RESUMO
The development of a portfolio of COVID-19 vaccines to vaccinate the global population remains an urgent public health imperative1. Here we demonstrate the capacity of a subunit vaccine, comprising the SARS-CoV-2 spike protein receptor-binding domain displayed on an I53-50 protein nanoparticle scaffold (hereafter designated RBD-NP), to stimulate robust and durable neutralizing-antibody responses and protection against SARS-CoV-2 in rhesus macaques. We evaluated five adjuvants including Essai O/W 1849101, a squalene-in-water emulsion; AS03, an α-tocopherol-containing oil-in-water emulsion; AS37, a Toll-like receptor 7 (TLR7) agonist adsorbed to alum; CpG1018-alum, a TLR9 agonist formulated in alum; and alum. RBD-NP immunization with AS03, CpG1018-alum, AS37 or alum induced substantial neutralizing-antibody and CD4 T cell responses, and conferred protection against SARS-CoV-2 infection in the pharynges, nares and bronchoalveolar lavage. The neutralizing-antibody response to live virus was maintained up to 180 days after vaccination with RBD-NP in AS03 (RBD-NP-AS03), and correlated with protection from infection. RBD-NP immunization cross-neutralized the B.1.1.7 SARS-CoV-2 variant efficiently but showed a reduced response against the B.1.351 variant. RBD-NP-AS03 produced a 4.5-fold reduction in neutralization of B.1.351 whereas the group immunized with RBD-NP-AS37 produced a 16-fold reduction in neutralization of B.1.351, suggesting differences in the breadth of the neutralizing-antibody response induced by these adjuvants. Furthermore, RBD-NP-AS03 was as immunogenic as a prefusion-stabilized spike immunogen (HexaPro) with AS03 adjuvant. These data highlight the efficacy of the adjuvanted RBD-NP vaccine in promoting protective immunity against SARS-CoV-2 and have led to phase I/II clinical trials of this vaccine (NCT04742738 and NCT04750343).
Assuntos
Adjuvantes Imunológicos , Anticorpos Neutralizantes/imunologia , Vacinas contra COVID-19/imunologia , COVID-19/imunologia , COVID-19/prevenção & controle , SARS-CoV-2/imunologia , Vacinas de Subunidades Antigênicas/imunologia , Compostos de Alúmen , Animais , Anticorpos Antivirais/imunologia , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/imunologia , COVID-19/virologia , Ensaios Clínicos Fase I como Assunto , Ensaios Clínicos Fase II como Assunto , Modelos Animais de Doenças , Imunidade Celular , Imunidade Humoral , Macaca mulatta/imunologia , Masculino , Oligodesoxirribonucleotídeos , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/imunologia , EsqualenoRESUMO
The novel coronavirus SARS-CoV-2 emerged in late 2019, rapidly reached pandemic status, and has maintained global ubiquity through the emergence of variants of concern. Efforts to develop animal models have mostly fallen short of recapitulating severe disease, diminishing their utility for research focusing on severe disease pathogenesis and life-saving medical countermeasures. We tested whether route of experimental infection substantially changes COVID-19 disease characteristics in two species of nonhuman primates (Macaca mulatta; rhesus macaques; RM, Chlorocebus atheiops; African green monkeys; AGM). Species-specific cohorts were experimentally infected with SARS-CoV-2 by either direct mucosal (intratracheal + intranasal) instillation or small particle aerosol in route-discrete subcohorts. Both species demonstrated analogous viral loads in all compartments by either exposure route although the magnitude and duration of viral loading was marginally greater in AGMs than RMs. Clinical onset was nearly immediate (+1dpi) in the mucosal exposure cohort whereas clinical signs and cytokine responses in aerosol exposure animals began +7dpi. Pathologies conserved in both species and both exposure modalities include pulmonary myeloid cell influx, development of pleuritis, and extended lack of regenerative capacity in the pulmonary compartment. Demonstration of conserved pulmonary pathology regardless of species and exposure route expands our understanding of how SARS-CoV-2 infection may lead to ARDS and/or functional lung damage and demonstrates the near clinical response of the nonhuman primate model for anti-fibrotic therapeutic evaluation studies.
