RESUMEN
Although helminth parasites have different life cycles, their hosts share similar immune responses involving Th2 cell-type. Here, we extracted proteins from the larvae of Anisakis simplex complex and Trichinella spiralis to identify common and specific antigens (or allergens) associated with the Th2 immune response. We performed two-dimensional electrophoresis analysis and Matrix-assisted laser desorption ionization-time of flight/time of flight (MALDI-TOF/TOF) experiments. We found 13 potentially immunogenic proteins, which included 5 spots specific to T. spiralis and 8 common to T. spiralis and A. simplex, by tandem mass spectrometry. These molecules were identified structurally as actin, tropomyosin, col cuticle N domain-containing protein, and heat shock proteins. We also identified molecules related to parasite-host immune modulation and interactions. Our results may contribute to reveal potential roles of immunological proteins in parasite-derived immune modulation.
Asunto(s)
Anisakis , Proteínas del Helminto , Proteoma , Trichinella spiralis , Animales , Proteoma/inmunología , Proteínas del Helminto/inmunología , Trichinella spiralis/inmunología , Anisakis/inmunología , Antígenos Helmínticos/inmunología , Antígenos Helmínticos/análisis , Electroforesis en Gel Bidimensional , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Interacciones Huésped-Parásitos/inmunología , Larva/inmunología , Espectrometría de Masas en TándemRESUMEN
Alveolar echinococcosis (AE) is a highly lethal helminth infection. Current chemotherapeutic strategies for AE primarily involve the use of benzimidazoles (BZs) such as mebendazole (MDZ) and albendazole (ABZ), which exhibit limited efficacy. In a previous study, the vaccine of recombinant Echinococcus granulosus P29 (rEgP29) showed significant immunoprotection against E. granulosus in both mice and sheep. In the current study, we utilized hybridoma technology to generate five monoclonal antibodies (mAbs) against P29, among which 4G10F4 mAb exhibited the highest antigen-specific binding capacity. This mAb was selected for further investigation of anti-AE therapy, both in vivo and in vitro. In vitro, 4G10F4 inhibited a noteworthy inhibition of E. multilocularis protoscoleces and primary cells viability through complement-dependent cytotoxicity (CDC) mechanism. In vivo, two experiments were conducted. In the first experiment, mice were intraperitoneally injected with Em protoscoleces, and subsequently treated with 4G10F4 mAb (2.5/5/10 mg/kg) at 12 weeks postinfection once per week for 8 times via tail vein injection. Mice that were treated with 4G10F4 mAb only in dosage of 5mg/kg exhibited a significant lower mean parasite burden (0.89±0.97 g) compared to isotype mAb treated control mice (2.21±1.30 g). In the second experiment, mice were infected through hepatic portal vein and treated with 4G10F4 mAb (5mg/kg) at one week after surgery once per week for 8 times. The numbers of hepatic metacestode lesions of the 4G10F4 treatment group were significantly lower in comparison to the isotype control group. Pathological analysis revealed severe disruption of the inner structure of the metacestode in both experiments, particularly affecting the germinal and laminated layers, resulting in the transformation into infertile vesicles after treatment with 4G10F4. In addition, the safety of 4G10F4 for AE treatment was confirmed through assessment of mouse weight and evaluation of liver and kidney function. This study presents antigen-specific monoclonal antibody immunotherapy as a promising therapeutic approach against E. multilocularis induced AE.
Asunto(s)
Anticuerpos Monoclonales , Equinococosis , Animales , Equinococosis/tratamiento farmacológico , Equinococosis/inmunología , Anticuerpos Monoclonales/farmacología , Ratones , Proteínas del Helminto/inmunología , Proteínas del Helminto/farmacología , Ratones Endogámicos BALB C , Echinococcus multilocularis/inmunología , Echinococcus multilocularis/efectos de los fármacos , Femenino , Echinococcus granulosus/inmunología , Ovinos , Anticuerpos Antihelmínticos/inmunología , Antígenos Helmínticos/inmunologíaRESUMEN
Fasciolosis, caused by the liver fluke Fasciola gigantica, is a major parasitic disease that affects livestock and therefore causes significant economic losses in tropical countries. Although anthelminthic drugs can kill the parasite, drug-resistant liver fluke populations are increasing. In this study, a recombinant F. gigantica chimeric protein (rFgCHI) consisting of cathepsin L1H (FgCL1H), cathepsin B3 (FgCB3), and Saposin-like protein 1 (FgSAP1) was designed and expressed in Escherichia coli (BL21). The molecular weight of rFgCHI was 61â¯kDa. To study the antibody response, male BALB/c mice were immunized via the subcutaneous injection of rFgCHI combined with Quil A. Immunization with rFgCHI showed the induction of IgG1 and IgG2a with a higher IgG1 isotype level, indicating the potential of mixed Th1/Th2 immune responses, with Th2 predominating. However, the results showed high levels of IgG against the single proteins, except for rFgSAP1. Through Western blotting, mouse anti-rFgCHI polyclonal antibodies could be detected to the native proteins obtained from the parasite at all stages. Immunolocalization also revealed that the anti-rFgCHI antibodies could detect targeted antigens in the cecal epithelium of the parasite. These results demonstrated that rFgCHI is immunogenic to the mouse immune system and may potentially be a protein candidate for the development of a fasciolosis vaccine.
