Age and sex influence antibody profiles associated with tuberculosis progression.

Antibody features vary with tuberculosis (TB) disease state. Whether clinical variables, such as age or sex, influence associations between Mycobacterium tuberculosis-specific antibody responses and disease state is not well explored. Here we profiled Mycobacterium tuberculosis-specific antibody responses in 140 TB-exposed South African individuals from the Adolescent Cohort Study. We identified distinct response features in individuals progressing to active TB from non-progressing, matched controls. A multivariate antibody score differentially associated with progression (SeroScore) identified progressors up to 2 years before TB diagnosis, earlier than that achieved with the RISK6 transcriptional signature of progression. We validated these antibody response features in the Grand Challenges 6-74 cohort. Both the SeroScore and RISK6 correlated better with risk of TB progression in adolescents compared with adults, and in males compared with females. This suggests that age and sex are important, underappreciated modifiers of antibody responses associated with TB progression.

Borrelia-specific antibody profiles and complement deposition in joint fluid distinguish antibiotic-refractory from -responsive Lyme arthritis.

Lyme arthritis, caused by the spirochete Borrelia burgdorferi, is the most common feature of late disseminated Lyme disease in the United States. While most Lyme arthritis resolves with antibiotics, termed "antibiotic-responsive", some individuals develop progressive synovitis despite antibiotic therapy, called "antibiotic-refractory" Lyme arthritis (LA). The primary drivers behind antibiotic-refractory arthritis remain incompletely understood. We performed a matched, cross-compartmental comparison of antibody profiles from blood and joint fluid of individuals with antibiotic-responsive (n = 11) or antibiotic-refractory LA (n = 31). While serum antibody profiles poorly discriminated responsive from refractory patients, a discrete profile of B.burgdorferi-specific antibodies in joint fluid discriminated antibiotic-responsive from refractory LA. Cross-compartmental comparison of antibody glycosylation, IgA1, and antibody-dependent complement deposition (ADCD) revealed more poorly coordinated humoral responses and increased ADCD in refractory disease. These data reveal B.burgdorferi-specific serological markers that may support early stratification and clinical management, and point to antibody-dependent complement activation as a key mechanism underlying persistent disease.

Understanding Fc function for rational vaccine design against pathogens.

Antibodies represent the primary correlate of immunity following most clinically approved vaccines. However, their mechanisms of action vary from pathogen to pathogen, ranging from neutralization, to opsonophagocytosis, to cytotoxicity. Antibody functions are regulated both by antigen specificity (Fab domain) and by the interaction of their Fc domain with distinct types of Fc receptors (FcRs) present in immune cells. Increasing evidence highlights the critical nature of Fc:FcR interactions in controlling pathogen spread and limiting the disease state. Moreover, variation in Fc-receptor engagement during the course of infection has been demonstrated across a range of pathogens, and this can be further influenced by prior exposure(s)/immunizations, age, pregnancy, and underlying health conditions. Fc:FcR functional variation occurs at the level of antibody isotype and subclass selection as well as post-translational modification of antibodies that shape Fc:FcR-interactions. These factors collectively support a model whereby the immune system actively harnesses and directs Fc:FcR interactions to fight disease. By defining the precise humoral mechanisms that control infections, as well as understanding how these functions can be actively tuned, it may be possible to open new paths for improving existing or novel vaccines.

Discrepancy between Mtb-specific IFN-γ and IgG responses in HIV-positive people with low CD4 counts.

