We enrolled 7 individuals with recurrent symptoms or antigen test conversion following nirmatrelvir-ritonavir treatment. High viral loads (median 6.1 log10 copies/mL) were detected after rebound for ...a median of 17 days after initial diagnosis. Three had culturable virus for up to 16 days after initial diagnosis. No known resistance-associated mutations were identified.
Several species of pathogenic bacteria replicate within an intracellular vacuolar niche. Bacteria that escape into the cytosol are captured by the autophagic pathway and targeted for lysosomal ...degradation, representing a defense against bacterial exploitation of the host cytosol. Autophagic capture of Salmonella Typhimurium occurs predominantly via generation of a polyubiquitin signal around cytosolic bacteria, binding of adaptor proteins, and recruitment of autophagic machinery. However, the components mediating bacterial target selection and ubiquitination remain obscure. We identify LRSAM1 as the E3 ligase responsible for anti-Salmonella autophagy-associated ubiquitination. LRSAM1 localizes to several intracellular bacterial pathogens and generates the bacteria-associated ubiquitin signal; these functions require LRSAM1's leucine-rich repeat and RING domains, respectively. Using cells from LRSAM1-deficient individuals, we confirm that LRSAM1 is required for ubiquitination associated with intracellular bacteria but dispensable for ubiquitination of aggregated proteins. LRSAM1 is therefore a bacterial recognition protein and ubiquitin ligase that defends the cytoplasm from invasive pathogens.
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► LRSAM1 localizes to bacteria via its LRR domain and restricts cytoplasmic replication ► LRSAM1 RING domain E3 ligase activity ubiquitinates bacteria and marks them for autophagy ► Antibacterial autophagy is decreased in cells from LRSAM1-deficient individuals
The principal components of both MHC class I and class II antigen processing and presentation pathways are well known. In dendritic cells, these pathways are tightly regulated by Toll-like-receptor ...signalling and include features, such as cross-presentation, that are not seen in other cell types. However, the exact mechanisms involved in the subcellular trafficking of antigens remain poorly understood and in some cases are controversial. Recent data suggest that diverse cellular machineries, including autophagy, participate in antigen processing and presentation, although their relative contributions remain to be fully elucidated. Here, we highlight some emerging themes of antigen processing and presentation that we think merit further attention.
•Fungal recognition by C-type lectin receptors are critical for proper immune response.•Innate effector pathways contribute to fungal killing and recruit immune cells.•CLR regulation and signaling ...can be leveraged for novel treatment strategies.
The innate immune system is critical to proper host defense against fungal pathogens, which is highlighted by increased susceptibility to invasive disease in immunocompromised patients. Innate cells (e.g. macrophages, neutrophils, dendritic cells, eosinophils) are equipped with intricate cell machinery to detect invading fungi and facilitate fungal killing, recruit additional immune cells, and direct the adaptive immune system responses. Understanding the mechanisms that govern a protective response will enable the development of novel treatment strategies. This review focuses on recent insights of signaling and regulation of C-type lectin receptors and their effector mechanisms enabling an effective host antifungal immunity.
Natural killer (NK) cells use the activating receptor NKp30 as a microbial pattern-recognition receptor to recognize, activate cytolytic pathways, and directly kill the fungi Cryptococcus neoformans ...and Candida albicans. However, the fungal pathogen-associated molecular pattern (PAMP) that triggers NKp30-mediated killing remains to be identified. Here we show that β-1,3-glucan, a component of the fungal cell wall, binds to NKp30. We further demonstrate that β-1,3-glucan stimulates granule convergence and polarization, as shown by live cell imaging. Through Src Family Kinase signaling, β-1,3-glucan increases expression and clustering of NKp30 at the microbial and NK cell synapse to induce perforin release for fungal cytotoxicity. Rather than blocking the interaction between fungi and NK cells, soluble β-1,3-glucan enhances fungal killing and restores defective cryptococcal killing by NK cells from HIV-positive individuals, implicating β-1,3-glucan to be both an activating ligand and a soluble PAMP that shapes NK cell host immunity.
