Infectious disease
Abstract
We developed a 29-color spectral cytometry panel to enhance nonhuman primate (NHP) models for cross-reactive immunophenotyping. This panel is suitable for biosafety level 4 (BSL-4) viruses and can be used with both human and NHP samples in BSL-2 research settings. Tissues from humans, rhesus monkeys (RhMs), crab-eating macaques (CEMs), and green monkeys (GMs) were stained with a 29-color immunophenotyping panel requiring only two clone substitutions. Comparable staining was observed for all samples. Unbiased analysis showed acceptable overlap in T-cell phenotypes across samples, with differences in human and NHP B cells and granulocytes. In CEMs, most circulating CD8+ T cells were from effector memory cells, with significantly higher levels than in humans (p<0.0001), RhMs (p<0.05), and GMs (p<0.01). Analysis of samples from various anatomical sites revealed distinct location-specific phenotypes. In Nipah-virus-exposed animals, splenocytes showed a substantial increase in IgM+ B cells (p<0.0001) and a reduction in effector memory CD8+ T cells (p<0.0001) compared to unexposed controls. Lymph nodes from Ebola-virus-exposed animals showed a loss of CXCR3+CD8+ T cells vs unexposed controls. This panel may guide the development of additional multi-color panels in preclinical and clinical settings and potentially increase understanding of the pathogenesis of diseases caused by emerging and re-emerging viruses.
Authors
Andrew P. Platt, Bobbi Barr, Anthony Marketon, Rebecca Bernbaum, Deja F.P. Rivera, Vincent J. Munster, Daniel S. Chertow, Michael R. Holbrook, Scott M. Anthony, Bapi Pahar
Abstract
Invasive aspergillosis is characterized by lung hemorrhage and release of extracellular heme, which promotes fungal growth. Heme can also mediate tissue injury directly, and both fungal growth and lung injury may induce hemorrhage. To assimilate these interdependent processes, we hypothesized that, during aspergillosis, heme mediates direct lung injury independent of fungal growth, leading to worse infection outcomes, and the scavenger protein, hemopexin, mitigates these effects. Mice with neutropenic aspergillosis were found to have a time-dependent increase in lung extracellular heme and a corresponding hemopexin induction. Hemopexin deficiency resulted in markedly increased lung injury, fungal growth, and lung hemorrhage. Using a computational model of the interactions of Aspergillus, heme, and the host, we predicted a critical role for heme-mediated generation of neutrophil-extracellular traps in this infection. We tested this prediction using a fungal strain unable to grow at body temperature, and found that extracellular heme and fungal exposure synergize to induce lung injury by promoting NET release, and disruption of NETs was sufficient to attenuate lung injury and fungal burden. These data implicate heme-mediated NETosis in both lung injury and fungal growth during aspergillosis, resulting in a detrimental positive feedback cycle that can be interrupted by scavenging heme or disrupting NETs.
Authors
Ganlin Qu, Henrique A.L. Ribeiro, Angelica L. Solomon, Luis Sordo Vieira, Yana Goddard, Nickolas G. Diodati, Arantxa V. Lazarte, Matthew Wheeler, Reinhard Laubenbacher, Borna Mehrad
Abstract
The SARS-CoV-2 pandemic highlighted the potential of mRNA vaccines in rapidly responding to emerging pathogens. However, immunity induced by conventional mRNA vaccines wanes quickly, requiring frequent boosters. Self-amplifying RNA (saRNA) vaccines, which extend antigen expression via self-replication, offer a promising strategy to induce more durable immune responses. In this study, we developed an saRNA vaccine encoding Zika virus (ZIKV) membrane and envelope (M/E) proteins and evaluated its efficacy in mice. A single vaccination elicited strong humoral and cellular immune responses and reduced viral loads, but only for 28 days. By day 84, antibody titers and T cell responses had significantly declined, resulting in reduced efficacy. To address this, we evaluated agonist antibodies targeting the T cell costimulatory molecules OX40 and 4-1BB. Co-administration of agonist antibodies enhanced CD8+ T cell responses to vaccination, resulting in sustained protection and reduced viral loads at day 84. Depletion and passive transfer studies confirmed that long-term protection was primarily CD8+ T cell-dependent, with minimal contributions from antibody responses. These findings suggest that agonists targeting members of the tumor necrosis receptor superfamily, such as OX40 and 4-1BB, might enhance the durability of saRNA vaccine-induced protection, addressing a key limitation of current mRNA vaccine platforms.
