Natural association between bovine leukemia virus and reproductive infectious diseases

Background: Enzootic bovine leukosis (EBL) is a widespread infectious disease caused by the bovine leukemia virus (BLV), which results in immune system dysfunction. The resulting immunosuppression may lead to an increased prevalence of other diseases. Dairy cows infected have altered immune function associated with decreased milk production and shortened lifespan and decreased immune response to immunization. BLV infection, however, is often asymptomatic, so any connection between subclinical infection and common reproductive diseases remains unknown. This study aimed to describe the relationship between naturally occurring subclinical BLV and infectious reproductive diseases seroconversion in the field. Materials, Methods & Results: The diseases investigated included Bovine viral diarrhoea (BVD), Bovine alfaherpesvirus 1 (BoHV-1), Bovine gammaherpesvirus 4 (BoHV-4), Chlamydiosis, Leptospirosis, Brucellosis and Neosporosis in dairy cattle. Six hundred fifty-five sera samples from the northern and south-central regions of Uruguay, from asymptomatic female Holstein and Holstein crosses without a history of vaccination against reproductive diseases were processed using reference diagnostic methods (Seronautalization, ELISA, MAT, Rose Bengal Plate test). The seroprevalence of BLV was 20.0%. Seroprevalence of reproductive diseases BVD, BoHV-1 and BoHV-4 were 99.3%, 41.2% and 27.3% of the populations, respectively, and the total seroprevalence of Leptospirosis, bovine Neospora caninum and Chlamydiosis were 19.8%, 29.8% and 33.0% respectively. The results revealed positive associations between naturally contracted BLV and the presence of antibodies against BoHV-1 (P = 0.002), as well as between naturally contracted BLV and presence of antibodies against Leptospira spp. (P = 0.028). Discussion: BLV infection can impact innate and adaptive immune system cells and alter the proper functioning of uninfected cells. BLV infection may also induce changes in the complex balance of cytokine expression, cell proliferation, and programmed cell death in T- and B-lymphocytes, which is critical for immune competence and effective response to infectious challenges. The progression of BLV infection has a substantial effect on host defense mechanisms. Indeed, lowmagnitude serologic responses to a commercial foot-and-mouth disease vaccine and a J5 Escherichia coli vaccine have been observed. These results are supported by recent trial studies showing a reduced immune response to vaccination against BoHV-1 and Leptospira spp. in asymptomatic animals infected with BLV. These are 2 of the most prevalent infectious reproductive diseases in cattle worldwide, and our results provide evidence that a link between BLV and susceptibility to these diseases may exist. Although there is evidence of the co-occurrence of these diseases, it remains unknown whether there is a direct or indirect effect of BLV on infertility, embryonic loss, or abortion. Another possibility is that natural infection with these reproductive pathogens (BoHV-1, Leptospira, or others) promotes BLV expression, negatively affecting the farms where these pathogens are endemic. Considering the high seroprevalence of BLV in dairy herds in North and South America where the infection is endemic, it was explored BLV's role as an immunosuppressant by quantifying its co-occurrence with diseases that affect reproductive performance in breeding herds. Future work should clarify the role of BLV and the co-occurring pathogens in causing infertility or abortions.

Cloning and expression of Neospora caninum rhoptry 2 and its potential as an immunobiological for neosporosis control

ABSTRACT: The protozoan Neospora caninum is known worldwide as one of the main causes of abortion in cattle. During infection, rhoptry proteins present in the apical complex of the parasite play important roles in adhesion and parasitophorous vacuole formation. The use of N. caninum ROP2 in experimental vaccines has shown promising protective results. In our study we performed cloning and expression in Escherichia coli of an antigenic portion of N. caninum ROP2. The recombinant protein (rROP2) was obtained in insoluble form, and the purified protein showed a size of approximately 18kDa. Even being a small truncate NcROP2 region, it was possible to conserve the antigenic epitopes which were recognized by bovine serum naturally infected with N. caninum. Vaccination with rROP2 on aluminum hydroxide adjuvant induced high levels of rROP2-specific IgG antibodies capable of recognizing native protein in tachyzoite lysates. In conclusion, our approaches were effective in obtaining the rROP2 protein, which induced specific mouse immune response and was also recognized by sera from N. caninum naturally infected cattle. These results suggest that it is a promising antigen for the development of neosporosis subunit vaccines as well as a suitable antigen for use in immunodiagnosis.

RESUMO: O protozoário Neospora caninum é conhecido mundialmente como uma das principais causas de aborto em bovinos. Durante a infecção, as proteínas rhoptry presentes no complexo apical do parasita desempenham papel importante na adesão e formação de vacúolos parasitóforos. O uso de ROP2 de N. caninum em vacinas experimentais tem mostrado resultados de proteção promissores. Em nosso estudo, realizamos a clonagem e expressão em Escherichia coli de uma porção antigênica de N. caninum ROP2. A proteína recombinante (rROP2) foi obtida na forma insolúvel, e a proteína purificada apresentou tamanho aproximado de 18kDa. Mesmo sendo uma pequena região truncada de NcROP2, foi possível conservar os epítopos antigênicos que foram reconhecidos pelo soro de bovinos naturalmente infectados com N. caninum. A vacinação com rROP2 adsorvida no adjuvante de hidróxido de alumínio induziu altos níveis de anticorpos IgG anti-rROP2, capazes de reconhecer a proteína nativa em lisados de taquizoítos. Em conclusão, nossas abordagens foram eficazes na obtenção da proteína rROP2, que induziu resposta imune específica em camundongos e também foi reconhecida por soros de bovinos naturalmente infectados com N. caninum. Estes resultados sugerem que rROP2 é um antígeno promissor para o desenvolvimento de vacinas de subunidades de neosporose, bem como um antígeno adequado para uso em imunodiagnóstico.

