Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 520
Filtrar
1.
Front Immunol ; 12: 674643, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34335572

RESUMO

Bovine tuberculosis is an important animal and zoonotic disease caused by Mycobacterium bovis. The innate immune response is the first line of defense against pathogens and is also crucial for the development of an efficient adaptive immune response. In this study we used an in vitro co-culture model of antigen presenting cells (APC) and autologous lymphocytes derived from peripheral blood mononuclear cells to identify the cell populations and immune mediators that participate in the development of an efficient innate response capable of controlling the intracellular replication of M. bovis. After M. bovis infection, bovine immune cell cultures displayed upregulated levels of iNOS, IL-22 and IFN-γ and the induction of the innate immune response was dependent on the presence of differentiated APC. Among the analyzed M. bovis isolates, only a live virulent M. bovis isolate induced an efficient innate immune response, which was increased upon stimulation of cell co-cultures with the M. bovis culture supernatant. Moreover, we demonstrated that an allelic variation of the early secreted protein ESAT-6 (ESAT6 T63A) expressed in the virulent strain is involved in this increased innate immune response. These results highlight the relevance of the compounds secreted by live M. bovis as well as the variability among the assessed M. bovis strains to induce an efficient innate immune response.


Assuntos
Imunidade Inata/imunologia , Mycobacterium bovis/imunologia , Tuberculose Bovina/imunologia , Animais , Antígenos de Bactérias/imunologia , Bovinos , Técnicas de Cocultura , Citocinas/metabolismo , Interferon gama/metabolismo , Macrófagos , Cultura Primária de Células
2.
Biomed Res Int ; 2021: 5532864, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33880367

RESUMO

Bovine tuberculosis (bTB) is a widespread zoonotic infection targeting the livestock sector, especially in developing countries, and posing a risk to humans and animal populations. Its recent prevalence in river buffaloes has been estimated as higher as 33.7%. In emergent countries like Pakistan, there is likeliness of human-livestock interfaces extensively and lacking of effective preventive measures that illustrate the risk of spreading the infection at a remarkable rate. The river buffalo (Bubalus bubalis) is an upkeep host of Mycobacterium bovis and is responsible for disease transmission among buffaloes and other livestock species. In this study, potential molecular biomarkers in the Interferon-gamma gene (IFNg) were identified after genomic screening of river buffaloes. Unique genomic loci in river buffalo proved the novelty of the genomic structure of this phenomenal animal but also highlighted its significance in natural immunity against the Mycobacterium. A total of eight single nucleotide polymorphisms were identified in the coding region of IFNg. The SNPs in the exonic region were all transitions, i.e., the conversion of purines to purines. These SNPs were analyzed for Hardy Weinberg Equilibrium, chi2 test, gene diversity, and protein structural conformation. Pathway analysis in tuberculosis revealed that IFNg inhibits the antigen-presenting cells (APC) through JAK and STAT pathways. Network analysis of IFNg proteins in both species showed strong associations among the immunity-related proteins (interleukins, tissue necrosis factors) and receptors of interferons. The identified polymorphic sites might be novel-potentiated markers for the selection of animals with superior immune response against bTB and can be exploited as promising genomic sites for breeding the resistant animal herds to combat Mycobacterium infection in a long run.


Assuntos
Búfalos/genética , Búfalos/imunologia , Interferon gama/genética , Tuberculose Bovina/genética , Tuberculose Bovina/imunologia , Animais , Bovinos , Frequência do Gene/genética , Redes Reguladoras de Genes , Genoma , Haplótipos/genética , Heterozigoto , Interferon gama/química , Razão de Chances , Estrutura Secundária de Proteína
3.
Res Vet Sci ; 136: 595-597, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33894619

RESUMO

H65, a fusion protein of three pairs of ESX-secreted antigens of Mycobacterium tuberculosis and Mycobacterium bovis, formulated with the liposomal adjuvant CAF01 has been shown to confer protection against M. tuberculosis infection in mice. In this study, we evaluated the impact of combining BCG with H65 + CAF01 immunization in a M. bovis mouse model of infection. We found that a BCG-H65 + CAF01/ H65 + CAF01 prime-boost scheme induced higher protection than BCG and H65 + CAF01 alone. Altogether, H65 antigen formulated in liposomal adjuvant improved the BCG-induced immune protection, thus making this vaccine strategy a promising tool to control bovine tuberculosis.


