Immunogenicity of Streptococcus equi subsp. equi recombinant SeM protein and bacterin in mice

ABSTRACT: The infection caused by Streptococcus equi, known as strangles, affects the respiratory system of horses, causing high morbidity and rapid spread among the herd. Bacterin vaccines, composed of inactivated whole cells of S. equi, have variable efficacy and duration. Infected animals produce specific antibodies against SeM, the immunodominant antigen of S. equi. This makes it a promising target for vaccine development. In this context, the objective of this work was to evaluate a vaccine combining S. equi bacterin and recombinant SeM protein. Mice were vaccinated with bacterin (S. equi ~1.2 × 108CFU/ml); rSeM protein (20g); bacterin-rSeM combination; or PBS (Control Group) and challenged with a suspension of S. equi, containing 10 × LD50. All vaccinated mice survived the challenge and produced anti-rSeM and anti-S. equi antibodies, which were assessed by indirect ELISA. The Control Group reached endpoint criteria 96 h after infection. These results demonstrate that a vaccine combining the S. equi bacterin with rSeM protein protects mice against strangles. This combination vaccine could potentially protect horses and overcome the limitations of currently available strangle vaccines.

RESUMO: A infecção causada por Streptococcus equi, denominada adenite, atinge o sistema respiratório de equinos, causando alta morbidade e rápida disseminação entre o rebanho. Vacinas bacterinas, compostas de células inteiras inativadas de S. equi apresentam eficácia e duração variáveis. Animais infectados apresentam anticorpos específicos à proteína SeM, antígeno imunodominante de S. equi, o que a torna um alvo promissor para o desenvolvimento de vacinas. Neste contexto, o objetivo deste trabalho foi avaliar uma vacina baseada na administração simultânea da bacterina e da proteína SeM recombinante. Camundongos foram vacinados com a bacterina (S. equi ~1.2 × 108CFU/ml); a proteína rSeM (20g); a bacterina e rSeM simultaneamente; ou PBS (Grupo Controle) e, posteriormente, foram desafiados com uma suspensão de S. equi contendo 10 × LD50. Todos os animais vacinados apresentaram anticorpos anti-rSeM e contra S. equi, avaliados através de ELISA indireto, e mantiveram-se e sobreviveram ao desafio letal. O Grupo Controle atingiu critérios de endpoint 96 h após a infecção. Estes resultados demonstram que uma vacina constituída de células inteiras de S. equi com rSeM protege camundongos contra adenite, sugerindo a capacidade de proteção a equinos e, possivelmente, superando as limitações das vacinas contra adenite atualmente disponíveis.

Immunogenicity of Streptococcus equi subsp. equi recombinant SeM protein and bacterin in mice / Imunogenicidade da proteína recombinante SeM de Streptococcus equi subsp. equi e da bacterina em camundongos

The infection caused by Streptococcus equi, known as strangles, affects the respiratory system of horses, causing high morbidity and rapid spread among the herd. Bacterin vaccines, composed of inactivated whole cells of S. equi, have variable efficacy and duration. Infected animals produce specific antibodies against SeM, the immunodominant antigen of S. equi. This makes it a promising target for vaccine development. In this context, the objective of this work was to evaluate a vaccine combining S. equi bacterin and recombinant SeM protein. Mice were vaccinated with bacterin (S. equi ~1.2 × 108CFU/ml); rSeM protein (20μg); bacterin-rSeM combination; or PBS (Control Group) and challenged with a suspension of S. equi, containing 10 × LD50. All vaccinated mice survived the challenge and produced anti-rSeM and anti-S. equi antibodies, which were assessed by indirect ELISA. The Control Group reached endpoint criteria 96 h after infection. These results demonstrate that a vaccine combining the S. equi bacterin with rSeM protein protects mice against strangles. This combination vaccine could potentially protect horses and overcome the limitations of currently available strangle vaccines.(AU)

A infecção causada por Streptococcus equi, denominada adenite, atinge o sistema respiratório de equinos, causando alta morbidade e rápida disseminação entre o rebanho. Vacinas bacterinas, compostas de células inteiras inativadas de S. equi apresentam eficácia e duração variáveis. Animais infectados apresentam anticorpos específicos à proteína SeM, antígeno imunodominante de S. equi, o que a torna um alvo promissor para o desenvolvimento de vacinas. Neste contexto, o objetivo deste trabalho foi avaliar uma vacina baseada na administração simultânea da bacterina e da proteína SeM recombinante. Camundongos foram vacinados com a bacterina (S. equi ~1.2 × 108CFU/ml); a proteína rSeM (20μg); a bacterina e rSeM simultaneamente; ou PBS (Grupo Controle) e, posteriormente, foram desafiados com uma suspensão de S. equi contendo 10 × LD50. Todos os animais vacinados apresentaram anticorpos anti-rSeM e contra S. equi, avaliados através de ELISA indireto, e mantiveram-se e sobreviveram ao desafio letal. O Grupo Controle atingiu critérios de endpoint 96 h após a infecção. Estes resultados demonstram que uma vacina constituída de células inteiras de S. equi com rSeM protege camundongos contra adenite, sugerindo a capacidade de proteção a equinos e, possivelmente, superando as limitações das vacinas contra adenite atualmente disponíveis.(AU)

