In medicine, virological diagnosis is mainly based on the detection of the viral genome and antigens, or on the identification of specific antibodies produced in response to infection. These strategies are suitable for characterizing an active infection or past contact with an already known virus. The recent development of tests for evaluating the host's cellular immune response opens new perspectives for personalized patient care based on immunomonitoring. The IGRA tests (Interferon Gamma Release Assay), measuring interferon gamma produced by T lymphocytes stimulated in vitro by antigenic peptides specific to infectious agents and the quantification of the blood viral load of Torque teno virus (TTV) thus constitute tools for assessing infectious risk, particularly usable to predict opportunistic viral reactivations in immunocompromised patients. The characterization of the expression profile of interferon stimulated genes (IGS) in a respiratory sample is also likely to provide significant assistance in diagnosis, discriminating a viral infection from a bacterial infection, an acute infection from a persistence of nucleic acids from non-replicating microorganisms or allowing, in case of viral emergence, to quickly identify infected subjects in the absence of specific PCR tests available. All of these new approaches, described in this review, have the potential to considerably improve patient care with the objective to correctly prescribe medical virology tests and anti-infective treatments.

Post-acute sequelae of COVID-19 (PASC) is a complex and multifaceted clinical challenge requiring to emphasize its underlying pathophysiological mechanisms. This study assessed hundreds of virological, serological, immunological, and tissue damage biomarkers in two patient cohorts who experienced mild (n=270) or severe (n=188) COVID-19, 6 to 9 months post-initial infection, and in which 40% and 57.4% of patients, respectively, developed PASC. Blood analysis showed that mains differences observed in humoral, viral, and biological biomarkers were associated with the initial COVID-19 severity, rather than being specifically linked to PASC. However, patients with PASC displayed altered CD4+ and CD8+ memory T-cell subsets, with higher cytokine-secreting cells and increased terminally differentiated CD45RA+ effector memory T cells (TEMRA). Elevated SARS-CoV-2-specific T cells responsive to nucleocapsid/membrane proteins with a TEMRA phenotype were also observed. A random forest model identified these features and initial symptom duration as top variables discriminating PASC, achieving over 80% classification accuracy.

Post-acute sequelae of COVID-19 (PASC) are a complex clinical condition that requires a better understanding of its underlying biological mechanisms. In this study, we assessed hundreds of virological, serological, immunological, and tissue damage biomarkers in two cohorts of patients who had experienced either mild (n = 270) or severe (n = 188) COVID-19, 6 to 9 months post-initial infection, and in which 40% and 57.4% of patients, respectively, developed PASC. Blood analysis showed that the main differences observed in humoral, viral, and biological biomarkers were associated with the initial COVID-19 severity, rather than being specifically linked to PASC. However, patients with PASC displayed altered CD4+ and CD8+ memory T cell subsets, with higher cytokine-secreting cells and increased terminally differentiated CD45RA+ effector memory T cells (TEMRA). Elevated SARS-CoV-2-specific T cells responsive to nucleocapsid/membrane proteins with a TEMRA phenotype were also observed. A random forest model identified these features and initial symptom duration as top variables discriminating PASC, achieving over 80% classification accuracy.

Torque teno virus (TTV) DNA load in plasma is suggested as a marker for immunosuppression post-transplantation. Crohn's disease (CD) arises from genetic susceptibility, environmental factors, and dysbiosis, causing immune responses. This study examines TTV DNA load in CD patients in remission and its correlation with relapse. Using quantitative real-time polymerase chain reaction (PCR) and metagenomic analysis, the dynamic of plasma TTV DNA load was analyzed from a cohort of CD patients (n = 39) over 1 year and compared with controls (n = 49). At inclusion, TTV DNA was significantly higher in CD patients compare to control (Median [IQR]: 3.27 [2.43-3.67] and 2.05 [1.28-2.80] Log copies/mL, p = 0.0004). Plasma TTV DNA load failed to predict disease relapse in CD patients. Augmented plasma TTV DNA levels in CD patients correlated with diminished circulating CD3 + T cells and especially CD4 + T cells. No preferential representation of TTV subspecies or Anelloviridae genera was detected in CD patients' plasma. This study revealed elevated TTV DNA levels in CD patients' plasma compared to healthy controls, underscoring the intriguing potential of TTV blood sampling as a biomarker in CD.