Assuntos
Vacina BCG/imunologia , Mycobacterium bovis/imunologia , Tuberculose Bovina/prevenção & controle , Adjuvantes Imunológicos/farmacologia , Animais , Bovinos , Modelos Animais de Doenças , Feminino , Camundongos , Camundongos Endogâmicos BALB C , Mycobacterium tuberculosis/imunologia , Tuberculose Bovina/imunologia , Vacinas de Subunidades/imunologia , Vacinas Sintéticas/imunologia
4.
Sci Rep ; 11(1): 2929, 2021 02 03.
Artigo em Inglês | MEDLINE | ID: mdl-33536465

RESUMO

Tuberculin Purified Protein Derivatives (PPDs) exhibit multiple limitations: they are crude extracts from mycobacterial cultures with largely unknown active components; their production depends on culture of mycobacteria requiring expensive BCL3 production facilities; and their potency depends on the technically demanding guinea pig assay. To overcome these limitations, we developed a molecularly defined tuberculin (MDT) by adding further antigens to our prototype reagent composed of ESAT-6, CFP-10 and Rv3615c (DIVA skin test, DST). In vitro screening using PBMC from infected and uninfected cattle shortlisted four antigens from a literature-based list of 18 to formulate the MDT. These four antigens plus the previously identified Rv3020c protein, produced as recombinant proteins or overlapping synthetic peptides, were formulated together with the three DST antigens into the MDT to test cattle experimentally and naturally infected with M. bovis, uninfected cattle and MAP vaccinated calves. We demonstrated significant increases in MDT-induced skin responses compared to DST in infected animals, whilst maintaining high specificity in unvaccinated or MAP vaccinated calves. Further, MDT can also be applied in in vitro blood-based interferon-gamma release assays. Thus, MDT promises to be a robust diagnostic skin and blood test reagent overcoming some of the limitations of PPDs and warrants full validation.


Assuntos
Mycobacterium bovis/isolamento & purificação , Paratuberculose/prevenção & controle , Teste Tuberculínico/veterinária , Tuberculina/imunologia , Tuberculose Bovina/diagnóstico , Animais , Bovinos , Leucócitos Mononucleares , Masculino , Mycobacterium avium subsp. paratuberculosis/imunologia , Mycobacterium bovis/imunologia , Paratuberculose/microbiologia , Teste Tuberculínico/métodos , Tuberculose Bovina/imunologia , Tuberculose Bovina/microbiologia , Vacinação/veterinária
5.
Int J Biol Macromol ; 171: 82-88, 2021 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-33418045

RESUMO

Translation engineering and bioinformatics have accelerated the rate at which gene sequences can be improved to generate multi-epitope proteins. Strong antigenic proteins for tuberculosis diagnosis include individual ESAT6 and CFP10 proteins or derived peptides. Obtention of heterologous multi-component antigens in E. coli without forming inclusion bodies remain a biotechnological challenge. The gene sequence for ESAT6-CFP10 fusion antigen was optimized by codon bias adjust for high-level expression as a soluble protein. The obtained fusion protein of 23.7 kDa was observed by SDS-PAGE and Western blot analysis after Ni-affinity chromatography and the yield of expressed soluble protein reached a concentration of approximately 67 mg/L in shake flask culture after IPTG induction. Antigenicity was evaluated at 4 µg/mL in whole blood cultures from bovines, and protein stimuli were assessed using a specific in vitro IFN-γ release assay. The hybrid protein was able to stimulate T-cell specific responses of bovine TB suspects. The results indicate that improved E. coli codon usage is a good and cost-effective strategy to potentialize large scale production of multi-epitope proteins with sustained antigenic properties for diagnostic purposes.


Assuntos
Antígenos de Bactérias/genética , Proteínas de Bactérias/genética , Mycobacterium bovis/imunologia , Fragmentos de Peptídeos/genética , Proteínas Recombinantes de Fusão/administração & dosagem , Vacinas contra a Tuberculose/administração & dosagem , Tuberculose Bovina/prevenção & controle , Sequência de Aminoácidos , Animais , Antígenos de Bactérias/imunologia , Proteínas de Bactérias/imunologia , Bovinos , Clonagem Molecular , Códon , Escherichia coli/genética , Escherichia coli/metabolismo , Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Histidina/genética , Histidina/metabolismo , Imunogenicidade da Vacina , Interferon gama/biossíntese , Mycobacterium bovis/química , Mycobacterium bovis/genética , Mycobacterium tuberculosis/química , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/imunologia , Oligopeptídeos/genética , Oligopeptídeos/metabolismo , Fragmentos de Peptídeos/imunologia , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologia , Alinhamento de Sequência , Vacinas contra a Tuberculose/genética , Vacinas contra a Tuberculose/imunologia , Tuberculose Bovina/imunologia , Tuberculose Bovina/microbiologia , Vacinação/métodos
6.
Immunology ; 162(2): 220-234, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33020922