BACKGROUND: Tuberculosis (TB) is a leading infectious cause of death worldwide and treating latent TB infection (LTBI) with TB preventative therapy is a global priority. This study aimed to measure interferon gamma (IFN-γ) release assay (IGRA) positivity (the current reference standard for LTBI diagnosis) and Mtb-specific IgG antibodies in otherwise healthy adults without HIV and those living with HIV (PLWH). METHODS: One-hundred and eighteen adults (65 without HIV and 53 antiretroviral-naïve PLWH), from a peri-urban setting in KwaZulu-Natal, South Africa were enrolled. IFN-γ released following stimulation with ESAT-6/CFP-10 peptides and plasma IgG antibodies specific for multiple Mtb antigens were measured using the QuantiFERON-TB Gold Plus (QFT) and customized Luminex assays, respectively. The relationships between QFT status, relative concentrations of anti-Mtb IgG, HIV-status, sex, age and CD4 count were analysed. FINDINGS: Older age, male sex and higher CD4 count were independently associated with QFT positivity (p = 0.045, 0.05 and 0.002 respectively). There was no difference in QFT status between people with and without HIV infection (58% and 65% respectively, p = 0.06), but within CD4 count quartiles, people with HIV had higher QFT positivity than people without HIV (p = 0.008 (2nd quartile), <0.0001 (3rd quartile)). Concentrations of Mtb-specific IFN-γ were lowest, and relative concentrations of Mtb-specific IgGs were highest in PLWH in the lowest CD4 quartile. INTERPRETATION: These results suggest that the QFT assay underestimates LTBI among immunosuppressed people with HIV and Mtb-specific IgG may be a useful alternative biomarker for Mtb infection. Further evaluation of how Mtb-specific antibodies can be leveraged to improve LTBI diagnosis is warranted, particularly in HIV-endemic areas. FUNDINGS: NIH, AHRI, SHIP: SA-MRC and SANTHE.

Distinguishing Severe Acute Respiratory Syndrome Coronavirus 2 Persistence and Reinfection: A Retrospective Cohort Study.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) reinfection is poorly understood, partly because few studies have systematically applied genomic analysis to distinguish reinfection from persistent RNA detection related to initial infection. We aimed to evaluate the characteristics of SARS-CoV-2 reinfection and persistent RNA detection using independent genomic, clinical, and laboratory assessments. METHODS: All individuals at a large academic medical center who underwent a SARS-CoV-2 nucleic acid amplification test (NAAT) ≥45 days after an initial positive test, with both tests between 14 March and 30 December 2020, were analyzed for potential reinfection. Inclusion criteria required having ≥2 positive NAATs collected ≥45 days apart with a cycle threshold (Ct) value <35 at repeat testing. For each included subject, likelihood of reinfection was assessed by viral genomic analysis of all available specimens with a Ct value <35, structured Ct trajectory criteria, and case-by-case review by infectious diseases physicians. RESULTS: Among 1569 individuals with repeat SARS-CoV-2 testing ≥45 days after an initial positive NAAT, 65 (4%) met cohort inclusion criteria. Viral genomic analysis characterized mutations present and was successful for 14/65 (22%) subjects. Six subjects had genomically supported reinfection, and 8 subjects had genomically supported persistent RNA detection. Compared to viral genomic analysis, clinical and laboratory assessments correctly distinguished reinfection from persistent RNA detection in 12/14 (86%) subjects but missed 2/6 (33%) genomically supported reinfections. CONCLUSIONS: Despite good overall concordance with viral genomic analysis, clinical and Ct value-based assessments failed to identify 33% of genomically supported reinfections. Scaling-up genomic analysis for clinical use would improve detection of SARS-CoV-2 reinfections.

Hybrid Immunity Shifts the Fc-Effector Quality of SARS-CoV-2 mRNA Vaccine-Induced Immunity.