Growing evidence suggests that conventional immunomonitoring alone may not be sufficient to fully capture the complexity of immune dysfunctions. Immune functional assays (IFAs) have therefore emerged as valuable complementary tools, offering functional insights that extend beyond traditional phenotypic or quantitative approaches. Nevertheless, although in vitro stimulation represents a central component of IFAs, its specific contribution has never been rigorously evaluated, raising the critical question of whether this step is truly essential for detecting clinically relevant immune dysfunctions. To address this question, the present study compared gene expression levels (Nanostring) obtained from samples stimulated (TruCulture) or unstimulated (PaxGene) using the same analytical pipeline, in two distinct clinical settings: immune reconstitution following allogeneic hematopoietic stem cell transplantation (allo-HSCT) and sepsis progression. In allo-HSCT patients, post-stimulation data revealed immune heterogeneity and alterations related to ongoing immunosuppressive treatment or infectious event, not detected using unstimulated transcriptomic or cellular profiles alone. Similarly, post-stimulation transcriptomic profiles in patients with sepsis revealed immune clusters linked to disease severity and outcomes, surpassing traditional markers like mHLA-DR, while analyses from the unstimulated datasets failed to generate clinically relevant stratification. These findings emphasize the value of IFAs in uncovering immune function alterations that unstimulated assessments may miss, which could offer deeper insights into immune dysfunction. This study supports the use of IFAs as complementary tools to current clinical practices to enhance patient management by offering a functional view of immune system dynamics.

Immune reconstitution (IR) following allogeneic hematopoietic stem cell transplantation (allo-HSCT) is currently monitored by measuring the absolute number of immune effectors. However, this approach does not capture functional immune capacities. In this study, we aimed to evaluate the temporal evolution of functional IR alongside traditional immune cell count measurements. Whole-blood stimulation with TruCulture tubes (Myriad Rbm, Austin, TX, USA) containing lipopolysaccharides or staphylococcal enterotoxin B was performed on 55 allo-HSCT recipients at 6 and 12 months post-transplant, and on 10 healthy volunteers. The expression of 144 immune-related genes was quantified using NanoString technology (NanoString Technologies, Seattle, WA, USA) . The temporal follow-up of functional immune profiles was analyzed over time according to demographics, clinical characteristics, and immune cell counts. The evaluation of IR in allo-HSCT recipients up to 12 months post-transplant showed a significant discrepancy between quantitative and qualitative assessments. While immune cell counts improved, eg. the proportion of recipients reaching normal CD4+ T cell values, increasing from 25% to 46%, transcriptomic profiles showed persistent functional alterations. More than 78% of less-induced genes observed at 6 months still exhibited a reduced expression at 12 months post-transplant. Transcriptomic immune profiling divulged diverse functional outcomes linked to clinical characteristics, which were not reflected by cell count assessments alone. Herein, we emphasize that quantitative assessment of immune effectors alone is not informative enough to classify allo-HSCT recipients regarding functional immune capacity. Our findings highlight the value of implementing immune functional assay (IFA) as an additional tool for a comprehensive understanding of functional IR post-allo-HSCT, which could serve as a straightforward and efficient method for enabling personalized post-transplant management.

OBJECTIVES:In the post-SARS-CoV-2 pandemic era, "breakthrough infections" are still documented, due to variants of concerns (VoCs) emergence and waning humoral immunity. Despite widespread utilization, the definition of the anti-Spike (S) immunoglobulin-G (IgG) threshold to define protection has unveiled several limitations. Here, we explore the advantages of incorporating T-cell response assessment to enhance the definition of immune memory profile. METHODS:SARS-CoV-2 interferon-gamma release assay test (IGRA) was performed on samples collected longitudinally from immunocompetent healthcare workers throughout their immunization by infection and/or vaccination, anti-receptor-binding domain IgG levels were assessed in parallel. The risk of symptomatic infection according to cellular/humoral immune capacities during Omicron BA.1 wave was then estimated. RESULTS:Close to 40% of our samples were exclusively IGRA-positive, largely due to time elapsed since their last immunization. This suggests that individuals have sustained long-lasting cellular immunity, while they would have been classified as lacking protective immunity based solely on IgG threshold. Moreover, the Cox regression model highlighted that Omicron BA.1 circulation raises the risk of symptomatic infection while increased anti-receptor-binding domain IgG and IGRA levels tended to reduce it. CONCLUSION:The discrepancy between humoral and cellular responses highlights the significance of assessing the overall adaptive immune response. This integrated approach allows the identification of vulnerable subjects and can be of interest to guide antiviral prophylaxis at an individual level.