RESUMO

TNF signalling through TNFRp55 and TNFRp75, and receptor shedding is important for immune activation and regulation. TNFRp75 deficiency leads to improved control of Mycobacterium tuberculosis (M. tuberculosis) infection, but the effects of early innate immune events in this process are unclear. We investigated the role of TNFRp75 on cell activation and apoptosis of alveolar macrophages and neutrophils during M. tuberculosis and M. bovis BCG infection. We found increased microbicidal activity against M. tuberculosis occurred independently of IFNy and NO generation, and displayed an inverse correlation with alveolar macrophages (AMs) apoptosis. Both M. tuberculosis and M. bovis BCG induced higher expression of MHC-II in TNFRp75-/- AMs; however, M bovis BCG infection did not alter AM apoptosis in the absence of TNFRp75. Pulmonary concentrations of CCL2, CCL3 and IL-1ß were increased in TNFRp75-/- mice during M, bovis BCG infection, but had no effect on neutrophil responses. Thus, TNFRp75-dependent regulation of mycobacterial replication is virulence dependent and occurs independently of early alveolar macrophage apoptosis and neutrophil responses.


Assuntos
Vacina BCG/imunologia , Macrófagos Alveolares/imunologia , Neutrófilos/imunologia , Receptores Tipo II do Fator de Necrose Tumoral/imunologia , Tuberculose Bovina/imunologia , Tuberculose/imunologia , Animais , Apoptose/imunologia , Bovinos , Células Cultivadas , Feminino , Pulmão/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mycobacterium bovis/imunologia , Mycobacterium tuberculosis/imunologia , Receptores Tipo I de Fatores de Necrose Tumoral/imunologia , Transdução de Sinais/imunologia , Receptores Chamariz do Fator de Necrose Tumoral/imunologia , Virulência/imunologia
7.
J Immunol Methods ; 491: 112941, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33321133

RESUMO

Detection of specific antibodies would be a useful test strategy for bovine tuberculosis (bTB) as a complement to the single skin test. We developed a lateral flow immunochromatography (LFIC) test for rapid bTB detection based on the use of a conjugate of gold nanoparticles with a recombinant G protein. After evaluating 3 Mycobacterium bovis (MB) antigens: ESAT-6, CFP-10 and MPB83 for the control line, we selected MPB83 given it was the most specific. The performance of the test was analyzed with 820 bovine sera, 40 sera corresponding to healthy animals, 5 sera from animals infected with Mycobacterium avium subsp. paratuberculosis (MAP) and 775 sera of animals from herds with bTB. All these sera were also submitted to a validated bTB-ELISA using whole-cell antigen from MB. From the 775 sera of animals from herds with bTB, 87 sera were positive by the bTB-ELISA, 45 were positive by LFIC and only 5 animals were positives by skin test (TST). To confirm bTB infection in the group of TST (-), bTB-ELISA (+) and LFIC (+) animals, we performed postmortem examination in 15 randomly selected animals. Macroscopically, these 15 animals had numerous small and large yellow-white granulomas, characteristic of bTB, and the infection was subsequently confirmed by PCR in these tissues with lesions (gold standard). No false positive test result was detected with the developed LFIC either with the sera from healthy animals or from animals infected with MAP demonstrating that it can be a useful technique for the rapid identification of animals infected with bTB.


Assuntos
Anticorpos Antibacterianos/sangue , Cromatografia de Afinidade/métodos , Tuberculose Bovina/diagnóstico , Animais , Antígenos de Bactérias/imunologia , Proteínas de Bactérias/imunologia , Bovinos , Ensaio de Imunoadsorção Enzimática , Proteínas de Membrana/imunologia , Tuberculose Bovina/imunologia
8.
PLoS One ; 15(11): e0239938, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33166313

RESUMO

Mycobacterium bovis, the causative agent of bovine tuberculosis, is a pathogen that impacts both animal and human health. Consequently, there is a need to improve understanding of disease dynamics, identification of infected animals, and characterization of the basis of immune protection. This study assessed the transcriptional changes occurring in cattle during the early weeks following a M. bovis infection. RNA-seq analysis of whole blood-cell transcriptomes revealed two distinct transcriptional clusters of infected cattle at both 4- and 10-weeks post-infection that correlated with disease severity. Cattle exhibiting more severe disease were transcriptionally divergent from uninfected animals. At 4-weeks post-infection, 25 genes had commonly increased expression in infected cattle compared to uninfected cattle regardless of disease severity. Ten weeks post-infection, differential gene expression was only observed when severely-affected cattle were compared to uninfected cattle. This indicates a transcriptional divergence based on clinical status following infection. In cattle with more severe disease, biological processes and cell type enrichment analyses revealed overrepresentation of innate immune-related processes and cell types in infected animals. Collectively, our findings demonstrate two distinct transcriptional profiles occur in cattle following M. bovis infection, which correlate to clinical status.