Authors
Hsueh-Han Lu, Rúbens Prince dos Santos Alves, Qin Hui Li, Luke Eder, Julia Timis, Henry Madany, Kantinan Chuensirikulchai, Krithik V. Varghese, Aditi Singh, Linda Le Tran, Audrey Street, Annie Elong Ngono, Michael Croft, Sujan Shresta
Abstract
The soluble variant of the ectopeptidase CD13 (sCD13), released from the cell surface by matrix metalloproteinase 14 (MMP14), is a potent pro-inflammatory mediator, displaying chemotactic, angiogenic, and arthritogenic properties through bradykinin receptor B1 (B1R). We reveal a link between sCD13 and amplified neutrophil-mediated inflammatory responses in SARS-CoV-2 infection. sCD13 was markedly elevated in COVID-19 patients and correlated with disease severity, variants, ethnicity, inflammation markers, and NETosis. Neutrophils treated with sCD13 showed heightened NETosis and chemotaxis which were inhibited by sCD13 receptor blockade. Meanwhile sCD13 did not induce platelet aggregation. Single-cell analysis of COVID-19 lungs revealed co-expression of CD13 and MMP14 by various cell types, and higher CD13 expression compared to controls. Neutrophils with high CD13 mRNA were enriched for genes associated with immaturity, though CD13 protein expression was lower. Histological examination of COVID-19 lungs revealed CD13-positive leukocytes trapped in vessels with fibrin thrombi. Flow cytometry confirmed the presence of B1R and a second sCD13 receptor, protease-activated receptor 4, on monocytes and neutrophils. These findings identify sCD13 as a potential instigator of COVID-19-associated NETosis, potentiating vascular stress and thromboembolic complications. The potent pro-inflammatory effects of sCD13 may contribute to severe COVID-19, suggesting that sCD13 and its receptors might be therapeutic targets.
Authors
Pei-Suen Tsou, Ramadan A. Ali, Chenyang Lu, Gautam Sule, Carmelo Carmona-Rivera, Serena Lucotti, Yuzo Ikari, Qi Wu, Phillip Campbell, Mikel Gurrea-Rubio, Kohei Maeda, Sharon E. Fox, William D. Brodie, Megan N. Mattichak, Caroline Foster, Ajay Tambralli, Srilakshmi Yalavarthi, M. Asif Amin, Katarina Kmetova, Bruna Mazetto Fonseca, Emily Chong, Yu Zuo, Michael Maile, Luisa Imberti, Arnaldo Caruso, Francesca Caccuri, Virginia Quaresima, Alessandra Sottini, Douglas B. Kuhns, Danielle L. Fink, Riccardo Castagnoli, Ottavia Delmonte, Heather Kenney, Yu Zhang, Mary Magliocco, Helen C. Su, Luigi D. Notarangelo, Rachel L. Zemans, Yang Mao-Draayer, Irina Matei, Mirella Salvatore, David C. Lyden, Yogendra Kanthi, Mariana J. Kaplan, Jason S. Knight, David A. Fox
Abstract
Urinary tract infections (UTIs) are one of the most commonly encountered infections in clinical practice, in which psychological stress is a critical pathological contributor to modulate immune function. However, mechanistic pathways linking stress networks in the brain to bladder infection remain poorly understood. In this study, we discovered that acute stress treatment suppressed bladder inflammation in mice with UTIs, and a significant number of neurons showing overlap between inflammation-associated markers and retrograde labeling were observed in the paraventricular nucleus (PVN) brain region of these mice. Activation of PVN alleviated UPEC-induced bladder inflammatory response. Moreover, blocked hypothalamic-pituitary-adrenal (HPA) axis reversed the anti-inflammatory reflex mediated by acute stress, suggesting that the potential of glucocorticoids levels through the brain-body circuits to ameliorate UTIs. Single cell-RNAseq of bladder immune cells revealed that type 2 innate lymphoid cells (ILC2) expressed abundant levels of glucocorticoid receptor (GR). The activation of PVN effectively inhibited the expression of the proinflammatory cytokine Csf2 by ILC2 through direct regulation of cell-intrinsic glucocorticoids signaling. Ultimately, our study has implications for the positioning of brain-body circuit for UTIs treatment.