Asunto(s)
Anticuerpos Antihelmínticos , Fasciola , Proteínas del Helminto , Ratones Endogámicos BALB C , Animales , Fasciola/inmunología , Fasciola/genética , Ratones , Anticuerpos Antihelmínticos/sangre , Masculino , Proteínas del Helminto/inmunología , Proteínas del Helminto/genética , Fascioliasis/veterinaria , Fascioliasis/prevención & control , Fascioliasis/inmunología , Inmunoglobulina G/sangre , Antígenos Helmínticos/inmunología , Antígenos Helmínticos/genética , Inmunización/veterinaria , Proteínas Recombinantes de Fusión/inmunología , Proteínas Recombinantes de Fusión/genética , Formación de AnticuerposRESUMEN
Opisthorchis viverrini is the etiological agent of the disease opisthorchiasis and related cholangiocarcinoma (CCA). It infects fish-eating mammals and more than 10 million people in Southeast Asia suffered from opisthorchiasis with a high fatality rate. The only effective drug against this parasite is Praziquantel, which has significant side effects. Due to the lack of appropriate treatment options and the high death rate, there is a dire need to develop novel therapies against this pathogen. In this study, we designed a multi-epitope chimeric vaccine design against O. viverrini by using immunoinformatics approaches. Non-allergenic and immunogenic MHC-1, MHC-2, and B cell epitopes of three candidate proteins thioredoxin peroxidase (Ov-TPx-1), cathepsin F1 (Ov-CF-1) and calreticulin (Ov-CALR) of O. viverrini, were predicted to construct a potent multiepitope vaccine. The coverage of the HLA-alleles of these selected epitopes was determined globally. Four vaccine constructs made by different adjuvants and linkers were evaluated in the context of their physicochemical properties, antigenicity, and allergenicity. Protein-protein docking and MD simulation found that vaccines 3 was more stable and had a higher binding affinity for TLR2 and TLR4 immune receptors. In-silico restriction cloning of vaccine model led to the formation of plasmid constructs for expression in a suitable host. Finally, the immune simulation showed strong immunological reactions to the engineered vaccine. These findings suggest that the final vaccine construct has the potential to be validated by in vivo and in vitro experiments to confirm its efficacy against the CCA causing O. viverrini.
Asunto(s)
Antígenos Helmínticos , Neoplasias de los Conductos Biliares , Colangiocarcinoma , Opistorquiasis , Opisthorchis , Vacunas de Subunidad , Opisthorchis/inmunología , Animales , Colangiocarcinoma/inmunología , Vacunas de Subunidad/inmunología , Opistorquiasis/inmunología , Opistorquiasis/prevención & control , Humanos , Neoplasias de los Conductos Biliares/inmunología , Antígenos Helmínticos/inmunología , Antígenos Helmínticos/química , Epítopos de Linfocito B/inmunología , Desarrollo de Vacunas , Biología Computacional/métodos , Simulación del Acoplamiento Molecular , Proteínas del Helminto/inmunología , Proteínas del Helminto/química , Epítopos de Linfocito T/inmunología , Receptor Toll-Like 4/inmunología , Receptor Toll-Like 4/metabolismo , Receptor Toll-Like 2/inmunologíaRESUMEN
Spirocerca lupi is a parasitic nematode affecting predominantly domestic dogs. It causes spirocercosis, a disease that is often fatal. The assembled draft genome of S. lupi consists of 13,627 predicted protein-coding genes and is approximately 150â¯Mb in length. Several known anthelmintic gene targets such as for ß-Tubulin, glutamate, and GABA receptors as well as known vaccine gene targets such as cysteine protease inhibitor and cytokines were identified in S. lupi by comparing orthologs of C. elegans anthelmintic gene targets as well as orthologs to known vaccine candidates. New anthelmintic targets were predicted through an inclusion-exclusion strategy and new vaccine targets were predicted through an immunoinformatics approach. New anthelminthic targets include DNA-directed RNA polymerases, chitin synthase, polymerases, and other enzymes. New vaccine targets include cuticle collagens. These gene targets provide a starting platform for new drug identification and vaccine design.
Asunto(s)
Antihelmínticos , Genoma de los Helmintos , Thelazioidea , Vacunas , Animales , Antihelmínticos/farmacología , Vacunas/inmunología , Vacunas/genética , Thelazioidea/genética , Thelazioidea/inmunología , Thelazioidea/efectos de los fármacos , Perros , Infecciones por Spirurida/parasitología , Infecciones por Spirurida/prevención & control , Infecciones por Spirurida/veterinaria , Infecciones por Spirurida/inmunología , Enfermedades de los Perros/parasitología , Enfermedades de los Perros/prevención & control , Proteínas del Helminto/genética , Proteínas del Helminto/inmunologíaRESUMEN
IL-33 plays a significant role in inflammation, allergy, and host defence against parasitic helminths. The model gastrointestinal nematode Heligmosomoides polygyrus bakeri secretes the Alarmin Release Inhibitor HpARI2, an effector protein that suppresses protective immune responses and asthma in its host by inhibiting IL-33 signalling. Here we reveal the structure of HpARI2 bound to mouse IL-33. HpARI2 contains three CCP-like domains, and we show that it contacts IL-33 primarily through the second and third of these. A large loop which emerges from CCP3 directly contacts IL-33 and structural comparison shows that this overlaps with the binding site on IL-33 for its receptor, ST2, preventing formation of a signalling complex. Truncations of HpARI2 which lack the large loop from CCP3 are not able to block IL-33-mediated signalling in a cell-based assay and in an in vivo female mouse model of asthma. This shows that direct competition between HpARI2 and ST2 is responsible for suppression of IL-33-dependent responses.
