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Vector-borne diseases are a leading cause of death worldwide and pose a substantial unmet medical need. Pathogens binding to host extracellular proteins (the "exoproteome") represents a crucial interface in the etiology of vector-borne disease. Here, we used bacterial selection to elucidate host-microbe interactions in high throughput (BASEHIT)-a technique enabling interrogation of microbial interactions with 3,324 human exoproteins-to profile the interactomes of 82 human-pathogen samples, including 30 strains of arthropod-borne pathogens and 8 strains of related non-vector-borne pathogens. The resulting atlas revealed 1,303 putative interactions, including hundreds of pairings with potential roles in pathogenesis, including cell invasion, tissue colonization, immune evasion, and host sensing. Subsequent functional investigations uncovered that Lyme disease spirochetes recognize epidermal growth factor as an environmental cue of transcriptional regulation and that conserved interactions between intracellular pathogens and thioredoxins facilitate cell invasion. In summary, this interactome atlas provides molecular-level insights into microbial pathogenesis and reveals potential host-directed targets for next-generation therapeutics.
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Interacciones Huésped-Patógeno , Humanos , Animales , Enfermedad de Lyme/microbiología , Enfermedades Transmitidas por Vectores , Interacciones Microbiota-Huesped , Borrelia burgdorferi/patogenicidad , Borrelia burgdorferi/metabolismoRESUMEN
The immune deficiency (IMD) pathway directs host defense in arthropods upon bacterial infection. In Pancrustacea, peptidoglycan recognition proteins sense microbial moieties and initiate nuclear factor-κB-driven immune responses. Proteins that elicit the IMD pathway in non-insect arthropods remain elusive. Here, we show that an Ixodes scapularis homolog of croquemort (Crq), a CD36-like protein, promotes activation of the tick IMD pathway. Crq exhibits plasma membrane localization and binds the lipid agonist 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol. Crq regulates the IMD and jun N-terminal kinase signaling cascades and limits the acquisition of the Lyme disease spirochete B. burgdorferi. Additionally, nymphs silenced for crq display impaired feeding and delayed molting to adulthood due to a deficiency in ecdysteroid synthesis. Collectively, we establish a distinct mechanism for arthropod immunity outside of insects and crustaceans.
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Artrópodos , Infecciones Bacterianas , Borrelia burgdorferi , Ixodes , Enfermedad de Lyme , Animales , Ixodes/microbiología , Borrelia burgdorferi/genética , FN-kappa B , Enfermedad de Lyme/microbiologíaRESUMEN
Production of IFN-γ contributes to host defense against Mycobacterium tuberculosis (Mtb) infection. We previously demonstrated that Signaling lymphocytic activation molecule-associated protein (SAP) expression on cells from tuberculosis (TB) patients was inversely correlated with IFN-γ production. Here we first investigated the role of NK, T- and B-cell antigen (NTB-A)/SAP pathway in the regulation of Th1 response against Mtb. Upon antigen stimulation, NTB-A phosphorylation rapidly increases and afterwards modulates IFN-γ and IL-17 secretion. To sustain a healthy immune system, controlled expansion and contraction of lymphocytes, both during and after an adaptive immune response, is essential. Besides, restimulation-induced cell death (RICD) results in an essential homeostatic mechanism for precluding excess T-cell accumulation and associated immunopathology during the course of certain infections. Accordingly, we found that the NTB-A/SAP pathway was required for RICD during active tuberculosis. In low responder (LR) TB patients, impaired RICD was associated with diminished FASL levels, IL-2 production and CD25high expression after cell-restimulation. Interestingly, we next observed that SAP mediated the recruitment of the Src-related kinase FYNT, only in T cells from LR TB patients that were resistant to RICD. Together, we showed that the NTB-A/SAP pathway regulates T-cell activation and RICD during human TB. Moreover, the NTB-A/SAP/FYNT axis promotes polarization to an unfavorable Th2-phenotype.
