Phosphatidylserine-Targeting Monoclonal Antibodies Exhibit Distinct Biochemical and Cellular Effects on Anti-CD3/CD28-Stimulated T Cell IFN-γ and TNF-α Production.

Phosphatidylserine (PS)-targeting monoclonal Abs (mAbs) that directly target PS and target PS via ß2-gp1 (ß2GP1) have been in preclinical and clinical development for over 10 y for the treatment of infectious diseases and cancer. Although the intended targets of PS-binding mAbs have traditionally included pathogens as well as stressed tumor cells and its associated vasculature in oncology, the effects of PS-targeting mAbs on activated immune cells, notably T cells, which externalize PS upon Ag stimulation, is not well understood. Using human T cells from healthy donor PBMCs activated with an anti-CD3 + anti-CD28 Ab mixture (anti-CD3/CD28) as a model for TCR-mediated PS externalization and T cell stimulation, we investigated effects of two different PS-targeting mAbs, 11.31 and bavituximab (Bavi), on TCR activation and TCR-mediated cytokine production in an ex vivo paradigm. Although 11.31 and Bavi bind selectivity to anti-CD3/28 activated T cells in a PS-dependent manner, surprisingly, they display distinct functional activities in their effect on IFN-γ and TNF-ɑ production, whereby 11.31, but not Bavi, suppressed cytokine production. This inhibitory effect on anti-CD3/28 activated T cells was observed on both CD4+ and CD8+ cells and independently of monocytes, suggesting the effects of 11.31 were directly mediated by binding to externalized PS on activated T cells. Imaging showed 11.31 and Bavi bind at distinct focal depots on the cell membrane. Collectively, our findings indicate that PS-targeting mAb 11.31 suppresses cytokine production by anti-CD3/28 activated T cells.

Dichotomy between the humoral and cellular responses elicited by mRNA and adenoviral vector vaccines against SARS-CoV-2.

BACKGROUND: Protection from severe disease and hospitalization by SARS-CoV-2 vaccination has been amply demonstrated by real-world data. However, the rapidly evolving pandemic raises new concerns. One pertains efficacy of adenoviral vector-based vaccines, particularly the single-dose Ad26.COV2.S, relative to mRNA vaccines. MAIN BODY: We investigated the immunogenicity of Ad26.COV2.S and mRNA vaccines in 33 subjects vaccinated with either vaccine class 5 months earlier on average. After controlling for the time since vaccination, Spike-binding antibody and neutralizing antibody levels were higher in the mRNA-vaccinated subjects, while no significant differences in antigen-specific B cell and T cell responses were observed between the two groups. CONCLUSIONS: A dichotomy exists between the humoral and cellular responses elicited by the two vaccine classes. Testing only for humoral responses to compare the durability of SARS-CoV-2 vaccine-induced responses, as typically performed for public health and research purposes, is insufficient.

Foam Cells: One Size Doesn't Fit All.

Chronic inflammation in many infectious and metabolic diseases, and some cancers, is accompanied by the presence of foam cells. These cells form when the intracellular lipid content of macrophages exceeds their capacity to maintain lipid homeostasis. Concurrently, critical macrophage immune functions are diminished. Current paradigms of foam cell formation derive from studies of atherosclerosis. However, recent studies indicate that the mechanisms of foam cell biogenesis during tuberculosis differ from those operating during atherogenesis. Here, we review how foam cell formation and function vary with disease context. Since foam cells are therapeutic targets in atherosclerosis, further research on the disease-specific mechanisms of foam cell biogenesis and function is needed to explore the therapeutic consequences of targeting these cells in other diseases.

Determinants and Dynamics of SARS-CoV-2 Infection in a Diverse Population: 6-Month Evaluation of a Prospective Cohort Study.