Asunto(s)
Asma , Proteínas del Helminto , Proteína 1 Similar al Receptor de Interleucina-1 , Interleucina-33 , Nematospiroides dubius , Animales , Interleucina-33/metabolismo , Interleucina-33/química , Nematospiroides dubius/inmunología , Proteínas del Helminto/metabolismo , Proteínas del Helminto/química , Proteínas del Helminto/inmunología , Ratones , Femenino , Proteína 1 Similar al Receptor de Interleucina-1/metabolismo , Asma/inmunología , Asma/metabolismo , Humanos , Transducción de Señal , Infecciones por Strongylida/inmunología , Infecciones por Strongylida/parasitología , Infecciones por Strongylida/metabolismo , Unión Proteica , Modelos Animales de Enfermedad , Sitios de Unión , Ratones Endogámicos BALB C , Ratones Endogámicos C57BLRESUMEN
Subcutaneous dirofilariasis, caused by the parasitic nematode Dirofilaria repens, is a growing concern in Europe, affecting both dogs and humans. This study focused on D. repens Dr20/22, a protein encoded by an alt (abundant larval transcript) gene family. While well-documented in L3 larvae of other filariae species, this gene family had not been explored in dirofilariasis. The research involved cloning Dr20/22 cDNA, molecular characterization, and evaluating its potential application in the diagnosis of dirofilariasis. Although Real-Time analysis revealed mRNA expression in both adult worms and microfilariae, the native protein remained undetected in lysates from both developmental stages. This suggests the protein's specificity for L3 larvae and may be related to a process called SLTS (spliced leader trans-splicing), contributing to stage-specific gene expression. The specificity of the antigen for invasive larvae positions it as a promising early marker for dirofilariasis. However, ELISA tests using sera from infected and uninfected dogs indicated limited diagnostic utility. While further research is required, our findings contribute to a deeper understanding of the molecular and immunological aspects of host-parasite interactions and could offer insights into the parasite's strategies for evading the immune system.
Asunto(s)
Dirofilaria repens , Dirofilariasis , Enfermedades de los Perros , Animales , Perros , Dirofilariasis/inmunología , Dirofilariasis/parasitología , Dirofilaria repens/genética , Dirofilaria repens/inmunología , Enfermedades de los Perros/parasitología , Enfermedades de los Perros/inmunología , Anticuerpos Antihelmínticos/inmunología , Anticuerpos Antihelmínticos/sangre , Proteínas del Helminto/genética , Proteínas del Helminto/inmunología , Proteínas del Helminto/metabolismo , Antígenos Helmínticos/inmunología , Antígenos Helmínticos/genética , Larva/inmunología , Formación de Anticuerpos/inmunologíaRESUMEN
BACKGROUND: For decades, studies have demonstrated the anti-inflammatory potential of proteins secreted by helminths in allergies and asthma. Previous studies have demonstrated the immunomodulatory capabilities of Succinate Coenzyme A ligase beta-like protein (SUCLA-ß) derived from Trichinella spiralis, a crucial excretory product of this parasite. OBJECTIVE: To explore the therapeutic potential of SUCLA-ß in alleviating and controlling ovalbumin (OVA)-induced allergic asthma, as well as its influence on host immune modulation. METHODS: In this research, we utilized the rTs-SUCLA-ß protein derived from T. spiralis to investigate its potential in mitigating airway inflammation in a murine model of asthma induced by OVA sensitization/stimulation, both pre- and post-challenge. The treatment's efficacy was assessed by quantifying the extent of inflammation in the lungs. RESULTS: Treatment with rTs-SUCLA-ß demonstrated efficacy in ameliorating OVA-induced airway inflammation, as evidenced by a reduction in eosinophil infiltration, levels of OVA-specific Immunoglobulin E, interferon-γ, interleukin (IL)-9, and IL-17A, along with an elevation in IL-10. The equilibrium between Th17 and Treg cells plays a pivotal role in modulating the abundance of inflammatory cells within the organism, thereby ameliorating inflammation and alleviating symptoms associated with allergic asthma. CONCLUSIONS AND CLINICAL RELEVANCE: Our data revealed that T. spiralis-derived Ts-SUCLA-ß protein may inhibit the allergic airway inflammation by regulating host immune responses.