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Mycobacterium tuberculosis/inmunología , Familia de Moléculas Señalizadoras de la Activación Linfocitaria/metabolismo , Células Th2/inmunología , Tuberculosis/inmunología , Adulto , Muerte Celular , Diferenciación Celular , Células Cultivadas , Femenino , Homeostasis , Humanos , Inmunidad , Terapia de Inmunosupresión , Interferón gamma/metabolismo , Interleucina-17/metabolismo , Activación de Linfocitos , Masculino , Proteínas de Neoplasias/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Transducción de SeñalRESUMEN
Although genetic manipulation is one of the hallmarks of model organisms, its applicability to non-model species has remained difficult due to our limited understanding of their fundamental biology. For instance, manipulation of a cell line originated from the black-legged tick Ixodes scapularis, an arthropod that serves as a vector for several human pathogens, has yet to be established. Here, we demonstrate the successful genetic modification of the commonly used tick ISE6 line through ectopic expression and clustered regularly interspaced palindromic repeats [(CRISPR)/CRISPR-associated protein 9 (Cas9)] genome editing. We performed ectopic expression using nucleofection and attained CRISPR-Cas9 editing via homology-dependent recombination. Targeting the E3 ubiquitin ligase x-linked inhibitor of apoptosis (xiap) and its substrate p47 led to an alteration in molecular signaling within the immune deficiency network and increased infection of the rickettsial agent Anaplasma phagocytophilum in I. scapularis ISE6 cells. Collectively, our findings complement techniques for the genetic engineering of I. scapularis ticks, which currently limit efficient and scalable molecular genetic screens in vivo.IMPORTANCEGenetic engineering in arachnids has lagged compared to insects, largely because of substantial differences in their biology. This study unveils the implementation of ectopic expression and CRISPR-Cas9 gene editing in a tick cell line. We introduced fluorescently tagged proteins in ISE6 cells and edited its genome via homology-dependent recombination. We ablated the expression of xiap and p47, two signaling molecules present in the immune deficiency (IMD) pathway of Ixodes scapularis. Impairment of the tick IMD pathway, an analogous network of the tumor necrosis factor receptor in mammals, led to enhanced infection of the rickettsial agent Anaplasma phagocytophilum. Altogether, our findings provide a critical technical resource to the scientific community to enable a deeper understanding of biological circuits in the black-legged tick I. scapularis.
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Anaplasma phagocytophilum , Borrelia burgdorferi , Ixodes , Rickettsia , Animales , Humanos , Borrelia burgdorferi/genética , Anaplasma phagocytophilum/genética , Línea Celular , MamíferosRESUMEN
Uncovering the complexity of systems in non-model organisms is critical for understanding arthropod immunology. Prior efforts have mostly focused on Dipteran insects, which only account for a subset of existing arthropod species in nature. Here we use and develop advanced techniques to describe immune cells (hemocytes) from the clinically relevant tick Ixodes scapularis at a single-cell resolution. We observe molecular alterations in hemocytes upon feeding and infection with either the Lyme disease spirochete Borrelia burgdorferi or the rickettsial agent Anaplasma phagocytophilum. We reveal hemocyte clusters exhibiting defined signatures related to immunity, metabolism, and proliferation. Depletion of phagocytic hemocytes affects hemocytin and astakine levels, two I. scapularis hemocyte markers, impacting blood-feeding, molting behavior, and bacterial acquisition. Mechanistically, astakine alters hemocyte proliferation, whereas hemocytin affects the c-Jun N-terminal kinase (JNK) signaling pathway in I. scapularis. Altogether, we discover a role for tick hemocytes in immunophysiology and provide a valuable resource for comparative biology in arthropods.