Immunogenicity of Streptococcus equi subsp. equi recombinant SeM protein and bacterin in mice / Imunogenicidade da proteína recombinante SeM de Streptococcus equi subsp. equi e da bacterina em camundongos

The infection caused by Streptococcus equi, known as strangles, affects the respiratory system of horses, causing high morbidity and rapid spread among the herd. Bacterin vaccines, composed of inactivated whole cells of S. equi, have variable efficacy and duration. Infected animals produce specific antibodies against SeM, the immunodominant antigen of S. equi. This makes it a promising target for vaccine development. In this context, the objective of this work was to evaluate a vaccine combining S. equi bacterin and recombinant SeM protein. Mice were vaccinated with bacterin (S. equi ~1.2 × 108CFU/ml); rSeM protein (20μg); bacterin-rSeM combination; or PBS (Control Group) and challenged with a suspension of S. equi, containing 10 × LD50. All vaccinated mice survived the challenge and produced anti-rSeM and anti-S. equi antibodies, which were assessed by indirect ELISA. The Control Group reached endpoint criteria 96 h after infection. These results demonstrate that a vaccine combining the S. equi bacterin with rSeM protein protects mice against strangles. This combination vaccine could potentially protect horses and overcome the limitations of currently available strangle vaccines.(AU)

A infecção causada por Streptococcus equi, denominada adenite, atinge o sistema respiratório de equinos, causando alta morbidade e rápida disseminação entre o rebanho. Vacinas bacterinas, compostas de células inteiras inativadas de S. equi apresentam eficácia e duração variáveis. Animais infectados apresentam anticorpos específicos à proteína SeM, antígeno imunodominante de S. equi, o que a torna um alvo promissor para o desenvolvimento de vacinas. Neste contexto, o objetivo deste trabalho foi avaliar uma vacina baseada na administração simultânea da bacterina e da proteína SeM recombinante. Camundongos foram vacinados com a bacterina (S. equi ~1.2 × 108CFU/ml); a proteína rSeM (20μg); a bacterina e rSeM simultaneamente; ou PBS (Grupo Controle) e, posteriormente, foram desafiados com uma suspensão de S. equi contendo 10 × LD50. Todos os animais vacinados apresentaram anticorpos anti-rSeM e contra S. equi, avaliados através de ELISA indireto, e mantiveram-se e sobreviveram ao desafio letal. O Grupo Controle atingiu critérios de endpoint 96 h após a infecção. Estes resultados demonstram que uma vacina constituída de células inteiras de S. equi com rSeM protege camundongos contra adenite, sugerindo a capacidade de proteção a equinos e, possivelmente, superando as limitações das vacinas contra adenite atualmente disponíveis.(AU)

Use of Staphylococcus aureus phage Lysate Staphage Lysate (SPL)® for the control of recurrent pyoderma eczema in dogs with atopic dermatitis

Background: Recurrent staphylococcal infections are frequent in dogs with atopic dermatitis (AD). Many factors seem to contribute to making bacterial pyoderma refractory to treatment. Short-term systemic antibiotic therapy is effective for the treatment of acute symptoms, and may, along with pulsatile therapy, contribute to the long-term control of the disease. However, microbial resistance has become a growing and alarming problem. The aim of this study was to evaluate whether the use of Staphylococcus aureus Phage Lysate Staphage Lysate (SPL)®, can minimize the symptoms of recurrent pyoderma and increase the interval between acute atopic manifestations in dogs. Materials, Methods & Results: Thirteen dogs with a history of Canine Atopic Dematitis (CAD) and recurrent bacterial pyoderma received SPL at increasing intervals for 23 weeks. The contents of an intact pustule of each dog was collected and submitted to microbiological analysis. Systemic antibiotic therapy was established for the first 4-6 weeks of SPL protocol, based on the antibiotic sensitivity tests. The animals included in the study underwent a therapeutic protocol receiving shots of 0.5 mL of SPL subcutaneously (SC) twice a week for the first 12 weeks; 1.0 mL of SPL (SC) once a week for four weeks; 1.0 mL of SPL (SC) once every 15 days; 1.0 mL of SPL (SC) after a three-week interval from the last dose on week [...](AU)

Use of Staphylococcus aureus phage Lysate Staphage Lysate (SPL)® for the control of recurrent pyoderma eczema in dogs with atopic dermatitis