Asunto(s)
Asma , Proteínas del Helminto , Ovalbúmina , Trichinella spiralis , Trichinella spiralis/inmunología , Animales , Asma/inmunología , Asma/tratamiento farmacológico , Ratones , Ovalbúmina/inmunología , Proteínas del Helminto/inmunología , Proteínas del Helminto/farmacología , Ratones Endogámicos BALB C , Modelos Animales de Enfermedad , Femenino , Citocinas/metabolismo , Citocinas/inmunología , Inmunoglobulina E/inmunología , Pulmón/inmunología , Pulmón/parasitología , Pulmón/patología , Linfocitos T Reguladores/inmunología , Hipersensibilidad/inmunología , Hipersensibilidad/tratamiento farmacológico , Células Th17/inmunologíaRESUMEN
Rheumatoid arthritis (RA) is an autoimmune disease that significantly impacts quality of life by disrupting CD4+ T cell immune homeostasis. The identification of a low-side-effect drug for RA treatment is urgently needed. Our previous study suggests that Trichinella spiralis paramyosin (Ts-Pmy) has immunomodulatory effects, but its potential effect on CD4+ T cell response in RA remains unclear. In this study, we used a murine model to investigate the role of rTs-Pmy in regulating CD4+ T cell differentiation in collagen-induced arthritis (CIA). Additionally, we assessed the impact of rTs-Pmy on CD4+ T cell differentiation towards the Th1 and Th17 phenotypes, which are associated with inflammatory responses in arthritis, using in vitro assays. The results demonstrated that rTs-Pmy administration reduced arthritis severity by inhibiting Th1 and Th17 response while enhancing Treg response. Prophylactic administration of Ts-Pmy showed superior efficacy on CIA compared to therapeutic administration. Furthermore, in vitro assays demonstrated that rTs-Pmy could inhibit the differentiation of CD4+ T cells into Th1 and Th17 while inducing the production of Tregs, suggesting a potential mechanism underlying its therapeutic effects. This study suggests that Ts-Pmy may ameliorate CIA by restoring the immune balance of CD4+ T cells and provides new insights into the mechanism through which helminth-derived proteins exert their effects on autoimmune diseases.
Asunto(s)
Artritis Experimental , Linfocitos T CD4-Positivos , Diferenciación Celular , Células Th17 , Trichinella spiralis , Tropomiosina , Animales , Trichinella spiralis/inmunología , Artritis Experimental/inmunología , Artritis Experimental/patología , Artritis Experimental/tratamiento farmacológico , Ratones , Diferenciación Celular/efectos de los fármacos , Tropomiosina/inmunología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/metabolismo , Células Th17/inmunología , Células Th17/metabolismo , Células TH1/inmunología , Masculino , Proteínas del Helminto/farmacología , Proteínas del Helminto/uso terapéutico , Proteínas del Helminto/inmunología , Artritis Reumatoide/inmunología , Artritis Reumatoide/tratamiento farmacológico , Linfocitos T Reguladores/inmunología , Modelos Animales de Enfermedad , Ratones Endogámicos DBARESUMEN
Lambs harboring the Hb-AA ß-globin haplotype present improved cell-mediated responses and increased resistance against Haemonchus contortus infection. The aim of the present study was to compare the effect of sex and ß-globin haplotypes on specific humoral responses and phenotypes of resistance during H. contortus infection in Morada Nova sheep. As expected, females displayed stronger resistance during the first and second experimental challenges. Differential systemic humoral immune responses were observed comparing sex groups, in which higher levels of specific antibodies targeting 24 kDa excretory-secretory (ES24) protein of H. contortus of IgG and IgM antibodies were respectively observed as predominant isotypes in males and females. The IgM levels were significantly correlated with phenotypes of resistance, evaluated by packed cell volume and fecal egg counts. To our knowledge this is the first study reporting divergent humoral responses profiles to H. contortus infection between male and female sheep. The impact of ß-globin haplotypes was less pronounced in females compared to males. Notably, only males showed significant weight differences across haplotypes, with Hb-AA lambs being the heaviest. Additionally, Hb-AA males had significantly higher PCV (indicating better red blood cell health) and lower FEC (indicating lower parasite burden). These findings suggest a more pronounced effect of ß-globin polymorphisms on H. contortus infection in males, potentially due to their generally weaker resistance compared to females. This study highlights the importance of sex and ß-globin haplotypes in shaping immune responses to H. contortus infection. Specifically, IgM antibodies targeting the ES24 protein appear to play a crucial role in host-parasite interactions and may hold promise for therapeutic development.
Asunto(s)
Hemoncosis , Haemonchus , Inmunidad Humoral , Polimorfismo Genético , Enfermedades de las Ovejas , Globinas beta , Animales , Femenino , Masculino , Anticuerpos Antihelmínticos/inmunología , Antígenos Helmínticos/inmunología , Globinas beta/genética , Globinas beta/inmunología , Resistencia a la Enfermedad/inmunología , Resistencia a la Enfermedad/genética , Hemoncosis/veterinaria , Hemoncosis/inmunología , Hemoncosis/parasitología , Haemonchus/inmunología , Haplotipos , Proteínas del Helminto/inmunología , Proteínas del Helminto/genética , Proteínas del Helminto/metabolismo , Inmunoglobulina G/inmunología , Inmunoglobulina G/metabolismo , Inmunoglobulina M/inmunología , Inmunoglobulina M/metabolismo , Factores Sexuales , Ovinos/inmunología , Enfermedades de las Ovejas/inmunología , Enfermedades de las Ovejas/parasitología , Enfermedades de las Ovejas/genéticaRESUMEN
Neurocysticercosis is the leading cause for acquired epilepsy worldwide, and it is caused by the larval stage of the parasite Taenia solium. Several proteins of this stage have been characterized and studied to understand the parasite-host interaction, however, the proteins from the early cysticercus stages (the postoncospheral form) have not yet been characterized. The study of the postoncospheral form proteins is important to understand the host-parasite relationship in the early stages of infection. The aim of this work was to identify postoncospheral form antigenic proteins using sera from neurocysticercosis patients. T. solium activated oncospheres were cultured in HCT-8 cells to obtain the postoncospheral form. Soluble total and excretory/secretory proteins were obtained from the postoncospheral form and were incubated with both pool sera and individual serum of neurocysticercosis positive human patients. Immunoblotting showed target antigenic proteins with apparent molecular weights of 23â¯kDa and 46-48â¯kDa. The 46-48â¯kDa antigen bands present in soluble total and excretory/secretory postoncospheral form proteins were analyzed by LC-MS/MS; proteins identified were: nuclear elongation factor 1 alpha, enolase, unnamed protein product/antigen diagnostic GP50, calcium binding protein calreticulin precursor and annexin. The postoncospheral form expresses proteins related to interaction with the host, some of these proteins are predicted to be exosomal proteins. In conclusion, postoncospheral proteins are consistent targets of the humoral immune response in human and may serve as targets for diagnosis and vaccines.
