predcited, which was further refined and validated by computational tools. After initial validation, molecular docking was performed between multi-epitope vaccine construct and chicken TLR-2 and 5 receptors, which predicted effective binding. The in silico results specify the structural stability, precise specificity, and immunogenic response of the designed multi-epitope vaccine, and it could be an appropriate vaccine candidate for the M. gallisepticum infection.

Enhanced Protection by Recombinant Newcastle Disease Virus Expressing Infectious Bronchitis Virus Spike Ectodomain and Chicken Granulocyte-Macrophage Colony-Stimulating Factor.

We previously reported that recombinant Newcastle disease virus LaSota (rLS) expressing infectious bronchitis virus (IBV) Arkansas (Ark)-type trimeric spike (S) ectodomain (Se; rLS/ArkSe) provides suboptimal protection against IBV challenge. We have now developed rLS expressing chicken granulocyte-macrophage colony-stimulating factor (GMCSF) and IBV Ark Se in an attempt to enhance vaccine effectiveness. In the current study, we first compared protection conferred by vaccination with rLS/ArkSe and rLS/ArkSe.GMCSF. Vaccinated chickens were challenged with virulent Ark, and protection was determined by clinical signs, viral load, and tracheal histomorphometry. Results showed that coexpression of GMCSF and the Se from rLS significantly reduced tracheal viral load and tracheal lesions compared with chickens vaccinated with rLS/ArkSe. In a second experiment, we evaluated enhancement of cross-protection of a Massachusetts (Mass) attenuated vaccine by priming or boosting with rLS/ArkSe.GMCSF. Vaccinated chickens were challenged with Ark, and protection was evaluated. Results show that priming or boosting with the recombinant virus significantly increased cross-protection conferred by Mass against Ark virulent challenge. Greater reductions of viral loads in both trachea and lachrymal fluids were observed in chickens primed with rLS/ArkSe.GMCSF and boosted with Mass. Consistently, Ark Se antibody levels measured with recombinant Ark Se protein-coated ELISA plates 14 days after boost were significantly higher in these chickens. Unexpectedly, the inverse vaccination scheme, that is, priming with Mass and boosting with the recombinant vaccine, proved somewhat less effective. We concluded that a prime and boost strategy by using rLS/ArkSe.GMCSF and the worldwide ubiquitous Mass attenuated vaccine provides enhanced cross-protection. Thus, rLS/GMCSF coexpressing the Se of regionally relevant IBV serotypes could be used in combination with live Mass to protect against regionally circulating IBV variant strains.

Protección incrementada por el virus recombinante de la enfermedad de Newcastle que expresa el ectodominio de la espícula del virus de la bronquitis infecciosa y el factor estimulante de colonias de granulocitos y macrófagos del pollo. Anteriormente se reportó que la cepa LaSota recombinante del virus de la enfermedad de Newcastle (rLS) que expresa el ectodominio de la espícula trimérica (S) de tipo Arkansas (Ark) del virus de la bronquitis infecciosa (IBV) (Se; rLS/ArkSe) proporciona una protección subóptima contra la exposición al virus de la bronquitis infecciosa. Ahora se ha desarrollado hemos desarrollado una cepa LaSota recombinante (rLS) que expresa el factor estimulante de colonias de granulocitos y macrófagos de pollo (GMCSF) y la espícula del virus de bronquitis Arkansas en un intento para mejorar la efectividad de la vacuna. En el estudio actual, primero se comparó la protección conferida por la vacunación con los virus rLS/ArkSe y rLS/ArkSe.GMCSF. Los pollos vacunados se desafiaron con un virus Arkansas virulento y la protección se determinó mediante los signos clínicos, la carga viral y la histomorfometría de la tráquea. Los resultados mostraron que la coexpresión del factor estimulante de colonias de granulocitos y macrófagos de pollo y la espícula de la cepa recombinante LaSota redujo significativamente la carga viral traqueal y las lesiones traqueales en comparación con los pollos vacunados con el virus rLS/ArkSe. En un segundo experimento, se evaluó el incremento en la protección cruzada por una vacuna atenuada de Massachusetts (Mass) mediante la primovacunación o la vacunación de refuerzo con rLS/ArkSe.GMCSF. Los pollos vacunados fueron desafiados con el virus Arkansas y se evaluó la protección. Los resultados mostraron que la primovacunación o la vacunación de refuerzo con el virus recombinante aumentó significativamente la protección cruzada conferida por el virus Massachusetts contra el desafío virulento con el virus Arkansas. Se observaron mayores reducciones de las cargas virales en los fluidos traqueales y lagrimales en pollos primovacunadoss con rLS/ArkSe.GMCSF y con vacunación de refuerzo con Massachusetts. De manera consistente, los niveles de anticuerpos Ark Se medidos con placas de ELISA recubiertas con proteína Ark Se recombinante a los 14 días después del refuerzo fueron significativamente más altos en estos pollos. De manera inesperada, el esquema de vacunación inverso, es decir, la primovacunación con Massachusetts y el refuerzo con la vacuna recombinante, resultó menos efectivo. Se concluye que una estrategia de primovacunación y refuerzo mediante el uso de rLS/ArkSe.GMCSF y la vacuna atenuada con Massachusetts usada en todo el mundo proporciona una protección cruzada aumentada. Por tanto, el virus rLS/GMCSF que coexpresa la proteína de la espícula de los serotipos regionales relevantes de bronquitis infecciosa podría usarse en combinación con una vacuna viva Massachusetts para proteger contra cepas variantes del virus de la bronquitis infecciosa que circulan regionalmente.