BackgroundExperimentally, ultra-protective ventilation (UPV, tidal volumes [VT] < 4 mL.kg−1) strategies in conjunction with veno-venous extracorporeal membrane oxygenation (VV-ECMO) are associated with lesser ventilator-induced lung injuries (VILI) during acute respiratory distress syndrome (ARDS). However, whether these strategies reduce lung inflammation more effectively than protective ventilation (PV) remains unclear. We aimed to demonstrate that a UPV strategy decreases acute lung inflammation in comparison with PV in an experimental swine model of ARDS.MethodsARDS was induced by tracheal instillation of chlorhydric acid in sedated and paralyzed animals under mechanical ventilation. Animals were randomized to receive either UPV (VT 1 mL.kg−1, positive end-expiration pressure [PEEP] set to obtain plateau pressure between 20 and 25 cmH2O and respiratory rate [RR] at 5 min−1 under VV-ECMO) or PV (VT 6 mL.kg−1, PEEP set to obtain plateau pressure between 28 and 30 cmH2O and RR at 25 min−1) during 4 h. After 4 h, a positron emission tomography with [11C](R)-PK11195 (ligand to TSPO-bearing macrophages) injection was realized, coupled with quantitative computerized tomography (CT). Pharmacokinetic multicompartment models were used to quantify regional [11C](R)-PK11195 lung uptake. [11C](R)-PK11195 lung uptake and CT-derived respiratory variables were studied regionally across eight lung regions distributed along the antero-posterior axis.ResultsFive pigs were randomized to each study group. Arterial O2 partial pressure to inspired O2 fraction were not significantly different between study groups after experimental ARDS induction (75 [68–80] mmHg in a PV group vs. 87 [69–133] mmHg in a UPV group, p = 0.20). Compared to PV animals, UPV animals exhibited a significant decrease in the regional non-aerated compartment in the posterior lung levels, in mechanical power, and in regional dynamic strain and no statistical difference in tidal hyperinflation after 4 h. UPV animals had a significantly lower [11C](R)-PK11195 uptake, compared to PV animals (non-displaceable binding potential 0.35 [IQR, 0.20–0.59] in UPV animals and 1.01 [IQR, 0.75–1.59] in PV animals, p = 0.01). Regional [11C](R)-PK11195 uptake was independently associated with the interaction of regional tidal hyperinflation and regional lung compliance.ConclusionIn an experimental model of ARDS, 4 h of UPV strategy significantly decreased lung inflammation, in relation to the control of VT-derived determinants of VILI.