Asunto(s)
Antígenos Helmínticos , Proteínas del Helminto , Neurocisticercosis , Taenia solium , Taenia solium/inmunología , Taenia solium/genética , Antígenos Helmínticos/inmunología , Animales , Humanos , Neurocisticercosis/inmunología , Neurocisticercosis/parasitología , Neurocisticercosis/diagnóstico , Proteínas del Helminto/inmunología , Proteínas del Helminto/genética , Proteínas del Helminto/química , Espectrometría de Masas en Tándem , Anticuerpos Antihelmínticos/sangre , Anticuerpos Antihelmínticos/inmunología , Cromatografía Liquida , Peso MolecularRESUMEN
Cystic echinococcosis, a zoonosis caused by cestodes belonging to the Echinococcus granulosus sensu lato (s.l.) genetic complex, affects humans and diverse livestock species. Although a veterinary vaccine exhibiting high levels of antibody-mediated protection has successfully reached the market, the large genetic diversity among parasite isolates and their particular host preferences, makes still necessary the search for novel vaccine candidates. Glutathione transferases (GSTs) constitute attractive targets for immunoprophylaxis due to their outstanding relevance in helminth detoxification processes, against both exogenous and endogenous stressors. Among the six GSTs known to be expressed in E. granulosus s.l., EgGST1 (Mu-class), EgGST2 (Sigma-class), and EgGST3 (a still non-classifiable isoenzyme), show the highest proteomic expression. Therefore, their recombinant forms -rEgGST1, rEgGST2 and rEgGST3- were herein analyzed regarding their potential to induce long-term antiparasite protection in mice. Only immunization with rEgGST1 induced long-lasting protection; and accordingly, rEgGST1-specific antibodies enhanced the parasite killing through both the classical activation of the host complement system and the antibody-dependent cellular cytotoxicity by macrophages. These results support further testing of rEgGST1 as a vaccine candidate in diverse hosts due to the broad expression of EgGST1 in different parasite stages and tissues.
Asunto(s)
Anticuerpos Antihelmínticos , Equinococosis , Echinococcus granulosus , Glutatión Transferasa , Echinococcus granulosus/inmunología , Echinococcus granulosus/genética , Echinococcus granulosus/enzimología , Animales , Equinococosis/prevención & control , Equinococosis/inmunología , Equinococosis/parasitología , Glutatión Transferasa/inmunología , Glutatión Transferasa/genética , Glutatión Transferasa/metabolismo , Ratones , Anticuerpos Antihelmínticos/inmunología , Formación de Anticuerpos/inmunología , Femenino , Ratones Endogámicos BALB C , Inmunización , Proteínas del Helminto/inmunología , Proteínas del Helminto/genéticaRESUMEN
Helminths produce calreticulin (CRT) to immunomodulate the host immune system as a survival strategy. However, the structure of helminth-derived CRT and the structural basis of the immune evasion process remains unclarified. Previous study found that the tissue-dwelling helminth Trichinella spiralis produces calreticulin (TsCRT), which binds C1q to inhibit activation of the complement classical pathway. Here, we used x-ray crystallography to resolve the structure of truncated TsCRT (TsCRTΔ), the first structure of helminth-derived CRT. TsCRTΔ was observed to share the same binding region on C1q with IgG based on the structure and molecular docking, which explains the inhibitory effect of TsCRT on C1q-IgG-initiated classical complement activation. Based on the key residues in TsCRTΔ involved in the binding activity to C1q, a 24 amino acid peptide called PTsCRT was constructed that displayed strong C1q-binding activity and inhibited C1q-IgG-initiated classical complement activation. This study is the first to elucidate the structural basis of the role of TsCRT in immune evasion, providing an approach to develop helminth-derived bifunctional peptides as vaccine target to prevent parasite infections or as a therapeutic agent to treat complement-related autoimmune diseases.