Macrophage Activation Assays to Evaluate the Immunostimulatory Capacity of Avibacterium paragallinarum in A Multivalent Poultry Vaccine.

High-quality vaccines are crucial to prevent infectious disease outbreaks in the poultry industry. In vivo vaccination tests are routinely used to test poultry vaccines for their potency, i.e., their capacity to induce protection against the targeted diseases. A better understanding of how poultry vaccines activate immune cells will facilitate the replacement of in vivo potency tests for in vitro assays. Using the chicken macrophage-like HD11 cell line as a model to evaluate innate immune responses, the current explorative study addresses the immunostimulatory capacity of an inactivated multivalent vaccine for infectious bronchitis, Newcastle disease, egg-drop syndrome, and infectious coryza. The vaccine stimulated HD11 cells to produce nitric oxide and to express pro-inflammatory cytokines IL-1ß, TNF, and IL-12p40, chemokines CXCLi1 and CXCLi2, and the anti-inflammatory cytokine IL-10, but only when inactivated Avibacterium paragallinarum, the causative agent of infectious coryza, was present. Lipopolysaccharides from Avibacterium paragallinarum were crucial for the production of nitric oxide and expression of IL-1ß and CXCLi1. The described immune parameters demonstrate the capacity of this multivalent vaccine to activate innate immune cells and may in the future, combined with antigen quantification methods, contribute to vaccine quality testing in vitro, hence the replacement of current in vivo vaccination tests.

Early Poultry Vaccine Company Development: the Era of Entrepreneurs.

Early events leading to the establishment of various branches of the U.S. poultry industry and the animal health industry in the United States are reviewed. Entrepreneurs see needs in a developing market and take action to meet these needs. Their skills are suited to organize people and processes to supply these needs. As the early poultry industry consolidated, this was followed by technical education as a marketing tool to expand access to the customer base. Focus will be on two entrepreneurial individuals not well covered by prior history articles, namely Hiram Lasher and Kenneth Eskelund. A discussion of how industries pass from an entrepreneurial phase to a corporate phase will follow, thus explaining the corporate- and biotechnology-oriented vaccine companies we see today.

Artículo histórico - Desarrollo temprano de las compañías de vacunas avícolas: la era de los empresarios. Se revisaron los primeros eventos que llevaron al establecimiento de varias ramas de la industria avícola y de la industria de la salud animal en los Estados Unidos. Los empresarios detectaron las necesidades en un mercado en desarrollo y tomaron medidas para satisfacer dichas necesidades. Sus habilidades fueron adecuadas para organizar personas y procesos para satisfacer estas necesidades. A medida que se consolidó la industria avícola temprana, esto fue seguido por la educación técnica como una herramienta de mercadeo para ampliar el acceso a la base de clientes. La atención se centró en dos personas emprendedoras que no han sido adecuadamente analizadas por los artículos históricos anteriores como son, Hiram Lasher y Kenneth Eskelund. Se explicó cómo las industrias pasaron de una fase empresarial a una fase corporativa, explicando así las compañías de vacunas orientadas a la biotecnología y las empresas actuales.

Evaluation of a toxoid fusion protein vaccine produced in plants to protect poultry against necrotic enteritis.