Cloning and expression of Neospora caninum rhoptry 2 and its potential as an immunobiological for neosporosis control / Clonagem e expressão de rhoptry 2 Neospora caninum e seu potencial como um imunobiológico para o controle de neosporose

The protozoan Neospora caninum is known worldwide as one of the main causes of abortion in cattle. During infection, rhoptry proteins present in the apical complex of the parasite play important roles in adhesion and parasitophorous vacuole formation. The use of N. caninum ROP2 in experimental vaccines has shown promising protective results. In our study we performed cloning and expression in Escherichia coli of an antigenic portion of N. caninum ROP2. The recombinant protein (rROP2) was obtained in insoluble form, and the purified protein showed a size of approximately 18kDa. Even being a small truncate NcROP2 region, it was possible to conserve the antigenic epitopes which were recognized by bovine serum naturally infected with N. caninum. Vaccination with rROP2 on aluminum hydroxide adjuvant induced high levels of rROP2-specific IgG antibodies capable of recognizing native protein in tachyzoite lysates. In conclusion, our approaches were effective in obtaining the rROP2 protein, which induced specific mouse immune response and was also recognized by sera from N. caninum naturally infected cattle. These results suggest that it is a promising antigen for the development of neosporosis subunit vaccines as well as a suitable antigen for use in immunodiagnosis.(AU)

O protozoário Neospora caninum é conhecido mundialmente como uma das principais causas de aborto em bovinos. Durante a infecção, as proteínas rhoptry presentes no complexo apical do parasita desempenham papel importante na adesão e formação de vacúolos parasitóforos. O uso de ROP2 de N. caninum em vacinas experimentais tem mostrado resultados de proteção promissores. Em nosso estudo, realizamos a clonagem e expressão em Escherichia coli de uma porção antigênica de N. caninum ROP2. A proteína recombinante (rROP2) foi obtida na forma insolúvel, e a proteína purificada apresentou tamanho aproximado de 18kDa. Mesmo sendo uma pequena região truncada de NcROP2, foi possível conservar os epítopos antigênicos que foram reconhecidos pelo soro de bovinos naturalmente infectados com N. caninum. A vacinação com rROP2 adsorvida no adjuvante de hidróxido de alumínio induziu altos níveis de anticorpos IgG anti-rROP2, capazes de reconhecer a proteína nativa em lisados de taquizoítos. Em conclusão, nossas abordagens foram eficazes na obtenção da proteína rROP2, que induziu resposta imune específica em camundongos e também foi reconhecida por soros de bovinos naturalmente infectados com N. caninum. Estes resultados sugerem que rROP2 é um antígeno promissor para o desenvolvimento de vacinas de subunidades de neosporose, bem como um antígeno adequado para uso em imunodiagnóstico.(AU)

Cloning and expression of Neospora caninum rhoptry 2 and its potential as an immunobiological for neosporosis control / Clonagem e expressão de rhoptry 2 Neospora caninum e seu potencial como um imunobiológico para o controle de neosporose

The protozoan Neospora caninum is known worldwide as one of the main causes of abortion in cattle. During infection, rhoptry proteins present in the apical complex of the parasite play important roles in adhesion and parasitophorous vacuole formation. The use of N. caninum ROP2 in experimental vaccines has shown promising protective results. In our study we performed cloning and expression in Escherichia coli of an antigenic portion of N. caninum ROP2. The recombinant protein (rROP2) was obtained in insoluble form, and the purified protein showed a size of approximately 18kDa. Even being a small truncate NcROP2 region, it was possible to conserve the antigenic epitopes which were recognized by bovine serum naturally infected with N. caninum. Vaccination with rROP2 on aluminum hydroxide adjuvant induced high levels of rROP2-specific IgG antibodies capable of recognizing native protein in tachyzoite lysates. In conclusion, our approaches were effective in obtaining the rROP2 protein, which induced specific mouse immune response and was also recognized by sera from N. caninum naturally infected cattle. These results suggest that it is a promising antigen for the development of neosporosis subunit vaccines as well as a suitable antigen for use in immunodiagnosis.(AU)

O protozoário Neospora caninum é conhecido mundialmente como uma das principais causas de aborto em bovinos. Durante a infecção, as proteínas rhoptry presentes no complexo apical do parasita desempenham papel importante na adesão e formação de vacúolos parasitóforos. O uso de ROP2 de N. caninum em vacinas experimentais tem mostrado resultados de proteção promissores. Em nosso estudo, realizamos a clonagem e expressão em Escherichia coli de uma porção antigênica de N. caninum ROP2. A proteína recombinante (rROP2) foi obtida na forma insolúvel, e a proteína purificada apresentou tamanho aproximado de 18kDa. Mesmo sendo uma pequena região truncada de NcROP2, foi possível conservar os epítopos antigênicos que foram reconhecidos pelo soro de bovinos naturalmente infectados com N. caninum. A vacinação com rROP2 adsorvida no adjuvante de hidróxido de alumínio induziu altos níveis de anticorpos IgG anti-rROP2, capazes de reconhecer a proteína nativa em lisados de taquizoítos. Em conclusão, nossas abordagens foram eficazes na obtenção da proteína rROP2, que induziu resposta imune específica em camundongos e também foi reconhecida por soros de bovinos naturalmente infectados com N. caninum. Estes resultados sugerem que rROP2 é um antígeno promissor para o desenvolvimento de vacinas de subunidades de neosporose, bem como um antígeno adequado para uso em imunodiagnóstico.(AU)

Effect of Infectious Bursal Disease (Ibd) Vaccines on Infection of Salmonella Heidelberg in Broiler Chickens