Asunto(s)
Calreticulina , Complemento C1q , Evasión Inmune , Trichinella spiralis , Trichinella spiralis/inmunología , Complemento C1q/inmunología , Complemento C1q/metabolismo , Complemento C1q/química , Animales , Calreticulina/inmunología , Calreticulina/química , Calreticulina/metabolismo , Cristalografía por Rayos X , Unión Proteica , Simulación del Acoplamiento Molecular , Proteínas del Helminto/inmunología , Proteínas del Helminto/química , Activación de Complemento/inmunología , Inmunoglobulina G/inmunología , Humanos , Antígenos Helmínticos/inmunología , Antígenos Helmínticos/química , Triquinelosis/inmunología , Triquinelosis/parasitología , Vía Clásica del Complemento/inmunología , Conformación ProteicaRESUMEN
The eggs of the blood fluke Schistosoma mansoni are the main cause of the clinical manifestations of chronic schistosomiasis. After laying, the egg "winners" attach to the endothelium of the mesenteric vein and, after a period of development, induce the growth of a small granuloma, which facilitates their passage to the intestinal lumen. Egg "losers" carried by the bloodstream to non-specific tissues also undergo full development and induce large granuloma formation, but their life ends there. Although these trapped eggs represent a dead end in the parasite life cycle, the vast majority of studies attempting to describe the biology of the S. mansoni eggs have studied these liver-trapped "losers" instead of migrating intestinal "winners". This raises the fundamental question of how these eggs differ. With robust comparative transcriptomic analysis performed on S. mansoni eggs isolated 7 weeks post infection, we show that gene expression is critically dependent on tissue localization, both in the early and late stages of development. While mitochondrial genes and venom allergen-like proteins are significantly upregulated in mature intestinal eggs, well-described egg immunomodulators IPSE/alpha-1 and omega-1, together with micro-exon genes, are predominantly expressed in liver eggs. In addition, several proteases and protease inhibitors previously implicated in egg-host interactions display clear tissue-specific gene expression patterns. These major differences in gene expression could be then reflected in the observed different ability of liver and intestinal soluble egg antigens to elicit host immune responses and in the shorter viability of miracidia hatched from liver eggs. Our comparative analysis provides a new perspective on the biology of parasite's eggs in the context of their development and tissue localization. These findings could contribute to a broader and more accurate understanding of parasite eggs interactions with the host, which have historically been often restricted to liver eggs and sometimes inaccurately generalized.
Asunto(s)
Hígado , Schistosoma mansoni , Esquistosomiasis mansoni , Animales , Schistosoma mansoni/inmunología , Schistosoma mansoni/genética , Hígado/parasitología , Hígado/inmunología , Hígado/metabolismo , Esquistosomiasis mansoni/inmunología , Esquistosomiasis mansoni/parasitología , Ratones , Óvulo/metabolismo , Óvulo/inmunología , Intestinos/parasitología , Intestinos/inmunología , Antígenos Helmínticos/inmunología , Proteínas del Helminto/genética , Proteínas del Helminto/metabolismo , Proteínas del Helminto/inmunología , Femenino , Proteínas del HuevoRESUMEN
Monogeneans are parasitic flatworms that represent a significant threat to the aquaculture industry. Species like Neobenedenia melleni (Capsalidae) and Rhabdosynochus viridisi (Diplectanidae) have been identified as causing diseases in farmed fish. In the past years, molecular research on monogeneans of the subclass Monopisthocotylea has focused on the generation of genomic and transcriptomic information and the identification in silico of some protein families of veterinary interest. Proteomic analysis has been suggested as a powerful tool to investigate proteins in parasites and identify potential targets for vaccine development and diagnosis. To date, the proteomic dataset for monogeneans has been restricted to a species of the subclass Polyopisthocotylea, while in monopisthocotyleans there is no proteomic data. In this study, we present the first proteomic data on two monopisthocotylean species, Neobenedenia sp. and R. viridisi, obtained from three distinct sample types: tissue, excretory-secretory products (ESPs), and eggs. A total of 1691 and 1846 expressed proteins were identified in Neobenedenia sp. and R. viridisi, respectively. The actin family was the largest protein family, followed by the tubulin family and the heat shock protein 70 (HSP70) family. We focused mainly on ESPs because they are important to modulate the host immune system. We identified proteins of the actin, tubulin, HSP70 and HSP90 families in both tissue and ESPs, which have been recognized for their antigenic activities in parasitic flatworms. Furthermore, our study uncovered the presence of proteins within ESPs, such as annexin, calcium-binding protein, fructose bisphosphate aldolase, glutamate dehydrogenase, myoferlin, and paramyosin, that are targets for immunodiagnostic and vaccine development and hold paramount relevance in veterinary medicine. This study expands our knowledge of monogeneans and identified proteins that, in other platyhelminths are potential targets for vaccines and drug discovery.
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Acuicultura , Enfermedades de los Peces , Proteómica , Animales , Enfermedades de los Peces/parasitología , Vacunas/inmunología , Proteínas del Helminto/genética , Proteínas del Helminto/inmunología , Proteínas del Helminto/análisis , Infecciones por Trematodos/veterinaria , Infecciones por Trematodos/parasitología , Infecciones por Trematodos/diagnóstico , Biomarcadores , Trematodos/genética , Trematodos/inmunología , Platelmintos/genética , Platelmintos/inmunologíaRESUMEN
Fatty acid binding proteins (FABPs) have emerged as attractive vaccination candidates for several platyhelminth species. To explore the physiological functions of Echinococcus multilocularis (E. multilocularis) FABP, the molecular characteristics of EmFABP1 were analyzed by online software, and the regulatory roles of rEmFABP1 protein in murine macrophages were further investigated. The emfabp1 gene encodes 133 amino acids with the characteristic ß-barrel shape of the cytoplasmic FABP family. Natural EmFABP1 protein is predominantly expressed in protoscoleces tegument and germinal layer cells and is also detected in cyst fluid and exosomes of E. multilocularis. rEmFABP1 protein demonstrated a notable suppression of phagocytic activity and nitric oxide production in murine macrophages. Additionally, the protein was observed to promote apoptosis and regulate cytokine expression in macrophages. These findings suggested that E. multilocularis FABP1 is critical in modifying macrophage physiological processes and that this protein may have immunomodulatory roles during infection.
