Respiratory disease complex is a major cause of mortality and economic losses in the commercial broiler industry. In 1991 a previously unidentified bacterium associated with respiratory disease and cranial cellulitis was isolated from broilers in the then Transvaal Province (van Beek, van Empel, van den Bosch, Storm, Bongers, du Preez, 1994. ). In 1994 the organism was named Ornithobacterium rhinotracheale (Vandamme, Segers, Vancanneyt, van Hove, Mutters, Hommez, Dewhirst, Paster, Kersters, Falsen, Devriese, Bisgaard, Hinz, Mannheim, 1994.). Since then Ornithobacterium rhinotracheale has been isolated worldwide from chickens and turkeys showing respiratory signs and has become well established as contributing to the respiratory disease complex in both species (van Empel, Hafez, 1999). In South Africa respiratory disease and Ornithobacterium rhinotracheale in particular is routinely controlled by the inclusion of antibiotics such as Oxtetracycline into the feed of broilers during rearing. Concerns about antibiotic residues in poultry meat for human consumption as well as evidence that suggests that Ornithobacterium rhinotracheale readily develops resistance to antibiotics (Devriese, Hommez, Vandamme, Kersters, Haesebrouck, 1995), make this strategy unsustainable. It was with a view to reducing producers’ dependence on long term prophylactic antibiotic therapy that this study to determine the safety and efficacy of an OR bacterin vaccine was carried out. Injection of the bacterin into broilers was deemed impractical on a commercial scale, so it was applied to broiler breeder parent stock in order that they could protect their progeny through vertically transmitted immunity developed as a result of vaccination. Breeder flocks were vaccinated intramuscularly at nine and 18 weeks with a monovalent bacterin based on OR serotype A with oil adjuvant. Vaccine safety was evaluated by palpation of vaccination sites and clinical observation of breeders for two weeks after vaccination. The serological response of breeders to vaccination was monitored using an ELISA test for Ornithobacterium rhinotracheale optimised for use under South African conditions. Vaccine efficacy was determined by monitoring of broiler progeny of vaccinated breeders raised under commercial conditions as well as through controlled challenge studies with Ornithobacterium rhinotracheale under laboratory conditions. In order to determine the financial consequences of using the test vaccine, a partial farm budget was drawn up from available broiler data and possible outcomes were modelled using a stochastic model. The vaccine proved to be safe for use in commercial broiler breeders and vaccinated birds developed a good humoral response to vaccination. As a result of cross-contamination of isolators with Ornithobacterium rhinotracheale the results of the challenge studies were inconclusive. No evidence of protection of broiler progeny of vaccinated breeder flocks could be detected through the challenge trials. In the absence of in-feed medication, broilers hatched from vaccinated breeders did, however, performed better under commercial conditions than those hatched from unvaccinated breeder flocks. The partial farm budget showed that broilers raised from OR vaccinated breeder flocks were more profitable than the negative control flocks. The quantitative risk analysis showed that the probability of making a relative profit from broilers as a result of OR vaccination of parent stock was 74%, from the use of in-feed medication in broilers from unvaccinated parents was 70% and from a combination of the interventions was 99%. It can be concluded that the last of these options was most profitable. / Dissertation (MSc (Veterinary Sciences))--University of Pretoria, 2003. / Veterinary Tropical Diseases / unrestricted

As piodermites, superficiais ou profundas, representam uma das dermatopatias caninas mais comuns no cotidiano da clínica dermatológica de cães. Dentre as classificadas como superficiais, destacam-se a foliculite bacteriana e a piodermite esfoliativa ou superficial disseminada, que representam a grande maioria dos casos. Em alguns cães, as piodermites superficiais são idiopáticas, apresentam evolução crônica e caráter recidivante. A despeito de criteriosa investigação acerca de sua possível etiologia, por vezes, não se consegue evidenciar a causa para a instalação e recorrência do quadro mórbido. Tais animais apresentam frequentes recidivas, em variáveis períodos de tempo, após o término da terapia antibiótica. Para estes caninos, na bibliografia médico-veterinária há recomendação de emprego das controvertidas imunoterapia ou de antibioticoterapia sob a forma de pulsos. Portanto, objetivou-se na casuística do Serviço de Dermatologia do Hospital Veterinário da USP, determinar as principais espécies bacterianas envolvidas, a susceptibilidade destas frente a diferentes antimicrobianos, bactericidas ou bacteriostáticos, assim como, comparar a eficácia e a segurança de três distintos protocolos terapêuticos no manejo a longo prazo, de piodermites superficiais recidivantes idiopáticas. Utilizaram-se 23 animais, de quaisquer dos sexos, de distintas raças e faixas etárias. A amostragem foi aleatoriamente disposta em três grupos de experimentação (GI, GII, G III), respectivamente submetidos à imunoterapia com bacterina comercial (Estafilin®) e às pulsoterapias de \"finais de semana\" ou em \"semanas alternadas\", empregando-se cefalexina \"per os\". O principal patógeno, em 67,6 % da totalidade de isolamentos bacterianos, foi Staphylococcus intermedius. Evidenciou-se (32,4 %), ainda, as espécies S. hyicus, S. schleiferi subespécie coagulans, S. warneri e Micrococcus sp. Segundo os antibiogramas, os fármacos mais efetivos in vitro foram a amoxicilina associada ao clavulanato de potássio, a cefalexina e o ceftiofur. Verificou-se resposta plena em percentis de, respectivamente, 50,0 %, 33,3 % e 28,6 % nos Grupos II, III e I; destarte, ao se agrupar as respostas plena e moderada evidenciaram-se valores relativos de, respectivamente, 83,3 %, 50,0 % e 42,9 %. À luz da estatística (Teste do Qui Quadrado), não se observaram quaisquer diferenças estatisticamente significativas (p > 0,05) entre os percentuais de resposta aos três protocolos empregados. Todos eles mostraram-se seguros, sem acarretar efeitos adversos farmacodérmicos. / Superficial and deep pyodermas are common canine skin disorders in the small animal dermatology practice. Among the superficial form, the bacterial folliculitis and superficial spreading pyoderma represent the great majority of cases. Some dogs are affected by idiopathic, chronic and recurrent type of superficial pyoderma. Despite a rigorous search for possible reasons for its etiology, sometimes it is not possible to determine an underlying cause for the disease recurrence. At variable time periods, these dogs show frequent recurrent episodes, after the interruption of antibiotic therapy. For these patients, the veterinary medicine literature recommends the use of the controversial immune therapy or antibiotic pulse therapy. Thus, using the University of Sao Paulo / Dermatology Service\'s casuistry the present study aimed to determine the major bacterial species responsible for pyodermas, their susceptibility to several bactericidal and bacteriostatic antibiotics as well as compare the efficacy and safety of three different therapeutics protocols for the long term management of idiopathic recurrent superficial pyoderma. A total of 23 male and female, purebred dogs from different ages were randomly allocated in three distinct experimental groups (G I, G II and G III), respectively treated with bacterin immune therapy, \"weekend\" antibiotic pulse therapy and \"week on - week off\" antibiotic pulse therapy, receiving cephalexin PO. S. intermedius was identified from 67,6 % of the totality of bacterial isolates and the remaining 32,4 % was composed of S. hyicus, S. schleiferi subspecie coagulans, S. warneri and Micrococcus sp. According to the antimicrobial susceptibility results the more effective drugs were amoxicillin plus potassium clavulanate, cephalexin and ceftioufur. In percentage values a full response of 50,0 %, 33,3 % and 28,6 % was achieved in Groups I, II and III, respectively. On the other hand, when the comparison was made considering full and moderate together the values were 83,3 %, 50,0 % and 42,9 % for Groups I, II and III, respectively. Through a statistical analysis (Qui score test) no significant difference (p > 0,05) among the response to treatment percentage values was observed for the three groups. All treatments were secure and no drug eruption side effects were observed.