Salmonella Heidelberg (SH) has represented a great concern to the Brazilian poultry industry in the last years. It is known that immunosuppression in poultry is a contributing factor to increase Salmonella faecal shedding and to disturb control programmes. Not only infectious bursal disease (IBD) virus but also some live vaccines have been reported to induce immunosuppression. In the present study we assessed the effects of two live vaccines against IBD on SH-infected broiler chicks. At 7 days of age, birds of three groups (vaccinated with recombinant HVT-IBD vector, with immune complex-IBD vaccine and unvaccinated) were orally challenged with 1 x 108 CFU of SH. A group of hatchmates remained unvaccinated/unchallenged to serve as negative controls. Caecal colonization and systemic invasion were evaluated by bacterial enumeration at 1, 3, 5, 7 and 14 days post-infection (Dpi) and SH faecal shedding assessed by cloacal swabs at 3, 7, 10 and 14 Dpi. The counts of SH in caecal contents were higher in birds vaccinated with immune complex-IBD than in those that received the HVT-IBD vector vaccine at 5, 7 and 14 dpi (p 0.01). There were no statistical differences in bacterial counts in liver and spleen among birds of different groups. Cloacal swabs also indicated that the birds vaccinated with immune complex-IBD shed more SH than those vaccinated with HVT-IBD vector or those unvaccinated (p 0.01). The results of the present study suggested that the immunosuppressive effect of the immune complex-IBD vaccine helped to increase the SH-faecal shedding in the infected birds.(AU)

Effect of Infectious Bursal Disease (Ibd) Vaccines on Infection of Salmonella Heidelberg in Broiler Chickens

Salmonella Heidelberg (SH) has represented a great concern to the Brazilian poultry industry in the last years. It is known that immunosuppression in poultry is a contributing factor to increase Salmonella faecal shedding and to disturb control programmes. Not only infectious bursal disease (IBD) virus but also some live vaccines have been reported to induce immunosuppression. In the present study we assessed the effects of two live vaccines against IBD on SH-infected broiler chicks. At 7 days of age, birds of three groups (vaccinated with recombinant HVT-IBD vector, with immune complex-IBD vaccine and unvaccinated) were orally challenged with 1 x 108 CFU of SH. A group of hatchmates remained unvaccinated/unchallenged to serve as negative controls. Caecal colonization and systemic invasion were evaluated by bacterial enumeration at 1, 3, 5, 7 and 14 days post-infection (Dpi) and SH faecal shedding assessed by cloacal swabs at 3, 7, 10 and 14 Dpi. The counts of SH in caecal contents were higher in birds vaccinated with immune complex-IBD than in those that received the HVT-IBD vector vaccine at 5, 7 and 14 dpi (p 0.01). There were no statistical differences in bacterial counts in liver and spleen among birds of different groups. Cloacal swabs also indicated that the birds vaccinated with immune complex-IBD shed more SH than those vaccinated with HVT-IBD vector or those unvaccinated (p 0.01). The results of the present study suggested that the immunosuppressive effect of the immune complex-IBD vaccine helped to increase the SH-faecal shedding in the infected birds.

Effect of parenteral vaccination of newborn Holstein calves against viral agents involved in bovine respiratory disease (BRD)

Background: Newborn calves are born immunosuppressed, hypogammaglobulinemic, immunologically immature, and therefore more vulnerable to many infectious diseases. During pregnancy, the fetal-placental environment is regulated by Th2-type cytokines that neutralize Th1 responses, an important factor for immune defense against viral agents. The ingestion and absorption of colostral immunoglobulins enhance the immunity of the neonate. However, the presence of maternal antibodies might negatively affect the success of parental vaccination in the first two months of life. This study aimed to evaluate the effecacy of parenteral vaccination in newborn calves with high titers of maternal antibodies against respiratory viruses. Materials, Methods & Results: Twenty-eight Holstein calves were allocated to the vaccinated group (VAC, n = 18) or an unvaccinated control group (NVAC, n = 10). The initial vaccination with 5 mL of a commercial vaccine occurred around the 14th day of life (D14) and the booster at D35. Respiratory and diarrhea symptoms were evaluated at D12, D14, D16, D20, D31, D36, D45, D53, and D60. Blood samples were taken for leukogram, haptoglobin, and seroneutralization of BVDV, BoHV-1, BRSV, and BPI3V, at the time of vaccination at D14 (T1), at booster (D35, T2), and 21 days after the booster (D56, T3). Despite the increased prevalence of BRD during the period of the study, no calves from either group exhibited respiratory disease at D12 or D14. In subsequent assessments, the frequency of BRD increased over time in the VAC group until it reached a maximum prevalence of 38.9% (7/18) at D31. In the NVAC group, the maximum prevalence observed was 40% at D45 and D60. A comparison of the frequencies for BRD cases showed a statistical trend at D36 (P = 0.07), with a higher prevalence for the NVAC group (30%) in relation to the VAC group (5.6%). For the NVAC group, a greater number of total...

Histopathology and serology reaction to an immune complex infectious bursal disease vaccine (V877 strain) in SPF and commercial birds / Reação histopatológica e sorológica a uma vacina de imunocomplexo contra gumboro (cepa V877) em aves SPF e comerciais

The purpose of this study was to investigate the effect of a new infectious bursal disease (IBD) immune complex vaccine on immune system response in both specific pathogen-free (SPF) and commercial birds. Evaluation of response to the vaccination in the two experiments was done by histopathological examination and serology. The results of this study have shown that immune complex vaccine with the V877 strain is quite safe in White Leghorn SPF birds in which there has been no participation of maternal antibodies. In commercial birds was also observed that the immune complex vaccine with the V877 strain acted synergistically with different levels of passive antibodies and the vaccine virus began to replicate as passive immunity decreased to provide the animal active immunological response.