Assuntos
COVID-19 , Aerossóis , Animais , Chlorocebus aethiops , Modelos Animais de Doenças , Humanos , Pulmão/patologia , Macaca mulatta , SARS-CoV-2RESUMO
The novel coronavirus SARS-CoV-2, the causative agent of COVID-19 disease, has killed over five million people worldwide as of December 2021 with infections rising again due to the emergence of highly transmissible variants. Animal models that faithfully recapitulate human disease are critical for assessing SARS-CoV-2 viral and immune dynamics, for understanding mechanisms of disease, and for testing vaccines and therapeutics. Pigtail macaques (PTM, Macaca nemestrina) demonstrate a rapid and severe disease course when infected with simian immunodeficiency virus (SIV), including the development of severe cardiovascular symptoms that are pertinent to COVID-19 manifestations in humans. We thus proposed this species may likewise exhibit severe COVID-19 disease upon infection with SARS-CoV-2. Here, we extensively studied a cohort of SARS-CoV-2-infected PTM euthanized either 6- or 21-days after respiratory viral challenge. We show that PTM demonstrate largely mild-to-moderate COVID-19 disease. Pulmonary infiltrates were dominated by T cells, including CD4+ T cells that upregulate CD8 and express cytotoxic molecules, as well as virus-targeting T cells that were predominantly CD4+. We also noted increases in inflammatory and coagulation markers in blood, pulmonary pathologic lesions, and the development of neutralizing antibodies. Together, our data demonstrate that SARS-CoV-2 infection of PTM recapitulates important features of COVID-19 and reveals new immune and viral dynamics and thus may serve as a useful animal model for studying pathogenesis and testing vaccines and therapeutics.
Assuntos
COVID-19 , Modelos Animais de Doenças , Macaca nemestrina , Doenças dos Macacos/virologia , Animais , COVID-19/imunologia , COVID-19/patologia , COVID-19/fisiopatologia , COVID-19/virologia , Humanos , Imunidade Humoral , Pulmão/imunologia , Pulmão/virologia , Masculino , Doenças dos Macacos/imunologia , Doenças dos Macacos/patologia , Doenças dos Macacos/fisiopatologia , Linfócitos T/imunologiaRESUMO
Rationale: Different Mycobacterium tuberculosis (Mtb) strains exhibit variable degrees of virulence in humans and animal models. Differing stress response strategies used by different strains of Mtb could influence virulence. Objectives: We compared the virulence of two strains of Mtb with use in animal model research: CDC1551 and Erdman. Methods: Rhesus macaques, which develop human-like tuberculosis attributes and pathology, were infected with a high dose of either strain via aerosol, and virulence was compared by bacterial burden and pathology. Measurements and Main Results: Infection with Erdman resulted in significantly shorter times to euthanasia and higher bacterial burdens and greater systemic inflammation and lung pathology relative to those infected with CDC1551. Macaques infected with Erdman also exhibited significantly higher early inflammatory myeloid cell influx to the lung, greater macrophage and T cell activity, and higher expression of lung remodeling (extracellular matrix) genes, consistent with greater pathology. Expression of NOTCH4 (neurogenic locus notch homolog 4) signaling, which is induced in response to hypoxia and promotes undifferentiated cellular state, was also higher in Erdman-infected lungs. The granulomas generated by Erdman, and not CDC1551, infection appeared to have larger regions of necrosis, which is strongly associated with hypoxia. To better understand the mechanisms of differential hypoxia induction by these strains, we subjected both to hypoxia in vitro. Erdman induced higher concentrations of DosR regulon relative to CDC1551. The DosR regulon is the global regulator of response to hypoxia in Mtb and critical for its persistence in granulomas. Conclusions: Our results show that the response to hypoxia is a critical mediator of virulence determination in Mtb, with potential impacts on bacillary persistence, reactivation, and efficiency of therapeutics.
Assuntos
Mycobacterium tuberculosis , Animais , Granuloma , Hipóxia , Inflamação/patologia , Pulmão/patologia , Macaca mulatta , Mycobacterium tuberculosis/genética , VirulênciaRESUMO
Breakthrough gastrointestinal COVID-19 was observed after experimental SARS-CoV-2 upper mucosal infection in a rhesus macaque undergoing low-dose monoclonal antibody prophylaxis. High levels of viral RNA were detected in intestinal sites contrasting with minimal viral replication in upper respiratory mucosa. Sequencing of virus recovered from tissue in 3 gastrointestinal sites and rectal swab revealed loss of furin cleavage site deletions present in the inoculating virus stock and 2 amino acid changes in spike that were detected in 2 colon sites but not elsewhere, suggesting compartmentalized replication and intestinal viral evolution. This suggests suboptimal antiviral therapies promote viral sequestration in these anatomies.