BACKGROUND: We studied risk factors, antibodies, and symptoms of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in a diverse, ambulatory population. METHODS: A prospective cohort (n = 831) previously undiagnosed with SARS-CoV-2 infection underwent serial testing (SARS-CoV-2 polymerase chain reaction, immunoglobulin G [IgG]) for 6 months. RESULTS: Ninety-three participants (11.2%) tested SARS-CoV-2-positive: 14 (15.1%) asymptomatic, 24 (25.8%) severely symptomatic. Healthcare workers (n = 548) were more likely to become infected (14.2% vs 5.3%; adjusted odds ratio, 2.1; 95% confidence interval, 1.4-3.3) and severely symptomatic (29.5% vs 6.7%). IgG antibodies were detected after 79% of asymptomatic infections, 89% with mild-moderate symptoms, and 96% with severe symptoms. IgG trajectories after asymptomatic infections (slow increases) differed from symptomatic infections (early peaks within 2 months). Most participants (92%) had persistent IgG responses (median 171 days). In multivariable models, IgG titers were positively associated with symptom severity, certain comorbidities, and hospital work. Dyspnea and neurologic changes (including altered smell/taste) lasted ≥ 120 days in ≥ 10% of affected participants. Prolonged symptoms (frequently more severe) corresponded to higher antibody levels. CONCLUSIONS: In a prospective, ethnically diverse cohort, symptom severity correlated with the magnitude and trajectory of IgG production. Symptoms frequently persisted for many months after infection.Clinical Trials Registration. NCT04336215.

Storage lipid studies in tuberculosis reveal that foam cell biogenesis is disease-specific.

Foam cells are lipid-laden macrophages that contribute to the inflammation and tissue damage associated with many chronic inflammatory disorders. Although foam cell biogenesis has been extensively studied in atherosclerosis, how these cells form during a chronic infectious disease such as tuberculosis is unknown. Here we report that, unlike the cholesterol-laden cells of atherosclerosis, foam cells in tuberculous lung lesions accumulate triglycerides. Consequently, the biogenesis of foam cells varies with the underlying disease. In vitro mechanistic studies showed that triglyceride accumulation in human macrophages infected with Mycobacterium tuberculosis is mediated by TNF receptor signaling through downstream activation of the caspase cascade and the mammalian target of rapamycin complex 1 (mTORC1). These features are distinct from the known biogenesis of atherogenic foam cells and establish a new paradigm for non-atherogenic foam cell formation. Moreover, they reveal novel targets for disease-specific pharmacological interventions against maladaptive macrophage responses.

Prevalence of SARS-CoV-2 infection in previously undiagnosed health care workers in New Jersey, at the onset of the U.S. COVID-19 pandemic.

BACKGROUND: Healthcare workers (HCW) are presumed to be at increased risk of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection due to occupational exposure to infected patients. However, there has been little epidemiological research to assess these risks. METHODS: We conducted a prospective cohort study of HCW (n = 546) and non-healthcare workers (NHCW; n = 283) with no known prior SARS-CoV-2 infection who were recruited from a large U.S. university and two affiliated university hospitals. In this cross-sectional analysis of data collected at baseline, we examined SARS-CoV-2 infection status (as determined by presence of SARS-CoV-2 RNA in oropharyngeal swabs) by healthcare worker status and role. RESULTS: At baseline, 41 (5.0%) of the participants tested positive for SARS-CoV-2 infection, of whom 14 (34.2%) reported symptoms. The prevalence of SARS-CoV-2 infection was higher among HCW (7.3%) than in NHCW (0.4%), representing a 7.0% greater absolute risk (95% confidence interval for risk difference 4.7, 9.3%). The majority of infected HCW (62.5%) were nurses. Positive tests increased across the two weeks of cohort recruitment in line with rising confirmed cases in the hospitals and surrounding counties. CONCLUSIONS: Overall, our results demonstrate that HCW had a higher prevalence of SARS-CoV-2 infection than NHCW. Continued follow-up of this cohort will enable us to monitor infection rates and examine risk factors for transmission.

Characterization of the Antigenic Heterogeneity of Lipoarabinomannan, the Major Surface Glycolipid of Mycobacterium tuberculosis, and Complexity of Antibody Specificities toward This Antigen.