Assuntos
Imunidade Inata/genética , Leucócitos Mononucleares/metabolismo , Mycobacterium bovis/imunologia , Transcriptoma/genética , Tuberculose Bovina/patologia , Animais , Bovinos , Expressão Gênica/genética , Perfilação da Expressão Gênica/veterinária , Leucócitos Mononucleares/citologia , Mycobacterium bovis/genética , Análise de Sequência com Séries de Oligonucleotídeos , Índice de Gravidade de Doença , Tuberculose Bovina/imunologia
9.
Vet Immunol Immunopathol ; 228: 110112, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32892112

RESUMO

Mycobacterium bovis, the causative agent of bovine tuberculosis (bTB), poses a risk of infection for livestock, humans, and wildlife. An interferon (IFN)-γ release assay has been used with tuberculin skin tests to detect bTB; however, infected animals may still be missed. Previous studies have suggested that bovine interleukin-2 (BoIL-2) may act as a potential biological marker for the diagnosis of bovine infectious diseases. However, a detailed evaluation of IL-2 as a diagnostic target for bTB is lacking. Therefore, we established hybridoma cell lines that produced monoclonal antibodies (mAbs) recognizing the native BoIL-2 and developed a flow cytometry assay, based on the BoIL-2 mAbs, for detecting M. bovis-specific IL-2. Subsequently, the method was utilized for a preliminary investigation of bTB in cattle; significantly (P < 0.0001) more CD4+IL-2+ T cells were detected in infected cattle than in healthy animals when a specific mycobacterial antigen CFP-10-ESAT-6 fusion protein was used. Moreover, our method demonstrated high coincidence rates with the BOVIGAM® test and an IFN-γ flow cytometry assay for the diagnosis of bTB. These findings show that the present method may be useful for detecting bTB.


Assuntos
Citometria de Fluxo/veterinária , Interleucina-2/análise , Mycobacterium bovis/isolamento & purificação , Tuberculose Bovina/diagnóstico , Animais , Anticorpos Monoclonais , Bovinos , Citometria de Fluxo/métodos , Hibridomas , Camundongos Endogâmicos BALB C , Mycobacterium bovis/imunologia , Teste Tuberculínico/métodos , Teste Tuberculínico/veterinária , Tuberculose Bovina/imunologia
10.
Infect Immun ; 88(12)2020 11 16.
Artigo em Inglês | MEDLINE | ID: mdl-32958527

RESUMO

Cell (CD3+ T cell and CD68+ macrophages), cytokine (interferon gamma-positive [IFN-γ+] and tumor necrosis factor alpha-positive [TNF-α+]), and effector molecule (inducible nitric oxide synthase-positive [iNOS+]) responses were evaluated in the lymph nodes and tissues of cattle naturally infected with Mycobacterium bovis Detailed postmortem and immunohistochemical examinations of lesions were performed on 16 cows that were positive by the single intradermal cervical comparative tuberculin (SICCT) test and that were identified from dairy farms located around the city of Addis Ababa, Ethiopia. The severity of the gross lesion was significantly higher (P = 0.003) in M. bovis culture-positive cows (n = 12) than in culture-negative cows (n = 4). Immunohistochemical techniques showed that in culture-positive cows, the mean immunolabeling fraction of CD3+ T cells decreased as the stage of granuloma increased from stage I to stage IV (P < 0.001). In contrast, the CD68+ macrophage, IFN-γ+, TNF-α+, and iNOS+ immunolabeling fractions increased from stage I to stage IV (P < 0.001). In the early stages, culture-negative cows showed a significantly higher fraction of CD68+ macrophage (P = 0.03) and iNOS+ (P = 0.007) immunolabeling fractions than culture-positive cows. Similarly, at advanced granuloma stages, culture-negative cows demonstrated significantly higher mean proportions of CD3+ T cells (P < 0.001) than culture-positive cows. Thus, this study demonstrates that, following natural infection of cows with M. bovis, as the stage of granuloma increases from stage I to stage IV, the immunolabeling fraction of CD3+ cells decreases, while the CD68+ macrophage, IFN-γ+, TNF-α+, and iNOS+ immunolabeling fractions increases.