Assuntos
COVID-19 , Animais , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus , Anticorpos Monoclonais , Macaca mulattaRESUMO
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) induces a wide range of disease severity, ranging from asymptomatic infection to a life-threating illness, particularly in the elderly population and individuals with comorbid conditions. Among individuals with serious coronavirus 2019 (COVID-19) disease, acute respiratory distress syndrome (ARDS) is a common and often fatal presentation. Animal models of SARS-CoV-2 infection that manifest severe disease are needed to investigate the pathogenesis of COVID-19-induced ARDS and evaluate therapeutic strategies. We report two cases of ARDS in two aged African green monkeys (AGMs) infected with SARS-CoV-2 that had pathological lesions and disease similar to severe COVID-19 in humans. We also report a comparatively mild COVID-19 phenotype characterized by minor clinical, radiographic, and histopathologic changes in the two surviving, aged AGMs and four rhesus macaques (RMs) infected with SARS-CoV-2. Notable increases in circulating cytokines were observed in three of four infected, aged AGMs but not in infected RMs. All the AGMs had increased levels of plasma IL-6 compared with baseline, a predictive marker and presumptive therapeutic target in humans infected with SARS-CoV-2. Together, our results indicate that both RMs and AGMs are capable of modeling SARS-CoV-2 infection and suggest that aged AGMs may be useful for modeling severe disease manifestations, including ARDS.
Assuntos
COVID-19/etiologia , Pulmão/virologia , SARS-CoV-2/patogenicidade , Envelhecimento , Animais , Chlorocebus aethiops/virologia , Infecções por Coronavirus/tratamento farmacológico , Citocinas/metabolismo , Humanos , Pulmão/patologia , Macaca mulatta/virologia , Carga Viral/métodosRESUMO
Preclinical mouse models that recapitulate some characteristics of coronavirus disease (COVID-19) will facilitate focused study of pathogenesis and virus-host responses. Human agniotensin-converting enzyme 2 (hACE2) serves as an entry receptor for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to infect people via binding to envelope spike proteins. Herein we report development and characterization of a rapidly deployable COVID-19 mouse model. C57BL/6J (B6) mice expressing hACE2 in the lung were transduced by oropharyngeal delivery of the recombinant human adenovirus type 5 that expresses hACE2 (Ad5-hACE2). Mice were infected with SARS-CoV-2 at Day 4 after transduction and developed interstitial pneumonia associated with perivascular inflammation, accompanied by significantly higher viral load in lungs at Days 3, 6, and 12 after infection compared with Ad5-empty control group. SARS-CoV-2 was detected in pneumocytes in alveolar septa. Transcriptomic analysis of lungs demonstrated that the infected Ad5-hACE mice had a significant increase in IFN-dependent chemokines Cxcl9 and Cxcl10, and genes associated with effector T-cell populations including Cd3 g, Cd8a, and Gzmb. Pathway analysis showed that several Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were enriched in the data set, including cytokine-cytokine receptor interaction, the chemokine signaling pathway, the NOD-like receptor signaling pathway, the measles pathway, and the IL-17 signaling pathway. This response is correlative to clinical response in lungs of patients with COVID-19. These results demonstrate that expression of hACE2 via adenovirus delivery system sensitized the mouse to SARS-CoV-2 infection and resulted in the development of a mild COVID-19 phenotype, highlighting the immune and inflammatory host responses to SARS-CoV-2 infection. This rapidly deployable COVID-19 mouse model is useful for preclinical and pathogenesis studies of COVID-19.
Assuntos
Células Epiteliais Alveolares/imunologia , COVID-19/imunologia , Expressão Gênica , SARS-CoV-2/imunologia , Transdução de Sinais/imunologia , Adenoviridae/genética , Adenoviridae/metabolismo , Células Epiteliais Alveolares/metabolismo , Células Epiteliais Alveolares/virologia , Enzima de Conversão de Angiotensina 2/biossíntese , Enzima de Conversão de Angiotensina 2/genética , Enzima de Conversão de Angiotensina 2/imunologia , Animais , COVID-19/genética , COVID-19/metabolismo , COVID-19/patologia , Citocinas/genética , Citocinas/imunologia , Modelos Animais de Doenças , Humanos , Camundongos , Camundongos Transgênicos , SARS-CoV-2/genética , SARS-CoV-2/metabolismo , Transdução de Sinais/genética , Transdução GenéticaRESUMO
Mycobacterium tuberculosis continues to cause devastating levels of mortality due to tuberculosis (TB). The failure to control TB stems from an incomplete understanding of the highly specialized strategies that M. tuberculosis utilizes to modulate host immunity and thereby persist in host lungs. Here, we show that M. tuberculosis induced the expression of indoleamine 2,3-dioxygenase (IDO), an enzyme involved in tryptophan catabolism, in macrophages and in the lungs of animals (mice and macaque) with active disease. In a macaque model of inhalation TB, suppression of IDO activity reduced bacterial burden, pathology, and clinical signs of TB disease, leading to increased host survival. This increased protection was accompanied by increased lung T cell proliferation, induction of inducible bronchus-associated lymphoid tissue and correlates of bacterial killing, reduced checkpoint signaling, and the relocation of effector T cells to the center of the granulomata. The enhanced killing of M. tuberculosis in macrophages in vivo by CD4+ T cells was also replicated in vitro, in cocultures of macaque macrophages and CD4+ T cells. Collectively, these results suggest that there exists a potential for using IDO inhibition as an effective and clinically relevant host-directed therapy for TB.