Lipoarabinomannan (LAM), the major antigenic glycolipid of Mycobacterium tuberculosis, is an important immunodiagnostic target for detecting tuberculosis (TB) infection in HIV-1-coinfected patients, and is believed to mediate a number of functions that promote infection and disease development. To probe the human humoral response against LAM during TB infection, several novel LAM-specific human mAbs were molecularly cloned from memory B cells isolated from infected patients and grown in vitro. The fine epitope specificities of these Abs, along with those of a panel of previously described murine and phage-derived LAM-specific mAbs, were mapped using binding assays against LAM Ags from several mycobacterial species and a panel of synthetic glycans and glycoconjugates that represented diverse carbohydrate structures present in LAM. Multiple reactivity patterns were seen that differed in their specificity for LAM from different species, as well as in their dependence on arabinofuranoside branching and nature of capping at the nonreducing termini. Competition studies with mAbs and soluble glycans further defined these epitope specificities and guided the design of highly sensitive immunodetection assays capable of detecting LAM in urine of TB patients, even in the absence of HIV-1 coinfection. These results highlighted the complexity of the antigenic structure of LAM and the diversity of the natural Ab response against this target. The information and novel reagents described in this study will allow further optimization of diagnostic assays for LAM and may facilitate the development of potential immunotherapeutic approaches to inhibit the functional activities of specific structural motifs in LAM.

Variations on a theme: evolution of the phage-shock-protein system in Actinobacteria.

The phage shock protein (Psp) stress-response system protects bacteria from envelope stress through a cascade of interactions with other proteins and membrane lipids to stabilize the cell membrane. A key component of this multi-gene system is PspA, an effector protein that is found in diverse bacterial phyla, archaea, cyanobacteria, and chloroplasts. Other members of the Psp system include the cognate partners of PspA that are part of known operons: pspF||pspABC in Proteobacteria, liaIHGFSR in Firmicutes, and clgRpspAMN in Actinobacteria. Despite the functional significance of the Psp system, the conservation of PspA and other Psp functions, as well as the various genomic contexts of PspA, remain poorly characterized in Actinobacteria. Here we utilize a computational evolutionary approach to systematically identify the variations of the Psp system in ~450 completed actinobacterial genomes. We first determined the homologs of PspA and its cognate partners (as reported in Escherichia coli, Bacillus subtilis, and Mycobacterium tuberculosis) across Actinobacteria. This survey revealed that PspA and most of its functional partners are prevalent in Actinobacteria. We then found that PspA occurs in four predominant genomic contexts within Actinobacteria, the primary context being the clgRpspAM system previously identified in Mycobacteria. We also constructed a phylogenetic tree of PspA homologs (including paralogs) to trace the conservation and evolution of PspA across Actinobacteria. The genomic context revealed that PspA shows changes in its gene-neighborhood. The presence of multiple PspA contexts or of other known Psp members in genomic neighborhoods that do not carry pspA suggests yet undiscovered functional implications in envelope stress response mechanisms.

Flow Cytometric Characterization of Antigen-Specific T Cells Based on RNA and Its Advantages in Detecting Infections and Immunological Disorders.

Fluorescence in situ hybridization coupled with flow cytometry (FISH-Flow) is a highly quantitative, high-throughput platform allowing precise quantification of total mRNA transcripts in single cells. In undiagnosed infections posing a significant health burden worldwide, such as latent tuberculosis or asymptomatic recurrent malaria, an important challenge is to develop accurate diagnostic tools. Antigen-specific T cells create a persistent memory to pathogens, making them useful for diagnosis of infection. Stimulation of memory response initiates T-cell transitions between functional states. Numerous studies have shown that changes in protein levels lag real-time T-cell transitions. However, analysis at the single-cell transcriptional level can determine the differences. FISH-Flow is a powerful tool with which to study the functional states of T-cell subsets and to identify the gene expression profiles of antigen-specific T cells during disease progression. Advances in instrumentation, fluorophores, and FISH methodologies will broaden and deepen the use of FISH-Flow, changing the immunological field by allowing determination of functional immune signatures at the mRNA level and the development of new diagnostic tools.

Profiling T cell activation using single-molecule fluorescence in situ hybridization and flow cytometry.

Flow cytometric characterization of Ag-specific T cells typically relies on detection of protein analytes. Shifting the analysis to detection of RNA would provide several significant advantages, which we illustrate by developing a new host immunity-based platform for detection of infections. Cytokine mRNAs synthesized in response to ex vivo stimulation with pathogen-specific Ags are detected in T cells with single-molecule fluorescence in situ hybridization followed by flow cytometry. Background from pre-existing in vivo analytes is lower for RNAs than for proteins, allowing greater sensitivity for detection of low-frequency cells. Moreover, mRNA analysis reveals kinetic differences in cytokine expression that are not apparent at the protein level but provide novel insights into gene expression programs expected to define different T cell subsets. The utility of probing immunological memory of infections is demonstrated by detecting T cells that recognize mycobacterial and viral Ags in donors exposed to the respective pathogens.