O objetivo desse estudo foi investigar o efeito de uma nova vacina de imunocomplexo contra a doença de Gumboro sobre o sistema imune de aves SPF e comerciais. A avaliação da resposta à vacinação foi realizada por meio de exame histopatológico e sorologia. Os resultados desse estudo demonstraram que a vacina de imunocomplexo com cepa V877 contra Gumboro é muito segura mesmo em aves SPF da linhagem White Leghorn nas quais não existia a participação de imunidade materna. Em aves comerciais também foi demonstrado que a vacina de imunocomplexo com a cepa V877 atuou sinergicamente com diferentes níveis de anticorpos passivos maternais, iniciando a replicação do vírus vacinal a partir do momento que a imunidade passiva diminui, para promover uma resposta imunológica ativa.

Histopathology and serology reaction to an immune complex infectious bursal disease vaccine (V877 strain) in SPF and commercial birds / Reação histopatológica e sorológica a uma vacina de imunocomplexo contra gumboro (cepa V877) em aves SPF e comerciais

The purpose of this study was to investigate the effect of a new infectious bursal disease (IBD) immune complex vaccine on immune system response in both specific pathogen-free (SPF) and commercial birds. Evaluation of response to the vaccination in the two experiments was done by histopathological examination and serology. The results of this study have shown that immune complex vaccine with the V877 strain is quite safe in White Leghorn SPF birds in which there has been no participation of maternal antibodies. In commercial birds was also observed that the immune complex vaccine with the V877 strain acted synergistically with different levels of passive antibodies and the vaccine virus began to replicate as passive immunity decreased to provide the animal active immunological response.(AU)

O objetivo desse estudo foi investigar o efeito de uma nova vacina de imunocomplexo contra a doença de Gumboro sobre o sistema imune de aves SPF e comerciais. A avaliação da resposta à vacinação foi realizada por meio de exame histopatológico e sorologia. Os resultados desse estudo demonstraram que a vacina de imunocomplexo com cepa V877 contra Gumboro é muito segura mesmo em aves SPF da linhagem White Leghorn nas quais não existia a participação de imunidade materna. Em aves comerciais também foi demonstrado que a vacina de imunocomplexo com a cepa V877 atuou sinergicamente com diferentes níveis de anticorpos passivos maternais, iniciando a replicação do vírus vacinal a partir do momento que a imunidade passiva diminui, para promover uma resposta imunológica ativa.(AU)

Effects of polysaccharide from Ophiopogon japonicus on immune response to Newcastle disease vaccine in chicken

In order to investigate the immune enhancement effects of Ophiopogon japonicus polysaccharide Ophiopogon japonicus (OJPS) on Newcastle disease (ND) live vaccine, chickens vaccinated against ND live vaccine was orally administered with the OJPS at high, medium and low concentrations respectively. In negative control group, chickens were given orally equal volume of physiological saline. On day 14, 21 and 28, the serum antibody titer, erythrocyte-C3b receptor rosette rate (E-C3bRR), erythrocyte-C3b immune complex rosette rate (E-ICRR) and peripheral lymphocyte proliferation were measured. The results showed that at most time points, the antibody titer, peripheral lymphocyte proliferation, E-C3bRR and elimination rate of immune complex of three OJPS administrating groups were significantly higher (P < 0.05) than those in negative control group. It indicated that OJPS could significantly improve the immune efficacy of Newcastle disease live vaccine, Ophiopogon japonicus polysaccharide possessed synergistical immunoenhancement.(AU)

Effects of polysaccharide from Ophiopogon japonicus on immune response to Newcastle disease vaccine in chicken

In order to investigate the immune enhancement effects of Ophiopogon japonicus polysaccharide Ophiopogon japonicus (OJPS) on Newcastle disease (ND) live vaccine, chickens vaccinated against ND live vaccine was orally administered with the OJPS at high, medium and low concentrations respectively. In negative control group, chickens were given orally equal volume of physiological saline. On day 14, 21 and 28, the serum antibody titer, erythrocyte-C3b receptor rosette rate (E-C3bRR), erythrocyte-C3b immune complex rosette rate (E-ICRR) and peripheral lymphocyte proliferation were measured. The results showed that at most time points, the antibody titer, peripheral lymphocyte proliferation, E-C3bRR and elimination rate of immune complex of three OJPS administrating groups were significantly higher (P<0.05) than those in negative control group. It indicated that OJPS could significantly improve the immune efficacy of Newcastle disease live vaccine, Ophiopogon japonicus polysaccharide possessed synergistical immunoenhancement.(AU)

Infectious Bursal Disease: Pathogenicity and Immunogenicity of Vaccines

The Infectious Bursal Disease (IBD) is a contagious viral disease that affects young chickens and may cause high morbidity and mortality. As the virus is very resistant to the environment, vaccination is required in case of high infection pressure. Due to variations in the virulence degree of the vaccines available to control IBD, this study aimed at evaluating the pathogenicity and immunogenicity of three types of vaccines. In total, 220 one-day-old specific pathogen free (SPF) chickens were immunized with recombinant, immune-complex and intermediate vaccines, or not vaccinated (55 birds per group) and challenged with IBD G11 strain on day 25. On days 25, 30, and 35, the Bursa of Fabricius (BF) were submitted to gross and histological examination, and serum samples were submitted to ELISA to determined anti-IBD antibody titers. On day 23, chickens were submitted to the test of hypersensitivity to phytohemagglutinin to evaluate the immunosuppressive effect of vaccines on the cell-mediated immunity. The results have indicated that the immune-complex vaccine induced the most severe BF lesions, whereas the recombinant vaccine preserved BF tissue and cell integrity. The three evaluated vaccines induced humoral immunity of similar intensity. The cellular reaction to phytohemagglutinin of the chickens immunized with recombinant and immune-complex vaccines was less severe compared with the unvaccinated chickens. In conclusion, these results indicate that the immune-complex vaccine was the most pathogenic and that all vaccines were effective in protecting SPF chickens against IBD.