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Anaplasma phagocytophilum , Artrópodos , Borrelia burgdorferi , Ixodes , Enfermedad de Lyme , Animales , Hemocitos , Ixodes/microbiología , Borrelia burgdorferi/fisiologíaRESUMEN
Arthropod-borne pathogens are responsible for hundreds of millions of infections in humans each year. The blacklegged tick, Ixodes scapularis, is the predominant arthropod vector in the United States and is responsible for transmitting several human pathogens, including the Lyme disease spirochete Borrelia burgdorferi and the obligate intracellular rickettsial bacterium Anaplasma phagocytophilum, which causes human granulocytic anaplasmosis. However, tick metabolic response to microbes and whether metabolite allocation occurs upon infection remain unknown. Here we investigated metabolic reprogramming in the tick ectoparasite I. scapularis and determined that the rickettsial bacterium A. phagocytophilum and the spirochete B. burgdorferi induced glycolysis in tick cells. Surprisingly, the endosymbiont Rickettsia buchneri had a minimal effect on bioenergetics. An unbiased metabolomics approach following A. phagocytophilum infection of tick cells showed alterations in carbohydrate, lipid, nucleotide and protein metabolism, including elevated levels of the pleiotropic metabolite ß-aminoisobutyric acid. We manipulated the expression of genes associated with ß-aminoisobutyric acid metabolism in I. scapularis, resulting in feeding impairment, diminished survival and reduced bacterial acquisition post haematophagy. Collectively, we discovered that metabolic reprogramming affects interspecies relationships and fitness in the clinically relevant tick I. scapularis.
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Arthropod-borne microbes rely on the metabolic state of a host to cycle between evolutionarily distant species. For instance, arthropod tolerance to infection may be due to redistribution of metabolic resources, often leading to microbial transmission to mammals. Conversely, metabolic alterations aids in pathogen elimination in humans, who do not ordinarily harbor arthropod-borne microbes. To ascertain the effect of metabolism on interspecies relationships, we engineered a system to evaluate glycolysis and oxidative phosphorylation in the tick Ixodes scapularis. Using a metabolic flux assay, we determined that the rickettsial bacterium Anaplasma phagocytophilum and the Lyme disease spirochete Borrelia burgdorferi, which are transstadially transmitted in nature, induced glycolysis in ticks. On the other hand, the endosymbiont Rickettsia buchneri, which is transovarially maintained, had a minimal effect on I. scapularis bioenergetics. Importantly, the metabolite ß-aminoisobutyric acid (BAIBA) was elevated during A. phagocytophilum infection of tick cells following an unbiased metabolomics approach. Thus, we manipulated the expression of genes associated with the catabolism and anabolism of BAIBA in I. scapularis and detected impaired feeding on mammals, reduced bacterial acquisition, and decreased tick survival. Collectively, we reveal the importance of metabolism for tick-microbe relationships and unveil a valuable metabolite for I. scapularis fitness.
RESUMEN
Uncovering the complexity of systems in non-model organisms is critical for understanding arthropod immunology. Prior efforts have mostly focused on Dipteran insects, which only account for a subset of existing arthropod species in nature. Here, we describe immune cells or hemocytes from the clinically relevant tick Ixodes scapularis using bulk and single cell RNA sequencing combined with depletion via clodronate liposomes, RNA interference, Clustered Regularly Interspaced Short Palindromic Repeats activation (CRISPRa) and RNA-fluorescence in situ hybridization (FISH). We observe molecular alterations in hemocytes upon tick infestation of mammals and infection with either the Lyme disease spirochete Borrelia burgdorferi or the rickettsial agent Anaplasma phagocytophilum. We predict distinct hemocyte lineages and reveal clusters exhibiting defined signatures for immunity, metabolism, and proliferation during hematophagy. Furthermore, we perform a mechanistic characterization of two I. scapularis hemocyte markers: hemocytin and astakine. Depletion of phagocytic hemocytes affects hemocytin and astakine levels, which impacts blood feeding and molting behavior of ticks. Hemocytin specifically affects the c-Jun N-terminal kinase (JNK) signaling pathway, whereas astakine alters hemocyte proliferation in I. scapularis. Altogether, we uncover the heterogeneity and pleiotropic roles of hemocytes in ticks and provide a valuable resource for comparative biology in arthropods.