BACKGROUND: Necrotic enteritis (NE) is caused by type A strains of the bacterium Clostridium perfringens. Total global economic losses to the poultry industry due to NE is estimated to be over two billion dollars annually. Traditionally, NE has been effectively controlled by inclusion of antibiotics in the diet of poultry. However, recent concerns regarding the impact of this practice on increasing antibiotic resistance in human pathogens have led us to consider alternative approaches, such as vaccination, for controlling this disease. NE strains of C. perfringens produce two major toxins, a-toxin and NetB. Immune responses against either toxin can provide partial protection against NE. METHODS: We have developed a fusion protein combining a non-toxic carboxyl-terminal domain of a-toxin (PlcC) and an attenuated, mutant form of NetB (NetB-W262A) for use as a vaccine antigen to immunize poultry against NE. We utilized a DNA sequence that was codon-optimized for Nicotiana benthamiana to enable high levels of expression. The 6-His tagged PlcC-NetB fusion protein was synthesized in N. benthamiana using a geminiviral replicon transient expression system, purified by metal affinity chromatography, and used to immunize broiler birds. RESULTS: Immunized birds produced a strong serum IgY response against both the plant produced PlcC-NetB protein and against bacterially produced His-PlcC and His-NetB. Immunized birds were significantly protected against a subsequent in-feed challenge with virulent C. perfringens when treated with the fusion protein. These results indicate that a plant-produced PlcC-NetB toxoid is a promising vaccine candidate for controlling NE in poultry.

Field evaluation of common poultry viral vaccinesin Egypt: a need for reassessment of the vaccine value chain.

Egypt has a large traditional and exotic poultry sector which is challenged regularly by poultry diseases in endemic and epidemic proportions. The household poultry in particular is a source of livelihoods and employment for millions of low income citizens. Highly pathogenic avian influenza (HPAI) H5N1 and Newcastle disease are the most important poultry diseases in this sector. Whereas poultry vaccines are available to reduce the incidence of disease in Egypt, their effectiveness is doubtful. We conducted a biological evaluation of selected viral vaccines of poultry in three governorates in Egypt. Fifty­four percent of the vaccines had reduced vaccine titres and the effect of secondary vaccine distributions was associated with the observed vaccine titres. External contamination was observed in some vaccines and break in cold chain was reported. Whereas no vaccine distributor used purpose­built vaccine refrigerator, none also had prescribed protocol for vaccine handling or kept record of vaccine. There is a need to review vaccine handling procedure, monitor of vaccine cold chain more critically and review the whole chain that support vaccine distributions in Egypt.

Immunization of Domestic Ducks with Live Nonpathogenic H5N3 Influenza Virus Prevents Shedding and Transmission of Highly Pathogenic H5N1 Virus to Chickens.

Wild ducks are known to be able to carry avian influenza viruses over long distances and infect domestic ducks, which in their turn infect domestic chickens. Therefore, prevention of virus transmission between ducks and chickens is important to control the spread of avian influenza. Here we used a low pathogenic wild aquatic bird virus A/duck/Moscow/4182/2010 (H5N3) for prevention of highly pathogenic avian influenza virus (HPAIV) transmission between ducks and chickens. We first confirmed that the ducks orally infected with H5N1 HPAIV A/chicken/Kurgan/3/2005 excreted the virus in feces. All chickens that were in contact with the infected ducks became sick, excreted the virus, and died. However, the ducks orally inoculated with 104 50% tissue culture infective doses of A/duck/Moscow/4182/2010 and challenged 14 to 90 days later with H5N1 HPAIV did not excrete the challenge virus. All contact chickens survived and did not excrete the virus. Our results suggest that low pathogenic virus of wild aquatic birds can be used for prevention of transmission of H5N1 viruses between ducks and chickens.

Full-length genome sequence analysis of an avian leukosis virus subgroup J (ALV-J) as contaminant in live poultry vaccine: The commercial live vaccines might be a potential route for ALV-J transmission.

One avian leukosis virus subgroup J (ALV-J) strain was isolated from 67 commercial live poultry vaccines produced by various manufacturers during 2013-2016 in China. The complete genomes of the isolate were sequenced and it was found that the genes gag and pol of the strain were relatively conservative, while the gp85 gene of the strain GX14YYA1 had the highest similarities with a field strain GX14ZS14, which was isolated from the chickens of a farm that had once used the same vaccine as the one found to be contaminated with the GX14YYA1. This is the first report of ALV-J contaminant in live poultry vaccine in China. Our finding demonstrates that vaccination of the commercial live vaccines might be a potential new route for ALV-J transmission in chickens and highlights the need for more extensive monitoring of the commercial live vaccines in China.