Recently, immune function assessment has gained prominence in clinical settings. Immune functional assays (IFAs), involving in vitro stimulation, offer a relevant approach to complement traditional immunomonitoring methods which, while widely used, do not fully capture functional immune capabilities. Despite growing interest in IFAs, their added value remains unclear. To address this gap, our study aimed to determine if insights from IFAs could be replicated with unstimulated immunoprofiling. Using the same analytical pipeline, we compared transcriptomic profiles (Nanostring®) between stimulated (TruCulture®) and unstimulated (PaxGene™) samples from i) patients with an overstimulated immune system 3-4 days post-sepsis onset, and ii) patients undergoing immune reconstitution 6-months post-allogeneic hematopoietic stem cell transplantation (allo-HSCT). In sepsis, post-stimulation transcriptomic profiles revealed immune clusters linked to disease severity and outcomes, surpassing traditional markers like mHLA-DR, while baseline analyses failed to generate clinically relevant stratification. Similarly, allo-HSCT patients’ post-stimulation data revealed immune heterogeneity and treatment-related alterations not detected using baseline transcriptomic or cellular profiles alone. Our findings emphasize the value of IFAs in uncovering functional immune alterations that unstimulated assessments may miss, which could offer deeper insights into immune dysfunction. This study supports IFAs as complementary tools to current clinical practices, enhancing patient management with a functional view of immune system dynamics. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement REALISM study :This work was supported by the French National Research Agency through a grant awarded to BIOASTER (Grant number #ANR-10-AIRT-03) and from bioMerieux, Sanofi and GSK. VaccHemInf FIGHT study : This work was supported by an internal grant from the Hospices Civils de Lyon (Appel d'Offre Jeune Chercheur 2018, to A.C.); the Region Auvergne Rhone Alpes (Pack Ambition Recherche 2019, to F.A.) This work was supported by the public grant overseen by the French National Research Agency (Chaires industrielles 2023) and supported by the Association Nationale de la Recherche et de la Technologie (ANRT) with a CIFRE fellowship granted to Marion Debombourg. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: The REALISM study protocol was approved by the regional ethics committee (Comite de Protection des Personnes Sud-Est II, number 2015 42 2). The VaccHemInf protocol was approved by the regional ethics committee (Comite de Protection des Personnes Sud-Est V, Grenoble, France, number 69HCL17 0769). I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes all data in the present study are available upon reasonable request to the authors

Staphylococcus aureus main internalization mechanism in osteoblasts relies on a tripartite interaction between bacterial fibronectin-binding proteins, extracellular matrix soluble fibronectin, and osteoblasts' β1 integrins. Caveolins, and particularly caveolin-1, have been shown to limit the plasma membrane microdomain mobility, and consequently reduce the uptake of S. aureus in keratinocytes. In this study, we aimed to deepen our understanding of the molecular mechanisms underlying S. aureus internalization in osteoblasts. Mechanistically, S. aureus internalization requires endosomal recycling of β1 integrins as well as downstream effectors such as Src, Rac1, and PAK1. Surprisingly, in β1 integrin deficient osteoblasts, S. aureus internalization is restored when Caveolin-1 is absent and requires αvβ3/5 integrins as backup fibronectin receptors. Altogether, our data support that β1 integrins regulate the level of detergent-resistant membrane at the plasma membrane in a an endosomal and Caveolin-1 dependent manner.

Background This study aimed to demonstrate the utility of the nasal Type I interferon (IFN-I) response as a marker for respiratory viral infections (RVIs) and its potential to enhance diagnosis when combined with fi rst-line PCR tests for Influenza A/B, RSV, and SARS-CoV-2. Methods Nasopharyngeal swabs (NPS) from patients at Hospices Civils de Lyon (November 2022-April 2024) suspected of viral infections (n = 788) and from healthy controls (n = 53) were analysed. The IFN-I score was measured using the FILMARRAY (R) IFN-I pouch prototype, which detects four interferon-stimulated genes. The study evaluated the performance of the IFN-I score in detecting samples positive for viruses by fi rst-line PCR and assessed its benefit in diagnosing RVIs in samples initially classified as negative by PCR. Findings Out of 788 NPS included, 504 (64%) were positive with the fi rst-line PCR tests, and IFN-I score was significantly higher in those samples (median [IQR]: 13.00 [2.76-45.40]) compared to ones collected from healthy controls (1.09 [0.67-1.30]; p < 0.0001), with an area under the curve (AUC; 95% CI) of 0.92 (0.90-0.92). Moreover, out of the 284 NPS negative with fi rst-line PCR tests, suspicion of viral infection according to IFN-I score was found in 63% of cases (178/284). Second-line test (BioFire (R) Respiratory Panel 2.1 plus) and viral metagenomic confirmed the presence of viruses 94% of cases. Interpretation The study highlights the potential of integrating nasal IFN-I score into clinical workflows to improve RVI diagnosis and enhance preparedness for emerging viruses. Copyright (c) 2024 The Author(s). Published by Elsevier B.V. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).

Prone position decreases acute lung macrophage inflammation quantified in vivo with [11C](R)-PK11195 positron emission tomography in an experimental acute respiratory distress syndrome. Regional macrophage inflammation is maximal in the most anterior and posterior lung section of supine animals, in relation with increased regional tidal strain and hyperinflation, and reduced regional lung compliance.