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Hematophagous ectoparasites, such as ticks, rely on impaired wound healing for skin attachment and blood feeding. Wound healing has been extensively studied through the lens of inflammatory disorders and cancer, but limited attention has been given to arthropod-borne diseases. Here, we used orthogonal approaches combining single-cell RNA sequencing (scRNAseq), flow cytometry, murine genetics, and intravital microscopy to demonstrate how tick extracellular vesicles (EVs) disrupt networks involved in tissue repair. Impairment of EVs through silencing of the SNARE protein vamp33 negatively impacted ectoparasite feeding and survival in three medically relevant tick species, including Ixodes scapularis. Furthermore, I. scapularis EVs affected epidermal γδ T cell frequencies and co-receptor expression, which are essential for keratinocyte function. ScRNAseq analysis of the skin epidermis in wildtype animals exposed to vamp33-deficient ticks revealed a unique cluster of keratinocytes with an overrepresentation of pathways connected to wound healing. This biological circuit was further implicated in arthropod fitness when tick EVs inhibited epithelial proliferation through the disruption of phosphoinositide 3-kinase activity and keratinocyte growth factor levels. Collectively, we uncovered a tick-targeted impairment of tissue repair via the resident γδ T cell-keratinocyte axis, which contributes to ectoparasite feeding.
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Alterations of myeloid cell populations have been reported in patients with tuberculosis (TB). In this work, we studied the relationship between myeloid-derived suppressor cells (MDSC) and monocytes subsets with the immunological responsiveness of TB patients. Individuals with active TB were classified as low responders (LR-TB) or high responders (HR-TB) according to their T cell responses against a cell lysate of Mycobacterium tuberculosis (Mtb-Ag). Thus, LR-TB, individuals with severe disease, display a weaker immune response to Mtb compare to HR-TB, subjects with strong immunity against the bacteria. We observed that LR-TB presented higher percentages of CD16 positive monocytes as compared to HR-TB and healthy donors. Moreover, monocyte-like (M-MDSC) and polymorphonuclear-like (PMN-MDSC) MDSC were increased in patients and the proportion of M-MDSC inversely correlated with IFN-γ levels released after Mtb-Ag stimulation in HR-TB. We also found that LR-TB displayed the highest percentages of circulating M-MDSC. These results demonstrate that CD16 positive monocytes and M-MDSC frequencies could be used as another immunological classification parameter. Interestingly, in LR-TB, frequencies of CD16 positive monocytes and M-MDSC were restored after only three weeks of anti-TB treatment. Together, our findings show a link between the immunological status of TB patients and the levels of different circulating myeloid cell populations.
Asunto(s)
Mycobacterium tuberculosis , Células Supresoras de Origen Mieloide , Tuberculosis , Humanos , Monocitos , Células MieloidesRESUMEN
Prostaglandin E2 (PGE2), an active lipid compound derived from arachidonic acid, regulates different stages of the immune response of the host during several pathologies such as chronic infections or cancer. In fact, manipulation of PGE2 levels was proposed as an approach for countering the Type I IFN signature of tuberculosis (TB). However, very limited information regarding the PGE2 pathway in patients with active TB is currently available. In the present work, we demonstrated that PGE2 exerts a potent immunosuppressive action during the immune response of the human host against Mycobacterium tuberculosis (Mtb) infection. Actually, we showed that PGE2 significantly reduced the surface expression of several immunological receptors, the lymphoproliferation and the production of proinflammatory cytokines. In addition, PGE2 promoted autophagy in monocytes and neutrophils cultured with Mtb antigens. These results suggest that PGE2 might be attenuating the excessive inflammatory immune response caused by Mtb, emerging as an attractive therapeutic target. Taken together, our findings contribute to the knowledge of the role of PGE2 in the human host resistance to Mtb and highlight the potential of this lipid mediator as a tool to improve anti-TB treatment.