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Echinococcus multilocularis , Proteínas de Unión a Ácidos Grasos , Proteínas del Helminto , Macrófagos , Fagocitosis , Animales , Echinococcus multilocularis/genética , Echinococcus multilocularis/inmunología , Macrófagos/inmunología , Macrófagos/parasitología , Ratones , Proteínas de Unión a Ácidos Grasos/genética , Proteínas de Unión a Ácidos Grasos/metabolismo , Proteínas del Helminto/genética , Proteínas del Helminto/metabolismo , Proteínas del Helminto/inmunología , Óxido Nítrico/metabolismo , Apoptosis , Citocinas/metabolismo , Células RAW 264.7RESUMEN
BACKGROUND: Opisthorchiasis and cholangiocarcinoma (CCA) continue to be public health concerns in many Southeast Asian countries. Although the prevalence of opisthorchiasis is declining, reported cases tend to have a light-intensity infection. Therefore, early detection by using sensitive methods is necessary. Several sensitive methods have been developed to detect opisthorchiasis. The immunological detection of antigenic proteins has been proposed as a sensitive method for examining opisthorchiasis. METHODS: The Opisthorchis viverrini antigenic proteins, including cathepsin B (OvCB), asparaginyl endopeptidase (OvAEP), and cathepsin F (OvCF), were used to construct multi-antigenic proteins. The protein sequences of OvCB, OvAEP, and OvCF, with a high probability of B cell epitopes, were selected using BepiPred 1.0 and the IEDB Analysis Resource. These protein fragments were combined to form OvCB_OvAEP_OvCF recombinant DNA, which was then used to produce a recombinant protein in Escherichia coli strain BL21(DE3). The potency of the recombinant protein as a diagnostic target for opisthorchiasis was assessed using immunoblotting and compared with that of the gold standard method, the modified formalin-ether concentration technique. RESULTS: The recombinant OvCB_OvAEP_OvCF protein showed strong reactivity with total immunoglobulin G (IgG) antibodies against light-intensity O. viverrini infections in the endemic areas. Consequently, a high sensitivity (100%) for diagnosing opisthorchiasis was reported. However, cross-reactivity with sera from other helminth and protozoan infections (including taeniasis, strongyloidiasis, giardiasis, E. coli infection, enterobiasis, and mixed infection of Echinostome spp. and Taenia spp.) and no reactivity with sera from patients with non-parasitic infections led to a reduced specificity of 78.4%. In addition, the false negative rate (FNR), false positive rate (FPR), positive predictive value (PPV), negative predictive value (NPV), and diagnostic accuracy were 0%, 21.6%, 81.4%, 100%, and 88.9%, respectively. CONCLUSIONS: The high sensitivity of the recombinant OvCB_OvAEP_OvCF protein in detecting opisthorchiasis demonstrates its potential as an opisthorchiasis screening target. Nonetheless, research on reducing cross-reactivity should be undertaken by detecting other antibodies in other sample types, such as saliva, urine, and feces.
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Antígenos Helmínticos , Opistorquiasis , Opisthorchis , Opistorquiasis/diagnóstico , Opisthorchis/inmunología , Opisthorchis/genética , Animales , Antígenos Helmínticos/genética , Antígenos Helmínticos/inmunología , Humanos , Anticuerpos Antihelmínticos/sangre , Proteínas Recombinantes/inmunología , Proteínas Recombinantes/genética , Sensibilidad y Especificidad , Proteínas del Helminto/inmunología , Proteínas del Helminto/genética , Epítopos/inmunología , Epítopos/genética , Catepsina B/genética , Catepsina B/inmunología , Escherichia coli/genética , Cisteína EndopeptidasasRESUMEN
PURPOSE: Fascioliasis is a common parasitic disease in humans and herbivores which is caused by Fasciola hepatica and Fasciola gigantica and has a worldwide distribution. Serological tests such as the enzyme-linked immunosorbent assay (ELISA) technique play a prominent role in the fast diagnosis of the disease. However, there are diagnostic limitations, including cross-reactivity with other worms, which decline the specificity of the results. This study aimed to evaluate the structure of a recombinant multi-epitope antigen produced from linear and conformational B-cell epitopes of three parasitic proteins with sera of individuals with fasciolosis, healthy controls, and those with other diseases to gain accurate sensitivity and specificity. METHODS: After designing the multi-epitope structure of cathepsin L1, FhTP16.5, and SAP-2 antigens and then synthesizing, cloning, and expressing, the extracted purified protein was evaluated by indirect ELISA to detect IgG antibodies against Fasciola hepatica parasite among the sera of 39 serum samples of Fasciola hepatica, 35 healthy individual samples, and 20 samples of other types of parasitic diseases. The synthesized multi-epitope produced from cathepsin L1, FhTP16.5, and SAP-2 antigens was evaluated using the indirect ELISA. RESULTS: The analysis of the samples mentioned for IgG antibody diagnosis against Fasciola hepatica showed 97.43% (95% confidence interval, 94.23-100%) sensitivity and 100% (95% confidence interval, 97-100%) specificity. CONCLUSION: The recombinant B-cell multi-epitope with high antigenic potency may increase the specificity of epitopic peptides and ultimately help improve and develop indirect ELISA commercial kits for the diagnosis of fascioliasis in humans.