Despite the robust immunogenicity of SARS-CoV-2 mRNA vaccines, emerging data have revealed enhanced neutralizing antibody and T cell cross-reactivity among individuals that previously experienced COVID-19, pointing to a hybrid immune advantage with infection-associated immune priming. Beyond neutralizing antibodies and T cell immunity, mounting data point to a potential role for additional antibody effector functions, including opsinophagocytic activity, in the resolution of symptomatic COVID-19. Whether hybrid immunity modifies the Fc-effector profile of the mRNA vaccine-induced immune response remains incompletely understood. Thus, here we profiled the SARS-CoV-2 specific humoral immune response in a group of individuals with and without prior COVID-19. As expected, hybrid Spike-specific antibody titers were enhanced following the primary dose of the mRNA vaccine but were similar to those achieved by naive vaccinees after the second mRNA vaccine dose. Conversely, Spike-specific vaccine-induced Fc-receptor binding antibody levels were higher after the primary immunization in individuals with prior COVID-19 and remained higher following the second dose compared to those in naive individuals, suggestive of a selective improvement in the quality, rather than the quantity, of the hybrid humoral immune response. Thus, while the magnitude of antibody titers alone may suggest that any two antigen exposures-either hybrid immunity or two doses of vaccine alone-represent a comparable prime/boost immunologic education, we find that hybrid immunity offers a qualitatively improved antibody response able to better leverage Fc-effector functions against conserved regions of the virus. IMPORTANCE Recent data indicates improved immunity to SARS-CoV-2 in individuals who experience a combination of two mRNA vaccine doses and infection, "hybrid immunity," compared to individuals who receive vaccination or experience infection alone. While previous infection accelerates the vaccine-induced immune response following the first dose of mRNA vaccination, subsequent doses demonstrate negligible increases in antibody titers or T cell immunity. Here, using systems serology, we observed a unique antibody profile induced by hybrid immunity, marked by the unique induction of robust Fc-recruiting antibodies directed at the conserved region of the viral Spike antigen, the S2-domain, induced at lower levels in individuals who only received mRNA vaccination. Thus, hybrid immunity clearly redirects vaccine-induced immunodominance, resulting in the induction of a robust functional humoral immune response to the most highly conserved region of the SARS-CoV-2 Spike antigen, which may be key to protection against existing and emerging variants of concern. Thus, next-generation vaccines able to mimic hybrid immunity and drive a balanced response to conserved regions of the Spike antigen may confer enhanced protection against disease.

The Kinetics of SARS-CoV-2 Antibody Development Is Associated with Clearance of RNAemia.

Persistent SARS-CoV-2 replication and systemic dissemination are linked to increased COVID-19 disease severity and mortality. However, the precise immune profiles that track with enhanced viral clearance, particularly from systemic RNAemia, remain incompletely defined. To define whether antibody characteristics, specificities, or functions that emerge during natural infection are linked to accelerated containment of viral replication, we examined the relationship of SARS-CoV-2-specific humoral immune evolution in the setting of SARS-CoV-2 plasma RNAemia, which is tightly associated with disease severity and death. On presentation to the emergency department, S-specific IgG3, IgA1, and Fc-γ-receptor (Fcγ R) binding antibodies were all inversely associated with higher baseline plasma RNAemia. Importantly, the rapid development of spike (S) and its subunit (S1/S2/receptor binding domain)-specific IgG, especially FcγR binding activity, were associated with clearance of RNAemia. These results point to a potentially critical and direct role for SARS-CoV-2-specific humoral immune clearance on viral dissemination, persistence, and disease outcome, providing novel insights for the development of more effective therapeutics to resolve COVID-19. IMPORTANCE We showed that persistent SARS-CoV-2 RNAemia is an independent predictor of severe COVID-19. We observed that SARS-CoV-2-targeted antibody maturation, specifically Fc-effector functions rather than neutralization, was strongly linked with the ability to rapidly clear viremia. This highlights the critical role of key humoral features in preventing viral dissemination or accelerating viremia clearance and provides insights for the design of next-generation monoclonal therapeutics. The main key points will be that (i) persistent SARS-CoV-2 plasma RNAemia independently predicts severe COVID-19 and (ii) specific humoral immune functions play a critical role in halting viral dissemination and controlling COVID-19 disease progression.

Serological Markers of SARS-CoV-2 Reinfection.