Background: Recurrent staphylococcal infections are frequent in dogs with atopic dermatitis (AD). Many factors seem to contribute to making bacterial pyoderma refractory to treatment. Short-term systemic antibiotic therapy is effective for the treatment of acute symptoms, and may, along with pulsatile therapy, contribute to the long-term control of the disease. However, microbial resistance has become a growing and alarming problem. The aim of this study was to evaluate whether the use of Staphylococcus aureus Phage Lysate Staphage Lysate (SPL)®, can minimize the symptoms of recurrent pyoderma and increase the interval between acute atopic manifestations in dogs. Materials, Methods & Results: Thirteen dogs with a history of Canine Atopic Dematitis (CAD) and recurrent bacterial pyoderma received SPL at increasing intervals for 23 weeks. The contents of an intact pustule of each dog was collected and submitted to microbiological analysis. Systemic antibiotic therapy was established for the first 4-6 weeks of SPL protocol, based on the antibiotic sensitivity tests. The animals included in the study underwent a therapeutic protocol receiving shots of 0.5 mL of SPL subcutaneously (SC) twice a week for the first 12 weeks; 1.0 mL of SPL (SC) once a week for four weeks; 1.0 mL of SPL (SC) once every 15 days; 1.0 mL of SPL (SC) after a three-week interval from the last dose on week [...]

Cross-protection between experimental anti-leptospirosis bacterins

We investigated the existence of cross-protection between two anti-leptospirosis monovalent experimental bacterins produced with two strains of Leptospira serogroup Pomona: Fromm strain of serovar Kennewicky, isolated from pigs in the United States, and strain GR6 of serovar Pomona isolated from pigs in Brazil. Both were added of aluminum hydroxide as an adjuvant. Experimental bacterins were tested with the hamster potency test in order to assess protection provided against the disease and against the establishment of kidney infection. Controls were polyvalent commercial vaccine produced with Leptospira strains isolated outside Brazil, which included a representative of Pomona serovar, or Sorensen solution added of aluminum hydroxide adjuvant. The challenge was performed with cross-strains of serogroup Pomona tested in accordance with international standards established for the potency test. After 21 days of the challenge, survivors were killed to evaluate the condition of Leptospira renal carrier. Experimental bacterins protected hamsters against homologous and heterologous strains, demonstrating the existence of cross-protection. The commercial vaccine protected the hamsters challenged with both strains, but there was a high proportion of animals diagnosed as renal carriers when the challenge was performed with strain GR6, isolated from pigs in Brazil.

Efficacy of bacterin-, outer membrane protein- and fimbriae extract-based vaccines for the control of Salmonella Enteritidis experimental infection in chickens / Eficácia de bactéria inativada (bacterina), proteína da membrana externa e extrato de fimbrias no controle de infecção experimental por Salmonella Enteritidis (SE) em galinhas

The efficacy of three vaccines was evaluated in chickens for the control of experimental infection with Salmonella Enteritidis (SE) phage type 4. The vaccines were produced with bacterin, outer membrane proteins (OMP) and fimbriae crude extract (FE). The chickens were vaccinated intramuscularly with two doses of each vaccine at 12 and 15 weeks of age. The chickens were then orally challenged with 10(9) CFU/chicken Salmonella Enteritidis phage type 4 at 18 weeks of age. Fecal swabs were performed for the recovery of shedding SE, and SE was recovered from the liver and spleen. Additionally, antibody titers were measured in the serum by micro-agglutination test. The results indicated that the vaccine produced with bacterin yielded better results and resulted in reduction of fecal shedding and organ invasion by SE after oral challenge, although no vaccine was 100% effective for the control of SE experimental infection.(AU)

A eficácia de três vacinas de Salmonella Enteritidis fagotipo 4, produzidas na forma de bacterina, proteínas de membrana externa (OMP) e extrato bruto de fímbrias (FE) foi avaliada para proteção de aves infectadas experimentalmente. As aves foram vacinadas por via intramuscular com duas doses de cada vacina as 12 e 15 semanas de idade e desafiadas com 10(9) UFCs de Salmonella Enteritidis fagotipo 4 às 18 semanas de idade, por via oral. A eficácia foi determinada através do reisolamento da bactéria nas fezes e no fígado e baço, e os anticorpos foram mensurados no soro. Os resultados demonstraram que a vacina produzida com a bacterina foi mais eficaz em comparação às outras vacinas examinadas, para reduzir a excreção fecal e a invasão de órgãos após o desafio por SE.(AU)

DESENVOLVIMENTO DE PROTÓTIPO DE VACINA CONTRA Pasteurella multocida SOROTIPO A EM SUÍNOS

LAZAROTO, Juliana. Desenvolvimento de protótipo de vacina contra Pasteurella multocida sorotipo A em suínos. 2015. 81f. Dissertação (Mestrado em Ciência Animal Área: Sanidade Animal) Universidade do Estado de Santa Catarina. Programa de Pós Graduação em Ciência Animal, Lages, 2015. 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 , IL2, TNF) 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 TNF. Também foram expressos IL-1 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.

LAZAROTO, Juliana. Evaluation of inactivated vaccine against Pasteurella multocida serotype A in swine. 2015. 81p. Dissertation (Master of Animal Science Field: Animal Sanity) Santa Catarina State University. Postgraduate program in Animal Science, Lages, 2015. 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 , IL2, TNF) 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 TNF. IL-1 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.