Background: The fungal cell-wall constituent 1,3-beta-D-glucan (BDG) is a pathogen-associated molecular pattern that can stimulate innate immunity. We hypothesized that BDG from colonizing fungi in critically-ill patients may translocate into the systemic circulation and be associated with host inflammation and outcomes. Methods: We enrolled 453 mechanically-ventilated patients with acute respiratory failure (ARF) without invasive fungal infection and measured BDG, innate immunity and epithelial permeability biomarkers in serially collected plasma samples. Results: Compared to healthy controls, ARF patients had significantly higher BDG levels (median [interquartile-range] 26 [15-49]pg/ml, p<0.001), whereas ARF patients with high BDG levels (≥40pg/ml, 31%) had higher odds for assignment to the prognostically adverse hyperinflammatory subphenotype (odds ratio [confidence interval] 2.88 [1.83-4.54], p<0.001). Baseline BDG levels were predictive of fewer ventilator-free days and worse 30-day survival (adjusted p<0.05). Integrative analyses of fungal colonization and epithelial barrier disruption suggested that BDG may translocate from either the lung or gut compartment. We validated the associations between plasma BDG and host inflammatory responses in 97 hospitalized patients with COVID-19. Conclusions: BDG measurements offered prognostic information in critically-ill patients without fungal infections. Further research in the mechanisms of translocation and innate immunity recognition and stimulation may offer new therapeutic opportunities in critical illness.
Georgios D. Kitsios, Daniel Kotok, Haopu Yang, Malcolm A. Finkelman, Yonglong Zhang, Noel Britton, Xiaoyun Li, Marina S. Levochkina, Amy K. Wagner, Caitlin Schaefer, John J. Villandre, Rui Guo, John W. Evankovich, William Bain, Faraaz Shah, Yingze Zhang, Barbara A. Methé, Panayiotis V. Benos, Bryan J. McVerry, Alison Morris
IFN-γ-driven responses to malaria have been shown to modulate the development and function of T follicular helper (TFH) cells and memory B cells (MBCs), with conflicting evidence in their involvement in the induction of antibody responses required to achieve clinical immunity and their association with disease outcomes. Using high-dimensional single cell mass cytometry, we identified distinct populations of TH1-polarized CD4+ T cells and MBCs expressing the TH1-defining transcription factor T-bet, associated with either increased or reduced risk of Plasmodium vivax malaria, demonstrating that inflammatory responses to malaria are not universally detrimental for infection. Furthermore, we found that whereas class-switched but not IgM+ MBCs were associated with reduced risk of symptomatic malaria, populations of TH1 cells with a stem central memory phenotype, TH17 cells and T regulatory cells were associated with protection from asymptomatic infection, suggesting that activation of cell mediated immunity might be also required to control persistent P. vivax infection of low parasite burden.
Lisa J. Ioannidis, Halina M. Pietrzak, Ann Ly, Retno Ayu Setya Utami, Emily M. Eriksson, Stephanie I. Studniberg, Waruni Abeysekera, Connie S.N. Li-Wai-Suen, Dylan Sheerin, Julie Healer, Agatha Mia Puspitasari, Dwi Apriyanti, Farah N. Coutrier, Jeanne Rini Poespoprodjo, Enny Kenangalem, Benediktus Andries, Pak Prayoga, Novita Sariyanti, Gordon K. Smyth, Leily Trianty, Alan F. Cowman, Ric N. Price, Rintis Noviyanti, Diana S. Hansen
BACKGROUND. The role of humoral immunity in the coronavirus disease 2019 (COVID-19) is not fully understood owing, in large part, to the complexity of antibodies produced in response to the SARS-CoV-2 infection. There is a pressing need for serology tests to assess patient-specific antibody response and predict clinical outcome. METHODS. Using SARS-CoV-2 proteome and peptide microarrays, we screened 146 COVID-19 patients plasma samples to identify antigens and epitopes. This enabled us to develop a master epitope array and an epitope-specific agglutination assay to gauge antibody responses systematically and with high resolution. RESULTS. We identified linear epitopes from the Spike (S) and Nucleocapsid (N) protein and showed that the epitopes enabled higher resolution antibody profiling than the S or N protein antigen. Specifically, we found that antibody responses to the S(811-825), S(881-895) and N(156-170) epitopes negatively or positively correlated with clinical severity or patient survival. Moreover, we found that the P681H and S235F mutations associated with the coronavirus variant of concern B.1.1.7 altered the specificity of the corresponding epitopes. CONCLUSIONS. Epitope-resolved antibody testing not only affords a high-resolution alternative to conventional immunoassays to delineate the complex humoral immunity to SARS-CoV-2 and differentiate between neutralizing and non-neutralizing antibodies, it may potentially be used to predict clinical outcome. The epitope peptides can be readily modified to detect antibodies against variants of concern (VOC) in both the peptide array and latex agglutination formats. FUNDING. Ontario Research Fund (ORF)-COVID-19 Rapid Research Fund, the Toronto COVID-19 Action Fund, Western University, the Lawson Health Research Institute, the London Health Sciences Foundation, and the AMOSO Innovation Fund.