The Psp system of Mycobacterium tuberculosis integrates envelope stress-sensing and envelope-preserving functions.

The bacterial envelope integrates essential stress-sensing and adaptive functions; thus, envelope-preserving functions are important for survival. In Gram-negative bacteria, envelope integrity during stress is maintained by the multi-gene Psp response. Mycobacterium tuberculosis was thought to lack the Psp system since it encodes only pspA and no other psp ortholog. Intriguingly, pspA maps downstream from clgR, which encodes a transcription factor regulated by the MprAB-σ(E) envelope-stress-signaling system. clgR inactivation lowered ATP concentration during stress and protonophore treatment-induced clgR-pspA expression, suggesting that these genes express Psp-like functions. We identified a four-gene set - clgR, pspA (rv2744c), rv2743c, rv2742c - that is regulated by clgR and in turn regulates ClgR activity. Regulatory and protein-protein interactions within the set and a requirement of the four genes for functions associated with envelope integrity and surface-stress tolerance indicate that a Psp-like system has evolved in mycobacteria. Among Actinobacteria, the four-gene module occurred only in tuberculous mycobacteria and was required for intramacrophage growth, suggesting links between its function and mycobacterial virulence. Additionally, the four-gene module was required for MprAB-σ(E) stress-signaling activity. The positive feedback between envelope-stress-sensing and envelope-preserving functions allows sustained responses to multiple, envelope-perturbing signals during chronic infection, making the system uniquely suited to tuberculosis pathogenesis.

Cutting edge: Vitamin D regulates lipid metabolism in Mycobacterium tuberculosis infection.

Vitamin D has long been linked to resistance to tuberculosis, an infectious respiratory disease that is increasingly hard to treat because of multidrug resistance. Previous work established that vitamin D induces macrophage antimicrobial functions against Mycobacterium tuberculosis. In this article, we report a novel, metabolic role for vitamin D in tuberculosis identified through integrated transcriptome and mechanistic studies. Transcriptome analysis revealed an association between vitamin D receptor (VDR) and lipid metabolism in human tuberculosis and infected macrophages. Vitamin D treatment of infected macrophages abrogated infection-induced accumulation of lipid droplets, which are required for intracellular M. tuberculosis growth. Additional transcriptomics results showed that vitamin D downregulates the proadipogenic peroxisome proliferator-activated receptor γ (PPARγ) in infected macrophages. PPARγ agonists reversed the antiadipogenic and the antimicrobial effects of VDR, indicating a link between VDR and PPARγ signaling in regulating both vitamin D functions. These findings suggest the potential for host-based, adjunct antituberculosis therapy targeting lipid metabolism.

Evidence for postinitiation regulation of mRNA biogenesis in tuberculosis.

Mycobacterium tuberculosis infection alters macrophage gene expression and macrophage response to IFN-γ, a critical host defense cytokine. However, regulation of these changes is poorly understood. We report discordance of changes in nascent transcript and total nuclear RNA abundance for the transcription factors STAT1 and IRF1, together with lack of effect on their RNA half-lives, in human THP-1 cells infected with M. tuberculosis and stimulated with IFN-γ. The results indicate that negative postinitiation regulation of mRNA biogenesis limits the expression of these factors, which mediate host defense against M. tuberculosis through the cellular response to IFN-γ. Consistent with the results for STAT1 and IRF1, transcriptome analysis reveals downregulation of postinitiation mRNA biogenesis processes and pathways by infection, with and without IFN-γ stimulation. Clinical relevance for regulation of postinitiation mRNA biogenesis is demonstrated by studies of donor samples showing that postinitiation mRNA biogenesis pathways are repressed in latent tuberculosis infection compared with cured disease and in active tuberculosis compared with ongoing treatment or with latent tuberculosis. For active disease and latent infection donors from two populations (London, U.K., and The Gambia), each analyzed using a different platform, pathway-related gene expression differences were highly correlated, demonstrating substantial specificity in the effect. Collectively, the molecular and bioinformatic analyses point toward downregulation of postinitiation mRNA biogenesis pathways as a means by which M. tuberculosis infection limits expression of immunologically essential transcription factors. Thus, negative regulation of postinitiation mRNA biogenesis can constrain the macrophage response to infection and overall host defense against tuberculosis.