Made available in DSpace on 2016-12-08T16:24:21Z (GMT). No. of bitstreams: 1 PGCA15MA170.pdf: 1407687 bytes, checksum: 6d77197fef831a6cbdc6fcf9e39c8629 (MD5) Previous issue date: 2015-07-23 / For a long time, Pasterella multocida has been considered only as a secondary agent in pneumonias in swine. However, studies developed in Brazil identified different isolates of P. multocida able to operate as primary disease agent in swine. The objective of his project was to study protection aspects of a vaccine with pathogenic samples of P. multocida in experimental model with high sanity status pigs. In addition, it aims to describe the expression of immune response in swine induced by P. multocida infection, and determinate the prevalence of P. multocida in tonsils of vaccinated and not vaccinated pigs through immunohistochemistry. The study was divided into three experiments: Immunization of mice with vaccines prepared with different adjuvants , Test of two adjuvants used in vaccine with a P. multocida homologous strain in swine and Experimental challenge in swine with a P. multocida heterologous strain to the vaccine . Four adjuvants were tested in mice: Al(OH)3, ISA 206, Montanide GEL 01 and ISA 760. Two of them were selected for testing in swine: Al(OH)3 which showed better results in mice-, and ISA 760 for being an oily adjuvant. At the second experiment, although two vaccines have been effective in the disease protection, the one with the adjuvant Al(OH)3 was selected, because showed lower vaccine reaction. Eighteen pigs were distributed into three groups for the specific test of the vaccine for pathogenic samples of P. multocida. Group 1 (G1) vaccine with Al(OH)3; group 2 (G2) infection control; and group 3 (G3) negative control. After the experiment, the animals were euthanized, and samples were collected in ice - for microbiological cultivation and relative quantification of cytokines gene expression (IL1 β, IL2, TNFα) and TLR4 , and in formaldehyde, for routine histopathology and immunohistochemistry. Results showed that the vaccine prototype with adjuvant Al(OH)3 was efficient in controlling pulmonary pasteurellosis. The immune response was mediated by TLR4, with increased expression of TNFα. IL-1β and IL2 were also expressed, however, there was no significant difference between the groups. The immunostaining of P. multocida in tonsil was positive in all pigs challenged with the bacteria / Por muito tempo, Pasteurella multocida foi considerada apenas como agente secundário nas pneumonias em suínos, todavia, estudos realizados no Brasil identificaram diferentes isolados de P. multocida capazes de atuar como agente primário de doença em suínos. Esse projeto teve como objetivo estudar aspectos de proteção de uma vacina com amostras patogênicas da P. multocida em modelo experimental com suínos de alto status sanitário . E de forma paralela descrever expressão da resposta imune dos suínos induzida pela infecção da P. multocida e determinar a prevalência dessa bactéria em tonsilas de suínos vacinados e não vacinados através de imunohistoquímica. O trabalho foi dividido em três experimentos: Imunização de camundongos com vacinas preparadas com diferentes adjuvantes ; Teste de dois adjuvantes utilizados na vacinação de suínos frente ao desafio com uma cepa homóloga de P. multocida ; e Desafio experimental em suínos com uma cepa de P. multocida heteróloga à da vacina . Em camundongo foram testados quatro adjuvantes: Al(OH)3, ISA 206, Montanide GEL 01 e ISA 760, dos quais, foram selecionados dois para teste em suínos: Al(OH)3, que apresentou melhor resultado nos camundongos e ISA 760, por ser um adjuvante oleoso. Embora as duas vacinas tenham sido efetivas na proteção da doença, no segundo experimento, selecionou-se àquela com adjuvante Al(OH)3, que apresentou menor reação vacinal. No terceiro experimento realizado em suínos, foram utilizados 18 suínos em três grupos: grupo 1 (G1) - vacina com Al(OH)3, e grupo 2 (G2) controle de infeção e grupo 3 (G3) controle negativo. Após o experimento os animais foram eutanasiados para avaliação patológica, coleta de amostras em gelo para cultivo microbiológico e quantificação relativa da expressão gênica das citocinas (IL1 β, IL2, TNFα) e TLR4, para histopatologia de rotina e imunohistoquimica. Os resultados demostraram que o protótipo de vacina com adjuvante Al(OH)3 foi eficiente no controle de pasteurelose pulmonar. O perfil da expressão da resposta imune foi mediado por TLR4, com aumento da expressão de TNFα. Também foram expressos IL-1β e IL2, porém sem diferença significativa entre os grupos. A imunomarcação de P. multocida na tonsila foi positiva em todos os suínos desafiados com a bactéria, independente de serem ou não vacinados

As piodermites, superficiais ou profundas, representam uma das dermatopatias caninas mais comuns no cotidiano da clínica dermatológica de cães. Dentre as classificadas como superficiais, destacam-se a foliculite bacteriana e a piodermite esfoliativa ou superficial disseminada, que representam a grande maioria dos casos. Em alguns cães, as piodermites superficiais são idiopáticas, apresentam evolução crônica e caráter recidivante. A despeito de criteriosa investigação acerca de sua possível etiologia, por vezes, não se consegue evidenciar a causa para a instalação e recorrência do quadro mórbido. Tais animais apresentam frequentes recidivas, em variáveis períodos de tempo, após o término da terapia antibiótica. Para estes caninos, na bibliografia médico-veterinária há recomendação de emprego das controvertidas imunoterapia ou de antibioticoterapia sob a forma de pulsos. Portanto, objetivou-se na casuística do Serviço de Dermatologia do Hospital Veterinário da USP, determinar as principais espécies bacterianas envolvidas, a susceptibilidade destas frente a diferentes antimicrobianos, bactericidas ou bacteriostáticos, assim como, comparar a eficácia e a segurança de três distintos protocolos terapêuticos no manejo a longo prazo, de piodermites superficiais recidivantes idiopáticas. Utilizaram-se 23 animais, de quaisquer dos sexos, de distintas raças e faixas etárias. A amostragem foi aleatoriamente disposta em três grupos de experimentação (GI, GII, G III), respectivamente submetidos à imunoterapia com bacterina comercial (Estafilin®) e às pulsoterapias de \"finais de semana\" ou em \"semanas alternadas\", empregando-se cefalexina \"per os\". O principal patógeno, em 67,6 % da totalidade de isolamentos bacterianos, foi Staphylococcus