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Dinoprostona/farmacología , Inmunosupresores/farmacología , Monocitos/inmunología , Mycobacterium tuberculosis/inmunología , Neutrófilos/inmunología , Tuberculosis/inmunología , Adulto , Dinoprostona/inmunología , Femenino , Humanos , Inmunosupresores/inmunología , MasculinoRESUMEN
Vector-borne illnesses comprise a significant portion of human maladies, representing 17% of global infections. Transmission of vector-borne pathogens to mammals primarily occurs by hematophagous arthropods. It is speculated that blood may provide a unique environment that aids in the replication and pathogenesis of these microbes. Lipids and their derivatives are one component enriched in blood and are essential for microbial survival. For instance, the malarial parasite Plasmodium falciparum and the Lyme disease spirochete Borrelia burgdorferi, among others, have been shown to scavenge and manipulate host lipids for structural support, metabolism, replication, immune evasion, and disease severity. In this Review, we will explore the importance of lipid hijacking for the growth and persistence of these microbes in both mammalian hosts and arthropod vectors.
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Metabolismo de los Lípidos , Enfermedades Transmitidas por Vectores/metabolismo , Animales , Borrelia/metabolismo , Culicidae/parasitología , Culicidae/virología , Humanos , Insectos/microbiología , Insectos/virología , Garrapatas/microbiología , Garrapatas/virología , Trypanosomatina/metabolismo , Enfermedades Transmitidas por Vectores/transmisiónRESUMEN
Trypanosoma rangeli is the second most common American trypanosome that infects man. It is vectored by triatomines from the genus Rhodnius, in which it invades the hemolymph and infects the salivary glands, avoiding the bug immune responses. In insects, these responses are initiated by well conserved pathways, mainly the IMD, Toll, and Jak/STAT. We hypothesize that long-term infection with T. rangeli in the gut or hemolymph of Rhodnius prolixus triggers different systemic immune responses, which influence the number of parasites that survive inside the vector. Thus, we investigated groups of insects with infections in the gut and/or hemolymph, and evaluated the parasite load and the expression in the fat body of transcription factors (Rp-Relish, Rp-Dorsal, and Rp-STAT) and inhibitors (Rp-Cactus and Rp-Caspar) of the IMD, Toll, and Jak/STAT pathways. We detected lower parasite counts in the gut of insects without hemolymph infection, compared to hemolymph-infected groups. Besides, we measured higher parasite numbers in the gut of bugs that were first inoculated with T. rangeli and then fed on infected mice, compared with control insects, indicating that hemolymph infection increases parasite numbers in the gut. Interestingly, we observed that genes from the three immune pathways where differentially modulated, depending on the region parasites were present, as we found (1) Rp-Relish downregulated in gut-and/or-hemolymph-infected insects, compared with controls; (2) Rp-Cactus upregulated in gut-infected insect, compared with controls and gut-and-hemolymph-infected groups; and (3) Rp-STAT downregulated in all groups of hemolymph-infected insects. Finally, we uncovered negative correlations between parasite loads in the gut and Rp-Relish and Rp-Cactus expression, and between parasite counts in the hemolymph and Rp-Relish levels, suggesting an association between parasite numbers and the IMD and Toll pathways. Overall, our findings reveal new players in R. prolixus-T. rangeli interactions that could be key for the capacity of the bug to transmit the pathogen.
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Rhodnius , Trypanosoma cruzi , Trypanosoma rangeli , Trypanosoma , Animales , Cuerpo Adiposo , Insectos Vectores , RatonesRESUMEN
Tuberculin skin test (TST) and IFN-γ release assays are currently used to detect Mycobacterium tuberculosis (Mtb) infection but none of them differentiate active from latent infection (LTBI). Since improved tests to diagnose Mtb infection are required, we studied the immune response to Mtb latency antigen Rv2626c in individuals exposed to the bacteria during different periods. Tuberculosis patients (TB), TB close contacts (CC: subjects exposed to Mtb for less than three months) and healthcare workers (HW: individuals exposed to Mtb at least two years) were recruited and QuantiFERON (QFT) assay, TST and IFN-γ secretion to Rv2626c were analyzed. Twenty-two percent of the individuals assessed had discordant results between QFT and TST tests. Furthermore, QFT negative and QFT positive individuals produced differential levels of IFN-γ against Rv2626c, in direct association with their exposure period to Mtb. Actually, 91% of CC QFT negative subjects secreted low levels of IFN-γ to Rv2626c, whereas 43% of HW QFT negative people produced elevated IFN-γ amounts against Rv2626c. Conversely, 69% of CC QFT positive subjects didn´t produce IFN-γ to Rv2626c. Interestingly, a similar pattern of IgG anti-Rv2626c plasma levels was observed. Therefore, determination of IFN-γ and IgG levels against the dormancy antigen Rv2626c allows to identify established LTBI.
