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1.
Medicine (Baltimore) ; 100(6): e24619, 2021 Feb 12.
Artículo en Inglés | MEDLINE | ID: mdl-33578573

RESUMEN

ABSTRACT: CD4+T cell epitopes plays a key role in anti-tuberculosis (TB) immunity, CD4+T cell epitopes suitable for the domestic population are lacking. Therefore, we predicted and identified novel CD4+T cell epitopes.The bioinformatics software, namely, DNAStar (DNASTAR of the United States), SYFPEITHI (INTERFACTORS INSTITUT Für ZELL Biologie of Germany), RANKPEP, and NetMHC IIpan (National Cancer Institute, United States of America), were used to comprehensively predict the CD4+T cell immune epitope of Mycobacterium TB, and the predicted epitope polypeptide was synthesized by the standard Fmoc scheme. The proliferation of PBMC and CD4+T cells stimulated by peptides was preliminarily detected by the CCK8 method. Then, the candidate polypeptides screened out by the CCK8 method were verified again by the BrdU assay, and flow cytometry was performed to analyze further the extent of their stimulation on the proliferation of CD4+T cells. The changes in the secreted cytokines IFN-γ, TNF-α, IL-2, and IL-10 before and after the candidate polypeptide stimulation of CD4+T lymphocytes were detected by ELISA. The preliminary humoral immunity test was conducted by indirect ELISA to evaluate the serological diagnostic value of the CD4+T cell epitope polypeptide.In this study, 5 novel candidate CD4+T cell epitope polypeptides with the amino acid sequences of LQGQWRGAAGTAAQA, PVTLAETGSTLLYPL, AAAWGGSGSEAYQGV, QFVYAGAMSGLLDPS, and KAALTRTASNMNAAA and others that have not been reported in the research were predicted. For convenience, the 5 candidates were successively named as P39, P50, P40, P185, and P62. P39, P62, and the mixed peptide P39+P62 could effectively induce the proliferation of CD4+T cells and increase the secretion of IFN-γ, TNF-α, and IL-2 from the CD4+T cells, while reducing the content of IL-10. The serological test showed that the sensitivity, specificity, and area under the receiver operating characteristic curve (AUC) of P39 were 75%, 67.71%, and 0.844, respectively. The sensitivity, specificity, and AUC of P62 were 91.66%, 46.87%, and 0.649, respectively. The sensitivity of the mixed peptide P39+P62 was 95.83%, the specificity was 97.91%, and the AUC was 0.793.The P39 and P62 polypeptides were predicted and identified as potential CD4+T cell immune epitope polypeptides of M. TB. The polypeptide had better diagnosis effect, which provided potential candidate epitope polypeptides for the development of TB-specific diagnosis reagents and novel TB epitope vaccines.


Asunto(s)
Antígenos Bacterianos/inmunología , Linfocitos T CD4-Positivos/inmunología , Epítopos de Linfocito T/inmunología , Mycobacterium tuberculosis/inmunología , Ensayo de Inmunoadsorción Enzimática , Humanos , Sensibilidad y Especificidad
2.
Proc Natl Acad Sci U S A ; 118(4)2021 01 26.
Artículo en Inglés | MEDLINE | ID: mdl-33468674

RESUMEN

The global incidence of tuberculosis remains unacceptably high, with new preventative strategies needed to reduce the burden of disease. We describe here a method for the generation of synthetic self-adjuvanted protein vaccines and demonstrate application in vaccination against Mycobacterium tuberculosis Two vaccine constructs were designed, consisting of full-length ESAT6 protein fused to the TLR2-targeting adjuvants Pam2Cys-SK4 or Pam3Cys-SK4 These were produced by chemical synthesis using a peptide ligation strategy. The synthetic self-adjuvanting vaccines generated powerful local CD4+ T cell responses against ESAT6 and provided significant protection in the lungs from virulent M. tuberculosis aerosol challenge when administered to the pulmonary mucosa of mice. The flexible synthetic platform we describe, which allows incorporation of adjuvants to multiantigenic vaccines, represents a general approach that can be applied to rapidly assess vaccination strategies in preclinical models for a range of diseases, including against novel pandemic pathogens such as SARS-CoV-2.


Asunto(s)
Mycobacterium tuberculosis/inmunología , Vacunas contra la Tuberculosis/farmacología , Tuberculosis/inmunología , Tuberculosis/prevención & control , Vacunas Conjugadas/farmacología , Adyuvantes Inmunológicos/farmacología , Animales , Antígenos Bacterianos/inmunología , Vacuna BCG/inmunología , Vacuna BCG/farmacología , Proteínas Bacterianas , Linfocitos T CD4-Positivos/inmunología , Modelos Animales de Enfermedad , Femenino , Ratones , Ratones Endogámicos C57BL , Receptor Toll-Like 2/inmunología , Vacunas contra la Tuberculosis/inmunología , Vacunas Conjugadas/inmunología , Vacunas Sintéticas/inmunología , Vacunas Sintéticas/farmacología
3.
J Nippon Med Sch ; 87(5): 299-303, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-33311009

RESUMEN

In Japan, pneumococcal vaccine has been routinely administered since 2010 to prevent invasive pneumococcal diseases such as Streptococcus pneumoniae meningitis. We describe a case of pneumococcal meningitis in a 7-month-old girl who had received three doses of 13-valent pneumococcal conjugate vaccine. Brain magnetic resonance imaging showed infarcts in the right frontal region, and she was treated with antibiotics, intravenous immunoglobulin, dexamethasone, and edaravone. On day 27, an enhanced brain CT scan showed improvement of abnormal findings in the frontal region, except for slight atrophy. The S. pneumoniae serotype was 12F, which is not included in the 13-valent pneumococcal conjugate vaccine. A future vaccine is expected to use cross-reactivity to target common antigens.