The phage shock protein (PSP) envelope stress response: discovery of novel partners and evolutionary history.

Bacterial phage shock protein (PSP) systems stabilize the bacterial cell membrane and protect against envelope stress. These systems have been associated with virulence, but despite their critical roles, PSP components are not well characterized outside proteobacteria. Using comparative genomics and protein sequence-structure-function analyses, we systematically identified and analyzed PSP homologs, phyletic patterns, domain architectures, and gene neighborhoods. This approach underscored the evolutionary significance of the system, revealing that its core protein PspA (Snf7 in ESCRT outside bacteria) was present in the last universal common ancestor and that this ancestral functionality has since diversified into multiple novel, distinct PSP systems across life. Several novel partners of the PSP system were identified: (i) the Toastrack domain, likely facilitating assembly of sub-membrane stress-sensing and signaling complexes, (ii) the newly defined HTH-associated α-helical signaling domain-PadR-like transcriptional regulator pair system, and (iii) multiple independent associations with ATPase, CesT/Tir-like chaperone, and Band-7 domains in proteins thought to mediate sub-membrane dynamics. Our work also uncovered links between the PSP components and other domains, such as novel variants of SHOCT-like domains, suggesting roles in assembling membrane-associated complexes of proteins with disparate biochemical functions. Results are available at our interactive web app, https://jravilab.org/psp.IMPORTANCEPhage shock proteins (PSP) are virulence-associated, cell membrane stress-protective systems. They have mostly been characterized in Proteobacteria and Firmicutes. We now show that a minimal PSP system was present in the last universal common ancestor that evolved and diversified into newly identified functional contexts. Recognizing the conservation and evolution of PSP systems across bacterial phyla contributes to our understanding of stress response mechanisms in prokaryotes. Moreover, the newly discovered PSP modularity will likely prompt new studies of lineage-specific cell envelope structures, lifestyles, and adaptation mechanisms. Finally, our results validate the use of domain architecture and genetic context for discovery in comparative genomics.

Pragmatic Return to Effective Dental Infection Control through Triage and Testing (PREDICT): an observational, feasibility study to improve dental office safety.

BACKGROUND: During the COVID-19 pandemic, there was a substantial interruption of care, with patients and workers fearful to return to the dental office. As dental practice creates a highly aerosolized environment, the potential for spread of airborne illness is magnified. As a means to increase safety and mitigate risk, pre-visit testing for SARS-CoV-2 has the potential to minimize disease transmission in dental offices. The Pragmatic Return to Effective Dental Infection Control through Testing (PREDICT) Feasibility Study examined the logistics and impact of two different testing mechanisms (laboratory-based PCR viral testing and point-of-care antigen testing) in dental offices. METHODS: Dental healthcare workers (DHCWs) and patients in four dental offices within the National Dental Practice-based Research Network participated in this prospective study. In addition to electronic surveys, participants in two offices completed POC testing, while participants in two offices used lab-based PCR methods to detect SARS-CoV-2 infection. Analysis was limited to descriptive measures, with median and interquartile ranges reported for Likert scale responses and mean and standard deviation for continuous variables. RESULTS: Of the total 72 enrolled, 28 DHCWs and 41 patients completed the protocol. Two patients (4.9%) tested positive prior to their visit, while 2 DHCWs (12.5%) tested positive for SARS-CoV-2 infection at the start of the study. DHCWs and patients shared similar degree of concern (69% and 63%, respectively) for contracting COVID-19 from patients, while patients feared contracting COVID-19 from DHCWs less (49%). Descriptive statistics calculations revealed that saliva, tongue epithelial cells, and nasal swabs were the most desirable specimen collection method; both testing (LAB and POC) protocols took similar amounts of total time to complete; and DHCWs and patients reported feeling more comfortable when both groups were tested. CONCLUSIONS: While a larger-scale, network study is necessary for generalizability of results, this feasibility study suggests that SARS-CoV-2 testing can be effectively implemented into dental practice workflows and positively impact perception of safety for DHCWs and patients. As new virulent infectious diseases emerge, preparing dental personnel to employ an entire toolbox of risk mitigation strategies, including testing, may have the potential to decrease dental practice closure time, maintaining continuity of dental care services for patients. TRIAL REGISTRATION: ClinicalTrials.gov: NCT05123742.