As public health guidelines throughout the world have relaxed in response to vaccination campaigns against SARS-CoV-2, it is likely that SARS-CoV-2 will remain endemic, fueled by the rise of more infectious SARS-CoV-2 variants. Moreover, in the setting of waning natural and vaccine immunity, reinfections have emerged across the globe, even among previously infected and vaccinated individuals. As such, the ability to detect reexposure to and reinfection by SARS-CoV-2 is a key component for global protection against this virus and, more importantly, against the potential emergence of vaccine escape mutations. Accordingly, there is a strong and continued need for the development and deployment of simple methods to detect emerging hot spots of reinfection to inform targeted pandemic response and containment, including targeted and specific deployment of vaccine booster campaigns. In this study, we identify simple, rapid immune biomarkers of reinfection in rhesus macaques, including IgG3 antibody levels against nucleocapsid and FcγR2A receptor binding activity of anti-RBD antibodies, that are recapitulated in human reinfection cases. As such, this cross-species analysis underscores the potential utility of simple antibody titers and function as price-effective and scalable markers of reinfection to provide increased resolution and resilience against new outbreaks. IMPORTANCE As public health and social distancing guidelines loosen in the setting of waning global natural and vaccine immunity, a deeper understanding of the immunological response to reexposure and reinfection to this highly contagious pathogen is necessary to maintain public health. Viral sequencing analysis provides a robust but unrealistic means to monitor reinfection globally. The identification of scalable pathogen-specific biomarkers of reexposure and reinfection, however, could significantly accelerate our capacity to monitor the spread of the virus through naive and experienced hosts, providing key insights into mechanisms of disease attenuation. Using a nonhuman primate model of controlled SARS-CoV-2 reexposure, we deeply probed the humoral immune response following rechallenge with various doses of viral inocula. We identified virus-specific humoral biomarkers of reinfection, with significant increases in antibody titer and function upon rechallenge across a range of humoral features, including IgG1 to the receptor binding domain of the spike protein of SARS-CoV-2 (RBD), IgG3 to the nucleocapsid protein (N), and FcγR2A receptor binding to anti-RBD antibodies. These features not only differentiated primary infection from reexposure and reinfection in monkeys but also were recapitulated in a sequencing-confirmed reinfection patient and in a cohort of putatively reinfected humans that evolved a PCR-positive test in spite of preexisting seropositivity. As such, this cross-species analysis using a controlled primate model and human cohorts reveals increases in antibody titers as promising cross-validated serological markers of reinfection and reexposure.

Fab and Fc contribute to maximal protection against SARS-CoV-2 following NVX-CoV2373 subunit vaccine with Matrix-M vaccination.

Recently approved vaccines have shown remarkable efficacy in limiting SARS-CoV-2-associated disease. However, with the variety of vaccines, immunization strategies, and waning antibody titers, defining the correlates of immunity across a spectrum of antibody titers is urgently required. Thus, we profiled the humoral immune response in a cohort of non-human primates immunized with a recombinant SARS-CoV-2 spike glycoprotein (NVX-CoV2373) at two doses, administered as a single- or two-dose regimen. Both antigen dose and boosting significantly altered neutralization titers and Fc-effector profiles, driving unique vaccine-induced antibody fingerprints. Combined differences in antibody effector functions and neutralization were associated with distinct levels of protection in the upper and lower respiratory tract. Moreover, NVX-CoV2373 elicited antibodies that functionally targeted emerging SARS-CoV-2 variants. Collectively, the data presented here suggest that a single dose may prevent disease via combined Fc/Fab functions but that two doses may be essential to block further transmission of SARS-CoV-2 and emerging variants.

Efficacy of Tocilizumab in Patients Hospitalized with Covid-19.