Authors
Yaxiao Liu, Jinhua Wang, Junyang Lin, Dingqi Sun, Kejia Zhu, Tongxiang Diao, Qiang Fu, Qingyu Ren
Abstract
Septic arthritis, the most severe joint disease, is frequently caused by Staphylococcus aureus (S. aureus). A substantial proportion of patients with septic arthritis experience poor joint outcomes, often necessitating joint replacement surgery. Here, we show that monocyte depletion confers full protection against bone erosion in a septic arthritis mouse model. In the infected synovium, Ly6Chigh monocytes exhibited increased expression of osteoclastogenesis-related molecules, including CCR2, c-Fms, and RANK. S. aureus lipoproteins induced elevated levels of RANKL, MCSF, and CCL-2 in joints, with synovial fibroblasts identified as the major RANKL producer. Anti-RANKL treatment prevented bone destruction in both local and hematogenous septic arthritis murine models. Importantly, combining anti-RANKL treatment with antibiotics provided robust protection against joint damage. Our results indicate that the infiltration and transformation of monocytes into bone-destructive, osteoclast-like cells are key mechanisms in septic arthritis. Combining anti-RANKL and antibiotic therapy represents a promising therapy against this devastating disease.
Authors
Zhicheng Hu, Meghshree Deshmukh, Anders Jarneborn, Miriam Bollmann, Carmen Corciulo, Pradeep Kumar Kopparapu, Abukar Ali, Mattias N.D. Svensson, Cecilia Engdahl, Rille Pullerits, Majd Mohammad, Tao Jin
Abstract
The effect of preexisting neutralizing antibodies (NAb) on SARS-CoV-2 shedding in postvaccination infection (PVI) is not well understood. We characterized viral shedding longitudinally in nasal specimens in relation to baseline (pre/periinfection) serum NAb titers in 125 participants infected with SARS-CoV-2 variants. Among 68 vaccinated participants, we quantified the effect of baseline NAb titers on maximum viral RNA titers and infectivity duration. Baseline NAbs were higher and targeted a broader range of variants in participants with monovalent ancestral booster vaccinations compared with those with a primary vaccine series. In Delta infections, baseline NAb titers targeting Delta or Wuhan-Hu-1 correlated negatively with maximum viral RNA. Per log10 increase in Delta-targeting baseline NAb IC50, maximum viral load was reduced –2.43 (95% CI: –3.76, –1.11) log10 nucleocapsid copies, and infectious viral shedding was reduced –2.79 (95% CI: –4.99, –0.60) days. Conversely, in Omicron infections (BA.1, BA.2, BA.4, or BA.5), baseline NAb titers against Omicron lineages or Wuhan-Hu-1 did not predict viral outcomes. Our results provide robust estimates of the effect of baseline NAbs on the magnitude and duration of nasal viral replication after PVI (albeit with an unclear effect on transmission) and show how immune escape variants efficiently evade these modulating effects.
Authors
Miguel A. Garcia-Knight, J. Daniel Kelly, Scott Lu, Michel Tassetto, Sarah A. Goldberg, Amethyst Zhang, Jesus Pineda-Ramirez, Khamal Anglin, Michelle C. Davidson, Jessica Y. Chen, Maya Fortes-Cobby, Sara Park, Ana Martinez, Matthew So, Aidan Donovan, Badri Viswanathan, Eugene T. Richardson, David R. McIlwain, Brice Gaudilliere, Rachel L. Rutishauser, Ahmed Chenna, Christos Petropoulos, Terri Wrin, Steven G. Deeks, Glen R. Abedi, Sharon Saydah, Jeffrey N. Martin, Melissa Briggs Hagen, Claire M. Midgley, Michael J. Peluso, Raul Andino
Abstract
The ACTH test diagnoses relative adrenal insufficiency (RAI) or critical illness-related corticosteroid insufficiency (CIRCI). Initially, guidelines recommended corticosteroid/glucocorticoid (GC) therapy for septic patients with RAI, but later trials did not show a survival benefit, leading to updated guidelines that abandon targeting RAI or CIRCI. Recent studies with an RAI mouse model showed a clear survival benefit from GC therapy in mice with RAI, suggesting that inconclusive GC clinical trials might be due to issues with the ACTH test rather than targeting RAI. To investigate, we performed the ACTH test in septic mice. Interestingly, the ACTH test identified most mice as having adrenal insufficiency in early and middle stages of sepsis, even those with a normal adrenal stress response. Surprisingly, the ACTH test increased inflammatory cytokine to lethal levels, moderately increasing mortality in septic mice. This study revealed significant flaws in the ACTH test for diagnosing RAI/CIRCI. It not only fails to correctly identify these conditions, leading to misguided use of GC, but also induces a lethal inflammatory response in sepsis. These findings suggest that inconclusive GC therapy trials may be due to the problematic nature of the ACTH test rather than ineffectiveness of targeting RAI/CIRCI.