Asunto(s)
Meningitis Bacterianas/etiología , Meningitis Bacterianas/microbiología , Infecciones Neumocócicas , Vacunas Neumococicas/efectos adversos , Streptococcus pneumoniae , Vacunas Conjugadas/efectos adversos , Antígenos Bacterianos/inmunología , Encéfalo/diagnóstico por imagen , Reacciones Cruzadas , Dexametasona/uso terapéutico , Edaravona/uso terapéutico , Femenino , Humanos , Inmunoglobulinas Intravenosas , Lactante , Imagen por Resonancia Magnética , Meningitis Bacterianas/diagnóstico por imagen , Meningitis Bacterianas/tratamiento farmacológico , Infecciones Neumocócicas/prevención & control , Serogrupo , Streptococcus pneumoniae/genética , Streptococcus pneumoniae/inmunología , Tomografía Computarizada por Rayos X
4.
Nihon Saikingaku Zasshi ; 75(2): 185-194, 2020.
Artículo en Japonés | MEDLINE | ID: mdl-33361654

RESUMEN

Countless numbers of bacteria inhabit the intestinal tract. One of the important functions of gut microbiota is the "colonization resistance" against infection by pathogenic microorganisms. However, detailed mechanism of the colonization resistance of intestinal bacteria is still largely unknown. We tried to identify molecular and cellular mechanism of it and found that antigen presentation by dendritic cells is required for the induction of intestinal segmented filamentous bacteria (SFB)-induced T helper 17 (Th17) cells that contribute to the protection against infection by Citrobacter rodentium. We further identified that gut Th17 cells selectively recognize antigens derived from SFB. We also revealed that SFB induce α1,2-fucose, one of carbohydrate chains, expressed on the intestinal epithelial cells mediated by group 3 innate lymphoid cells. Epithelial α1,2-fucose protected against infection by pathogenic bacterium Salmonella typhimurium. Furthermore, it was found that intestinal bacteria inhibit colonization of the pathogenic fungus Candida albicans as well as pathogenic bacteria. From these studies, detailed mechanism of "colonization resistance" against pathogenic microorganisms by intestinal bacteria has been clarified.


Asunto(s)
Candida albicans/patogenicidad , Citrobacter rodentium/patogenicidad , Microbioma Gastrointestinal/inmunología , Microbioma Gastrointestinal/fisiología , Interacciones Microbiota-Huesped/inmunología , Sistema Inmunológico/inmunología , Mucosa Intestinal/microbiología , Salmonella typhimurium/patogenicidad , Células Th17/inmunología , Animales , Presentación de Antígeno , Antígenos Bacterianos/inmunología , Adhesión Bacteriana/inmunología , Candida albicans/inmunología , Citrobacter rodentium/inmunología , Células Dendríticas/inmunología , Fucosa/metabolismo , Humanos , Mucosa Intestinal/inmunología , Mucosa Intestinal/metabolismo , Ratones , Salmonella typhimurium/inmunología
5.
BMC Infect Dis ; 20(1): 822, 2020 Nov 10.
Artículo en Inglés | MEDLINE | ID: mdl-33172393

RESUMEN

BACKGROUND: Plague, a fatal disease caused by the bacillus, Yersinia pestis, still affects resources-limited countries. Information on antibody response to plague infection in human is scarce. Anti-F1 Ig G are among the known protective antibodies against Y. pestis infection. As a vaccine preventable disease, knowledge on antibody response is valuable for the development of an effective vaccine to reduce infection rate among exposed population in plague-endemic regions. In this study, we aim to describe short and long-term humoral immune responses against Y. pestis in plague-confirmed patients from Madagascar, the most affected country in the world. METHODS: Bubonic (BP) and pneumonic plague (PP) patients were recruited from plague- endemic foci in the central highlands of Madagascar between 2005 and 2017. For short-term follow-up, 6 suspected patients were enrolled and prospectively investigated for kinetics of the anti-F1 IgG response, whereas the persistence of antibodies was retrospectively studied in 71 confirmed convalescent patients, using an ELISA which was validated for the detection of plague in human blood samples in Madagascar. RESULTS: Similarly to previous findings, anti-F1 IgG rose quickly during the first week after disease onset and increased up to day 30. In the long-term study, 56% of confirmed cases remained seropositive, amongst which 60 and 40% could be considered as high- and low-antibody responders, respectively. Antibodies persisted for several years and up to 14.8 years for one individual. Antibody titers decreased over time but there was no correlation between titer and time elapsed between the disease onset and serum sampling. In addition, the seroprevalence rate was not significantly different between gender (P = 0.65) nor age (P = 0.096). CONCLUSION: Our study highlighted that the circulating antibody response to F1 antigen, which is specific to Y. pestis, may be attributable to individual immune responsiveness. The finding that a circulating anti-F1 antibody titer could persist for more than a decade in both BP and PP recovered patients, suggests its probable involvement in patients' protection. However, complementary studies including analyses of the cellular immune response to Y. pestis are required for the better understanding of long-lasting protection and development of a potential vaccine against plague.