Assuntos
Indolamina-Pirrol 2,3,-Dioxigenase/imunologia , Pulmão/imunologia , Mycobacterium tuberculosis/imunologia , Triptofano/imunologia , Tuberculoma/imunologia , Tuberculose Pulmonar/imunologia , Animais , Linfócitos T CD4-Positivos/imunologia , Proliferação de Células , Granuloma/imunologia , Granuloma/patologia , Pulmão/patologia , Macaca mulatta , Macrófagos/imunologia , Macrófagos/patologia , Mycobacterium tuberculosis/patogenicidade , Tuberculoma/patologia , Tuberculose Pulmonar/patologiaRESUMO
The synergy between Mycobacterium tuberculosis (Mtb) and HIV in coinfected patients has profoundly impacted global mortality because of tuberculosis (TB) and AIDS. HIV significantly increases rates of reactivation of latent TB infection (LTBI) to active disease, with the decline in CD4(+) T cells believed to be the major causality. In this study, nonhuman primates were coinfected with Mtb and simian immunodeficiency virus (SIV), recapitulating human coinfection. A majority of animals exhibited rapid reactivation of Mtb replication, progressing to disseminated TB and increased SIV-associated pathology. Although a severe loss of pulmonary CD4(+) T cells was observed in all coinfected macaques, a subpopulation of the animals was still able to prevent reactivation and maintain LTBI. Investigation of pulmonary immune responses and pathology in this cohort demonstrated that increased CD8(+) memory T-cell proliferation, higher granzyme B production, and expanded B-cell follicles correlated with protection from reactivation. Our findings reveal mechanisms that control SIV- and TB-associated pathology. These CD4-independent protective immune responses warrant further studies in HIV coinfected humans able to control their TB infection. Moreover, these findings will provide insight into natural immunity to Mtb and will guide development of novel vaccine strategies and immunotherapies.
Assuntos
Infecções por HIV/imunologia , Tuberculose Latente/imunologia , Mycobacterium tuberculosis/patogenicidade , Vírus da Imunodeficiência Símia/patogenicidade , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/microbiologia , Linfócitos T CD4-Positivos/patologia , Linfócitos T CD4-Positivos/virologia , Proliferação de Células/genética , Coinfecção/virologia , HIV/imunologia , HIV/patogenicidade , Infecções por HIV/fisiopatologia , Infecções por HIV/virologia , Humanos , Memória Imunológica/genética , Tuberculose Latente/microbiologia , Tuberculose Latente/patologia , Tuberculose Latente/virologia , Ativação Linfocitária/imunologia , Macaca mulatta/imunologia , Macaca mulatta/microbiologia , Macaca mulatta/virologia , Mycobacterium tuberculosis/imunologia , Vírus da Imunodeficiência Símia/imunologiaRESUMO
Although it is accepted that the environment within the granuloma profoundly affects Mycobacterium tuberculosis (Mtb) and infection outcome, our ability to understand Mtb gene expression in these niches has been limited. We determined intragranulomatous gene expression in human-like lung lesions derived from nonhuman primates with both active tuberculosis (ATB) and latent TB infection (LTBI). We employed a non-laser-based approach to microdissect individual lung lesions and interrogate the global transcriptome of Mtb within granulomas. Mtb genes expressed in classical granulomas with central, caseous necrosis, as well as within the caseum itself, were identified and compared with other Mtb lesions in animals with ATB (n = 7) or LTBI (n = 7). Results were validated using both an oligonucleotide approach and RT-PCR on macaque samples and by using human TB samples. We detected approximately 2,900 and 1,850 statistically significant genes in ATB and LTBI lesions, respectively (linear models for microarray analysis, Bonferroni corrected, P < 0.05). Of these genes, the expression of approximately 1,300 (ATB) and 900 (LTBI) was positively induced. We identified the induction of key regulons and compared our results to genes previously determined to be required for Mtb growth. Our results indicate pathways that Mtb uses to ensure its survival in a highly stressful environment in vivo. A large number of genes is commonly expressed in granulomas with ATB and LTBI. In addition, the enhanced expression of the dormancy survival regulon was a key feature of lesions in animals with LTBI, stressing its importance in the persistence of Mtb during the chronic phase of infection.