IntroductionSevere Legionnaires’ disease (LD) can lead to multi-organ failure or death in 10%–30% of patients. Although hyper-inflammation and immunoparalysis are well described in sepsis and are associated with high disease severity, little is known about the immune response in LD. This study aimed to evaluate the immune status of patients with LD and its association with disease severity.MethodsA total of 92 hospitalized LD patients were included; 19 plasmatic cytokines and pulmonary Legionella DNA load were measured in 84 patients on the day of inclusion (day 0, D0). Immune functional assays (IFAs) were performed from whole blood samples collected at D2 and stimulated with concanavalin A [conA, n = 19 patients and n = 21 healthy volunteers (HV)] or lipopolysaccharide (LPS, n = 14 patients and n = 9 HV). A total of 19 cytokines (conA stimulation) and TNF-α (LPS stimulation) were quantified from the supernatants. The Sequential Organ Failure Assessment (SOFA) severity score was recorded at D0 and the mechanical ventilation (MV) status was recorded at D0 and D8.ResultsAmong the 84 patients, a higher secretion of plasmatic MCP-1, MIP1-β, IL-6, IL-8, IFN-γ, TNF-α, and IL-17 was observed in the patients with D0 and D8 MV. Multiparametric analysis showed that these seven cytokines were positively associated with the SOFA score. Upon conA stimulation, LD patients had a lower secretion capacity for 16 of the 19 quantified cytokines and a higher release of IL-18 and MCP-1 compared to HV. IL-18 secretion was higher in D0 and D8 MV patients. TNF-α secretion, measured after ex vivo LPS stimulation, was significantly reduced in LD patients and was associated with D8 MV status.DiscussionThe present findings describe a hyper-inflammatory phase at the initial phase of Legionella pneumonia that is more pronounced in patients with severe LD. These patients also present an immunoparalysis for a large number of cytokines, except IL-18 whose secretion is increased. An assessment of the immune response may be relevant to identify patients eligible for future innovative host-directed therapies.

Immune reconstitution after allogeneic-hematopoietic-stem-cell transplantation (allo-HSCT) is a complex and individual process. In this cross-sectional study, whole-blood (WB) immune functional assay (IFA) was used to characterize immune function by assessing immune-related gene/pathway alterations. The usefulness of this tool in the context of infection, 6 months after transplantation, was evaluated. Sixty allo-HSCT recipients at 6 months after transplantation and 10 healthy volunteers (HV) were included. WB was stimulated in standardized TruCulture tubes using lipopolysaccharides and Staphylococcal enterotoxin B. Gene expression was quantified using a custom 144-gene panel using NanoString nCounter technology and analyzed using Ingenuity Pathway Analysis. The relationships between immune function and clinical characteristics, immune cell counts, and post-transplantation infections were assessed. Allo-HSCT recipients were able to activate similar networks of the innate and adaptive immune response compared to HV, with, nevertheless, a lower intensity. A reduced number and a lower expression of genes associated with immunoregulatory and inflammatory processes were observed in allo-HSCT recipients. The use of immunosuppressive treatments was associated with a protracted immune reconstitution revealed by transcriptomic immunoprofiling. No difference in immune cell counts was observed among patients receiving or not receiving immunosuppressive treatments using a large immunophenotyping panel. Moreover, the expression of a set of genes, including CCL3/CCL4, was significantly lower in patients with Herpesviridae reactivation (32%, 19/60), which once again was not identified using classical immune cell counts. Transcriptional IFA revealed the heterogeneity among allo-HSCT recipients with a reduced immune function, a result that could not be captured by circulating immune cell counts. This highlights the potential added value of this tool for the personalized care of immunocompromised patients.