Assuntos
Citocinas/metabolismo , Granuloma/metabolismo , Macrófagos/imunologia , Mycobacterium bovis/isolamento & purificação , Linfócitos T/imunologia , Tuberculose Bovina/metabolismo , Animais , Antígenos CD/metabolismo , Antígenos de Diferenciação Mielomonocítica/metabolismo , Doenças Assintomáticas , Complexo CD3/metabolismo , Bovinos , Etiópia , Feminino , Granuloma/imunologia , Granuloma/microbiologia , Granuloma/patologia , Imuno-Histoquímica , Interferon gama/metabolismo , Pulmão/imunologia , Pulmão/metabolismo , Pulmão/microbiologia , Pulmão/patologia , Linfonodos/imunologia , Linfonodos/metabolismo , Linfonodos/microbiologia , Linfonodos/patologia , Macrófagos/metabolismo , Óxido Nítrico Sintase/metabolismo , Índice de Gravidade de Doença , Linfócitos T/metabolismo , Tuberculose Bovina/imunologia , Tuberculose Bovina/microbiologia , Tuberculose Bovina/patologia , Fator de Necrose Tumoral alfa/metabolismo
11.
Tuberculosis (Edinb) ; 124: 101979, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32814303

RESUMO

Bovine tuberculosis is an important animal health problem and the predominant cause of zoonotic tuberculosis worldwide. It results in serious economic burden due to losses in productivity and the cost of control programmes. Control could be greatly improved by the introduction of an efficacious cattle vaccine but the most likely candidate, BCG, has several limitations including variable efficacy. Augmentation of BCG with a subunit vaccine booster has been shown to increase protection but the selection of antigens has hitherto been left largely to serendipity. In the present study, we take a rational approach to identify the protective antigens of BCG, selecting a BCG transposon mutant library in naïve and BCG-vaccinated cattle. Ten mutants had increased relative survival in vaccinated compared to naïve cattle, consistent with loss of protective antigen targets making the mutants less visible to the BCG immune response. The immunogenicity of three putative protective antigens, BCG_0116, BCG_0205 (YrbE1B) and BCG_1448 (PPE20) was investigated using peptide pools and PBMCs from BCG vaccinated cattle. BCG vaccination induced PBMC to release elevated levels of IP10, IL-17a and IL-10 in response to all three antigens. Taken together, the data supports the further study of these antigens for use in subunit vaccines.


Assuntos
Antígenos de Bactérias/administração & dosagem , Antígenos de Bactérias/genética , Vacina BCG/administração & dosagem , Imunogenicidade da Vacina , Leucócitos Mononucleares/imunologia , Mycobacterium tuberculosis/genética , Tuberculose Bovina/prevenção & controle , Vacinação/veterinária , Animais , Antígenos de Bactérias/imunologia , Vacina BCG/imunologia , Bovinos , Citocinas/imunologia , Citocinas/metabolismo , Elementos de DNA Transponíveis , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/microbiologia , Mutação , Mycobacterium tuberculosis/imunologia , Tuberculose Bovina/imunologia , Tuberculose Bovina/metabolismo , Tuberculose Bovina/microbiologia
12.
Res Vet Sci ; 132: 416-425, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32768870

RESUMO

Bovine tuberculosis (bTB) caused by Mycobacterium bovis has a significant economic impact worldwide each year. Control of bTB is based on skin testing and removal of reactors. However, additional strategies are required to control this disorder. Natural disease resistance has been defined as the inherent capacity of an individual to resist disease when exposed to pathogens without previous exposure or immunization. However, little is known about natural disease resistance against Mycobacterium bovis in cattle. In this study, we aimed to identify candidate biomarkers to detect host resistance to M. bovis. We used a microbicidal assay to identify the resistance phenotype. A genomic microarray analysis was carried out on RNA from 2 resistant (R) and 2 susceptible (S) cows. Our results evidenced 69 differentially expressed genes. A subset of six genes that showed differential up (IL1RN), and down-regulation (VNN, GATM, ARHGEF11, NAAA and HSPA2) were selected for further analysis. To further validate the candidate biomarkers, we identified the R phenotype in 31 cattle (9 R and 22 S). Macrophage mRNA was isolated from this group of cattle. Expression of candidate biomarkers was evaluated by qPCR 2-ΔCt and ROC curves to determine their diagnostic potential. Candidates IL1RN and ARHGEF11 discriminates between R and S cattle. Furthermore, combination of candidates ARHGEF11: VNN: HSPA2 discriminate between R from S with AUC 0.7993 and agreement index of 0.853 (p ≤ 0.01). Our data suggest that candidate biomarkers may support the preliminary screening to identify natural resistance in herds against Mycobacterium bovis in Holstein-Friesian cattle.