Assuntos
Regulação Bacteriana da Expressão Gênica , Genes Bacterianos , Granuloma/microbiologia , Viabilidade Microbiana/genética , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/fisiologia , Anaerobiose , Animais , Perfilação da Expressão Gênica , Granuloma/patologia , Pulmão/microbiologia , Pulmão/patologia , Macaca , Reação em Cadeia da Polimerase em Tempo Real , Regulon/genética , Reprodutibilidade dos Testes , Transcriptoma/genética , Tuberculose/genética , Tuberculose/microbiologia , Tuberculose/patologiaRESUMO
Mycobacterium tuberculosis (Mtb) is the leading cause of death from an infectious disease worldwide and is the causative agent of tuberculosis (Chao, M. C., and Rubin, E. J. (2010) Annu. Rev. Microbiol. 64, 293-311). Throughout infection, Mtb encounters a variety of host pressures. Thus, responding to these host stresses via the induction of multiple regulatory networks is needed for survival within the host. The Clp protease gene regulator, Rv2745c (clgR), is induced in response to environmental stress conditions, implicating its potential role in Mtb pathogenesis. Transcriptional activation of genes downstream of Rv2745c occurs in a condition-dependent manner. Our isogenic Mtb:ΔRv2745c mutant expresses a significantly different phenotype upon reaeration conditions. Transcriptional analysis revealed differential gene expression profiles relative to wild-type Mtb. Rv2745c is strongly induced in response to hypoxic and reaeration conditions, implicating a role of Rv2745c in vivo during both establishment of infection and reactivation. We found dysregulation of downstream genes within both the σ(H)/σ(E) regulon as well as the dosR regulon in the isogenic mutant, Mtb:ΔRv2745c. Upon hypoxic and reaeration conditions, Clp protease induction occurred within wild-type Mtb, indicating that activation of clgR, which subsequently leads to Clp protease induction, is crucial for degradation of misfolded proteins and ultimately survival of Mtb upon specific stress conditions. Our data indicate the diverse response of Rv2745c, σ(H) and σ(E) in response to a variety of stress conditions. Activation of Rv2745c in response to various stress conditions leads to differential activation of downstream genes, indicating the diverse role of Rv2745c and its importance for Mtb survival in vivo.
Assuntos
Proteínas de Bactérias/metabolismo , Regulação Bacteriana da Expressão Gênica , Genes Reguladores , Mycobacterium tuberculosis/metabolismo , Oxigênio/metabolismo , Fatores de Transcrição/metabolismo , Ácidos Graxos/metabolismo , Genes Bacterianos , Pulmão/microbiologia , Mutação , Análise de Sequência com Séries de Oligonucleotídeos , Oxirredução , Processamento Pós-Transcricional do RNA , RNA Bacteriano/metabolismo , Fator sigma/metabolismo , Estresse Fisiológico , Transcrição GênicaRESUMO
RATIONALE: Hypoxia promotes dormancy by causing physiologic changes to actively replicating Mycobacterium tuberculosis. DosR controls the response of M. tuberculosis to hypoxia. OBJECTIVES: To understand DosR's contribution in the persistence of M. tuberculosis, we compared the phenotype of various DosR regulon mutants and a complemented strain to M. tuberculosis in macaques, which faithfully model M. tuberculosis infection. METHODS: We measured clinical and microbiologic correlates of infection with M. tuberculosis relative to mutant/complemented strains in the DosR regulon, studied lung pathology and hypoxia, and compared immune responses in lung using transcriptomics and flow cytometry. MEASUREMENTS AND MAIN RESULTS: Despite being able to replicate initially, mutants in DosR regulon failed to persist or cause disease. On the contrary, M. tuberculosis and a complemented strain were able to establish infection and tuberculosis. The attenuation of pathogenesis in animals infected with the mutants coincided with the appearance of a Th1 response and organization of hypoxic lesions wherein M. tuberculosis expressed dosR. The lungs of animals infected with the mutants (but not the complemented strain) exhibited early transcriptional signatures of T-cell recruitment, activation, and proliferation associated with an increase of T cells expressing homing and proliferation markers. CONCLUSIONS: Delayed adaptive responses, a hallmark of M. tuberculosis infection, not only lead to persistence but also interfere with the development of effective antituberculosis vaccines. The DosR regulon therefore modulates both the magnitude and the timing of adaptive immune responses in response to hypoxia in vivo, resulting in persistent infection. Hence, DosR regulates key aspects of the M. tuberculosis life cycle and limits lung pathology.