Background: Respiratory viral infections (RVI) represent a major public health issue. We aimed to evaluate a new diagnostic marker, the nasal type I interferon (IFN-I) score, by comparing it to classical and recently described methods.Methods: Nasopharyngeal swabs (NPS) positive for at least one respiratory virus were collected from patients admitted to the Hospices Civils de Lyon (n=421) and from healthy controls (HCs; n=30) between November 2022 and May 2023. IFN-I response was assessed using a FILMARRAY® IFN-I pouch prototype detecting four interferon-stimulated genes summarised in a nasal IFN-I score. The performance was compared to virus detection by multiplex PCR, normalised nasal viral load, viral culture, and nasal IFN-y-inducible protein 10 (IP-10).Findings: Among the 12 identified viruses, the most prevalent were SARS-CoV-2 (31·6%, 104/329), IAV (19·5%, 64/329), RSV (17·6%, 58/329), and HRV/EV (16·4%, 54/329). There was a significantly higher nasal IFN-I score in infected patients (median [IQR], 13·63 [3·21-44·00]) compared to HCs (0·84 [0·55-1·34]; p<0·0001) with an area under the curve (AUC; 95%CI) of 0·93 (0·91-0·96). The median [IQR] nasal IFN-I score was significantly higher in positive viral culture samples (n=163, 31·08 [10·71-87·62]) than in negative ones (n=150; 3·48 [1·64-10·57]; p<0·0001). The nasal IFN-I score was as effective as viral load in determining viruses replicative capacity (p>0·435). Although IP-10 can diagnose respiratory viral infections as well as the nasal IFN-I score, the AUC was lower for distinguishing replicative from non-replicative viruses [0·71, 95%CI (0·66-0·77) vs 0·83, 95%CI (0·79-0·88), respectively].Interpretation: The nasal IFN-I score is a relevant and reliable technique both in terms of detection and prediction of virus infectivity. It represents real hope for better care and isolation of infectious patients in the prevention and management of future epidemics.Funding: Public grant overseen by the French National Research Agency (Chaires industrielles 2023 and Recherche Hospitalo-Universitaire; ANR-21-RHUS-08).Declaration of Interest: CG, KBP, VC, GO, MM, AuG, SP, LG, and AF are employees of bioMérieux SA, an in vitro diagnostic company. BioMérieux kindly provided kits for the study. This funder of the study played a role in study design, data collection, data analysis, data interpretation, writing of the report, and decision to submit the manuscript for publication.Ethical Approval: The data processing associated with this study complies with the General Data Protection Regulation (GDPR) and is registered in the data controller's data processing register (HCL) under the number 21_5647, it was also approved by the scientific and ethics committee of the HCL (IRB n°00013204) under the number N°21_647. In agreement with the GDPR [Regulation (EU) 2016/679 and Directive 95/46/EC] and French data protection law (Law n°78-17 on 06/01/1978 and Decret n °2019-536 on 29/05/2019), a non-opposition consent form was sent to all patients.

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), along with extensive nonpharmacological interventions, have profoundly altered the epidemiology of major respiratory viruses. Some studies have described virus-virus interactions, particularly manifested by viral interference mechanisms at different scales. However, our knowledge of the interactions between SARS-CoV-2 and other respiratory viruses remains incomplete. Here, we studied the interactions between SARS-CoV-2 and several respiratory viruses (influenza, respiratory syncytial virus, human metapneumovirus, and human rhinovirus) in a reconstituted human epithelial airway model, exploring different scenarios affecting the sequence and timing of coinfections. We show that the virus type and sequence of infections are key factors in virus-virus interactions, the primary infection having a determinant role in the immune response to the secondary infection.