intermedius. Evidenciou-se (32,4 %), ainda, as espécies S. hyicus, S. schleiferi subespécie coagulans, S. warneri e Micrococcus sp. Segundo os antibiogramas, os fármacos mais efetivos in vitro foram a amoxicilina associada ao clavulanato de potássio, a cefalexina e o ceftiofur. Verificou-se resposta plena em percentis de, respectivamente, 50,0 %, 33,3 % e 28,6 % nos Grupos II, III e I; destarte, ao se agrupar as respostas plena e moderada evidenciaram-se valores relativos de, respectivamente, 83,3 %, 50,0 % e 42,9 %. À luz da estatística (Teste do Qui Quadrado), não se observaram quaisquer diferenças estatisticamente significativas (p > 0,05) entre os percentuais de resposta aos três protocolos empregados. Todos eles mostraram-se seguros, sem acarretar efeitos adversos farmacodérmicos. / Superficial and deep pyodermas are common canine skin disorders in the small animal dermatology practice. Among the superficial form, the bacterial folliculitis and superficial spreading pyoderma represent the great majority of cases. Some dogs are affected by idiopathic, chronic and recurrent type of superficial pyoderma. Despite a rigorous search for possible reasons for its etiology, sometimes it is not possible to determine an underlying cause for the disease recurrence. At variable time periods, these dogs show frequent recurrent episodes, after the interruption of antibiotic therapy. For these patients, the veterinary medicine literature recommends the use of the controversial immune therapy or antibiotic pulse therapy. Thus, using the University of Sao Paulo / Dermatology Service\'s casuistry the present study aimed to determine the major bacterial species responsible for pyodermas, their susceptibility to several bactericidal and bacteriostatic antibiotics as well as compare the efficacy and safety of three different therapeutics protocols for the long term management of idiopathic recurrent superficial pyoderma. A total of 23 male and female, purebred dogs from different ages were randomly allocated in three distinct experimental groups (G I, G II and G III), respectively treated with bacterin immune therapy, \"weekend\" antibiotic pulse therapy and \"week on - week off\" antibiotic pulse therapy, receiving cephalexin PO. S. intermedius was identified from 67,6 % of the totality of bacterial isolates and the remaining 32,4 % was composed of S. hyicus, S. schleiferi subspecie coagulans, S. warneri and Micrococcus sp. According to the antimicrobial susceptibility results the more effective drugs were amoxicillin plus potassium clavulanate, cephalexin and ceftioufur. In percentage values a full response of 50,0 %, 33,3 % and 28,6 % was achieved in Groups I, II and III, respectively. On the other hand, when the comparison was made considering full and moderate together the values were 83,3 %, 50,0 % and 42,9 % for Groups I, II and III, respectively. Through a statistical analysis (Qui score test) no significant difference (p > 0,05) among the response to treatment percentage values was observed for the three groups. All treatments were secure and no drug eruption side effects were observed.

O controle da eficiência de bacterinas antileptospirose de uso animal é o teste de potência com desafio em hamsters, contudo, na atualidade, tem sido estimulada a busca de alternativas que dispensem o uso de animais de laboratório. O teste de inibição de crescimento de leptospiras in vitro (ICLIV) tem sido proposto como possível alternativa. O presente trabalho empregou os testes de ICLIV, soroaglutinação microscópica (SAM) e ELISA anti IgG para avaliar a intensidade e a duração da imunidade passiva em leitões em aleitamento e ativa em matrizes suínas e leitões desmamados imunizados com bacterina experimental antileptospirose aprovada no teste de potência em hamster. Foi produzida uma bacterina experimental antileptospirose com estirpe patogênica de Leptospira interrogans, sorovar Kennewicki, estirpe Pomona Fromm (LPF), padronizada para conter 109 leptospiras por mL e associada ao adjuvante de hidróxido de alumínio na proporção de 10% do volume da dose final. Para a avaliação da eficácia da concentração mínima de leptospiras a ser utilizada na bacterina, diluições seriadas de razão dez de cultivo de leptospiras variando de 105 a 109 leptospiras/mL, foram submetidas ao teste de potência com desafio em hamsters (Experimento A), apenas a bacterina produzida na concentração de 109 leptospiras/mL foi capaz de proteger os hamsters contra a infecção induzida pela estirpe LPF, quando a vacina foi testada na diluição de 1:800, critério internacional de aprovação. A bacterina na concentração de 109 leptospiras/mL foi submetida ao teste de potência em hamsters (Experimento B), imunizados com a vacina pura e em diluições seriadas de razão dois (200 a 25600). A bacterina foi aprovada no teste de desafio em hamster até a diluição de 1:6400. O controle do inóculo de desafio foi constituído por 100 DL50. Fêmeas suínas que nunca haviam sido vacinadas contra a leptospirose e que foram não reagentes no teste de soroaglutinação microscópica aplicado a leptospirose efetuado com 24 estirpes de referência e no teste ICLIV com a estirpe LPF (Experimento 1), receberam duas aplicações intervaladas de 30 dias e um reforço aos 210 da primeira dose da bacterina pura e em diluições seriadas de razão dois (400 a 3200). Estes animais foram 16 monitorados com colheitas de sangue efetuadas a cada 30 dias. Os picos máximos de anticorpos avaliados pelos testes de SAM e de ICLIV foram observados aos 30 dias da segunda aplicação da vacina, com maior magnitude para a vacina pura, contudo aos 120 dias da segunda aplicação da vacina houve um declínio acentuado nos níveis de anticorpos. Para encontrar um melhor intervalo entre as imunizações (Experimento 2), fêmeas suínas receberam duas aplicações da bacterina pura intervaladas de 30 dias e o reforço aos 150 da primeira dose. A redução do intervalo de revacinação após as duas doses iniciais determinou a persistência dos títulos de aglutininas e de anticorpos neutralizantes com níveis sempre superiores a 0,4 log. Nos leitões em aleitamento filhos das matrizes imunizadas com bacterina na concentração de 109 leptospiras/mL foi constatada a transferência de imunidade passiva, confirmada pelos títulos de anticorpos aglutinantes detectáveis no quinto dia de vida e de neutralizantes no quinto e décimo dia de vida. Nos ensaios realizados em leitões desmamados imunizados com uma série de diluições de razão dez variando