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Anticuerpos Antihelmínticos , Antígenos Helmínticos , Ensayo de Inmunoadsorción Enzimática , Fasciola hepatica , Fascioliasis , Inmunoglobulina G , Proteínas Recombinantes , Sensibilidad y Especificidad , Pruebas Serológicas , Fascioliasis/diagnóstico , Fascioliasis/inmunología , Animales , Humanos , Antígenos Helmínticos/inmunología , Antígenos Helmínticos/genética , Ensayo de Inmunoadsorción Enzimática/métodos , Fasciola hepatica/inmunología , Fasciola hepatica/genética , Anticuerpos Antihelmínticos/sangre , Pruebas Serológicas/métodos , Proteínas Recombinantes/inmunología , Proteínas Recombinantes/genética , Inmunoglobulina G/sangre , Epítopos de Linfocito B/inmunología , Epítopos de Linfocito B/genética , Proteínas del Helminto/inmunología , Proteínas del Helminto/genética , Epítopos/inmunología , Catepsina L/inmunología , Catepsina L/genéticaRESUMEN
The mouse intestinal helminth Heligmosomoides polygyrus modulates host immune responses by secreting a transforming growth factor (TGF)-ß mimic (TGM), to expand the population of Foxp3+ Tregs. TGM comprises five complement control protein (CCP)-like domains, designated D1-D5. Though lacking homology to TGF-ß, TGM binds directly to the TGF-ß receptors TßRI and TßRII and stimulates the differentiation of naïve T-cells into Tregs. However, the molecular determinants of binding are unclear. Here, we used surface plasmon resonance, isothermal calorimetry, NMR spectroscopy, and mutagenesis to investigate how TGM binds the TGF-ß receptors. We demonstrate that binding is modular, with D1-D2 binding to TßRI and D3 binding to TßRII. D1-D2 and D3 were further shown to compete with TGF-ß(TßRII)2 and TGF-ß for binding to TßRI and TßRII, respectively. The solution structure of TGM-D3 revealed that TGM adopts a CCP-like fold but is also modified to allow the C-terminal strand to diverge, leading to an expansion of the domain and opening potential interaction surfaces. TGM-D3 also incorporates a long structurally ordered hypervariable loop, adding further potential interaction sites. Through NMR shift perturbations and binding studies of TGM-D3 and TßRII variants, TGM-D3 was shown to occupy the same site of TßRII as bound by TGF-ß using both a novel interaction surface and the hypervariable loop. These results, together with the identification of other secreted CCP-like proteins with immunomodulatory activity in H. polygyrus, suggest that TGM is part of a larger family of evolutionarily plastic parasite effector molecules that mediate novel interactions with their host.
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Proteínas del Helminto , Interacciones Huésped-Patógeno , Nematospiroides dubius , Receptores de Factores de Crecimiento Transformadores beta , Factor de Crecimiento Transformador beta , Animales , Evolución Biológica , Proteínas del Helminto/genética , Proteínas del Helminto/inmunología , Interacciones Huésped-Patógeno/inmunología , Ratones , Nematospiroides dubius/clasificación , Nematospiroides dubius/genética , Nematospiroides dubius/inmunología , Nematospiroides dubius/metabolismo , Unión Proteica , Receptores de Factores de Crecimiento Transformadores beta/metabolismo , Linfocitos T Reguladores/metabolismo , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
Thioredoxin is a low molecular weight redox-active protein of filarial parasite that plays a crucial role in downregulating the host immune response to prolong the survival of the parasite within the host body. It has the ability to cope up with the oxidative challenges posed by the host. Hence, the antioxidant protein of the filarial parasite has been suggested to be a useful target for immunotherapeutic intervention of human filariasis. In this study, we have designed a multi-epitope peptide-based vaccine using thioredoxin of Wuchereria bancrofti. Different MHC-I and MHC-II epitopes were predicted using various web servers to construct the vaccine model as MHC-I and MHC-II epitopes are crucial for the development of both humoral and cellular immune responses. Moreover, TLRs specific adjuvants were also incorporated into the vaccine candidates as TLRs are the key immunomodulator to execute innate immunity. Protein-protein molecular docking and simulation analysis between the vaccine and human TLR was performed. TLR5 is the most potent receptor to convey the vaccine-mediated inductive signal for eliciting an innate immune response. A satisfactory immunogenic report from an in-silico immune simulation experiment directed us to propose our vaccine model for experimental and clinical validation. The reverse translated vaccine sequence was also cloned in pET28a(+) to apply the concept in a wet lab experiment in near future. Taken together, this in-silico study on the design of a vaccine construct to target W. bancrofti thioredoxin is predicted to be a future hope in saving human-being from the threat of filariasis.