Asunto(s)
Inmunidad Humoral , Peste/inmunología , Yersinia pestis/inmunología , Adolescente , Adulto , Animales , Anticuerpos Antibacterianos/sangre , Antígenos Bacterianos/inmunología , Proteínas Bacterianas/inmunología , Niño , Femenino , Estudios de Seguimiento , Humanos , Inmunoglobulina G/sangre , Madagascar/epidemiología , Masculino , Peste/epidemiología , Peste/microbiología , Estudios Prospectivos , Estudios Retrospectivos , Estudios Seroepidemiológicos , Adulto Joven
6.
Nat Commun ; 11(1): 4994, 2020 10 05.
Artículo en Inglés | MEDLINE | ID: mdl-33020485

RESUMEN

Serogroup B meningococcus (MenB) is a leading cause of meningitis and sepsis across the world and vaccination is the most effective way to protect against this disease. 4CMenB is a multi-component vaccine against MenB, which is now licensed for use in subjects >2 months of age in several countries. In this study, we describe the development and use of an ad hoc protein microarray to study the immune response induced by the three major 4CMenB antigenic components (fHbp, NHBA and NadA) in individual sera from vaccinated infants, adolescents and adults. The resulting 4CMenB protein antigen fingerprinting allowed the identification of specific human antibody repertoire correlating with the bactericidal response elicited in each subject. This work represents an example of epitope mapping of the immune response induced by a multicomponent vaccine in different age groups with the identification of protective signatures. It shows the high flexibility of this microarray based methodology in terms of high-throughput information and minimal volume of biological samples needed.


Asunto(s)
Anticuerpos Antibacterianos/inmunología , Antígenos Bacterianos/inmunología , Infecciones Meningocócicas/inmunología , Vacunas Meningococicas/inmunología , Neisseria meningitidis Serogrupo B/inmunología , Adolescente , Adulto , Anticuerpos Antibacterianos/sangre , Niño , Preescolar , Mapeo Epitopo , Humanos , Lactante , Infecciones Meningocócicas/prevención & control , Biblioteca de Péptidos , Análisis por Matrices de Proteínas , Determinación de Anticuerpos Séricos Bactericidas , Adulto Joven
7.
PLoS One ; 15(9): e0239453, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32977328

RESUMEN

BACKGROUND: Cerebrospinal fluid (CSF) oligoclonal bands (OCB) occur in chronic or post-acute phase of inflammatory diseases of the central nervous system. OBJECTIVE: To determine whether CSF OCB in patients with neuroborreliosis (NB) are specific for borrelia burgdorferi senso lato. METHODS: We performed isoelectric focusing followed by immunoblotting in CSF of 10 NB patients and 11 controls (7 patients with multiple sclerosis, 2 patients with neuromyelitis optica spectrum disease, 1 patient with dementia and 1 patient with monoclonal gammopathy). Immunoblotting was performed using an uncoated as well as a borrelia antigen pre-coated nitrocellulose membrane (NCM). OCB were counted by visual inspection and photometric analysis. OCB were compared between uncoated und pre-coated NCM both in the NB and control group. For validation purposes inter-assay precision was determined by calculating the coefficient of variation (CV). RESULTS: Borrelia-specific OCB were found in the CSF of 9 NB patients and in none of the control subjects resulting in a sensitivity of 90% and a specificity of 100%. Number of NB specific OCB were 11±7 bands by photometric analyses compared to 9±5 bands by visual inspection. Validation experiments revealed an inconsistent inter-assay precision between visual and photometric analyses (NB uncoated: visual 28% versus photometric 14%, control subject uncoated: visual 16% versus photometric 24%). CONCLUSIONS: In CSF samples with positive OCB, Borrelia-specific bands were detected in almost all NB patients and in none of the control subjects. Inconsistent inter-assay precision may be explained by a poor comparability of visual and photometric approach.


Asunto(s)
Borrelia burgdorferi/inmunología , Líquido Cefalorraquídeo/inmunología , Líquido Cefalorraquídeo/microbiología , Neuroborreliosis de Lyme/líquido cefalorraquídeo , Neuroborreliosis de Lyme/inmunología , Bandas Oligoclonales/líquido cefalorraquídeo , Bandas Oligoclonales/inmunología , Adulto , Antígenos Bacterianos/inmunología , Estudios de Casos y Controles , Estudios Transversales , Demencia/inmunología , Femenino , Humanos , Immunoblotting/métodos , Neuroborreliosis de Lyme/microbiología , Masculino , Persona de Mediana Edad , Esclerosis Múltiple/inmunología , Enfermedades del Sistema Nervioso/inmunología , Enfermedades del Sistema Nervioso/microbiología , Neuromielitis Óptica/inmunología , Paraproteinemias/inmunología , Estudios Retrospectivos , Sensibilidad y Especificidad
8.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 36(8): 680-686, 2020 Aug.
Artículo en Chino | MEDLINE | ID: mdl-32958123

RESUMEN

Objective To investigate the role of butyrophilin 3A1 (BTN3A1) in the activation and proliferation of human peripheral blood Vγ9Vδ2 T cells induced by M. tuberculosis heat resistant antigen (MTB-HAg). Methods Human peripheral blood mononuclear cells (PBMCs) were treated with BTN3A1 blocking antibody for 3 hours and then stimulated with MTB-HAg or phosphoantigen (PAg). At 24 hours of stimulation, the cells were collected to detect the expression of CD69 in Vγ9Vδ2 T cells by flow cytometry. At 20 hours of stimulation, the cells were collected to detect the proportions of cells producing helper T cell type I (Th1) cytokines IFN-γ and tumor necrosis factor α (TNF-α) in the Vγ9Vδ2 T cells. The PBMCs were also stimulated and cultured in IL-2-containing medium for 10 days, and the expansion and proliferation activity of Vγ9Vδ2 T cells were detected. Results After stimulated with MTB-HAg, the average fluorescence intensity of CD69 and the proportion of CD69 positive cells in Vγ9Vδ2 T cells decreased significantly in BTN3A1 blocked group, being 13.84% and 43.00% of those in the stimulated group, respectively. However, the average fluorescence intensity of CD69 molecules and the proportion of positive cells in PAg blocked group were significantly inhibited (3.10%, 4.47% and 9.53%, 10.91% of those in the stimulated group). The proportions of IFN-γ and TNF-α producing Vγ9Vδ2 T cells stimulated with MTB-HAg decreased significantly in the BTN3A1 blocked group, and the expansion number and cell proliferation activity of Vγ9Vδ2 T cells were also reduced significantly in the BTN3A1 blocked group. The results were similar to those of the PAg blocked group. Conclusion BTN3A1 promotes activation and proliferation of peripheral blood Vγ9Vδ2 T cells induced by MTB-HAg.