de a 109 leptospiras/mL os picos máximos de anticorpos foram observados aos 30 dias da segunda imunização, com maior magnitude para a bacterina testada na concentração de 109 leptospiras/mL. Os parâmetros finalmente obtidos foram que matrizes suínas e leitões desmamados, primovacinados com duas aplicações intervaladas de 30 dias da bacterina aprovada no teste de potência com desafio em hamster, apresentaram no teste de ICLIV efetuado aos 60 dias da primo-vacinação, intervalos de títulos de anticorpos (95%) expressos em log variando, respectivamente de (0,87 a 1,35) e de (1,22 a 1,58). Os valores máximos (12.800) para o teste de ELISA anti IgG das matrizes suínas foram obtidos após o reforço efetuado aos 210 dias. / The potency of antileptospirosis bacterins is controlled by in vivo experimental assay performed in hamsters; however, nowadays the search for in vitro methodologies that could replace the use of laboratory animals has been stimulated. For this purpose, the In vitro leptospiral growth inhibition test (IVLGIT) has been proposed as an alternative test. In this work, the intensity and duration of the passive immunity in suckling piglets and active immunity in weaned piglets and adult sows were evaluated, after vaccination with an experimental leptospirosis bacterin, which had been approved previously on the potency test in hamsters. The in vitro tests applied to swine sera were the growth inhibition test (IVLGIT), microscopic agglutination (MAT) and anti-IgG ELISA. The bacterin was produced with a pathogenic Leptospira interrogans serovar Kennewicki strain identified as Pomona Fromm (LPF), and added with aluminum hydroxide as adjuvant at a concentration of 10% of the final volume dose. In order to evaluate the efficacy of the minimum concentration of the leptospires to be used in the bacterin, ten-fold serial dilutions of a culture of leptospira varying from 105 to 109 leptospires/mL were submitted to potency-challenge test in hamsters (Trial A). Only the bacterin produced at the concentration of 109 leptospires/mL protected the hamsters against the infection by LPF strain, when the vaccine was tested at the dilution of 1:800, which is the international criterion for the approval. The bacterin at the concentration of 109 leptospires/mL was submitted to the potency test in hamsters (Trial B), which had been immunized with an undiluted and with two-fold serially diluted bacterin (from 1:200 to 1:25,600). In the challenge test in hamsters, the bacterin has passed till the dilution of 1:6,400. The control of the challenge inoculum presented a titer of 100 LD50. Adult sows, never been vaccinated against leptospirosis, and that showed negative MAT results using 24 reference serovars and with negative IVLGIT with the LPF strain (Trial 1) had administered two doses of bacterin with 30 day interval and a booster dose at 210th day after the first application of the undiluted bacterin and the two-fold diluted ones (1:400 to :3,200). The animals were 18 monitored with bleeding performed each 30 days. The levels of antibodies evaluated by MAT and IVLGIT showed the maximum peak at the 30th day after the second vaccination of the bacterin, with high magnitude found with the undiluted bacterin, however, at the 120th day after the second vaccination there were found a marked decline in antibodies levels. To find a better immunization scheme (Trial 2), the sows received two doses of undiluted bacterin with 30 day interval and the booster dose at the 150th day after the first dose. The reduction on the interval of revaccination after the two initial doses determined the persistence of a higher levels agglutinins and neutralizing antibodies with titers 0.4 log. In suckling piglets born from sows immunized with the bacterin at the concentration of 109 leptospires/mL, the passive transference of antibodies was confirmed by MAT titers detected on the fifth day, and by IVLGIT titers, at the fifth and tenth days after birth. In the study of weaned piglets immunized with the bacterin at the concentrations of 106, 107, 108 and 109 leptospires/mL (Trial 3), the highest antibodies levels were observed at the 30th day after the second immunization, with greater magnitude for the bacterin tested at the concentration of 109 leptospires/mL. The final results indicate that sows and weaned piglets, primo-vaccinated and revaccinated with 30 day interval with bacterin that had passed in the potency-challenge test in hamsters, presented the IVLGIT results varying (95% CI) from 0.87 log to 1.35 log for sows and 1.22 to 1.58 log for weaned piglets, at the 60th day after the first vaccination. In adult sows the highest anti-IgG ELISA titer reached 12,800, obtained after the booster vaccine dose performed at 210th day from the first vaccination.

Made available in DSpace on 2015-03-26T13:47:20Z (GMT). No. of bitstreams: 1 texto completo.pdf: 198280 bytes, checksum: 2e22264df93097f0e349bfa39abd6bcd (MD5) Previous issue date: 2003-09-05 / The present work was developed with the objective to determine the efficiency of an autogenous bacterin against Streptococcus suis serotype 2 and to test route and inoculation doses. Two experiments were accomplished with 64 animals in each experiment. In the Experiment I, 32 animals were vaccinated and 32 animals received placebo, being both groups challenged at the 60 days of age. In the Experiment II, the animals were divided in four groups: NVIP - 16 animals not vaccinated and challenged by the intraperitoneal route; VIP - 16 vaccinated animals and challenged by the intraperitoneal route; NVIV - 16 animals not vaccinated and challenged by the intravenous route; and VIV - 16 vaccinated animals and challenged by the intravenous route. All the groups were challenged at the 88 days of age. In the Experiment I, it was not possible to detect significant difference in the protection, clinical signs and lesions (P>0.05) among the groups not vaccinated and vaccinated. However, the isolation of S. suis and the average of the bacterial count was higher (P <0.01) in the group not vaccinated. The results observed in Experiment II in the groups NVIV and VIV presented protection, clinical signs and lesions similar (P>0.05), and the number of animals with positive isolation of S. suis and the average of the bacterial count was higher in the group NVIV (P<0.05). The groups NVIP and VIP presented significant difference for protection, clinical signs, lesions, number of positive isolations and average of