Courtney Voss, Sally Esmail, Xuguang Liu, Michael J. Knauer, Suzanne Ackloo, Tomonori Kaneko, Lori E. Lowes, Peter J. Stogios, Almagul Seitova, Ashley Hutchinson, Farhad Yusifov, Tatiana Skarina, Elena Evdokimova, Peter Loppnau, Pegah Ghiabi, Taraneh Hajian, Shanshan Zhong, Husam Abdoh, Benjamin D. Hedley, Vipin Bhayana, Claudio M. Martin, Marat Slessarev, Benjamin Chin-Yee, Douglas D. Fraser, Ian Chin-Yee, Shawn S.C. Li
The skin lesion erythema migrans (EM) is an initial sign of the Ixodes-tick transmitted Borreliella spirochetal infection known as Lyme disease. T cells and innate immune cells have previously been shown to predominate the EM lesion and promote the reaction. Despite the established importance of B cells and antibodies in preventing infection, the role of B cells in the skin immune response to Borreliella is unknown. Here, we used single-cell RNA-Seq in conjunction with B cell receptor (BCR) sequencing to immunophenotype EM lesions and their associated B cells and BCR repertoires. We found that B cells were more abundant in EM in comparison to autologous uninvolved skin; many were clonally expanded and had circulating relatives. EM-associated B cells upregulated expression of MHC class II genes and exhibited preferential IgM isotype usage. A subset also exhibited low levels of somatic hypermutation despite a gene expression profile consistent with memory B cells. Our study demonstrates that single-cell gene expression with paired BCR sequencing can be used to interrogate the sparse B cell populations in human skin and reveals that B cells in the skin infection site in early Lyme disease express a phenotype consistent with local antigen presentation and antibody production.
Ruoyi Jiang, Hailong Meng, Khadir Raddassi, Ira Fleming, Kenneth B. Hoehn, Kenneth R. Dardick, Alexia A. Belperron, Ruth R. Montgomery, Alex K. Shalek, David A. Hafler, Steven H. Kleinstein, Linda K. Bockenstedt
There is an emerging need for accurate and rapid identification of bacteria in the human body to achieve diverse biomedical objectives. Copper homeostasis is vital for the survival of bacterial species owing to the roles of the metal as a nutrient, respiratory enzyme cofactor, and a toxin. Here, we report the development of a copper-64–labeled bacterial metal chelator, yersiniabactin, to exploit a highly conserved metal acquisition pathway for noninvasive and selective imaging of bacteria. Compared with traditional techniques used to manufacture probes, our strategy simplifies the process considerably by combining the function of metal attachment and cell recognition to the same molecule. We demonstrate, for the first time to our knowledge, how a copper-64 PET probe can be used to identify specific bacterial populations, monitor antibiotic treatment outcomes, and track bacteria in diverse niches in vivo.