Assuntos
Proteínas de Bactérias/genética , Hipóxia/metabolismo , Mycobacterium tuberculosis/genética , Proteínas Quinases/genética , Regulon/genética , Tuberculose/genética , Animais , Proteínas de Bactérias/imunologia , Proteínas de Ligação a DNA , Modelos Animais de Doenças , Macaca mulatta , Mycobacterium tuberculosis/imunologia , Proteínas Quinases/imunologia , Regulon/imunologia , Linfócitos T/imunologia , Tuberculose/imunologia , Tuberculose/prevenção & controleRESUMO
Mycobacterium tuberculosis (Mtb) must counter hypoxia within granulomas to persist. DosR, in concert with sensor kinases DosS and DosT, regulates the response to hypoxia. Yet Mtb lacking functional DosR colonize the lungs of C57Bl/6 mice, presumably owing to the lack of organized lesions with sufficient hypoxia in that model. We compared the phenotype of the Δ-dosR, Δ-dosS, and Δ-dosT mutants to Mtb using C3HeB/FeJ mice, an alternate mouse model where lesions develop hypoxia. C3HeB/FeJ mice were infected via aerosol. The progression of infection was analyzed by tissue bacterial burden and histopathology. A measure of the comparative global immune responses was also analyzed. Although Δ-dosR and Δ-dosT grew comparably to wild-type Mtb, Δ-dosS exhibited a significant defect in bacterial burden and pathology in vivo, accompanied by ablated proinflammatory response. Δ-dosS retained the ability to induce DosR. The Δ-dosS mutant was also attenuated in murine macrophages ex vivo, with evidence of reduced expression of the proinflammatory signature. Our results show that DosS, but not DosR and DosT, is required by Mtb to survive in C3HeB/FeJ mice. The attenuation of Δ-dosS is not due to its inability to induce the DosR regulon, nor is it a result of the accumulation of hypoxia. That the in vivo growth restriction of Δ-dosS could be mimicked ex vivo suggested sensitivity to macrophage oxidative burst. Anoxic caseous centers within tuberculosis lesions eventually progress to cavities. Our results provide greater insight into the molecular mechanisms of Mtb persistence within host lungs.
Assuntos
Proteínas de Bactérias/genética , Granuloma do Sistema Respiratório/microbiologia , Mycobacterium tuberculosis/patogenicidade , Protamina Quinase/genética , Tuberculose Pulmonar/microbiologia , Animais , Proteínas de Bactérias/metabolismo , Hipóxia Celular , Células Cultivadas , Regulação Bacteriana da Expressão Gênica , Macrófagos/microbiologia , Masculino , Camundongos Endogâmicos C3H , Viabilidade Microbiana , Mycobacterium tuberculosis/genética , Protamina Quinase/metabolismo , Regulon , VirulênciaRESUMO
Hyperglycemia, and exacerbation of pre-existing deficits in glucose metabolism, are manifestations of the post-acute sequelae of SARS-CoV-2. Our understanding of metabolic decline after acute COVID-19 remains unclear due to the lack of animal models. Here, we report a non-human primate model of metabolic post-acute sequelae of SARS-CoV-2 using SARS-CoV-2 infected African green monkeys. Using this model, we identify a dysregulated blood chemokine signature during acute COVID-19 that correlates with elevated and persistent hyperglycemia four months post-infection. Hyperglycemia also correlates with liver glycogen levels, but there is no evidence of substantial long-term SARS-CoV-2 replication in the liver and pancreas. Finally, we report a favorable glycemic effect of the SARS-CoV-2 mRNA vaccine, administered on day 4 post-infection. Together, these data suggest that the African green monkey model exhibits important similarities to humans and can be utilized to assess therapeutic candidates to combat COVID-related metabolic defects.
Assuntos
COVID-19 , Modelos Animais de Doenças , Hiperglicemia , Fígado , SARS-CoV-2 , Animais , Hiperglicemia/imunologia , COVID-19/imunologia , COVID-19/virologia , COVID-19/sangue , Chlorocebus aethiops , SARS-CoV-2/imunologia , Fígado/virologia , Fígado/metabolismo , Fígado/imunologia , Glicogênio/metabolismo , Glicemia/metabolismo , Humanos , Masculino , Pâncreas/virologia , Pâncreas/imunologia , Pâncreas/patologia , Pâncreas/metabolismo , Quimiocinas/metabolismo , Quimiocinas/sangue , Feminino , Replicação ViralRESUMO
The effects of immunodeficiency associated with chronic HIV infection on COVID-19 disease and viral persistence have not been directly addressed in a controlled setting. In this pilot study, we exposed two pigtail macaques (PTMs) chronically infected with SIVmac239, exhibiting from very low to no CD4 T cells across all compartments, to SARS-CoV-2. We monitored the disease progression, viral replication, and evolution, and compared these outcomes with SIV-naïve PTMs infected with SARS-CoV-2. No overt signs of COVID-19 disease were observed in either animal, and the SARS-CoV-2 viral kinetics and evolution in the SIVmac239 PTMs were indistinguishable from those in the SIV-naïve PTMs in all sampled mucosal sites. However, the single-cell RNA sequencing of bronchoalveolar lavage cells revealed an infiltration of functionally inert monocytes after SARS-CoV-2 infection. Critically, neither of the SIV-infected PTMs mounted detectable anti-SARS-CoV-2 T-cell responses nor anti-SARS-CoV-2 binding or neutralizing antibodies. Thus, HIV-induced immunodeficiency alone may not be sufficient to drive the emergence of novel viral variants but may remove the ability of infected individuals to mount adaptive immune responses against SARS-CoV-2.