A genetically modulated Toll-like-receptor-tolerant phenotype in peripheral blood cells of children with multisystem inflammatory syndrome.

Dysregulated innate immune responses contribute to multisystem inflammatory syndrome in children (MIS-C), characterized by gastrointestinal, mucocutaneous, and/or cardiovascular injury occurring weeks after SARS-CoV-2 exposure. To investigate innate immune functions in MIS-C, we stimulated ex vivo peripheral blood cells from MIS-C patients with agonists of Toll-like receptors (TLR), key innate immune response initiators. We found severely dampened cytokine responses and elevated gene expression of negative regulators of TLR signaling. Increased plasma levels of zonulin, a gut leakage marker, were also detected. These effects were also observed in children enrolled months after MIS-C recovery. Moreover, cells from MIS-C children carrying rare genetic variants of lysosomal trafficking regulator (LYST) were less refractory to TLR stimulation and exhibited lysosomal and mitochondrial abnormalities with altered energy metabolism. Our results strongly suggest that MIS-C hyperinflammation and/or excessive or prolonged stimulation with gut-originated TLR ligands drive immune cells to a lasting refractory state. TLR hyporesponsiveness is likely beneficial, as suggested by excess lymphopenia among rare LYST variant carriers. Our findings point to cellular mechanisms underlying TLR hyporesponsiveness; identify genetic determinants that may explain the MIS-C clinical spectrum; suggest potential associations between innate refractory states and long COVID; and highlight the need to monitor long-term consequences of MIS-C.

Researching COVID to enhance recovery (RECOVER) pediatric study protocol: Rationale, objectives and design.

IMPORTANCE: The prevalence, pathophysiology, and long-term outcomes of COVID-19 (post-acute sequelae of SARS-CoV-2 [PASC] or "Long COVID") in children and young adults remain unknown. Studies must address the urgent need to define PASC, its mechanisms, and potential treatment targets in children and young adults. OBSERVATIONS: We describe the protocol for the Pediatric Observational Cohort Study of the NIH's REsearching COVID to Enhance Recovery (RECOVER) Initiative. RECOVER-Pediatrics is an observational meta-cohort study of caregiver-child pairs (birth through 17 years) and young adults (18 through 25 years), recruited from more than 100 sites across the US. This report focuses on two of four cohorts that comprise RECOVER-Pediatrics: 1) a de novo RECOVER prospective cohort of children and young adults with and without previous or current infection; and 2) an extant cohort derived from the Adolescent Brain Cognitive Development (ABCD) study (n = 10,000). The de novo cohort incorporates three tiers of data collection: 1) remote baseline assessments (Tier 1, n = 6000); 2) longitudinal follow-up for up to 4 years (Tier 2, n = 6000); and 3) a subset of participants, primarily the most severely affected by PASC, who will undergo deep phenotyping to explore PASC pathophysiology (Tier 3, n = 600). Youth enrolled in the ABCD study participate in Tier 1. The pediatric protocol was developed as a collaborative partnership of investigators, patients, researchers, clinicians, community partners, and federal partners, intentionally promoting inclusivity and diversity. The protocol is adaptive to facilitate responses to emerging science. CONCLUSIONS AND RELEVANCE: RECOVER-Pediatrics seeks to characterize the clinical course, underlying mechanisms, and long-term effects of PASC from birth through 25 years old. RECOVER-Pediatrics is designed to elucidate the epidemiology, four-year clinical course, and sociodemographic correlates of pediatric PASC. The data and biosamples will allow examination of mechanistic hypotheses and biomarkers, thus providing insights into potential therapeutic interventions. CLINICAL TRIALS.GOV IDENTIFIER: Clinical Trial Registration: http://www.clinicaltrials.gov. Unique identifier: NCT05172011.

Immunodiagnosis of tuberculosis: a dynamic view of biomarker discovery.