Infectious Bursal Disease: Pathogenicity and Immunogenicity of Vaccines

The Infectious Bursal Disease (IBD) is a contagious viral disease that affects young chickens and may cause high morbidity and mortality. As the virus is very resistant to the environment, vaccination is required in case of high infection pressure. Due to variations in the virulence degree of the vaccines available to control IBD, this study aimed at evaluating the pathogenicity and immunogenicity of three types of vaccines. In total, 220 one-day-old specific pathogen free (SPF) chickens were immunized with recombinant, immune-complex and intermediate vaccines, or not vaccinated (55 birds per group) and challenged with IBD G11 strain on day 25. On days 25, 30, and 35, the Bursa of Fabricius (BF) were submitted to gross and histological examination, and serum samples were submitted to ELISA to determined anti-IBD antibody titers. On day 23, chickens were submitted to the test of hypersensitivity to phytohemagglutinin to evaluate the immunosuppressive effect of vaccines on the cell-mediated immunity. The results have indicated that the immune-complex vaccine induced the most severe BF lesions, whereas the recombinant vaccine preserved BF tissue and cell integrity. The three evaluated vaccines induced humoral immunity of similar intensity. The cellular reaction to phytohemagglutinin of the chickens immunized with recombinant and immune-complex vaccines was less severe compared with the unvaccinated chickens. In conclusion, these results indicate that the immune-complex vaccine was the most pathogenic and that all vaccines were effective in protecting SPF chickens against IBD. (AU)

NANOVACINA rMSP1a ASSOCIADA A ANTÍGENO INATIVADO CONTRA Anaplasma marginale: AVALIAÇÃO CLÍNICA E IMUNOLÓGICA EM BEZERROS SUBMETIDOS A DESAFIO EXPERIMENTAL COM AMOSTRA HETERÓLOGA

O complexo tristeza parasitária bovina (TPB) é composto por doenças provocadas por patógenos sanguíneos que ocorrem em todo o Brasil, sendo a anaplasmose a mais relatada. Recentemente não houve mudanças expressivas nas medidas de controle da anaplasmose. A vacinação seria uma alternativa econômica e eficaz para o controle da anaplasmose, porém hoje não é comercializada nenhuma vacina inativada que promova resposta imune efetiva. O presente grupo de pesquisa já mostrou que os nanotubos de carbono de paredes múltiplas (MWNT) como molécula carreadora para a proteína rMSP1a na imunização de camundongos e bovinos é capaz de induzir respostas imune celular e humoral, o que vem a colaborar com a eficácia da vacina inativada AmUFMG2 produzida in vitro. Com isso, o presente trabalho tem como objetivo avaliar as respostas imunológica e clínica induzida por uma vacina inativada contra A. marginale produzida in vitro associada à rMSP1a funcionalizada a MWNT em bezerros imunizados e desafiados experimentalmente. Foram utilizados 12 bezerros machos da raça Holandês, sabidamente negativos ao PCR para A. marginale, B. bovis e B. bigemina e soronegativos à RIFI para estes patógenos. Os animais foram distribuídos em dois grupos: vacinados (AmUFMG2 + MWNT+rMSP1a + adjuvante) e controle (PBS + adjuvante + MWNT não-carboxilado). Durante as fases de vacinação e de desafio, os animais foram avaliados em relação à resposta imune celular (CD4+CD25+, CD8+CD25+, CD21+ e CD335+), resposta imune humoral (IgG, IgG1 e IgG2 anti-AmUFMG2 e anti-rMSP1a), hemograma, rickettsemia, VG e temperatura. A vacina induziu resposta imune celular e humoral, porém após desafio com 5x106 eritrócitos infectados por A. marginale com a amostra UFMG1, os animais vacinados apresentaram quadro clínico de anaplasmose, mas com menores rickettsemia e febre. Os resultados mostram que a vacina foi capaz de gerar resposta imune nos animais e diminuir a intensidade da doença, resultado este promissor, já que foi utilizado desafio artificial, o que não condiz com o desafio natural

The tick fever complex is composed of diseases caused by blood pathogens that occur throughout Brazil, with anaplasmosis being the most reported. Recently, there have been no significant changes in anaplasmosis control measures. Vaccination would be an economical and effective alternative for the control of anaplasmosis, but today no inactivated vaccine that promotes an effective immune response is commercialized. The present research group has already shown that MWNT as a carrier molecule for the rMSP1a protein in the immunization of mice and cattle is able to induce cellular and humoral immune responses, which comes to collaborate with the effectiveness of the inactivated AmUFMG2 vaccine produced in vitro. Thus, the present work aims to evaluate the immunological and clinical response induced by an inactivated vaccine against A. marginale produced in vitro associated with rMSP1a functionalized to multi-walled carbon nanotubes in immunized and experimentally challenged calves. Twelve male Holstein calves were used, known to be negative for PCR for A. marginale, B. bovis and B. bigemina and seronegative to RIFI for these pathogens. The animals were distributed into two groups: vaccinated (AmUFMG2 + MWNT+rMSP1a + adjuvant) and control (PBS + adjuvant + non-carboxylated MWNT). During the vaccination and challenge phases, the animals were evaluated for cellular immune response (CD4+CD25+, CD8+CD25+, CD21+ and CD335+), humoral immune response (IgG, IgG1 and IgG2 anti-AmUFMG2 and anti-rMSP1a), blood count, rickettsemia, VG and temperature. The vaccine induced cellular and humoral immune responses, however, after challenge with 5x106 erythrocytes infected by A. marginale with the UFMG1 sample, the vaccinated animals showed clinical signs of anaplasmosis, but with less rickettsemia and fever. The results show that the vaccine was able to induce an immune response in animals and decrease the intensity of the disease, a promising result, since an artificial challenge was used, which does not match the natural challenge.