Assuntos
COVID-19 , Coinfecção , Modelos Animais de Doenças , SARS-CoV-2 , Síndrome de Imunodeficiência Adquirida dos Símios , Vírus da Imunodeficiência Símia , Animais , Vírus da Imunodeficiência Símia/imunologia , COVID-19/imunologia , COVID-19/virologia , Síndrome de Imunodeficiência Adquirida dos Símios/imunologia , Síndrome de Imunodeficiência Adquirida dos Símios/virologia , SARS-CoV-2/imunologia , Coinfecção/imunologia , Coinfecção/virologia , Replicação Viral , Macaca nemestrina , Projetos Piloto , Anticorpos Antivirais/imunologia , Anticorpos Antivirais/sangue , Carga Viral , Linfócitos T CD4-Positivos/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/sangueRESUMO
The continued evolution of SARS-CoV-2 variants capable of subverting vaccine and infection-induced immunity suggests the advantage of a broadly protective vaccine against betacoronaviruses (ß-CoVs). Recent studies have isolated monoclonal antibodies (mAbs) from SARS-CoV-2 recovered-vaccinated donors capable of neutralizing many variants of SARS-CoV-2 and other ß-CoVs. Many of these mAbs target the conserved S2 stem region of the SARS-CoV-2 spike protein, rather the receptor binding domain contained within S1 primarily targeted by current SARS-CoV-2 vaccines. One of these S2-directed mAbs, CC40.8, has demonstrated protective efficacy in small animal models against SARS-CoV-2 challenge. As the next step in the pre-clinical testing of S2-directed antibodies as a strategy to protect from SARS-CoV-2 infection, we evaluated the in vivo efficacy of CC40.8 in a clinically relevant non-human primate model by conducting passive antibody transfer to rhesus macaques (RM) followed by SARS-CoV-2 challenge. CC40.8 mAb was intravenously infused at 10mg/kg, 1mg/kg, or 0.1 mg/kg into groups (n=6) of RM, alongside one group that received a control antibody (PGT121). Viral loads in the lower airway were significantly reduced in animals receiving higher doses of CC40.8. We observed a significant reduction in inflammatory cytokines and macrophages within the lower airway of animals infused with 10mg/kg and 1mg/kg doses of CC40.8. Viral genome sequencing demonstrated a lack of escape mutations in the CC40.8 epitope. Collectively, these data demonstrate the protective efficiency of broadly neutralizing S2-targeting antibodies against SARS-CoV-2 infection within the lower airway while providing critical preclinical work necessary for the development of pan-ß-CoV vaccines.
RESUMO
BACKGROUND: Sigma H (sigH) is a major Mycobacterium tuberculosis (Mtb) stress response factor. It is induced in response to heat, oxidative stress, cell wall damage, and hypoxia. Infection of macrophages with the Δ-sigH mutant generates more potent innate immune response than does infection with Mtb. The mutant is attenuated for pathology in mice. METHODS: We used a nonhuman primate (NHP) model of acute tuberculosis, to better understand the phenotype of the Δ-sigH mutant in vivo. NHPs were infected with high doses of Mtb or the mutant, and the progression of tuberculosis was analyzed in both groups using clinical, pathological, microbiological, and immunological parameters. RESULTS: Animals exposed to Mtb rapidly progressed to acute pulmonary tuberculosis as indicated by worsening clinical correlates, high lung bacterial burden, and granulomatous immunopathology. All the animals rapidly succumbed to tuberculosis. On the other hand, the NHPs exposed to the Mtb:Δ-sigH mutant did not exhibit acute tuberculosis, instead showing significantly blunted disease. These NHPs survived the entire duration of the study. CONCLUSIONS: The Mtb:Δ-sigH mutant is completely attenuated for bacterial burden as well as immunopathology in NHPs. SigH and its regulon are required for complete virulence in primates. Further studies are needed to identify the molecular mechanism of this attenuation.