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Anticuerpos Antibacterianos , Antígenos Bacterianos , Inmunoglobulina G , Interferón gamma , Tuberculosis Latente , Mycobacterium tuberculosis , Adolescente , Adulto , Anciano , Anticuerpos Antibacterianos/sangre , Anticuerpos Antibacterianos/inmunología , Antígenos Bacterianos/sangre , Antígenos Bacterianos/inmunología , Femenino , Humanos , Inmunoglobulina G/sangre , Inmunoglobulina G/inmunología , Interferón gamma/sangre , Interferón gamma/inmunología , Tuberculosis Latente/sangre , Tuberculosis Latente/diagnóstico , Tuberculosis Latente/inmunología , Masculino , Persona de Mediana Edad , Mycobacterium tuberculosis/inmunología , Mycobacterium tuberculosis/metabolismoRESUMEN
Th17 lymphocytes, that produce IL17A, IL17F, and IL22, play a crucial role during the immune response against Mycobacterium tuberculosis (Mtb) infection. Whereas, the contribution of IL17A in immunity to tuberculosis is usually accepted, the role of IL17F has been scarcely studied so far. The aim of this work was to evaluate the existence of a potential association of the non-synonymous variant rs763780 SNP of the IL17F gene with human tuberculosis. Accordingly, by comparing healthy donors (HD) and tuberculosis patients (TB) populations we demonstrated an association between the C allele of the SNP and the susceptibility to tuberculosis disease in Argentina. Furthermore, we found that peripheral blood mononuclear cells (PBMCs) from individuals with a more effective immune response against Mtb secreted the highest levels of IL17F when stimulated with a lysate of Mtb (Mtb-Ag). Besides, we evidenced that Mtb-Ag-stimulated PBMCs from HD carrying the C variant of the SNP displayed the lowest IFNG secretion, proliferation index, and SLAM expression as compared to TT carriers. Moreover, Mtb-Ag-stimulated PBMCs from TB carrying the C allele produced the lowest levels of IFNG, the highest level of IL17A, and the minimum proliferation indexes as compared to TT TB, suggesting a relationship between the C allele and tuberculosis severity. In fact, TB carrying the C allele presented a more severe disease, with the highest bacilli burden in sputum. Together, our findings identify the IL17F rs763780 SNP as a biomarker of tuberculosis susceptibility and advanced disease severity in Argentina, suggesting that IL17F could be a critical cytokine in tuberculosis immunity.
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Predisposición Genética a la Enfermedad/genética , Interleucina-17/genética , Polimorfismo de Nucleótido Simple/genética , Tuberculosis/genética , Adulto , Alelos , Argentina , Estudios de Casos y Controles , Femenino , Frecuencia de los Genes/genética , Genotipo , Heterocigoto , Humanos , Leucocitos Mononucleares , Masculino , Mycobacterium tuberculosis/patogenicidadRESUMEN
Interferon gamma (IFNG) plays a key role during Mycobacterium tuberculosis (Mtb) infection, and several polymorphisms located in its gene are associated with risk of tuberculosis in diverse populations. Nevertheless, the genetic resistance/susceptibility to tuberculosis in Argentina is unknown. The IFNG rs1861494 polymorphism (GâA) was reported to alter the binding of transcription factors to this region, influencing IFNG production. Using a case-control study, we found an association between the AA and AG genotypes and tuberculosis resistance (AA vs. GG: odds ratio (OR) = 0.235, p-value = 0.012; AG vs. GG: OR = 0.303, p-value = 0.044; AA vs. AG: OR = 0.776, p-value = 0.427; AA + AG vs. GG: OR = 0.270, p-value = 0.022). Moreover, Mtb-antigen stimulated peripheral blood mononuclear cells (PBMCs) from healthy donors and AA carriers secreted the highest amounts of IFNG in culture supernatants (p-value = 0.034) and presented the greatest percentage of CD4âºIFNG⺠lymphocytes (p-value = 0.035), in comparison with GG carriers. No association between the polymorphism and clinical parameters of tuberculosis severity was detected. However, our findings indicate that the rs1861494 single nucleotide polymorphism (SNP) could be considered as a biomarker of tuberculosis resistance in the Argentinean population.