Assuntos
Biomarcadores/metabolismo , Mycobacterium bovis/imunologia , Tuberculose Bovina/imunologia , Animais , Bovinos , Feminino , Genômica , Imunidade Inata , Masculino , Análise em Microsséries/veterinária , Tuberculose Bovina/microbiologia
13.
BMC Immunol ; 21(1): 26, 2020 05 12.
Artigo em Inglês | MEDLINE | ID: mdl-32397995

RESUMO

BACKGROUND: Understanding pathogenic mechanisms is imperative for developing novel treatment to the tuberculosis, an important public health burden worldwide. Recent studies demonstrated that host cholesterol levels have implications in the establishment of Mycobacterium tuberculosis (M. tuberculosis, Mtb) infection in host cells, in which the intracellular cholesterol-mediated ATP-binding cassette transporters (ABC-transporters) and cholesterol acyltransferase1 (ACAT1) exhibited abilities to regulate macrophage autophagy induced by Mycobacterium bovis bacillus Calmette-Guérin (BCG). RESULTS: The results showed that a down-regulated expression of the ABC-transporters and ACAT1 in primary bovine alveolar macrophages (AMs) and murine RAW264.7 cells in response to a BCG infection. The inhibited expression of ABC-transporters and ACAT1 was associated with the reduction of intracellular free cholesterol, which in turn induced autophagy in macrophages upon to the Mycobacterial infection. These results strongly suggest an involvement of ABC-transporters and ACAT1 in intracellular cholesterol-mediated autophagy in AMs in response to BCG infection. CONCLUSION: This study thus provides an insight into into a mechanism by which the cholesterol metabolism regulated the autophagy in macrophages in response to mycobacterial infections.


Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Autofagia/fisiologia , Colesterol/metabolismo , Macrófagos Alveolares/metabolismo , Esterol O-Aciltransferase/metabolismo , Tuberculose Bovina/metabolismo , Animais , Vacina BCG/imunologia , Bovinos , Linhagem Celular , Regulação para Baixo/fisiologia , Macrófagos Alveolares/imunologia , Camundongos , Mycobacterium bovis/imunologia , Mycobacterium bovis/patogenicidade , Mycobacterium tuberculosis/imunologia , Mycobacterium tuberculosis/patogenicidade , Células RAW 264.7 , Tuberculose/imunologia , Tuberculose/metabolismo , Tuberculose Bovina/imunologia
14.
Comp Immunol Microbiol Infect Dis ; 69: 101424, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31972498

RESUMO

Cattle, bison and buffaloes are susceptible to Mycobacterium bovis, the causative agent for bovine tuberculosis. Accurate and timely identification of infected animals is critical for improved management and control of disease in these species. Bovids develop humoral immune responses to M. bovis infection making serological tests attractive for tuberculosis screening. However, optimization and validation of antibody assays designed for various animal species require understanding of antigen recognition patterns in each target host. The objective of this study was to characterize serological reactivity profiles generated by cattle, American bison, and African buffaloes in tuberculosis. Serum samples from M. bovis-infected animals were tested for the presence of IgM and IgG antibodies to MPB70/MPB83 and CFP10/ESAT6 chimeric proteins using Dual-Path Platform technology. All three host species showed IgG responses of higher magnitude and frequency than IgM responses; further, IgM seroreactivity was limited to MPB70/MPB83, whereas IgG antibodies recognized both test antigens. In cattle, the IgM and IgG responses were elicited mainly by MPB70/MPB83, whereas bison and buffaloes showed similar IgG seroreactivity rates for MPB70/MPB83 and CFP10/ESAT6 antigens. The findings demonstrate distinct patterns of predominant antigen recognition by different bovid species in M. bovis infection.


Assuntos
Anticorpos Antibacterianos/imunologia , Antígenos de Bactérias/imunologia , Interações Hospedeiro-Patógeno/imunologia , Mycobacterium bovis/imunologia , Tuberculose Bovina/imunologia , Tuberculose Bovina/microbiologia , Animais , Anticorpos Antibacterianos/sangue , Bovinos , Imunoensaio , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Imunoglobulina M/sangue , Imunoglobulina M/imunologia
15.
Comp Immunol Microbiol Infect Dis ; 70: 101369, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31718809

RESUMO

The endemic presence of bovine tuberculosis (BTB) in African buffaloes in South Africa has severe consequences for BTB control in domestic cattle, buffalo ranching and wildlife conservation, and poses a potential risk to public health. This study determined the BTB prevalence in free-ranging buffaloes in two game reserves and assessed the influence of the prevalence of mycobacterial infections on the performance of a commercial cattle-specific serological assay for BTB (TB ELISA). Buffaloes (n = 997) were tested with the tuberculin skin test and TB ELISA; a subset (n = 119) was tested longitudinally. Culture, PCR and sequencing were used to confirm infection with M. bovis and/or non-tuberculous mycobacteria (NTM). Prevalence of BTB, but not NTM, influenced the TB ELISA performance. Multiple testing did not increase test confidence. The findings strongly illustrate the need for development of novel assays that can supplement existing assays for a more comprehensive testing scheme for BTB in African buffaloes.