Immune response in holstein calves vaccinated at six months of age

Background: Calves are agammaglobulinemic and immature at birth and their immunological defense must be improved by colostrum, although, maternal antibodies decrease after two months of age and calves are susceptible to Bovine Respiratory Disease Complex (BRDC). Then, this research evaluated the immune response to BVDV and BoHV-1 in young calves to prevent BRDC.Materials, Methods & Results: Ten male Holstein calves were distributed in two groups with five animals on each: nonvaccinated VAC (-) and vaccinated VAC (+). Calves were vaccinated twice at 180 and 210 days of age. It was selected a commercial multivalent vaccine containing inactivated isolated of BVDV type-1 (strains 5960) and BVDV type-2 (strains 53637), and modified-live BoHV-1 (strains RLB103), with Quil A, cholesterol and Amphigen as adjuvant. The immune response (IR) was evaluated at 180 (T0), 210 (T1) and 240 (T2) days of life by serum neutralization (SN) and immunophenotyping. Specific antibodies to BVDV were detected in 40% (2/5) and 60% (3/5) of vaccinated calves at T1 and T2, respectively. Specific antibodies (Abs) to BoHV-1 were observed in 40% (2/5) at T1 and 100% (5/5) at T2 in the VAC (+) group. Titers of Abs to BoHV-1 increase from T0 to T2 (P = 0.034) in VAC (+). Moreover, geometric mean titer (GMT) to BoHV-1 in VAC (+) group was higher than VAC (-) after secondary IR in T2 (P = 0.006). The proportion of the markers T lymphocytes subpopulation (CD3+ , CD4+ , CD8+ and WC1+ ) and B lymphocytes (CD21+ ) were similar in the two experimental groups, while the expression of the CD25+ marker by gamma-delta (WC1+ ) was higher in VAC (+) at T1.Discussion: Low titers of circulating Abs could be observed just for BoHV-1 in the calves from VAC (-) at T0, probably due to remaining Abs from dams transferred by colostrum intake. This fact indicates the need of appropriate vaccination schedules to prevent the virosis in dairy heifers.[...](AU)

Immune response in holstein calves vaccinated at six months of age

Background: Calves are agammaglobulinemic and immature at birth and their immunological defense must be improved by colostrum, although, maternal antibodies decrease after two months of age and calves are susceptible to Bovine Respiratory Disease Complex (BRDC). Then, this research evaluated the immune response to BVDV and BoHV-1 in young calves to prevent BRDC.Materials, Methods & Results: Ten male Holstein calves were distributed in two groups with five animals on each: nonvaccinated VAC (-) and vaccinated VAC (+). Calves were vaccinated twice at 180 and 210 days of age. It was selected a commercial multivalent vaccine containing inactivated isolated of BVDV type-1 (strains 5960) and BVDV type-2 (strains 53637), and modified-live BoHV-1 (strains RLB103), with Quil A, cholesterol and Amphigen as adjuvant. The immune response (IR) was evaluated at 180 (T0), 210 (T1) and 240 (T2) days of life by serum neutralization (SN) and immunophenotyping. Specific antibodies to BVDV were detected in 40% (2/5) and 60% (3/5) of vaccinated calves at T1 and T2, respectively. Specific antibodies (Abs) to BoHV-1 were observed in 40% (2/5) at T1 and 100% (5/5) at T2 in the VAC (+) group. Titers of Abs to BoHV-1 increase from T0 to T2 (P = 0.034) in VAC (+). Moreover, geometric mean titer (GMT) to BoHV-1 in VAC (+) group was higher than VAC (-) after secondary IR in T2 (P = 0.006). The proportion of the markers T lymphocytes subpopulation (CD3+ , CD4+ , CD8+ and WC1+ ) and B lymphocytes (CD21+ ) were similar in the two experimental groups, while the expression of the CD25+ marker by gamma-delta (WC1+ ) was higher in VAC (+) at T1.Discussion: Low titers of circulating Abs could be observed just for BoHV-1 in the calves from VAC (-) at T0, probably due to remaining Abs from dams transferred by colostrum intake. This fact indicates the need of appropriate vaccination schedules to prevent the virosis in dairy heifers.[...]

Expressão gênica intestinal de uma vacina de DNA via oral utilizando dois métodos de proteção em tilápia-do-Nilo