BACKGROUND: The efficacy of interleukin-6 receptor blockade in hospitalized patients with coronavirus disease 2019 (Covid-19) who are not receiving mechanical ventilation is unclear. METHODS: We performed a randomized, double-blind, placebo-controlled trial involving patients with confirmed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, hyperinflammatory states, and at least two of the following signs: fever (body temperature >38°C), pulmonary infiltrates, or the need for supplemental oxygen in order to maintain an oxygen saturation greater than 92%. Patients were randomly assigned in a 2:1 ratio to receive standard care plus a single dose of either tocilizumab (8 mg per kilogram of body weight) or placebo. The primary outcome was intubation or death, assessed in a time-to-event analysis. The secondary efficacy outcomes were clinical worsening and discontinuation of supplemental oxygen among patients who had been receiving it at baseline, both assessed in time-to-event analyses. RESULTS: We enrolled 243 patients; 141 (58%) were men, and 102 (42%) were women. The median age was 59.8 years (range, 21.7 to 85.4), and 45% of the patients were Hispanic or Latino. The hazard ratio for intubation or death in the tocilizumab group as compared with the placebo group was 0.83 (95% confidence interval [CI], 0.38 to 1.81; P = 0.64), and the hazard ratio for disease worsening was 1.11 (95% CI, 0.59 to 2.10; P = 0.73). At 14 days, 18.0% of the patients in the tocilizumab group and 14.9% of the patients in the placebo group had had worsening of disease. The median time to discontinuation of supplemental oxygen was 5.0 days (95% CI, 3.8 to 7.6) in the tocilizumab group and 4.9 days (95% CI, 3.8 to 7.8) in the placebo group (P = 0.69). At 14 days, 24.6% of the patients in the tocilizumab group and 21.2% of the patients in the placebo group were still receiving supplemental oxygen. Patients who received tocilizumab had fewer serious infections than patients who received placebo. CONCLUSIONS: Tocilizumab was not effective for preventing intubation or death in moderately ill hospitalized patients with Covid-19. Some benefit or harm cannot be ruled out, however, because the confidence intervals for efficacy comparisons were wide. (Funded by Genentech; ClinicalTrials.gov number, NCT04356937.).

Fecal microbiota transplantation: current clinical efficacy and future prospects.

Fecal microbiota transplantation (FMT) has gained mainstream attention with its remarkable efficacy in treating recurrent Clostridium difficile infection (RCDI) when there are no other effective therapies. Methods of selecting donors and routes of administration vary among studies, but there are now randomized controlled trials showing efficacy of FMT in treating RCDI. Ongoing trials of FMT for other disease such as inflammatory bowel disease are underway; this therapy should not be used for these conditions unless there is strong evidence for efficacy. Long-term safety data are sorely needed, as well as clarification of regulatory concerns.

Therapeutic vaccination expands and improves the function of the HIV-specific memory T-cell repertoire.

BACKGROUND: The licensing of herpes zoster vaccine has demonstrated that therapeutic vaccination can help control chronic viral infection. Unfortunately, human trials of immunodeficiency virus (HIV) vaccine have shown only marginal efficacy. METHODS: In this double-blind study, 17 HIV-infected individuals with viral loads of <50 copies/mL and CD4(+) T-cell counts of >350 cells/µL were randomly assigned to the vaccine or placebo arm. Vaccine recipients received 3 intramuscular injections of HIV DNA (4 mg) coding for clade B Gag, Pol, and Nef and clade A, B, and C Env, followed by a replication-deficient adenovirus type 5 boost (10(10) particle units) encoding all DNA vaccine antigens except Nef. Humoral, total T-cell, and CD8(+) cytotoxic T-lymphocyte (CTL) responses were studied before and after vaccination. Single-copy viral loads and frequencies of latently infected CD4(+) T cells were determined. RESULTS: Vaccination was safe and well tolerated. Significantly stronger HIV-specific T-cell responses against Gag, Pol, and Env, with increased polyfunctionality and a broadened epitope-specific CTL repertoire, were observed after vaccination. No changes in single-copy viral load or the frequency of latent infection were observed. CONCLUSIONS: Vaccination of individuals with existing HIV-specific immunity improved the magnitude, breadth, and polyfunctionality of HIV-specific memory T-cell responses but did not impact markers of viral control. CLINICAL TRIALS REGISTRATION: NCT00270465.