Assuntos
Búfalos/microbiologia , Ensaio de Imunoadsorção Enzimática/normas , Kit de Reagentes para Diagnóstico/normas , Testes Sorológicos/normas , Teste Tuberculínico/veterinária , Tuberculose Bovina/diagnóstico , Animais , Animais Selvagens/microbiologia , Bovinos , Mycobacterium bovis , Prevalência , África do Sul/epidemiologia , Tuberculose Bovina/epidemiologia , Tuberculose Bovina/imunologia
16.
Vet Immunol Immunopathol ; 220: 109988, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31846797

RESUMO

Whole blood based assays, particularly interferon gamma (IFN-γ) release assays (IGRAs), are used for the diagnosis of both bovine and human tuberculosis (TB). The aim of the current study was to evaluate a panel of cytokines and chemokines for potential use as diagnostic readouts indicative of Mycobacterium bovis (M. bovis) infection in cattle. A gene expression assay was used to determine the kinetics of the response to M. bovis purified protein derivative and a fusion protein consisting of ESAT-6, CFP10, and Rv3615c upon aerosol infection with ∼104 cfu of M. bovis. The panel of biomarkers included: IFN-γ, CXCL9, CXCL10, CCL2, CCL3, TNF-α, IL-1α, IL-1ß, IL-1Ra, IL-22, IL-21 and IL-13. Protein levels of IFN-γ, CXCL9, and CXCL10 were determined by ELISA. Findings suggest that CXCL9, CXCL10, IL-21, IL-13, and several acute phase cytokines may be worth pursuing as diagnostic biomarkers of M. bovis infection in cattle.


Assuntos
Doenças dos Bovinos/diagnóstico , Citocinas/genética , Imunidade Celular , Tuberculose Bovina/diagnóstico , Tuberculose Bovina/imunologia , Animais , Biomarcadores/sangue , Bovinos , Doenças dos Bovinos/sangue , Doenças dos Bovinos/imunologia , Quimiocina CXCL10/sangue , Quimiocina CXCL9/sangue , Citocinas/imunologia , Expressão Gênica , Interferon gama , Testes de Liberação de Interferon-gama , Masculino , Mycobacterium bovis , Tuberculose Bovina/sangue
18.
Sci Rep ; 9(1): 18012, 2019 11 29.
Artigo em Inglês | MEDLINE | ID: mdl-31784649

RESUMO

Tuberculosis (TB) disease still kills 1-person every 21-seconds. Few TB diagnostic tests are considered truly appropriate for point of care settings. The WHO-endorsed immunodiagnostic Alere Determine Lipoarabinomannan Ag-test (LAM-test) detects Mycobacterium tuberculosis complex LAM in urine, and its use is recommended for TB diagnosis among HIV co-infected individuals with low CD4 T-cell counts. Here we found that a simple 15-minute enzymatic treatment at room temperature of LAM-spiked urine with α-mannosidase (for human TB), and LAM-spiked milk with combined lactase and caseinase (for bovine TB), enhanced 10-fold the detection levels of the LAM-test and thus, improved the detection of LAM by the LAM-test in urine and milk that otherwise could be missed in the field. Future separate clinical research studies specifically designed to address the potential of these findings are required.


Assuntos
Antígenos de Bactérias/isolamento & purificação , Testes Imunológicos/métodos , Lipopolissacarídeos/isolamento & purificação , Sistemas Automatizados de Assistência Junto ao Leito , Tuberculose Bovina/diagnóstico , Tuberculose/diagnóstico , Animais , Antígenos de Bactérias/imunologia , Bovinos , Feminino , Humanos , Lipopolissacarídeos/imunologia , Leite/microbiologia , Mycobacterium bovis/imunologia , Mycobacterium bovis/isolamento & purificação , Mycobacterium tuberculosis/imunologia , Mycobacterium tuberculosis/isolamento & purificação , Sensibilidade e Especificidade , Tuberculose/imunologia , Tuberculose/microbiologia , Tuberculose/urina , Tuberculose Bovina/imunologia , Tuberculose Bovina/microbiologia , Urina/microbiologia
19.
Int J Mol Sci ; 20(23)2019 Nov 29.
Artigo em Inglês | MEDLINE | ID: mdl-31795474