Nabil A. Siddiqui, Hailey A. Houson, Nitin S. Kamble, Jose R. Blanco, Robert E. O’Donnell, Daniel J. Hassett, Suzanne E. Lapi, Nalinikanth Kotagiri
BACKGROUND. Little is known about pathogen-specific humoral immunity in individuals with long-term remission after treatment with chimeric antigen receptor-modified T-cells (CAR-T-cells) for B-cell lineage malignancies. METHODS. We conducted a prospective cross-sectional study of CD19-targeted or BCMA-targeted CAR-T-cell therapy recipients ≥6 months post-treatment and in remission. We measured lymphocyte subsets, immunoglobulins, pathogen-specific IgG for 12 vaccine-preventable infections, and the total number of viral and bacterial epitopes to which IgG was detected (‘epitope hits’) using a serological profiling assay. The primary outcome was the proportion of participants with IgG levels above a threshold correlated with seroprotection for vaccine-preventable infections. RESULTS. We enrolled 65 children and adults a median of 20 months after CD19- (n=54) or BCMA- (n=11) CAR-T-cell therapy. Among 30 adults without IgG replacement therapy (IGRT) in the prior 16 weeks, 27 (90%) had hypogammaglobulinemia. Despite this, these individuals had seroprotection to a median of 67% (IQR, 59-73%) of tested vaccine-preventable infections. Proportions of participants with seroprotection per-pathogen were comparable to population-based studies, but most individuals lacked seroprotection to specific pathogens. Compared to CD19-CAR-T-cell recipients, BCMA-CAR-T-cell recipients were half as likely to have seroprotection to vaccine-preventable infections (prevalence ratio, 0.47; 95% CI, 0.18-1.25) and had fewer pathogen-specific epitope hits (mean difference, -90 epitope hits; 95% CI, -157 to -22). CONCLUSIONS. Seroprotection for vaccine-preventable infections in adult CD19-CAR-T-cell recipients was comparable to the general population, but BCMA-CAR-T-cell recipients have fewer pathogen-specific antibodies. Deficits in both groups support the need for randomized vaccine and IGRT trials to determine efficacy and risk-benefit.
Carla S. Walti, Elizabeth M. Krantz, Joyce Maalouf, Jim Boonyaratanakornkit, Jacob Keane-Candib, Laurel Joncas-Schronce, Terry Stevens-Ayers, Sayan Dasgupta, Justin J. Taylor, Alexandre V. Hirayama, Merav Bar, Rebecca A. Gardner, Andrew J. Cowan, Damian J. Green, Michael J. Boeckh, David G. Maloney, Cameron J. Turtle, Joshua A. Hill
Emerging coronaviruses from zoonotic reservoirs including severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV), and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have been associated with human-to-human transmission and significant morbidity and mortality. Here we study both intradermal (ID) and intramuscular (IM) two-dose delivery regimens of an advanced synthetic DNA vaccine candidate encoding a full-length MERS-CoV Spike (S) protein, which induced potent binding and neutralizing antibodies, as well as cellular immune responses in rhesus macaques. In a MERS-CoV challenge, all immunized rhesus macaques exhibited reduced clinical symptoms, lowered viral lung load, and decreased severity of pathological signs of disease compared to controls. ID vaccination was dose sparing and more effective in this model at protecting animals from disease. The data support the further study of this vaccine for preventing MERS-CoV infection and transmission, including investigation of such vaccines and simplified delivery routes against emerging coronaviruses.