the bacterial agent count (P <0.01). The intravenous challenge route was not appropriate for that experiment type and the intraperitoneal route presented favorable aspects. The vaccine was efficient for prevention of the infection with high virulent S. suis serotype 2, protecting the animals with an efficiency of 87.5 percent. / O presente trabalho foi desenvolvido com o objetivo de determinar a eficácia de uma bacterina autógena na proteção de suínos desafiados com Streptococcus suis sorotipo 2 de alta virulência e testar vias e doses de inoculação do agente. Foram realizados dois experimentos com 64 animais em cada experimento. No Experimento I, 32 animais foram vacinados aos 21 e 45 dias de idade, através de injeção intramuscular de 2 e 3 mL da bacterina, respectivamente. Os 32 animais restantes constituíram o grupo controle não vacinado. Aos 60 dias de idade os 64 animais foram desafiados via intravenosa, na dosagem de 8,6 x 109 UFC. No experimento II, 64 animais foram divididos em dois grupos de 32 animais. Um grupo foi vacinado aos 25 (2 mL) e 50 (3 mL) dias de idade, por via intramuscular, e o outro constituiu o grupo controle não vacinado. Os animais foram desafiados aos 88 dias de idade na dosagem de 7,2 x 109 UFC. Para efeito de desafio, os animais foram divididos em quatro grupos: NVIP - 16 animais não vacinados e desafiados pela via intraperitoneal; NVIV - 16 animais não vacinados e desafiados pela via intravenosa; VIP 16 animais vacinados e desafiados pela via intraperitoneal; e VIV - 16 animais vacinados e desafiados pela via intravenosa. No Experimento I, não foi possível detectar diferença significativa na proteção, sinais clínicos e lesões (p>0.05) entre os grupos não vacinado e vacinado. Só foi possível detectar diferença significativa no número de animais com isolamento positivo do S. suis e na média da contagem de Unidades Formadoras de Colônias (UFC) de S. suis (P<0.01). Os resultados observados no Experimento II, foram: 1) os grupos NVIV e VIV apresentaram proteção, sinais clínicos e lesões similares (P>0.05), e o número de animais com isolamento positivo do S. suis e a média da contagem de colônias de S. suis foi mais elevada no grupo NVIV (P<0.01), confirmando os resultados apresentados no Experimento I; e 2) os grupos NVIP e VIP apresentaram diferenças significativas para proteção, sinais clínicos, lesões, número de isolamentos positivos e média da contagem bacteriana do agente (P<0.01). O grupo NVIP apresentou 5/16 mortes e 11/16 sobreviventes doentes, enquanto que o grupo VIP teve 14/16 sobreviventes sadios e 2/16 de sobreviventes doentes, não apresentando nenhuma morte. A temperatura retal no grupo NVIP foi, em média, 0,5o C superior à do grupo VIP (P<0.01). O escore médio dos sinais clínicos nos grupos NVIP e VIP foi de 1,10±0,10 e 0,34±0,05, respectivamente, sendo mais elevado no grupo não vacinado (P<0.05). O escore médio das lesões também foi mais elevado no grupo NVIP (0,89±0,19) em relação ao grupo VIP (0,20±0,08) (P<0.05). O isolamento do S. suis foi positivo em 9/16 animais do grupo NVIP, com média da contagem bacteriana de 295,25±91,62 UFC/mL, sendo que no grupo VIP não foi possível isolar o agente em qualquer dos animais. A proteção da vacina para os animais desafiados pela via intraperitoneal foi calculada em 87,5%. Portanto, a via de desafio utilizada influenciou nos sinais clínicos, lesões e isolamento bacteriano dos animais vacinados e não vacinados. A via intravenosa não se mostrou adequada para esse tipo de experimento e a via intraperitoneal apresentou aspectos favoráveis. Finalmente, a vacina mostrou ser eficaz na prevenção da infecção pelo S. suis sorotipo 2 de alta virulência, protegendo os animais com uma eficiência estimada de 87,5%.

O controle da eficiência de bacterinas antileptospirose de uso animal é o teste de potência com desafio em hamsters, contudo, na atualidade, tem sido estimulada a busca de alternativas que dispensem o uso de animais de laboratório. O teste de inibição de crescimento de leptospiras in vitro (ICLIV) tem sido proposto como possível alternativa. O presente trabalho empregou os testes de ICLIV, soroaglutinação microscópica (SAM) e ELISA anti IgG para avaliar a intensidade e a duração da imunidade passiva em leitões em aleitamento e ativa em matrizes suínas e leitões desmamados imunizados com bacterina experimental antileptospirose aprovada no teste de potência em hamster. Foi produzida uma bacterina experimental antileptospirose com estirpe patogênica de Leptospira interrogans, sorovar Kennewicki, estirpe Pomona Fromm (LPF), padronizada para conter 109 leptospiras por mL e associada ao adjuvante de hidróxido de alumínio na proporção de 10% do volume da dose final. Para a avaliação da eficácia da concentração mínima de leptospiras a ser utilizada na bacterina, diluições seriadas de razão dez de cultivo de leptospiras variando de 105 a 109 leptospiras/mL, foram submetidas ao teste de potência com desafio em hamsters (Experimento A), apenas a bacterina produzida na concentração de 109 leptospiras/mL foi capaz de proteger os hamsters contra a infecção induzida pela estirpe LPF, quando a vacina foi testada na diluição de 1:800, critério internacional de aprovação. A bacterina na concentração de 109 leptospiras/mL foi submetida ao teste de potência em hamsters (Experimento B), imunizados com a vacina pura e em diluições seriadas de razão dois (200 a 25600). A bacterina foi aprovada no teste de desafio em hamster até a diluição de 1:6400. O controle do inóculo de desafio foi constituído por 100 DL50. Fêmeas suínas que nunca haviam sido vacinadas contra a leptospirose e que foram não reagentes no teste de soroaglutinação microscópica aplicado a leptospirose efetuado com 24 estirpes de referência e no teste ICLIV com a estirpe LPF (Experimento 1), receberam duas aplicações intervaladas de 30 dias e um reforço aos 210 da primeira dose da bacterina pura e em diluições seriadas de razão dois (400 a 3200). Estes animais foram 16 monitorados com colheitas de sangue efetuadas a cada 30 dias. Os picos máximos de anticorpos avaliados pelos testes de SAM e de ICLIV foram observados aos 30 dias da segunda aplicação da vacina, com maior magnitude para a vacina pura, contudo aos 120 dias da segunda aplicação da vacina houve um declínio acentuado nos níveis de anticorpos. Para encontrar um melhor intervalo entre as imunizações (Experimento 2), fêmeas suínas receberam duas aplicações da bacterina pura intervaladas de 30 dias e o reforço aos 150 