O controle e manejo de doenças que acometem a piscicultura mundial, a exemplo das causadas por protozoários, parasitas, vermes, bactérias oportunistas e fungos, é um pré-requisito para a sustentabilidade desse setor produtivo e, um desafio constante, muito complexo e atualmente, limitado de produtos eficazes de tratamento. Além disso, alevinos e juvenis são ainda mais suscetíveis, de maneira geral, a surtos dessas doenças, por terem o sistema imunológico ainda em formação, de modo que, a sobrevivência desses animais depende de estratégias multidisciplinares de manejo adequado de sua saúde, nutrição, da qualidade da água e de medidas de biossegurança. Portanto, se faz necessária a ênfase em prevenção e não somente em tratamento. Por esses motivos, vacinas de DNA ganham atenção crescente, devido a superarem desvantagens de outros tratamentos preventivos, em geral de maneira mais eficiente. Elas funcionam expressando no hospedeiro, diferentes proteínas de patógenos, codificadas em vetores plasmideais. As vacinas comerciais licenciadas atualmente são relativamente caras, o que não gera estratégias amplas de vacinação. Foi avaliada a funcionalidade do plasmídeo vetorial pExu, transformado na linhagem Lactococcus lactis MG 1363, em peixes. Esta vacina gênica L. lactis MG1363 (pExu:mCherry) foi fornecida, por via oral, através de microencapsulamento ou liofilização, para a espécie Tilápia-do-Nilo (Oreochromis niloticus), de elevado interesse comercial. Para tanto, 108 juvenis machos de Tilápia-do-Nilo com peso de 250 ± 20 g foram utilizados. O equivalente a 1 dose (1x109 UFC) foi administrado por via oral [microencapsulado (administração direta) e liofilizado (incorporado à ração comercial)] aos animais e, a área que apresentava fluorescência, foi comparada, nas três secções intestinais (Anterior, Média e Posterior), nos períodos de 6, 18, 24, 48, 72, 96, 120, 144, e 168 horas após a administração. Ambos os métodos avaliados permitiram a expressão do gene repórter em todas as regiões de interesse. Houve, no entanto, diferença significativa (p < 0.001) na área de expressão média por seção intestinal, de modo que a seção Posterior apresentou menor expressão em relação às outras duas seções, tanto quando avaliado o microencapsulamento quanto a liofilização (redução de 40% e 87%, respectivamente). No entanto, a expressão da proteína mCherry se mostrou mais eficiente pelo método de liofilização que pelo microencapsulamento (p < 0.001), ao apresentar, aproximadamente, áreas de fluorescência 25 mil, 13 mil e 4 mil porcento maiores, nas seções Anterior, Média e Posterior, respectivamente. O método de microencapsulamento e liofilização apresentaram amplitude máxima da expressão no intervalo de 24 horas e 96 horas, respectivamente. Tais descobertas são importantes pois irão guiar pesquisas sobre novas estratégias com vacinas de DNA, em especial, as fornecidas oralmente.

The control and management of diseases that affect global pisciculture, such as those caused by protozoa, parasites, worms, opportunistic bacteria and fungi, is a prerequisite for the sustainability of this productive sector and a constant challenge, very complex and currently, limited of effective treatment products. In addition, fingerling and juvenile are even more susceptible, in general, to outbreaks of these diseases, as their immune systems are still in formation, so their survival relies on multidisciplinary strategies for the proper management of their health, nutrition, water quality and biosafety measures. Therefore, emphasis on prevention and not only just treatment is necessary. For these reasons, DNA vaccines are gaining increasing attention, due to overcoming disadvantages of other preventive treatments, generally more efficiently. They work by expressing different pathogen proteins in the host, encoded in plasmid vectors. Licensed commercial vaccines today are relatively expensive, which does not generate extensive vaccination strategies. It was evaluated the functionality of the plasmid pExu vector transformed in Lactococcus lactis MG 1363 strains, for fish. This genic L. lactis MG1363 (pExu:mCherry) vaccine was offered orally by either microencapsulation or lyophilization, for the Nile tilapia (Oreochromis niloticus), of elevated commercial interest. 108 Nile tilapia male juveniles, weighing 250 ± 20 g were utilized. 1 dose (equivalent of 1x109 CFU) were administrated orally [microencapsulated (direct administration) and lyophilized (incorporated into fish feed)] to the animals, and the area that presents fluorescence were compared in three intestinal sections (Anterior, Medium, Posterior) at 6, 18, 24, 48, 72, 96, 120, 144, e 168 hours after administration. Both methods allowed the expression of the reporter gene in all regions of interest. Statistical difference was observed (p < 0.001) in mean área of expression from different sections, so that the Posterior section presented inferior expression in relation to others, either by microencapsulation or lyophilization (40% and 87% reduction, respectively). mCherry protein expression was more efficient with lyophilization than microencapsulation (p < 0.001), by express, in average, fluorescence areas 25 thousand, 13 thousand and 4 thousand percent higher, on Anterior, Medium and Posterior sections, respectively. Microencapsulation and lyophilization presented maximum amplitude of expression in the 24 hours and 96 hours intervals, respectively. Such findings are important because they will guide research on new strategies with DNA vaccines, especially those given orally.

Suplementação dietética de levedura de cerveja e de minerais orgânicos sobre o desempenho e resposta imune em frangos de corte desafiados com a vacina de coccidiose

The objective of this study was to evaluate the effects of beer yeast and organic minerals supplementations on the performance, intestinal morphometry and humoral immune response of broilers that were challenged with coccidiosis vaccine. Three-hundred-eighty-four chicks were used, randomly distributed in an entirely random design total with four treatments (1. control, 2. organic minerals on top, 3. beer yeast, 4. association between organic minerals and beer yeast) and four treatments with 24-bird repetition each. At housing, 12 birds of each repetition were identified and received the eye-drop vaccine against coccidiosis. At 7, 21, 35 and 42 days it was collected blood to analyze the total serum proteins and the eletrophoretic profile of IgA and IgG and a duodenal segment for morphometric analysis. Neither the beer yeast nor the organic minerals supplementation affected the broiler performance or the antibodies production. The immunological challenge like the coccidiosis vaccine was not efficacious enough to provide an immune response able to modify the duodenal mucosa morphometry and the antibodies production.