Asunto(s)
Antígenos Bacterianos , Antígenos CD , Butirofilinas , Activación de Linfocitos , Linfocitos T , Antígenos Bacterianos/inmunología , Antígenos CD/genética , Antígenos CD/metabolismo , Butirofilinas/genética , Butirofilinas/metabolismo , Proliferación Celular/genética , Humanos , Activación de Linfocitos/genética , Receptores de Antígenos de Linfocitos T gamma-delta/metabolismo , Linfocitos T/citología , Linfocitos T/inmunología
9.
J Med Microbiol ; 69(10): 1249-1252, 2020 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-32924920

RESUMEN

Erysipelothrix rhusiopathiae is a zoonotic pathogen that causes erysipelas in a variety of animals. In humans, in contrast to the cutaneous form called erysipeloid, which is an occupational disease and common in individuals who handle raw meat and fish, invasive systemic infections are unusual. E. rhusiopathiae expresses an immunogenic surface protein, Spa (surface protective antigen), which is involved in virulence. Among the antigenically different Spa proteins (SpaA, B and C), which are mostly associated with serovars, SpaA is by far the most prevalent in E. rhusiopathiae isolates from diseased animals. However, the Spa type has not been examined for human isolates, and it is unknown whether SpaB- or SpaC-possessing isolates can cause disease in humans. A Gram-positive, rod-shaped bacterium isolated from a case of human pyogenic spondylitis was analysed. The bacterium was identified as E. rhusiopathiae by a routine biochemical test and MS, and ultimately confirmed by an E. rhusiopathiae-specific PCR assay. Spa typing by sequencing revealed the SpaB type, and the serovar of the strain was identified as untypeable by a conventional agar gel precipitation test, but determined to be serovar 6 by a serotyping PCR assay. Sequence analysis of the serovar-defining chromosomal region revealed that the isolate displayed the same gene organization as the serovar 6 reference strain, but the region was disrupted by an insertion sequence element, suggesting that the isolate originated from a serovar 6 strain. These results highlight that unusual, spaB-possessing E. rhusiopathiae strains can potentially pose serious risks to humans.


Asunto(s)
Antígenos de Superficie/inmunología , Infecciones por Erysipelothrix/microbiología , Erysipelothrix/genética , Anciano de 80 o más Años , Antígenos Bacterianos/inmunología , Antígenos Bacterianos/metabolismo , Antígenos de Superficie/metabolismo , Proteínas Bacterianas/inmunología , Proteínas Bacterianas/metabolismo , Erysipelothrix/metabolismo , Erysipelothrix/patogenicidad , Femenino , Humanos , Proteínas de la Membrana/genética , Reacción en Cadena de la Polimerasa/métodos , Serogrupo , Serotipificación , Virulencia
10.
BMC Infect Dis ; 20(1): 677, 2020 Sep 17.
Artículo en Inglés | MEDLINE | ID: mdl-32942991

RESUMEN

BACKGROUND: Approximately 80% - 90% of individuals infected with latent Mycobacterium tuberculosis (Mtb) remain protected throughout their life-span. The release of unique, latent-phase antigens are known to have a protective role in the immune response against Mtb. Although the BCG vaccine has been administered for nine decades to provide immunity against Mtb, the number of TB cases continues to rise, thereby raising doubts on BCG vaccine efficacy. The shortcomings of BCG have been associated with inadequate processing and presentation of its antigens, an inability to optimally activate T cells against Mtb, and generation of regulatory T cells. Furthermore, BCG vaccination lacks the ability to eliminate latent Mtb infection. With these facts in mind, we selected six immunodominant CD4 and CD8 T cell epitopes of Mtb expressed during latent, acute, and chronic stages of infection and engineered a multi-epitope-based DNA vaccine (C6). RESULT: BALB/c mice vaccinated with the C6 construct along with a BCG vaccine exhibited an expansion of both CD4 and CD8 T cell memory populations and augmented IFN-γ and TNF-α cytokine release. Furthermore, enhancement of dendritic cell and macrophage activation was noted. Consequently, illustrating the elicitation of immunity that helps in the protection against Mtb infection; which was evident by a significant reduction in the Mtb burden in the lungs and spleen of C6 + BCG administered animals. CONCLUSION: Overall, the results suggest that a C6 + BCG vaccination approach may serve as an effective vaccination strategy in future attempts to control TB.


Asunto(s)
Vacuna BCG/inmunología , Epítopos de Linfocito T , Tuberculosis/prevención & control , Vacunas de ADN/inmunología , Animales , Antígenos Bacterianos/inmunología , Vacuna BCG/genética , Vacuna BCG/farmacología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Epítopos de Linfocito T/genética , Femenino , Memoria Inmunológica , Interferón gamma/metabolismo , Tuberculosis Latente/prevención & control , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Mycobacterium tuberculosis/inmunología , Tuberculosis/inmunología , Factor de Necrosis Tumoral alfa/metabolismo , Vacunas de ADN/farmacología
11.
Proc Natl Acad Sci U S A ; 117(34): 20848-20859, 2020 08 25.
Artículo en Inglés | MEDLINE | ID: mdl-32778586

RESUMEN

Tuberculosis (TB) claims 1.5 million lives per year. This situation is largely due to the low efficacy of the only licensed TB vaccine, Bacillus Calmette-Guérin (BCG) against pulmonary TB. The metabolic disease type 2 diabetes (T2D) is a risk factor for TB and the mechanisms underlying increased TB susceptibility in T2D are not well understood. Furthermore, it is unknown if new TB vaccines will provide protection in the context of T2D. Here we used a diet-induced murine model of T2D to investigate the underlying mechanisms of TB/T2D comorbidity and to evaluate the protective capacity of two experimental TB vaccines in comparison to conventional BCG. Our data reveal a distinct immune dysfunction that is associated with diminished recognition of mycobacterial antigens in T2D. More importantly, we provide compelling evidence that mucosal delivery of recombinant BCG strains expressing the Mycobacterium tuberculosis (Mtb) ESX-1 secretion system (BCG::RD1 and BCG::RD1 ESAT-6 ∆92-95) are safe and confer superior immunity against aerosol Mtb infection in the context of T2D. Our findings suggest that the remarkable anti-TB immunity by these recombinant BCG strains is achieved via augmenting the numbers and functional capacity of antigen presenting cells in the lungs of diabetic mice.