da primeira dose. A redução do intervalo de revacinação após as duas doses iniciais determinou a persistência dos títulos de aglutininas e de anticorpos neutralizantes com níveis sempre superiores a 0,4 log. Nos leitões em aleitamento filhos das matrizes imunizadas com bacterina na concentração de 109 leptospiras/mL foi constatada a transferência de imunidade passiva, confirmada pelos títulos de anticorpos aglutinantes detectáveis no quinto dia de vida e de neutralizantes no quinto e décimo dia de vida. Nos ensaios realizados em leitões desmamados imunizados com uma série de diluições de razão dez variando de a 109 leptospiras/mL os picos máximos de anticorpos foram observados aos 30 dias da segunda imunização, com maior magnitude para a bacterina testada na concentração de 109 leptospiras/mL. Os parâmetros finalmente obtidos foram que matrizes suínas e leitões desmamados, primovacinados com duas aplicações intervaladas de 30 dias da bacterina aprovada no teste de potência com desafio em hamster, apresentaram no teste de ICLIV efetuado aos 60 dias da primo-vacinação, intervalos de títulos de anticorpos (95%) expressos em log variando, respectivamente de (0,87 a 1,35) e de (1,22 a 1,58). Os valores máximos (12.800) para o teste de ELISA anti IgG das matrizes suínas foram obtidos após o reforço efetuado aos 210 dias. / The potency of antileptospirosis bacterins is controlled by in vivo experimental assay performed in hamsters; however, nowadays the search for in vitro methodologies that could replace the use of laboratory animals has been stimulated. For this purpose, the In vitro leptospiral growth inhibition test (IVLGIT) has been proposed as an alternative test. In this work, the intensity and duration of the passive immunity in suckling piglets and active immunity in weaned piglets and adult sows were evaluated, after vaccination with an experimental leptospirosis bacterin, which had been approved previously on the potency test in hamsters. The in vitro tests applied to swine sera were the growth inhibition test (IVLGIT), microscopic agglutination (MAT) and anti-IgG ELISA. The bacterin was produced with a pathogenic Leptospira interrogans serovar Kennewicki strain identified as Pomona Fromm (LPF), and added with aluminum hydroxide as adjuvant at a concentration of 10% of the final volume dose. In order to evaluate the efficacy of the minimum concentration of the leptospires to be used in the bacterin, ten-fold serial dilutions of a culture of leptospira varying from 105 to 109 leptospires/mL were submitted to potency-challenge test in hamsters (Trial A). Only the bacterin produced at the concentration of 109 leptospires/mL protected the hamsters against the infection by LPF strain, when the vaccine was tested at the dilution of 1:800, which is the international criterion for the approval. The bacterin at the concentration of 109 leptospires/mL was submitted to the potency test in hamsters (Trial B), which had been immunized with an undiluted and with two-fold serially diluted bacterin (from 1:200 to 1:25,600). In the challenge test in hamsters, the bacterin has passed till the dilution of 1:6,400. The control of the challenge inoculum presented a titer of 100 LD50. Adult sows, never been vaccinated against leptospirosis, and that showed negative MAT results using 24 reference serovars and with negative IVLGIT with the LPF strain (Trial 1) had administered two doses of bacterin with 30 day interval and a booster dose at 210th day after the first application of the undiluted bacterin and the two-fold diluted ones (1:400 to :3,200). The animals were 18 monitored with bleeding performed each 30 days. The levels of antibodies evaluated by MAT and IVLGIT showed the maximum peak at the 30th day after the second vaccination of the bacterin, with high magnitude found with the undiluted bacterin, however, at the 120th day after the second vaccination there were found a marked decline in antibodies levels. To find a better immunization scheme (Trial 2), the sows received two doses of undiluted bacterin with 30 day interval and the booster dose at the 150th day after the first dose. The reduction on the interval of revaccination after the two initial doses determined the persistence of a higher levels agglutinins and neutralizing antibodies with titers 0.4 log. In suckling piglets born from sows immunized with the bacterin at the concentration of 109 leptospires/mL, the passive transference of antibodies was confirmed by MAT titers detected on the fifth day, and by IVLGIT titers, at the fifth and tenth days after birth. In the study of weaned piglets immunized with the bacterin at the concentrations of 106, 107, 108 and 109 leptospires/mL (Trial 3), the highest antibodies levels were observed at the 30th day after the second immunization, with greater magnitude for the bacterin tested at the concentration of 109 leptospires/mL. The final results indicate that sows and weaned piglets, primo-vaccinated and revaccinated with 30 day interval with bacterin that had passed in the potency-challenge test in hamsters, presented the IVLGIT results varying (95% CI) from 0.87 log to 1.35 log for sows and 1.22 to 1.58 log for weaned piglets, at the 60th day after the first vaccination. In adult sows the highest anti-IgG ELISA titer reached 12,800, obtained after the booster vaccine dose performed at 210th day from the first vaccination.

Bacterial pathogens cause severe infections worldwide in livestock and in humans, and antibiotic resistance further increases the importance of prophylactic vaccines. Inactivated bacterial vaccines (bacterins) are usually produced via incubation of the pathogen with chemicals such as formaldehyde, which is time consuming and may cause loss of immunogenicity due to the modification of structural components. We evaluated low-energy electron irradiation (LEEI) as an alternative method to generate a bacterin. Rodentibacter pneumotropicus, an invasive Gram-negative murine pathogen, was inactivated with LEEI and formaldehyde. LEEI resulted in high antigen conservation, and LPS activity was significantly better maintained when compared with formaldehyde treatment. Immunization of mice with LEEI-inactivated R. pneumotropicus elicited a strong immune response with no detectable bacterial burden upon sublethal challenge. The results of this study suggest the inactivation of bacteria with LEEI as an alternative, fast and efficient method to generate bacterial vaccines with increased efficacy.