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During mycobacterial infection, macroautophagy/autophagy, a process modulated by cytokines, is essential for mounting successful host responses. Autophagy collaborates with human immune responses against Mycobacterium tuberculosis (Mt) in association with specific IFNG secreted against the pathogen. However, IFNG alone is not sufficient to the complete bacterial eradication, and other cytokines might be required. Actually, induction of Th1 and Th17 immune responses are required for protection against Mt. Accordingly, we showed that IL17A and IFNG expression in lymphocytes from tuberculosis patients correlates with disease severity. Here we investigate the role of IFNG and IL17A during autophagy in monocytes infected with Mt H37Rv or the mutant MtΔRD1. Patients with active disease were classified as high responder (HR) or low responder (LR) according to their T cell responses against Mt. IL17A augmented autophagy in infected monocytes from HR patients through a mechanism that activated MAPK1/ERK2-MAPK3/ERK1 but, during infection of monocytes from LR patients, IL17A had no effect on the autophagic response. In contrast, addition of IFNG to infected monocytes, increased autophagy by activating MAPK14/p38 α both in HR and LR patients. Interestingly, proteins codified in the RD1 region did not interfere with IFNG and IL17A autophagy induction. Therefore, in severe tuberculosis patients' monocytes, IL17A was unable to augment autophagy because of a defect in the MAPK1/3 signaling pathway. In contrast, both IFNG and IL17A increased autophagy levels in patients with strong immunity to Mt, promoting mycobacterial killing. Our findings might contribute to recognize new targets for the development of novel therapeutic tools to fight the pathogen.
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Autofagia , Interleucina-17/fisiología , Monocitos/inmunología , Tuberculosis/inmunología , Células Cultivadas , Humanos , Interferón gamma/fisiología , Monocitos/microbiología , Mycobacterium tuberculosis/fisiología , Transducción de Señal , Tuberculosis/diagnóstico , Tuberculosis/microbiologíaRESUMEN
IFN-γ release assays (IGRAs) are better indicators of Mycobacterium tuberculosis infection than the tuberculin skin test (TST) in Bacillus Calmette-Guérin (BCG)-vaccinated populations. However, IGRAs do not discriminate active and latent infections (LTBI) and no gold standard for LTBI diagnosis is available. Thus, since improved tests to diagnose M. tuberculosis infection are required, we assessed the efficacy of several M. tuberculosis latency antigens. BCG-vaccinated healthy donors (HD) and tuberculosis (TB) patients were recruited. QuantiFERON-TB Gold In-Tube, TST and clinical data were used to differentiate LTBI. IFN-γ production against CFP-10, ESAT-6, Rv2624c, Rv2626c and Rv2628 antigens was tested in peripheral blood mononuclear cells. LTBI subjects secreted significantly higher IFN-γ levels against Rv2626c than HD. Additionally, Rv2626c peptide pools to which only LTBI responded were identified, and their cumulative IFN-γ response improved LTBI discrimination. Interestingly, whole blood stimulation with Rv2626c allowed the discrimination between active and latent infections, since TB patients did not secrete IFN-γ against Rv2626c, in contrast to CFP-10 + ESAT-6 stimulation that induced IFN-γ response from both LTBI and TB patients. ROC analysis confirmed that Rv2626c discriminated LTBI from HD and TB patients. Therefore, since only LTBI recognizes specific epitopes from Rv2626c, this antigen could improve LTBI diagnosis, even in BCG-vaccinated people.