Candida glabrata currently ranks as the second most frequent cause of invasive candidiasis. Our previous work has shown that C. glabrata is adapted to intracellular survival in macrophages and ...replicates within non-acidified late endosomal-stage phagosomes. In contrast, heat killed yeasts are found in acidified matured phagosomes. In the present study, we aimed at elucidating the processes leading to inhibition of phagosome acidification and maturation. We show that phagosomes containing viable C. glabrata cells do not fuse with pre-labeled lysosomes and possess low phagosomal hydrolase activity. Inhibition of acidification occurs independent of macrophage type (human/murine), differentiation (M1-/M2-type) or activation status (vitamin D3 stimulation). We observed no differential activation of macrophage MAPK or NFκB signaling cascades downstream of pattern recognition receptors after internalization of viable compared to heat killed yeasts, but Syk activation decayed faster in macrophages containing viable yeasts. Thus, delivery of viable yeasts to non-matured phagosomes is likely not triggered by initial recognition events via MAPK or NFκB signaling, but Syk activation may be involved. Although V-ATPase is abundant in C. glabrata phagosomes, the influence of this proton pump on intracellular survival is low since blocking V-ATPase activity with bafilomycin A1 has no influence on fungal viability. Active pH modulation is one possible fungal strategy to change phagosome pH. In fact, C. glabrata is able to alkalinize its extracellular environment, when growing on amino acids as the sole carbon source in vitro. By screening a C. glabrata mutant library we identified genes important for environmental alkalinization that were further tested for their impact on phagosome pH. We found that the lack of fungal mannosyltransferases resulted in severely reduced alkalinization in vitro and in the delivery of C. glabrata to acidified phagosomes. Therefore, protein mannosylation may play a key role in alterations of phagosomal properties caused by C. glabrata.
Despite vaccination and antiviral therapies, immunocompromised individuals are at risk for prolonged severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, but the immune defects ...that predispose an individual to persistent coronavirus disease 2019 (COVID-19) remain incompletely understood. In this study, we performed detailed viro-immunologic analyses of a prospective cohort of participants with COVID-19. The median times to nasal viral RNA and culture clearance in individuals with severe immunosuppression due to hematologic malignancy or transplant (S-HT) were 72 and 40 days, respectively, both of which were significantly longer than clearance rates in individuals with severe immunosuppression due to autoimmunity or B cell deficiency (S-A), individuals with nonsevere immunodeficiency, and nonimmunocompromised groups (
< 0.01). Participants who were severely immunocompromised had greater SARS-CoV-2 evolution and a higher risk of developing resistance against therapeutic monoclonal antibodies. Both S-HT and S-A participants had diminished SARS-CoV-2-specific humoral responses, whereas only the S-HT group had reduced T cell-mediated responses. This highlights the varied risk of persistent COVID-19 across distinct immunosuppressive conditions and suggests that suppression of both B and T cell responses results in the highest contributing risk of persistent infection.
Data are conflicting regarding an association between treatment of acute COVID-19 with nirmatrelvir-ritonavir (N-R) and virologic rebound (VR).
To compare the frequency of VR in patients with and ...without N-R treatment for acute COVID-19.
Observational cohort study.
Multicenter health care system in Boston, Massachusetts.
Ambulatory adults with acute COVID-19 with and without use of N-R.
Receipt of 5 days of N-R treatment versus no COVID-19 therapy.
The primary outcome was VR, defined as either a positive SARS-CoV-2 viral culture result after a prior negative result or 2 consecutive viral loads above 4.0 log
copies/mL that were also at least 1.0 log
copies/mL higher than a prior viral load below 4.0 log
copies/mL.
Compared with untreated persons (
= 55), those taking N-R (
= 72) were older, received more COVID-19 vaccinations, and more commonly had immunosuppression. Fifteen participants (20.8%) taking N-R had VR versus 1 (1.8%) who was untreated (absolute difference, 19.0 percentage points 95% CI, 9.0 to 29.0 percentage points;
= 0.001). All persons with VR had a positive viral culture result after a prior negative result. In multivariable models, only N-R use was associated with VR (adjusted odds ratio, 10.02 CI, 1.13 to 88.74;
= 0.038). Virologic rebound was more common among those who started therapy within 2 days of symptom onset (26.3%) than among those who started 2 or more days after symptom onset (0%) (
= 0.030). Among participants receiving N-R, those who had VR had prolonged shedding of replication-competent virus compared with those who did not have VR (median, 14 vs. 3 days). Eight of 16 participants (50% CI, 25% to 75%) with VR also reported symptom rebound; 2 were completely asymptomatic. No post-VR resistance mutations were detected.
Observational study design with differences between the treated and untreated groups; positive viral culture result was used as a surrogate marker for risk for ongoing viral transmission.
Virologic rebound occurred in approximately 1 in 5 people taking N-R, often without symptom rebound, and was associated with shedding of replication-competent virus.
National Institutes of Health.
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