Les infections à Salmonella Enteritidis chez les humains sont associées à la consommation d’œufs ou d’ovoproduits contaminés. La vaccination est un outil utilisé pour diminuer les risques d’infection à SE chez la volaille, mais avec des résultats variables. Au Canada deux bactérines, MBL SE4C et Layermune, sont couramment utilisées pour lutter contre SE. Cependant, leur efficacité n’a pas été complètement déterminée chez les poules pondeuses plus âgées. Par ailleurs, la capacité de ces vaccins à prévenir la transmission verticale et horizontale n’a pas encore été étudiée. L’objectif principal de cette étude était d’évaluer l’effet des deux bactérines sur la réponse immunitaire chez les poules pondeuses, de vérifier la protection conférée par ces vaccins contre l’infection expérimentale à SE, et d’identifier des protéines immunogènes afin de développer un vaccin sous-unitaire. Les oiseaux ont été vaccinés avec deux protocoles d’immunisation en cours d’élevage (soit à 12 et 18, ou à 16 semaines d’âge). Le groupe contrôle a été injecté avec la solution saline. Les oiseaux ont été inoculés per os avec 2 x 109 CFU de la souche SE lysotype 4 à 55 ou à 65 semaines d’âge. Les anticorps (IgG et IgA) ont été mesurés à différents temps avec un ELISA maison en utilisant l’antigène entier de SE. La phagocytose, flambée oxydative, les populations des splénocytes B et T ont été analysées en utilisant la cytométrie en flux. Les signes cliniques, l’excrétion fécale, la contamination des jaunes d’œufs et l’invasion des salmonelles dans les organes ont été étudiés pour évaluer l’efficacité de protection. La transmission horizontale a aussi été étudiée en évaluant l’infection à SE chez les oiseaux mis en contact avec les oiseaux inoculés. Les protéines immunogènes ont été identifiées par SDS-PAGE et Western blot à l’aide d’antisérums prélevés suite à la vaccination et/ou à l’infection expérimentale/naturelle, puis caractérisées par la spectrométrie de masse. Le protocole de vaccination avec deux immunisations a généré un niveau élevé de séroconversion à partir de 3 jusqu’à 32-34 semaines post-vaccination par rapport à celui avec une seule immunisation (p < 0.02), mais il n’y avait plus de différence entre les groupes à 54 et 64 semaines d’âge. Il n’y a pas eu de corrélation entre les niveaux d’IgG et les taux d’isolement des salmonelles dans les organes et des jaunes d’œuf. La production des IgA n’a été observée que chez les oiseaux vaccinés avec 2 injections de MBL SE4C (p ≤ 0.04). Après l’infection expérimentale, la production des IgA a été significativement plus élevée aux jours 1 et 7 p.i dans l’oviducte des oiseaux vaccinés (sauf pour le groupe vacciné avec 2 injections de Layermune) par comparaison avec le groupe contrôle (p ≤ 0.03). Seule la bactérine MBL SE4C a eu un effet protecteur contre la contamination des jaunes d’œuf chez les oiseaux infectés. Ce vaccin réduit partiellement en utilisant deux immunisations, le taux d’excrétion fécale des salmonelles chez les oiseaux inoculés et les oiseaux horizontalement infectés (p ≤ 0.02). Cinq des protéines identifiées par la spectrométrie de masse sont considérées comme des protéines potentiellement candidates pour une étude plus approfondie de leur immonogénicité: Lipoamide dehydrogenase, Enolase (2-phosphoglycerate dehydratase) (2-phospho-D-glycerate hydro-lyase), Elongation factor Tu (EF-Tu), Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) et DNA protection during starvation protein. En général, les bactérines ont induit une immunité humorale (IgG et IgA) chez les poules pondeuses. Cette réponse immunitaire a protégé partiellement les oiseaux quant à l’élimination des salmonelles, la contamination des jaunes d’œuf, ainsi que la transmission horizontale. Dans cette étude, la bactérine MBL SE4C (avec deux immunisations) s’est montrée plus efficace pour protéger les oiseaux que la bactérine Layermune. Nos résultats apportent des informations objectives et complémentaires sur le potentiel de deux bactérines pour lutter contre SE chez les poules pondeuses. Étant donné la protection partielle obtenue en utilisant ces vaccins, l’identification des antigènes immunogènes a permis de sélectionner des protéines spécifiques pour l’élaboration éventuelle d’un vaccin plus efficace contre SE chez les volailles. / Contaminated eggs and egg products have been associated with outbreaks of human Salmonella Enteritidis (SE) infections. Killed bacteria (bacterins) have been used to control Salmonella infections in poultry but variation in the conferred protection has been observed. In Canada the bacterins MBL SE4C and Layermune are currently used to control SE. However, their efficacy in protecting older layers has not been fully determined. Furthermore, the capacity of these bacterins to prevent vertical and horizontal transmissions has not yet been investigated. The main objectives of this study were to evaluate the effect of two available commercial bacterins on the immune response of laying hens, to verify the protection conferred by these vaccines against SE challenge and to identify immunogenic proteins to develop an oral subunit vaccine. Laying hens were vaccinated with two immunization schedules prior to the lay cycle (either at 12 and 18, or 16 weeks of age). The control group was injected with a saline solution. Laying hens were later inoculated per os with 2 x 109 CFU of SE PT4 strain either at 55 or 65 weeks of age. Serum IgG and mucosal IgA antibodies were measured with an in-house SE whole cell antigen ELISA. The phagocytosis, oxidative burst, splenic T and B cells populations were analyzed using flow cytometry. Clinical signs, fecal shedding, egg yolks contamination and organ invasion by SE were assessed to evaluate vaccine protection. Potential horizontal transmission from inoculated laying hens to non-inoculated laying hens, housed in the same isolator unit, was also evaluated. Immunogenic proteins were identified by SDS-PAGE and Western blot with sampled antisera during vaccination and/or infection of poultry with SE and then subjected to mass spectrometry. The vaccination protocol with two immunizations showed a higher seroconversion level than the single vaccination at 3 until 32-34 weeks post vaccination (p < 0.02) but no difference before challenge (54 and 64 old weeks). There was no relationship between high IgG level and SE isolation rates in organs and egg yolks. Only the MBL SE4C vaccine elicited IgA antibody production at 3 weeks post vaccination in both immunization protocols (p ≤ 0.04). Significant higher mucosal IgA levels were observed at day 1 and 7 post challenge in oviduct of vaccinated birds (except for the twice vaccinated Layermune group) compared to the control group (p ≤ 0.03). Humoral efficacy to protect from SE contamination of egg yolk was only observed in MBL SE4C vaccinated group and only this bacterin administered twice reduced SE shedding rate in inoculated birds and their exposed cagemates (p ≤ 0.02). A set of 5 proteins were considered as putative protein candidates to further detailed study on their immunogenicity: Lipoamide dehydrogenase; Enolase (2-phosphoglycerate dehydratase) (2-phospho-D-glycerate hydro-lyase); Elongation factor Tu (EF-Tu); Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and DNA protection during starvation protein. Overall, the commercial bacterins induced humoral immunity (IgG and IgA antibodies) in laying hens. This immune response partially protected for SE clearance, egg yolks contamination as well as horizontal transmission. In this study, MBL SE4C bacterin appeared to be more efficient in comparison to Layermune for protection of hens with a vaccination protocol comprising two immunizations. Our results provide additional and objective information on the potential of these vaccines for the control of SE in laying hens. Considering the partial protection achieved with the use of these bacterins, the identification of immunogenic antigens could help in the selection of specific proteins to elaborate a more efficient vaccine against SE in poultry.