RESUMO

Mycobacterium bovis (M. bovis) is the causative agent of bovine tuberculosis in cattle population across the world. Human beings are at equal risk of developing tuberculosis beside a wide range of M. bovis infections in animal species. Autophagic sequestration and degradation of intracellular pathogens is a major innate immune defense mechanism adopted by host cells for the control of intracellular infections. It has been reported previously that the catalytic subunit of protein phosphatase 2A (PP2Ac) is crucial for regulating AMP-activated protein kinase (AMPK)-mediated autophagic signaling pathways, yet its role in tuberculosis is still unclear. Here, we demonstrated that M. bovis infection increased PP2Ac expression in murine macrophages, while nilotinib a tyrosine kinase inhibitor (TKI) significantly suppressed PP2Ac expression. In addition, we observed that TKI-induced AMPK activation was dependent on PP2Ac regulation, indicating the contributory role of PP2Ac towards autophagy induction. Furthermore, we found that the activation of AMPK signaling is vital for the regulating autophagy during M. bovis infection. Finally, the transient inhibition of PP2Ac expression enhanced the inhibitory effect of TKI-nilotinib on intracellular survival and multiplication of M. bovis in macrophages by regulating the host's immune responses. Based on these observations, we suggest that PP2Ac should be exploited as a promising molecular target to intervene in host-pathogen interactions for the development of new therapeutic strategies towards the control of M. bovis infections in humans and animals.


Assuntos
Proteínas Quinases Ativadas por AMP/imunologia , Macrófagos/imunologia , Mycobacterium bovis/imunologia , Proteína Fosfatase 2/imunologia , Tuberculose/veterinária , Animais , Autofagia , Bovinos , Interações Hospedeiro-Patógeno , Humanos , Macrófagos/microbiologia , Camundongos , Mycobacterium bovis/fisiologia , Fagocitose , Células RAW 264.7 , Tuberculose/imunologia , Tuberculose/microbiologia , Tuberculose Bovina/imunologia , Tuberculose Bovina/microbiologia
20.
BMC Vet Res ; 15(1): 359, 2019 Oct 22.
Artigo em Inglês | MEDLINE | ID: mdl-31640672

RESUMO

BACKGROUND: Bovine tuberculosis (bTB) is prevalent in dairy cattle in Ethiopia. Currently used diagnostic tools such as the single intradermal comparative tuberculin test (SICTT) are time consuming and labor intensive. A rapid, easy-to-use and cost-effective diagnostic test would greatly contribute to the control of bTB in developing countries like Ethiopia. In the present study, two point-of-care diagnostic tests were evaluated for the detection of bTB: LIONEX® Animal TB Rapid test, a membrane-based test for the detection of antibodies to Mycobacterium bovis in blood and ALERE® Determine TB Lipoarabinomannan (LAM) Ag, an immunoassay for the detection of lipoarabinomannan (LAM) antigen (Ag) of mycobacteria in urine. A combination of the SICTT and gamma interferon (IFN-γ) test was used as the gold standard for the validation of these point-of-care tests, as it was not feasible to slaughter the study animals to carry out the historical gold standard of mycobacterial culture. A total of 175 heads of cattle having three different bTB infection categories (positive SICTT, negative SICTT, and unknown SICTT status) were used for this study. RESULT: The sensitivity and specificity of TB LAM Ag were 72.2% (95% CI = 62.2, 80.4) and 98.8% (95% CI = 93.6, 99.7), respectively, while the sensitivity and specificity of the LIONEX Animal TB rapid test assay were 54% (95% CI = 44.1 64.3) and 98.8% (95% CI = 93.6, 99.7) respectively. The agreement between TB LAM Ag and SICTT was higher (κ = 0.85; 95% CI = 0.65-0.94) than between TB LAM Ag and IFN-γ (κ = 0.67; 95% CI = 0.52-0.81). The agreement between LIONEX Animals TB Rapid blood test and SICTT was substantial, (κ = 0.63; 95% CI = 0.49-0.77) while the agreement between LIONEX Animal TB rapid blood test and IFN-γ test was moderate (κ = 0.53; 95% CI = 0.40-0.67). Analysis of receiver operating curve (ROC) indicated that the area under the ROC curve (AUC) for TB LAM Ag was 0.85 (95% CI = 0.79-0.91) while it was 0.76 (95% CI; =0.69-0.83) for LIONEX Animal TB rapid test assay. CONCLUSION: This study showed that TB LAM Ag had a better diagnostic performance and could potentially be used as ancillary either to SICTT or IFN-γ test for diagnosis of bTB.


Assuntos
Imunoensaio/veterinária , Lipopolissacarídeos/sangue , Mycobacterium tuberculosis/isolamento & purificação , Tuberculose Bovina/diagnóstico , Animais , Anticorpos Antibacterianos/sangue , Bovinos , Etiópia , Interferon gama/sangue , Lipopolissacarídeos/imunologia , Mycobacterium tuberculosis/imunologia , Sensibilidade e Especificidade , Teste Tuberculínico/veterinária , Tuberculose Bovina/sangue , Tuberculose Bovina/imunologia
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...