Assuntos
Proteínas de Bactérias/metabolismo , Regulação Bacteriana da Expressão Gênica/fisiologia , Pulmão/imunologia , Pulmão/microbiologia , Mycobacterium tuberculosis/metabolismo , Fator sigma/metabolismo , Tuberculose Pulmonar/microbiologia , Animais , Proteínas de Bactérias/genética , Perfilação da Expressão Gênica , Granuloma , Imuno-Histoquímica , Macaca mulatta , Mycobacterium tuberculosis/genética , Fator sigma/genética , Tuberculose Pulmonar/patologiaRESUMO
Persistent and uncontrolled SARS-CoV-2 replication in immunocompromised individuals has been observed and may be a contributing source of novel viral variants that continue to drive the pandemic. Importantly, the effects of immunodeficiency associated with chronic HIV infection on COVID-19 disease and viral persistence have not been directly addressed in a controlled setting. Here we conducted a pilot study wherein two pigtail macaques (PTM) chronically infected with SIVmac239 were exposed to SARS-CoV-2 and monitored for six weeks for clinical disease, viral replication, and viral evolution, and compared to our previously published cohort of SIV-naïve PTM infected with SARS-CoV-2. At the time of SARS-CoV-2 infection, one PTM had minimal to no detectable CD4+ T cells in gut, blood, or bronchoalveolar lavage (BAL), while the other PTM harbored a small population of CD4+ T cells in all compartments. Clinical signs were not observed in either PTM; however, the more immunocompromised PTM exhibited a progressive increase in pulmonary infiltrating monocytes throughout SARS-CoV-2 infection. Single-cell RNA sequencing (scRNAseq) of the infiltrating monocytes revealed a less activated/inert phenotype. Neither SIV-infected PTM mounted detectable anti-SARS-CoV-2 T cell responses in blood or BAL, nor anti-SARS-CoV-2 neutralizing antibodies. Interestingly, despite the diminished cellular and humoral immune responses, SARS-CoV-2 viral kinetics and evolution were indistinguishable from SIV-naïve PTM in all sampled mucosal sites (nasal, oral, and rectal), with clearance of virus by 3-4 weeks post infection. SIV-induced immunodeficiency significantly impacted immune responses to SARS-CoV-2 but did not alter disease progression, viral kinetics or evolution in the PTM model. SIV-induced immunodeficiency alone may not be sufficient to drive the emergence of novel viral variants.
RESUMO
Omicron SARS-CoV-2 variants escape vaccine-induced neutralizing antibodies and cause nearly all current COVID-19 cases. Here, we compared the efficacy of three booster vaccines against Omicron BA.5 challenge in rhesus macaques: mRNA-1273, the Novavax ancestral spike protein vaccine (NVX-CoV2373), or Omicron BA.1 spike protein version (NVX-CoV2515). All three booster vaccines induced a strong BA.1 cross-reactive binding antibody and changed immunoglobulin G (Ig) dominance from IgG1 to IgG4 in the serum. All three booster vaccines also induced strong and comparable neutralizing antibody responses against multiple variants of concern, including BA.5 and BQ.1.1, along with long-lived plasma cells in the bone marrow. The ratio of BA.1 to WA-1 spike-specific antibody-secreting cells in the blood was higher in NVX-CoV2515 animals compared with NVX-CoV2373 animals, suggesting a better recall of BA.1-specific memory B cells by the BA.1 spike-specific vaccine compared with the ancestral spike-specific vaccine. Further, all three booster vaccines induced low levels of spike-specific CD4 but not CD8 T cell responses in the blood. After challenge with SARS-CoV-2 BA.5 variant, all three vaccines showed strong protection in the lungs and controlled virus replication in the nasopharynx. In addition, both Novavax vaccines blunted viral replication in nasopharynx at day 2. The protection against SARS-CoV-2 BA.5 infection in the upper respiratory airways correlated with binding, neutralizing, and ADNP activities of the serum antibody. These data have important implications for COVID-19 vaccine development, because vaccines that lower nasopharyngeal virus may help to reduce transmission.
Assuntos
Vacina de mRNA-1273 contra 2019-nCoV , COVID-19 , Animais , Humanos , Vacinas contra COVID-19 , COVID-19/prevenção & controle , Macaca mulatta , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus/genética , Imunoglobulina GRESUMO
Tuberculosis (TB) is a major infectious disease problem: 1.7 million people annually die due to TB. Emergence of drug-resistant Mycobacterium tuberculosis and the lack of new antibiotics have exacerbated the situation. There is an urgent need to develop or repurpose drugs against TB. We evaluated inhaled gentamicin as direct respiratory system-targeted therapy in a murine model of TB. Aerosolized-gentamicin-treated mice showed significantly reduced lung M. tuberculosis loads and fewer granulomas relative to untreated controls. These results suggest that direct delivery of antibiotics to the respiratory system may provide therapeutic benefit to conventional treatment regimes for treatment of pulmonary TB.