Development and Evaluation of the Protective Efficacy of Novel Marek's Disease Virus Rispens Vector Vaccines Against Infectious Bursal Disease.

Infectious bursal disease (IBD) is a major disease affecting the poultry industry and is caused by infection with IBD virus (IBDV). To develop a novel vaccine to prevent IBD in chickens, recombinant Marek's disease virus Rispens viruses carrying the VP2 gene of IBDV driven by five different promoters (Rispens/IBD) were constructed using homologous recombination and a bacterial artificial chromosome (BAC). Rispens/IBD driven by the chicken beta-actin (Bac) promoter (Rispens/Bac-IBD), Rous sarcoma virus promoter, or simian virus 40 promoter were administered to 1-day-old SPF chicks, and the protective efficacy against IBDV was evaluated by challenging chicks with virulent IBDV. As a result, Rispens/Bac-IBD showed the best protection (87%). Next, we constructed the virus driven by the Bac-derived Coa5 promoter (Rispens/Coa5-IBD) for a secondary in vivo trial using commercial layer chickens since Rispens/Bac-IBD was thought to be genetically unstable. Rispens/Coa5-IBD showed stability in vitro and exhibited better antibody production and protection during challenge against virulent IBDV at both 5 (95%) and 7 wk of age (91%) compared with that of Rispens/Bac-IBD (90% at 5 wk of age and 84% at 7 wk of age). Thus, Rispens/Coa5-IBD may be a novel promising vaccine against IBD and virulent Marek's disease.

Combination of Two Marek's Disease Virus Vectors Shows Effective Vaccination Against Marek's Disease, Infectious Bursal Disease, and Newcastle Disease.

Herpesvirus of turkeys (HVT) is a widely used vector for poultry vaccines. However, different HVTs expressing different foreign antigens cannot always be used simultaneously because of the risk of recombination and interference. In this study, we inoculated a mixture of an HVT-expressing the antigen of Newcastle disease virus (NDV; HVT/ND) and Marek's disease virus (MDV) serotype 1 Rispens virus expressing the antigen of infectious bursal disease virus (IBD; Ripens/IBD) into chickens. This mixture showed 94%, 100%, or 94% protection against MDV, IBDV, or NDV challenge, respectively. In conclusion, the combination of Rispens/IBD and HVT/ND is effective for vaccination against MDV, IBDV, and NDV without significant interference.

Pulmonary immunization of chickens using non-adjuvanted spray-freeze dried whole inactivated virus vaccine completely protects against highly pathogenic H5N1 avian influenza virus.

Highly pathogenic avian influenza (HPAI) H5N1 virus is a major threat to public health as well as to the global poultry industry. Most fatal human infections are caused by contact with infected poultry. Therefore, preventing the virus from entering the poultry population is a priority. This is, however, problematic in emergency situations, e.g. during outbreaks in poultry, as there are currently no mass application methods to effectively vaccinate large numbers of birds within a short period of time. To evaluate the suitability of needle-free pulmonary immunization for mass vaccination of poultry against HPAI H5N1, we performed a proof-of-concept study in which we investigated whether non-adjuvanted spray-freeze-dried (SFD) whole inactivated virus (WIV) can be used as a dry powder aerosol vaccine to immunize chickens. Our results show that chickens that received SFD-WIV vaccine as aerosolized powder directly at the syrinx (the site of the tracheal bifurcation), mounted a protective antibody response after two vaccinations and survived a lethal challenge with HPAI H5N1. Furthermore, both the number of animals that shed challenge virus, as well as the level of virus shedding, were significantly reduced. Based on antibody levels and reduction of virus shedding, pulmonary vaccination with non-adjuvanted vaccine was at least as efficient as intratracheal vaccination using live virus. Animals that received aerosolized SFD-WIV vaccine by temporary passive inhalation showed partial protection (22% survival) and a delay in time-to-death, thereby demonstrating the feasibility of the method, but indicating that the efficiency of vaccination by passive inhalation needs further improvement. Altogether our results provide a proof-of-concept that pulmonary vaccination using an SFD-WIV powder vaccine is able to protect chickens from lethal HPAI challenge. If the efficacy of pulmonary vaccination by passive inhalation can be improved, this method might be suitable for mass application.