Asunto(s)
Antígenos Bacterianos/farmacología , Proteínas Bacterianas/farmacología , Diabetes Mellitus Tipo 2/metabolismo , Animales , Antígenos Bacterianos/inmunología , Antígenos Bacterianos/metabolismo , Vacuna BCG , Proteínas Bacterianas/inmunología , Proteínas Bacterianas/metabolismo , Diabetes Mellitus Experimental , Modelos Animales de Enfermedad , Pulmón/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Membrana Mucosa/inmunología , Mycobacterium bovis/inmunología , Mycobacterium tuberculosis/inmunología , Tuberculosis/inmunología , Vacunas contra la Tuberculosis/inmunología , Vacunación
12.
Nat Commun ; 11(1): 3763, 2020 07 28.
Artículo en Inglés | MEDLINE | ID: mdl-32724132

RESUMEN

In both animals and plants, the perception of bacterial flagella by immune receptors elicits the activation of defence responses. Most plants are able to perceive the highly conserved epitope flg22 from flagellin, the main flagellar protein, from most bacterial species. However, flagellin from Ralstonia solanacearum, the causal agent of the bacterial wilt disease, presents a polymorphic flg22 sequence (flg22Rso) that avoids perception by all plants studied to date. In this work, we show that soybean has developed polymorphic versions of the flg22 receptors that are able to perceive flg22Rso. Furthermore, we identify key residues responsible for both the evasion of perception by flg22Rso in Arabidopsis and the gain of perception by the soybean receptors. Heterologous expression of the soybean flg22 receptors in susceptible plant species, such as tomato, enhances resistance to bacterial wilt disease, demonstrating the potential of these receptors to enhance disease resistance in crop plants.


Asunto(s)
Flagelina/inmunología , Inmunidad de la Planta , Proteínas de Plantas/inmunología , Receptores Inmunológicos/inmunología , Soja/inmunología , Antígenos Bacterianos/genética , Antígenos Bacterianos/inmunología , Antígenos Bacterianos/metabolismo , Arabidopsis/genética , Arabidopsis/inmunología , Arabidopsis/metabolismo , Resistencia a la Enfermedad/genética , Resistencia a la Enfermedad/inmunología , Epítopos/inmunología , Flagelina/genética , Flagelina/metabolismo , Evasión Inmune/genética , Enfermedades de las Plantas/inmunología , Enfermedades de las Plantas/microbiología , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas Modificadas Genéticamente , Polimorfismo Genético/inmunología , Ralstonia solanacearum/inmunología , Ralstonia solanacearum/patogenicidad , Receptores Inmunológicos/genética , Receptores Inmunológicos/metabolismo , Soja/genética , Soja/metabolismo , Soja/microbiología
13.
Mol Immunol ; 125: 123-130, 2020 09.
Artículo en Inglés | MEDLINE | ID: mdl-32659597

RESUMEN

The development of a more efficient vaccine is needed to improve tuberculosis control. One of the current approaches is to identify immunogenic T-cell peptides that can elicit a protective and specific immune response. These peptides come from immunogenic proteins of the pathogen. The PE_PGRS33 protein of Mycobacterium tuberculosis has been proved immunogenic. However, little is known about immunogenic T-cell peptides of PE_PGRS33 and their interactions with MHC-II molecules. Therefore, we used the SYFPHEITHI database to determine the immunogenic PE_PGRS33 T-cell peptides. Next, we built homology models by using MOE v2018.1 software in order to obtain information about the specific interactions between the peptides and I-Ak. The AlgPred server was employed to look for allergenic sites in PE_PGRS33. We developed a sequence alignment between PE_PGRS33 and all the human proteins by using BLAST. Three peptides were commercially synthesized, and their activity was evaluated in vitro by the stimulation of PBMC from household contacts of TB patients. Our in silico results showed five immunogenic T-cell peptides. BLAST analysis showed low homology of PE_PGRS33 with human proteins and AlgPred did not reveal allergenic sites in PE_PGRS33. The three peptides triggered the activation of CD4+ T cells from the households contacts, showed by the production of IFN-γ. We identified three immunogenic peptides of PE_PGRS33 that demonstrated activity in vitro which allows to deepen into the immune response towards mycobacterial antigens, moving forward to the identification of new vaccine candidates.