O objetivo desta pesquisa foi avaliar o efeito da suplementação de levedura de cerveja e minerais orgânicos sobre o desempenho, morfometria intestinal e resposta imune humoral de frangos de corte, desafiados com a vacina de coccidiose. Foram utilizados 384 pintos de corte, distribuídos aleatoriamente em um delineamento inteiramente casualizado com quatro tratamentos (1. controle, 2. minerais orgânicos 3. levedo de cerveja, 4. associação de minerais orgânicos e levedo de cerveja) e 4 repetições de 24 aves cada. No alojamento, 12 aves de cada repetição foram identificadas com anilha e receberam a vacina contra coccidiose via ocular. Aos 7, 21, 35 e 42 dias, foi coletado sangue para análise de proteínas séricas totais e perfil eletroforético de IgA e IgG e um segmento do duodeno para análise morfométrica. A suplementação de levedo de cerveja ou de minerais orgânicos não afetou o desempenho de frangos de corte ou a produção de anticorpos. O desafio imunológico com a vacina de coccidiose não foi suficientemente eficaz em provocar uma resposta imune capaz de alterar a morfometria da mucosa duodenal e a produção de anticorpos.

Suplementação dietética de levedura de cerveja e de minerais orgânicos sobre o desempenho e resposta imune em frangos de corte desafiados com a vacina de coccidiose

The objective of this study was to evaluate the effects of beer yeast and organic minerals supplementations on the performance, intestinal morphometry and humoral immune response of broilers that were challenged with coccidiosis vaccine. Three-hundred-eighty-four chicks were used, randomly distributed in an entirely random design total with four treatments (1. control, 2. organic minerals on top, 3. beer yeast, 4. association between organic minerals and beer yeast) and four treatments with 24-bird repetition each. At housing, 12 birds of each repetition were identified and received the eye-drop vaccine against coccidiosis. At 7, 21, 35 and 42 days it was collected blood to analyze the total serum proteins and the eletrophoretic profile of IgA and IgG and a duodenal segment for morphometric analysis. Neither the beer yeast nor the organic minerals supplementation affected the broiler performance or the antibodies production. The immunological challenge like the coccidiosis vaccine was not efficacious enough to provide an immune response able to modify the duodenal mucosa morphometry and the antibodies production.

O objetivo desta pesquisa foi avaliar o efeito da suplementação de levedura de cerveja e minerais orgânicos sobre o desempenho, morfometria intestinal e resposta imune humoral de frangos de corte, desafiados com a vacina de coccidiose. Foram utilizados 384 pintos de corte, distribuídos aleatoriamente em um delineamento inteiramente casualizado com quatro tratamentos (1. controle, 2. minerais orgânicos 3. levedo de cerveja, 4. associação de minerais orgânicos e levedo de cerveja) e 4 repetições de 24 aves cada. No alojamento, 12 aves de cada repetição foram identificadas com anilha e receberam a vacina contra coccidiose via ocular. Aos 7, 21, 35 e 42 dias, foi coletado sangue para análise de proteínas séricas totais e perfil eletroforético de IgA e IgG e um segmento do duodeno para análise morfométrica. A suplementação de levedo de cerveja ou de minerais orgânicos não afetou o desempenho de frangos de corte ou a produção de anticorpos. O desafio imunológico com a vacina de coccidiose não foi suficientemente eficaz em provocar uma resposta imune capaz de alterar a morfometria da mucosa duodenal e a produção de anticorpos.

Effects of polysaccharide from Chuanminshen violaceum on immune response of newcastle disease vaccine in chicken

Background: Newcastle disease (ND), caused by avian paramyxovirus serotype 1 (APMV-1), also known as Newcastle disease virus (NDV), is regarded as one of the two most devastating diseases of poultry with the characteristics of serious disease and high flock mortality. It causes severe economic losses in domestic poultry, especially in chickens. At present, there were no effective drugs to treat this disease and the main method to control ND was vaccination. Moreover, new strains of virus resistant to chemotherapy continue to emerge, so does the need for a safe and effective vaccine. The immune adjuvant can make vaccine generate a strong immune response providing long-term protection against infection. With commonly usage of some immune adjuvants (e.g. mineral oil and aluminium hydroxide), many problems were occurred, such as side effects, strong local stimulation and carcinogenesis, together with complicated preparation, or failure to increase immunogenicity of weak antigen and so on. Botanical polysaccharides, as a new type of adjuvant or immunopotentiator, had become the hot research area because of their less side effects and no toxicity. The purpose of this research was to observe whether Chuanminshen violaceum polysaccharide (CVPS) possessed synergistical immunoenhancement, and offer the theoretical evidence for developing potential new-type adjuvant. Materials, Methods & Results: 200 three-yellow chickens at one day of age were randomly divided into four groups and reared in separated pens. On 7 days old, the average maternal serum hemaglutination inhibition (HI) antibody titer was less than 3 log2, all chickens of each group (the average body weight (BW) was 120 g) were vaccinated with ND live vaccine through nasal drip and eye-drop. At the same time, the chickens in three CVPS groups (high, medium and low doses of CVPS) were orally administered with 0.5 mL of CVPS at concentrations of 100 mg/kg BW, 50 mg/kg BW, 25 mg/kg BW respectively, once a day for fi ve successive day, in negative control group (NC), with 0.5 mL of physiological saline, once a day for fi ve successive day. On days 14, 21 and 28, the serum antibody titer, erythrocyte-C3b receptor rosette rate (E-C3bRR), erythrocyte-C3b immune complex rosette rate (E-ICRR), peripheral lymphocyte proliferation and peripheral CD4+/CD8+ ratio were measured. The results showed that the antibody titer, E-C3bRR, elimination rate of immune complex and peripheral lymphocyte proliferation in three CVPS groups and peripheral CD4+/CD8+ ratio in medium dosage of CVPS group were significantly higher (P < 0.05) than those in control group throughout the process of whole experiment almost. Discussion: The CVPS not only improved the E-C3bRR and accelerated the elimination rate of CIC, but also induced higher antibody titer, peripheral lymphocyte proliferation and peripheral CD4+ /CD8+ ratio in chickens vaccinated against ND live vaccine. This indicated that CVPS possessed immune-enhancement efficacy of ND live vaccine and might be expected as a candidate of new-type adjuvant.