Ami Patel, Emma L. Reuschel, Ziyang Xu, Faraz I. Zaidi, Kevin Y. Kim, Dana P. Scott, Janess Mendoza, Stephanie Ramos, Regina Stoltz, Friederike Feldmann, Atsushi Okumura, Kimberly Meade-White, Elaine Haddock, Tina Thomas, Rebecca Rosenke, Jamie Lovaglio, Patrick W. Hanley, Greg Saturday, Kar Muthumani, Heinz Feldmann, Laurent M. Humeau, Kate E. Broderick, David B. Weiner
Background: COVID-19 is more benign in children compared to adults for unknown reasons. This contrasts with other respiratory viruses where disease manifestations are often more severe in children. We hypothesize that a more robust early innate immune response to SARS-CoV-2 protects against severe disease. Methods: Clinical outcomes, SARS-CoV-2 viral copies and cellular gene expression were compared in nasopharyngeal swabs obtained at the time of presentation to the Emergency Department from 12 children and 27 adults using bulk RNA sequencing and quantitative reverse transcription PCR. Total protein, cytokines and anti-SARS-CoV-2 IgG and IgA were quantified in nasal fluid. Results: SARS-CoV-2 copies, ACE2 and TMPRSS2 gene expression were similar in children and adults, but children displayed higher expression of genes associated with interferon signaling, NLRP3 inflammasome, and other innate pathways. Higher levels of IFN-α2, IFN-γ, IP-10, IL-8, and IL-1β protein were detected in nasal fluid in children versus adults. Children also expressed higher levels of genes associated with immune cells whereas expression of those associated with epithelial cells did not differ in children versus adults. Anti-SARS-CoV-2 IgA and IgG were detected at similar levels in nasal fluid from both groups. None of the children required supplemental oxygen whereas 7 adults did (p=0.03); four adults died. Conclusions: These findings provide direct evidence of a more vigorous early mucosal immune response in children compared to adults and suggest that this contributes to favorable clinical outcomes.
Carl A. Pierce, Sharlene Sy, Benjamin Galen, Doctor Y. Goldstein, Erika P. Orner, Marla J. Keller, Kevan C. Herold, Betsy C. Herold
Mounting evidence suggests that the balance of T cell costimulatory and coinhibitory signals contributes to mortality during sepsis. Here, we identified a critical role of the coinhibitory molecule T cell Ig and ITIM domain (TIGIT) in regulating sepsis mortality. Because TIGIT is significantly upregulated on memory T cells, we developed a “memory mouse” model to study the role of TIGIT during sepsis in a more physiologically relevant context. Mice received sequential pathogen exposure and developed memory T cell frequencies, similar to those observed in adult humans, and were then subjected to sepsis induction via cecal ligation and puncture. Our results show that targeting the TIGIT pathway during sepsis is fundamentally different in previously naive versus memory mice, in that αTIGIT Ab had no effect on survival in previously naive septic mice but sharply worsened survival in memory septic mice. Mechanistically, αTIGIT increased apoptosis of memory T cells, decreased T cell function, and downregulated the costimulatory receptor DNAM on memory CD8+ T cells in memory septic mice, but not in previously naive septic mice. Additionally, αTIGIT diminished Helios expression in Tregs in memory but not previously naive septic mice. These data highlight fundamental differences in the pathophysiological impact of targeting TIGIT in immunologically experienced versus previously naive hosts during sepsis.
Yini Sun, Jerome C. Anyalebechi, He Sun, Tetsuya Yumoto, Ming Xue, Danya Liu, Zhe Liang, Craig M. Coopersmith, Mandy L. Ford
Studies of human hepatitis B virus (HBV) immune pathogenesis are hampered by limited access to liver tissues and technologies for detailed analyses. Here, utilizing imaging mass cytometry (IMC) to simultaneously detect 30 immune, viral and structural markers in liver biopsies from patients with HBeAg+ chronic hepatitis B, we provide novel comprehensive visualization, quantitation and phenotypic characterizations of hepatic adaptive and innate immune subsets that correlated with hepatocellular injury, histological fibrosis and age. We further show marked correlations between adaptive and innate immune cell frequencies and phenotype, highlighting complex immune interactions within the hepatic microenvironment with relevance to HBV pathogenesis.
Daniel Traum, Yue J. Wang, Kathleen B. Schwarz, Jonathan Schug, David K.H. Wong, Harry L.A. Janssen, Norah A. Terrault, Mandana Khalili, Abdus S. Wahed, Karen F. Murray, Philip Rosenthal, Simon C. Ling, Norberto Rodriguez-Baez, Richard K. Sterling, Daryl T.Y. Lau, Timothy M. Block, Michael D. Feldman, Emma E. Furth, William M. Lee, David E. Kleiner, Anna S. Lok, Klaus H. Kaestner, Kyong-Mi Chang
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