Asunto(s)
Antígenos Bacterianos/inmunología , Linfocitos T CD4-Positivos/inmunología , Mycobacterium tuberculosis/inmunología , Vacunas contra la Tuberculosis/inmunología , Humanos , Activación de Linfocitos/inmunología , Péptidos/inmunología , Vacunas de Subunidad/inmunología
14.
BMC Infect Dis ; 20(1): 469, 2020 Jul 02.
Artículo en Inglés | MEDLINE | ID: mdl-32615981

RESUMEN

BACKGROUND: Interferon-γ release assays (IGRA) with Resuscitation promoting factor (Rpf) proteins enhanced tuberculosis (TB) screening and diagnosis in adults but have not been evaluated in children. Children often develop paucibacillary TB and their immune response differs from that of adults, which together affect TB disease diagnostics and immunodiagnostics. We assessed the ability of Rpf to identify infection among household TB-exposed children in The Gambia and investigated their ability to discriminate Mycobacterium tuberculosis complex (MTBC) infection from active TB disease in children. METHODS: Detailed clinical investigations were done on 93 household TB-exposed Gambian children and a tuberculin skin test (TST) was administered to asymptomatic children. Venous blood was collected for overnight stimulation with ESAT-6/CFP-10-fusion protein (EC), purified protein derivative and RpfA, B, C, D and E. Interferon gamma (IFN-γ) production was measured by ELISA in supernatants and corrected for the background level. Infection status was defined by IGRA with EC and TB disease by mycobacterial confirmation and/or clinical diagnosis. We compared IFN-γ levels between infected and uninfected children and between infected and TB diseased children using a binomial logistic regression model while correcting for age and sex. A Receiver Operating Characteristics analysis was done to find the best cut-off for IFN-γ level and calculate sensitivity and specificity. RESULTS: Interferon gamma production was significantly higher in infected (IGRA+, n = 45) than in uninfected (IGRA-, n = 20) children after stimulation with RpfA, B, C, and D (P = 0.03; 0.007; 0.03 and 0.003, respectively). Using RpfB and D-specific IFN-γ cut-offs (33.9 pg/mL and 67.0 pg/mL), infection was classified with a sensitivity-specificity combination of 73-92% and 77-72% respectively, which was similar to and better than 65-75% for TST. Moreover, IFN-γ production was higher in infected than in TB diseased children (n = 28, 5 bacteriologically confirmed, 23 clinically diagnosed), following RpfB and D stimulation (P = 0.02 and 0.03, respectively). CONCLUSION: RpfB and RpfD show promising results for childhood MTBC infection screening, and both performed similar to and better than the TST in our study population. Additionally, both antigens appear to discriminate between infection and disease in children and thus warrant further investigation as screening and diagnostic antigens for childhood TB.


Asunto(s)
Antígenos Bacterianos/inmunología , Proteínas Bacterianas/inmunología , Citocinas/inmunología , Ensayos de Liberación de Interferón gamma/métodos , Tuberculosis Latente/diagnóstico , Tuberculosis Latente/epidemiología , Tamizaje Masivo/métodos , Mycobacterium tuberculosis/inmunología , Niño , Preescolar , Ensayo de Inmunoadsorción Enzimática , Composición Familiar , Femenino , Gambia/epidemiología , Humanos , Interferón gamma/inmunología , Interferón gamma/metabolismo , Tuberculosis Latente/microbiología , Masculino , Sensibilidad y Especificidad , Prueba de Tuberculina
15.
BMC Infect Dis ; 20(1): 459, 2020 Jul 01.
Artículo en Inglés | MEDLINE | ID: mdl-32611401

RESUMEN

BACKGROUND: Extra pulmonary manifestation of tuberculosis (TB) accounts for approximately one-half of TB cases in HIV-infected individuals with pleural TB as the second most common location. Even though mycobacteria are cleared, mycobacterial antigens may persist in infected tissues, causing sustained inflammation and chronicity of the disease. The aim of this study was to explore various mycobacterial antigens in pleural effusions, the impact of HIV infection and CD4+ T-cell depletion on the presence of antigens, and the diagnostic potential of antigens for improved and rapid diagnosis of pleural TB. METHODS: Pleural fluid specimens were collected from patients presenting with clinically suspected pleural TB, and processed routinely for culture, cytology, and adenosine deaminase activity analysis. HIV status and CD4+ T-cell counts were recorded. Pleural fluid mononuclear cells (PFMC) were isolated, and cell smears were stained with acid-fast staining and immunocytochemistry for various mycobacterial antigens. Real-time and nested-PCR were performed. Patients were categorized as pleural TB or non-TB cases using a composite reference standard. Performance of the mycobacterial antigens as diagnostic test was assessed. RESULTS: A total of 41 patients were enrolled, of which 32 were classified as pleural TB and 9 as non-TB. Thirteen patients had culture confirmed pleural TB, 26 (81%) were HIV-TB co-infected, and 64% had < 100 CD4+ T-cells/microL. Both secreted and cell-wall mycobacterial antigens were detected in PFMC. Lipoarabinomannan (LAM) was the most frequently detected antigen. There was no direct correlation between positive culture and antigens. Cases with low CD4+ T-cell counts had higher bacterial and antigen burden. By combining detection of secreted antigen or LAM, the sensitivity and specificity to diagnose pleural TB was 56 and 78%, respectively, as compared to 41 and 100% for culture, 53 and 89% for nested PCR, and 6 and 100% for real-time PCR. CONCLUSION: Mycobacterial antigens were detectable in PFMC from tuberculous pleural effusions, even in cases where viable mycobacteria or bacterial DNA were not always detected. Thus, a combination of secreted antigen and LAM detection by immunocytochemistry may be a complement to acid-fast staining and contribute to rapid and accurate diagnosis of pleural TB.