Authors
Dan Hao, Qian Wang, Misa Ito, Jianyao Xue, Ling Guo, Bin Huang, Chieko Mineo, Philip W. Shaul, Xiang-An Li
Abstract
Chlamydia trachomatis (CT) is the most common bacterial sexually transmitted infection globally. Understanding natural immunity to CT will inform vaccine design. This study aimed to profile immune cells and associated functional features in CT-infected women, and determine immune profiles associated with reduced risk of ascended endometrial CT infection and CT reinfection. PBMCs from CT-exposed women were profiled by mass cytometry and random forest models identified key features that distinguish outcomes. CT+ participants exhibited higher frequencies of CD4+ Th2, Th17, and Th17 DN CD4 T effector memory (TEM) cells than uninfected participants with decreased expression of T cell activation and differentiation markers. Minimal differences were detected between women with or without endometrial CT infection. Participants who remained follow-up negative (FU-) showed higher frequencies of CD4 T central memory (TCM) Th1, Th17, Th1/17, and Th17 DN but reduced CD4 TEM Th2 cells than FU+ participants. Expression of markers associated with central memory and Th17 lineage were increased on T cell subsets among FU- participants. These data indicate that peripheral T cells exhibit distinct features associated with resistance to CT reinfection. The highly plastic Th17 lineage appears to contribute to protection. Addressing these immune nuances could promote efficacy of CT vaccines.
Authors
Kacy S. Yount, Chi-Jane Chen, Avinash Kollipara, Chuwen Liu, Neha Vivek Mokashi, Xiaojing Zheng, C Bruce Bagwell, Taylor B. Poston, Harold C. Wiesenfeld, Sharon L. Hillier, Catherine M. O'Connell, Natalie Stanley, Toni Darville
Abstract
Dengue is widespread in tropical and subtropical regions globally and imposes a considerable disease burden. Annually, dengue virus (DENV) causes up to 400 million infections, of which approximately 25% present with clinical manifestations ranging from mild to fatal. Despite its significance as a growing public health concern, developing effective DENV vaccines has been challenging. One reason is the lack of comprehensive understanding of the influence exerted by prior DENV infections and immune responses with cross-reactive properties. To investigate this, we collected samples from a pediatric cohort study in dengue-endemic Managua, Nicaragua. We characterized T cell responses in 71 healthy children who had previously experienced 1 or more natural DENV infections and who, within 1 year after sample collection, had a subsequent DENV infection that was either symptomatic or inapparent. Our study investigated the effect of preexisting DENV-specific T cell responses on clinical outcomes of subsequent DENV infection. We assessed DENV-specific T cell responses using an activation-induced marker assay. Children with only 1 prior DENV infection displayed heterogeneous DENV-specific CD4+ and CD8+ T cell frequencies. In contrast, children with 2 or more prior DENV infections showed significantly higher DENV-specific CD4+ and CD8+ T cell frequencies associated with inapparent rather than symptomatic outcomes in subsequent infection. These findings demonstrate the protective role of DENV-specific T cells against symptomatic DENV infection and advance efforts to identify protective immune correlates against dengue.
Authors
Rosa Isela Gálvez, Amparo Martínez-Pérez, E. Alexandar Escarrega, Tulika Singh, José Victor Zambrana, Ángel Balmaseda, Eva Harris, Daniela Weiskopf
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