Infection with Mycobacterium tuberculosis causes a variety of clinical conditions ranging from life-long asymptomatic infection to overt disease with increasingly severe tissue damage and a heavy bacillary burden. Immune biomarkers should follow the evolution of infection and disease because the host immune response is at the core of protection against disease and tissue damage in M. tuberculosis infection. Moreover, levels of immune markers are often affected by the antigen load. We review how the clinical spectrum of M. tuberculosis infection correlates with the evolution of granulomatous lesions and how granuloma structural changes are reflected in the peripheral circulation. We also discuss how antigen-specific, peripheral immune responses change during infection and how these changes are associated with the physiology of the tubercle bacillus. We propose that a dynamic approach to immune biomarker research should overcome the challenges of identifying those asymptomatic and symptomatic stages of infection that require antituberculosis treatment. Implementation of such a view requires longitudinal studies and a systems immunology approach leading to multianalyte assays.

Heterogeneity of foam cell biogenesis across diseases.

Foam cells are dysfunctional, lipid-laden macrophages associated with chronic inflammation of infectious and non-infectious origin. For decades, the paradigm underlying foam cell biology has been based on atherogenesis, a disease in which macrophages are cholesterol-enriched. Our previous work showed that foam cells in tuberculous lung lesions surprisingly accumulate triglycerides, suggesting multiple modalities of foam cell biogenesis. In the present study, we used matrix-assisted laser desorption/ionization mass spectrometry imaging to assess the spatial distribution of storage lipids relative to foam-cell-rich areas in murine lungs infected with the fungal pathogen Cryptococcus neoformans and in human papillary renal cell carcinoma resection tissues. We also analyzed neutral lipid content and the transcriptional program of lipid-laden macrophages generated under corresponding in vitro conditions. The in vivo data were consistent with in vitro findings showing that C. neoformans-infected macrophages accumulated triglycerides, while macrophages exposed to human renal cell carcinoma-conditioned medium accumulated both triglycerides and cholesterol. Moreover, macrophage transcriptome analyses provided evidence for condition-specific metabolic remodeling. The in vitro data also showed that although both Mycobacterium tuberculosis and C. neoformans infections induced triglyceride accumulation in macrophages, they did so by different molecular mechanisms, as evidenced by different sensitivity of lipid accumulation to the drug rapamycin and the characteristics of macrophage transcriptome remodeling. Collectively, these data demonstrate that the mechanisms of foam cell formation are specific to the disease microenvironment. Since foam cells have been regarded as targets of pharmacological intervention in several diseases, recognizing that their formation is disease-specific opens new research directions of biomedical significance.

Saliva microbiome in relation to SARS-CoV-2 infection in a prospective cohort of healthy US adults.

BACKGROUND: The clinical outcomes of SARS-CoV-2 infection vary in severity, potentially influenced by the resident human microbiota. There is limited consensus on conserved microbiome changes in response to SARS-CoV-2 infection, with many studies focusing on severely ill individuals. This study aimed to assess the variation in the upper respiratory tract microbiome using saliva specimens in a cohort of individuals with primarily mild to moderate disease. METHODS: In early 2020, a cohort of 831 adults without known SARS-CoV-2 infection was followed over a six-month period to assess the occurrence and natural history of SARS-CoV-2 infection. From this cohort, 81 participants with a SARS-CoV-2 infection, along with 57 unexposed counterparts were selected with a total of 748 serial saliva samples were collected for analysis. Total bacterial abundance, composition, population structure, and gene function of the salivary microbiome were measured using 16S rRNA gene and shotgun metagenomic sequencing. FINDINGS: The salivary microbiome remained stable in unexposed individuals over the six-month study period, as evidenced by all measured metrics. Similarly, participants with mild to moderate SARS-CoV-2 infection showed microbiome stability throughout and after their infection. However, there were significant reductions in microbiome diversity among SARS-CoV-2-positive participants with severe symptoms early after infection. Over time, the microbiome diversity in these participants showed signs of recovery. INTERPRETATION: These findings demonstrate the resilience of the salivary microbiome in relation to SARS-CoV-2 infection. Mild to moderate infections did not significantly disrupt the stability of the salivary microbiome, suggesting its ability to maintain its composition and function. However, severe SARS-CoV-2 infection was associated with temporary reductions in microbiome diversity, indicating the limits of microbiome resilience in the face of severe infection. FUNDING: This project was supported in part by Danone North America and grants from the National Institutes of Health, United States.