Asunto(s)
Infecciones Oportunistas Relacionadas con el SIDA/microbiología , Linfocitos T CD4-Positivos/inmunología , Coinfección/diagnóstico , Pruebas Diagnósticas de Rutina/métodos , Lipopolisacáridos/genética , Lipopolisacáridos/inmunología , Mycobacterium tuberculosis/inmunología , Derrame Pleural/microbiología , Tuberculosis Pleural/diagnóstico , Adulto , Anciano , Antígenos Bacterianos/genética , Antígenos Bacterianos/inmunología , Proteínas Bacterianas/genética , Proteínas Bacterianas/inmunología , Recuento de Linfocito CD4 , Coinfección/microbiología , Femenino , Humanos , Inmunohistoquímica , Masculino , Persona de Mediana Edad , Mycobacterium tuberculosis/aislamiento & purificación , Derrame Pleural/patología , Reacción en Cadena en Tiempo Real de la Polimerasa , Sensibilidad y Especificidad , Adulto Joven
16.
J Vis Exp ; (159)2020 05 19.
Artículo en Inglés | MEDLINE | ID: mdl-32510489

RESUMEN

The opsono-adherence assay is a functional assay that enumerates the attachment of bacterial pathogens to professional phagocytes. Because adherence is requisite to phagocytosis and killing, the assay is an alternative method to opsono-phagocytic killing assays. An advantage of the opsono-adherence assay is the option of using inactivated pathogens and mammalian cell lines, which allows standardization across multiple experiments. The use of an inactivated pathogen in the assay also facilitates work with biosafety level 3 infectious agents and other virulent pathogens. In our work, the opsono-adherence assay was used to assess the functional ability of antibodies, from sera of animals immunized with an anthrax capsule-based vaccine, to induce adherence of fixed Bacillus anthracis to a mouse macrophage cell line, RAW 264.7. Automated fluorescence microscopy was used to capture images of bacilli adhering to macrophages. Increased adherence was correlated with the presence of anti-capsule antibodies in the serum. Non-human primates that exhibited high serum anti-capsule antibody concentrations were protected from anthrax challenge. Thus, the opsono-adherence assay can be used to elucidate the biological functions of antigen specific antibodies in sera, to evaluate the efficacy of vaccine candidates and other therapeutics, and to serve as a possible correlate of immunity.


Asunto(s)
Vacunas contra el Carbunco/inmunología , Carbunco/inmunología , Anticuerpos Antibacterianos/inmunología , Bacillus anthracis/inmunología , Adhesión Bacteriana , Proteínas Opsoninas/inmunología , Animales , Carbunco/microbiología , Carbunco/prevención & control , Antígenos Bacterianos/inmunología , Fluoresceína-5-Isotiocianato/metabolismo , Fluorescencia , Humanos , Macrófagos/inmunología , Ratones , Primates/inmunología , Primates/microbiología , Células RAW 264.7
17.
PLoS One ; 15(6): e0234130, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32497095

RESUMEN

Better triage tests for screening tuberculosis (TB) disease are needed for people living with HIV (PLHIV). We performed the first evaluation of a previously-validated 8-antigen serological panel to screen PLHIV for pulmonary TB in Kampala, Uganda. We selected a random 1:1 sample with and without TB (defined by sputum culture) from a cohort of PLHIV initiating antiretroviral therapy. We used a multiplex microbead immunoassay and an ensemble machine learning classifier to determine the area under the receiver operating characteristic curve (AUC) for Ag85A, Ag85B, Ag85C, Rv0934-P38, Rv3881, Rv3841-BfrB, Rv3873, and Rv2878c. We then assessed the performance with the addition of four TB-specific antigens ESAT-6, CFP-10, Rv1980-MPT64, and Rv2031-HSPX, and every antigen combination. Of 262 participants (median CD4 cell-count 152 cells/µL [IQR 65-279]), 138 (53%) had culture-confirmed TB. The 8-antigen panel had an AUC of 0.53 (95% CI 0.40-0.66), and the additional 4 antigens did not improve performance (AUC 0.51, 95% CI 0.39-0.64). When sensitivity was restricted to ≥90% for the 8- and 12-antigen panel, specificity was 2.2% (95% CI 0-17.7%) and 8.1% (95% CI 0-23.9%), respectively. A three-antigen combination (Rv0934-P38, Ag85A, and Rv2031-HSPX) outperformed both panels, with an AUC of 0.60 (95% CI 0.48-0.73), 90% sensitivity (95% CI 78.2-96.7%) and 29.7% specificity (95% CI 15.9-47%). The multi-antigen panels did not achieve the target accuracy for a TB triage test among PLHIV. We identified a new combination that improved performance for TB screening in an HIV-positive sample compared to an existing serological panel in Uganda, and suggests an approach to identify novel antigen combinations specifically for screening TB in PLHIV.


Asunto(s)
Antígenos Bacterianos/inmunología , Infecciones por VIH/complicaciones , Tuberculosis/complicaciones , Tuberculosis/diagnóstico , Adulto , Fármacos Anti-VIH/uso terapéutico , Estudios de Casos y Controles , Femenino , Infecciones por VIH/tratamiento farmacológico , Humanos , Inmunoensayo , Masculino , Pruebas Serológicas , Tuberculosis/inmunología
18.
PLoS One ; 15(6): e0235139, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32574205

RESUMEN

Viral infections complicated by a bacterial infection are typically referred to as coinfections or superinfections. Streptococcus pyogenes, the group A streptococcus (GAS), is not the most common bacteria associated with influenza A virus (IAV) superinfections but did cause significant mortality during the 2009 influenza pandemic even though all isolates are susceptible to penicillin. One approach to improve the outcome of these infections is to use passive immunization targeting GAS. To test this idea, we assessed the efficacy of passive immunotherapy using antisera against either the streptococcal M protein or streptolysin O (SLO) in a murine model of IAV-GAS superinfection. Prophylactic treatment of mice with antiserum to either SLO or the M protein decreased morbidity compared to mice treated with non-immune sera; however, neither significantly decreased mortality. Therapeutic use of antisera to SLO decreased morbidity compared to mice treated with non-immune sera but neither antisera significantly reduced mortality. Overall, the results suggest that further development of antibodies targeting the M protein or SLO may be a useful adjunct in the treatment of invasive GAS diseases, including IAV-GAS superinfections, which may be particularly important during influenza pandemics.