Herpes zoster, which is due to the reactivation of Varicella zoster virus (VZV), is a leading cause of morbidity after allogeneic hematopoietic stem cell transplantation (allo-HSCT). While cell-mediated immunity (CMI) is critical to inhibiting VZV reactivation, CMI is not routinely assessed due to a lack of reliable tests. In this study, we aimed to evaluate VZV-specific CMI among allo-HSCT recipients (n = 60) and healthy individuals (HI, n = 17) through a panel of three immune functional assays after ex vivo stimulation by VZV antigen: quantification of (i) IFN-γ release in the supernatants, (ii) T-cell proliferation after a 7-day stimulation of peripheral blood mononuclear cells (PBMC), and (iii) measurement of the ifn - γ mRNA gene expression level after 24 h of stimulation of a whole-blood sample. VZV responsiveness was defined according to IFN-γ release from VZV-stimulated PBMC. Upon VZV stimulation, we found that allo-HSCT recipients at a median time of 6 [5-8] months post-transplant had lower IFN-γ release (median [IQR], 0.34 [0.12–8.56] vs. 409.5 [143.9–910.2] pg/ml, P <.0001) and fewer proliferating T cells (0.05 [0.01–0.57] % vs. 8.74 [3.12–15.05] %, P <.0001) than HI. A subset of allo-HSCT recipients (VZV-responders, n = 15/57, 26%) distinguished themselves from VZV-non-responders (n = 42/57, 74%; missing data, n = 3) by higher IFN-γ release (80.45 [54.3–312.8] vs. 0.22 [0.12–0.42] pg/ml, P <.0001) and T-cell proliferation (2.22 [1.18–7.56] % vs. 0.002 [0.001–0.11] %, P <.0001), suggesting recovery of VZV-specific CMI. Interestingly, VZV responders had a significant fold increase in ifn-γ gene expression, whereas ifn-γ mRNA was not detected in whole blood of VZV-non-responders ( P <.0001). This study is the first to suggest that measurement of ifn-γ gene expression in 24-h-stimulated whole blood could be an accurate test of VZV-specific CMI. The routine use of this immune functional assay to guide antiviral prophylaxis at an individual level remains to be evaluated.

IFN-I and IFN-III immunity in the nasal mucosa is poorly characterized during SARS-CoV-2 infection. We analyze the nasal IFN-I/III signature, namely the expression of ISGF-3-dependent IFN-stimulated genes, in mildly symptomatic COVID-19 patients and show its correlation with serum IFN-α2 levels, which peak at symptom onset and return to baseline from day 10 onward. Moreover, the nasal IFN-I/III signature correlates with the nasopharyngeal viral load and is associated with the presence of infectious viruses. By contrast, we observe low nasal IFN-I/III scores despite high nasal viral loads in a subset of critically ill COVID-19 patients, which correlates with the presence of autoantibodies (auto-Abs) against IFN-I in both blood and nasopharyngeal mucosa. In addition, functional assays in a reconstituted human airway epithelium model of SARS-CoV-2 infection confirm the role of such auto-Abs in abrogating the antiviral effects of IFN-I, but not those of IFN-III. Thus, IFN-I auto-Abs may compromise not only systemic but also local antiviral IFN-I immunity at the early stages of SARS-CoV-2 infection.

Antigen-specific T-cells are essential for protective immunity against SARS-CoV-2. We set up a semi-automated whole-blood Interferon-gamma release assay (WB IGRA) to monitor the T-cell response after stimulation with SARS-CoV-2 peptide pools. We report that the WB IGRA is complementary to serological assays to assess SARS-CoV-2 immunity.

Methicillin-resistant Staphylococcus aureus (MRSA) of the ST1-SCCmecIV lineage has been associated with community-acquired (CA) infections in North America and Australia. In Brazil, multi-drug resistant ST1-SCCmecIV MRSA has emerged in hospital-associated (HA) diseases in Rio de Janeiro. To understand these epidemiological differences, genomic and phylogenetic analyses were performed. In addition, virulence assays were done for representative CA - and HA-MRSA strains. Despite the conservation of the virulence repertoire, some genes were missing in Brazilian ST1-SCCmecIV including lukSF-PV, fnbB, and several superantigen-encoded genes. Additionally, CA-MRSA lost the splDE while HA-MRSA strains conserved the complete operon. Most of these variable genes were located in mobile genetic elements (MGE). However, conservation and maintenance of MGEs were often observed despite the absence of their associated virulence markers. A Bayesian phylogenetic tree revealed the occurrence of more than one entrance of ST1 strains in Rio de Janeiro. The tree shape and chronology allowed us to infer that the hospital-associated ST1-SCCmecIV from Brazil and the community-acquired USA400 from North America are not closely related and that they might have originated from different MSSA strains that independently acquired SCCmecIV cassettes. As expected, representatives of ST1 strains from Brazil showed lower cytotoxicity and a greater ability to survive inside human host cells. We suggest that Brazilian ST1-SCCmecIV strains have adapted to the hospital setting by reducing virulence and gaining the ability to persist and survive inside host cells. Possibly, these evolutionary strategies may balance the biologic cost of retaining multiple antibiotic resistance genes.