Les infections à Salmonella Enteritidis chez les humains sont associées à la consommation d’œufs ou d’ovoproduits contaminés. La vaccination est un outil utilisé pour diminuer les risques d’infection à SE chez la volaille, mais avec des résultats variables. Au Canada deux bactérines, MBL SE4C et Layermune, sont couramment utilisées pour lutter contre SE. Cependant, leur efficacité n’a pas été complètement déterminée chez les poules pondeuses plus âgées. Par ailleurs, la capacité de ces vaccins à prévenir la transmission verticale et horizontale n’a pas encore été étudiée. L’objectif principal de cette étude était d’évaluer l’effet des deux bactérines sur la réponse immunitaire chez les poules pondeuses, de vérifier la protection conférée par ces vaccins contre l’infection expérimentale à SE, et d’identifier des protéines immunogènes afin de développer un vaccin sous-unitaire. Les oiseaux ont été vaccinés avec deux protocoles d’immunisation en cours d’élevage (soit à 12 et 18, ou à 16 semaines d’âge). Le groupe contrôle a été injecté avec la solution saline. Les oiseaux ont été inoculés per os avec 2 x 109 CFU de la souche SE lysotype 4 à 55 ou à 65 semaines d’âge. Les anticorps (IgG et IgA) ont été mesurés à différents temps avec un ELISA maison en utilisant l’antigène entier de SE. La phagocytose, flambée oxydative, les populations des splénocytes B et T ont été analysées en utilisant la cytométrie en flux. Les signes cliniques, l’excrétion fécale, la contamination des jaunes d’œufs et l’invasion des salmonelles dans les organes ont été étudiés pour évaluer l’efficacité de protection. La transmission horizontale a aussi été étudiée en évaluant l’infection à SE chez les oiseaux mis en contact avec les oiseaux inoculés. Les protéines immunogènes ont été identifiées par SDS-PAGE et Western blot à l’aide d’antisérums prélevés suite à la vaccination et/ou à l’infection expérimentale/naturelle, puis caractérisées par la spectrométrie de masse. Le protocole de vaccination avec deux immunisations a généré un niveau élevé de séroconversion à partir de 3 jusqu’à 32-34 semaines post-vaccination par rapport à celui avec une seule immunisation (p < 0.02), mais il n’y avait plus de différence entre les groupes à 54 et 64 semaines d’âge. Il n’y a pas eu de corrélation entre les niveaux d’IgG et les taux d’isolement des salmonelles dans les organes et des jaunes d’œuf. La production des IgA n’a été observée que chez les oiseaux vaccinés avec 2 injections de MBL SE4C (p ≤ 0.04). Après l’infection expérimentale, la production des IgA a été significativement plus élevée aux jours 1 et 7 p.i dans l’oviducte des oiseaux vaccinés (sauf pour le groupe vacciné avec 2 injections de Layermune) par comparaison avec le groupe contrôle (p ≤ 0.03). Seule la bactérine MBL SE4C a eu un effet protecteur contre la contamination des jaunes d’œuf chez les oiseaux infectés. Ce vaccin réduit partiellement en utilisant deux immunisations, le taux d’excrétion fécale des salmonelles chez les oiseaux inoculés et les oiseaux horizontalement infectés (p ≤ 0.02). Cinq des protéines identifiées par la spectrométrie de masse sont considérées comme des protéines potentiellement candidates pour une étude plus approfondie de leur immonogénicité: Lipoamide dehydrogenase, Enolase (2-phosphoglycerate dehydratase) (2-phospho-D-glycerate hydro-lyase), Elongation factor Tu (EF-Tu), Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) et DNA protection during starvation protein. En général, les bactérines ont induit une immunité humorale (IgG et IgA) chez les poules pondeuses. Cette réponse immunitaire a protégé partiellement les oiseaux quant à l’élimination des salmonelles, la contamination des jaunes d’œuf, ainsi que la transmission horizontale. Dans cette étude, la bactérine MBL SE4C (avec deux immunisations) s’est montrée plus efficace pour protéger les oiseaux que la bactérine Layermune. Nos résultats apportent des informations objectives et complémentaires sur le potentiel de deux bactérines pour lutter contre SE chez les poules pondeuses. Étant donné la protection partielle obtenue en utilisant ces vaccins, l’identification des antigènes immunogènes a permis de sélectionner des protéines spécifiques pour l’élaboration éventuelle d’un vaccin plus efficace contre SE chez les volailles. / Contaminated eggs and egg products have been associated with outbreaks of human Salmonella Enteritidis (SE) infections. Killed bacteria (bacterins) have been used to control Salmonella infections in poultry but variation in the conferred protection has been observed. In Canada the bacterins MBL SE4C and Layermune are currently used to control SE. However, their efficacy in protecting older layers has not been fully determined. Furthermore, the capacity of these bacterins to prevent vertical and horizontal transmissions has not yet been investigated. The main objectives of this study were to evaluate the effect of two available commercial bacterins on the immune response of laying hens, to verify the protection conferred by these vaccines against SE challenge and to identify immunogenic proteins to develop an oral subunit vaccine. Laying hens were vaccinated with two immunization schedules prior to the lay cycle (either at 12 and 18, or 16 weeks of age). The control group was injected with a saline solution. Laying hens were later inoculated per os with 2 x 109 CFU of SE PT4 strain either at 55 or 65 weeks of age. Serum IgG and mucosal IgA antibodies were measured with an in-house SE whole cell antigen ELISA. The phagocytosis, oxidative burst, splenic T and B cells populations were analyzed using flow cytometry. Clinical signs, fecal shedding, egg yolks contamination and organ invasion by SE were assessed to evaluate vaccine protection. Potential horizontal transmission from inoculated laying hens to non-inoculated laying hens, housed in the same isolator unit, was also evaluated. Immunogenic proteins were identified by SDS-PAGE and Western blot with sampled antisera during vaccination and/or infection of poultry with SE and then subjected to mass spectrometry. The vaccination protocol with two immunizations showed a higher seroconversion level than the single vaccination at 3 until 32-34 weeks post vaccination (p < 0.02) but no difference before challenge (54 and 64 old weeks). There was no relationship between high IgG level and SE isolation rates in organs and egg yolks. Only the MBL SE4C vaccine elicited IgA antibody production at 3 weeks post vaccination in both immunization protocols (p ≤ 0.04). Significant higher mucosal IgA levels were observed at day 1 and 7 post challenge in oviduct of vaccinated birds (except for the twice vaccinated Layermune group) compared to the control group (p ≤ 0.03). Humoral efficacy to protect from SE contamination of egg yolk was only observed in MBL SE4C vaccinated group and only this bacterin administered twice reduced SE shedding rate in inoculated birds and their exposed cagemates (p ≤ 0.02). A set of 5 proteins were considered as putative protein candidates to further detailed study on their immunogenicity: Lipoamide dehydrogenase; Enolase (2-phosphoglycerate dehydratase) (2-phospho-D-glycerate hydro-lyase); Elongation factor Tu (EF-Tu); Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and DNA protection during starvation protein. Overall, the commercial bacterins induced humoral immunity (IgG and IgA antibodies) in laying hens. This immune response partially protected for SE clearance, egg yolks contamination as well as horizontal transmission. In this study, MBL SE4C bacterin appeared to be more efficient in comparison to Layermune for protection of hens with a vaccination protocol comprising two immunizations. Our results provide additional and objective information on the potential of these vaccines for the control of SE in laying hens. Considering the partial protection achieved with the use of these bacterins, the identification of immunogenic antigens could help in the selection of specific proteins to elaborate a more efficient vaccine against SE in poultry.