Asunto(s)
Antígenos Bacterianos/inmunología , Proteínas de la Membrana Bacteriana Externa/inmunología , Proteínas Portadoras/inmunología , Inmunoterapia/métodos , Virus de la Influenza A/inmunología , Infecciones por Orthomyxoviridae/inmunología , Infecciones Estreptocócicas/inmunología , Streptococcus pyogenes/inmunología , Estreptolisinas/inmunología , Animales , Anticuerpos Antibacterianos/sangre , Anticuerpos Antibacterianos/inmunología , Antígenos Bacterianos/metabolismo , Proteínas de la Membrana Bacteriana Externa/antagonistas & inhibidores , Proteínas de la Membrana Bacteriana Externa/metabolismo , Proteínas Bacterianas/antagonistas & inhibidores , Proteínas Bacterianas/inmunología , Proteínas Bacterianas/metabolismo , Proteínas Portadoras/antagonistas & inhibidores , Proteínas Portadoras/metabolismo , Coinfección/microbiología , Coinfección/terapia , Coinfección/virología , Femenino , Interacciones Huésped-Patógeno/efectos de los fármacos , Interacciones Huésped-Patógeno/inmunología , Humanos , Sueros Inmunes/inmunología , Sueros Inmunes/farmacología , Virus de la Influenza A/fisiología , Ratones Endogámicos BALB C , Infecciones por Orthomyxoviridae/terapia , Infecciones por Orthomyxoviridae/virología , Conejos , Infecciones Estreptocócicas/microbiología , Infecciones Estreptocócicas/terapia , Streptococcus pyogenes/metabolismo , Streptococcus pyogenes/fisiología , Estreptolisinas/antagonistas & inhibidores , Estreptolisinas/metabolismo , Sobreinfección/microbiología , Sobreinfección/terapia , Sobreinfección/virología
19.
BMC Infect Dis ; 20(1): 444, 2020 Jun 23.
Artículo en Inglés | MEDLINE | ID: mdl-32576149

RESUMEN

BACKGROUND: The syphilis epidemic continues to cause substantial morbidity and mortality worldwide, particularly in low- and middle-income countries, despite several recent disease control initiatives. Though our understanding of the pathogenesis of this disease and the biology of the syphilis agent, Treponema pallidum subsp. pallidum has improved over the last two decades, further research is necessary to improve clinical diagnosis and disease management protocols. Additionally, such research efforts could contribute to the identification of possible targets for the development of an effective vaccine to stem syphilis spread. METHODS: This study will recruit two cohorts of participants with active syphilis infection, one with de novo infection, one with repeat infection. Whole blood specimens will be collected from each study participant at baseline, 4, 12, 24, 36, and 48 weeks, to track specific markers of their immunological response, as well as to compare humoral reactivity to Treponema pallidum antigens between the two groups. Additionally, we will use serum specimens to look for unique cytokine patterns in participants with early syphilis. Oral and blood samples, as well as samples from any syphilitic lesions present, will also be collected to sequence any Treponema pallidum DNA found. DISCUSSION: By furthering our understanding of syphilis pathogenesis and human host immune response to Treponema pallidum, we will provide important data that will help in development of new point-of-care tests that could better identify active infection, leading to improved syphilis diagnosis and management. Findings could also contribute to vaccine development efforts.


Asunto(s)
Vacunas Bacterianas/uso terapéutico , Sífilis/epidemiología , Sífilis/prevención & control , Treponema pallidum/inmunología , Vacunación , Antígenos Bacterianos/inmunología , Secuencia de Bases , Estudios de Cohortes , Citocinas/análisis , ADN Bacteriano/genética , Estudios de Seguimiento , Humanos , Tipificación Molecular , Perú/epidemiología , Sífilis/sangre , Sífilis/inmunología , Treponema pallidum/genética
20.
Nat Commun ; 11(1): 2465, 2020 05 18.
Artículo en Inglés | MEDLINE | ID: mdl-32424289

RESUMEN

Nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL) is a rare lymphoma of B-cell origin with frequent expression of functional B-cell receptors (BCRs). Here we report that expression cloning followed by antigen screening identifies DNA-directed RNA polymerase beta' (RpoC) from Moraxella catarrhalis as frequent antigen of BCRs of IgD+ LP cells. Patients show predominance of HLA-DRB1*04/07 and the IgVH genes encode extraordinarily long CDR3s. High-titer, light-chain-restricted anti-RpoC IgG1/κ-type serum-antibodies are additionally found in these patients. RpoC and MID/hag, a superantigen co-expressed by Moraxella catarrhalis that is known to activate IgD+ B cells by binding to the Fc domain of IgD, have additive activation effects on the BCR, the NF-κB pathway and the proliferation of IgD+ DEV cells expressing RpoC-specific BCRs. This suggests an additive antigenic and superantigenic stimulation of B cells with RpoC-specific IgD+ BCRs under conditions of a permissive MHC-II haplotype as a model of NLPHL lymphomagenesis, implying future treatment strategies.


Asunto(s)
Antígenos Bacterianos/inmunología , Linfocitos B/inmunología , Enfermedad de Hodgkin/inmunología , Enfermedad de Hodgkin/microbiología , Moraxella catarrhalis/inmunología , Adolescente , Adulto , Anciano , Autoantígenos/inmunología , Línea Celular Tumoral , Proliferación Celular , Niño , ARN Polimerasas Dirigidas por ADN/metabolismo , Antígenos de Histocompatibilidad Clase II/metabolismo , Enfermedad de Hodgkin/sangre , Humanos , Inmunoglobulina D/metabolismo , Fragmentos Fab de Inmunoglobulinas/inmunología , Región Variable de Inmunoglobulina/genética , Masculino , Persona de Mediana Edad , Modelos Biológicos , Receptores de Antígenos de Linfocitos B/metabolismo
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