Simultaneous Dengue and Chikungunya Coinfection in Endemic Area in Brazil: Clinical Presentation and Implications for Public Health.

Background: Dengue virus (DENV) and Chikungunya virus (CHIKV) pose significant public health threats in Brazil, where favorable conditions facilitated the proliferation of Aedes mosquitoes. Since the mid-1980s, Brazil has experienced annual outbreaks of DENV, with recent increases in confirmed cases. In addition, CHIKV, which was first reported in 2014, has spread across the country. The concurrent presence of these viruses has triggered public health alerts in endemic regions, underscoring the complexity of managing vector-borne diseases. Case Presentation: This report details a case of simultaneous DENV and CHIKV infections. A 77-year-old female patient who has diabetes and arrhythmia exhibited symptoms including fever, myalgia, and severe arthralgia. Laboratory tests confirmed the coinfection through RNA detection. The patient received supportive care, showed gradual improvement, and was eventually discharged. Conclusions: Coinfection with DENV and CHIKV cases reported here developed with mild outcomes. However, one of the patients did not recover from the arthralgia after presenting diagnostic challenges, which underscores the need for accurate differentiation to manage symptoms effectively. The reported cases, amidst increasing DENV outbreaks, highlight the urgency for preparedness in the healthcare system. The Ribeirão Preto region's endemicity for DENV, coupled with the rising incidence of CHIKV, emphasizes the evolving landscape of arbovirus transmission. Studies on Aedes mosquitoes suggest potential implications for human infection dynamics, warranting further investigation into arbovirus transmission efficacy and coinfection dynamics.

Exploring the Chikungunya virus landscape in a dengue-endemic Brazilian area.

We aimed to describe the landscape, including molecular, epidemiological, and clinical aspects of CHIKV infections in the Ribeirao Preto region, an area endemic to dengue. We randomly screened 3744 plasma samples that had undergone DENV diagnosis to evaluate CHIKV-RNA using an in-house RT-PCR assay. Positive samples were followed clinically, and RNA samples were submitted to whole genome sequencing. Seventeen cases (0.5 %) were positive for CHIKV-RNA despite being negative for DENV-RNA. Notably, half of the patients experienced prolonged arthralgia lasting more than 90 days. Compared with the healthy control group, leukopenia and thrombocytopenia were observed in all CHIKV-positive individuals with statistically significant P values (P < 0.0001 and P = 0.0003, respectively). The genomic analysis revealed that the CHIKV strains being studied are classified within the East-Central-South-African (ECSA) genotype. This analysis identified new mutations, E1: K211E and E2: V264A, while the previously known mutation E1: A226V was not detected among these strains. This study highlights the need for epidemiological surveillance and preparedness for potential CHIKV epidemics in Brazil, particularly where other arboviruses co-circulate.

Age-related Morphofunctional Changes in Sickle Cell Mice Bone Marrow Mesenchymal Stromal Cells.

BACKGROUND AND OBJECTIVES: Bone marrow mesenchymal stromal cells (BM-MSCs) are key elements of the hematopoietic niche and participate in the regulatory mechanisms of hematopoietic stem cells (HSCs). Hematological diseases can affect MSCs and their functions. However, the dysregulations caused by sickle cell disease (SCD) are not fully elucidated. This work explored changes in BM-MSCs and their relationship with age using sickle cell mice (Townes-SS). MATERIALS AND METHODS: BM-MSCs were isolated from Townes-SS, and control groups 30- and 60-day-old Townes-AA and C57BL/6 J. RESULTS: The BM-MSCs showed no morphological differences in culture and demonstrated a murine MSC-like immunophenotypic profile (Sca-1+, CD29+, CD44+, CD90.2+, CD31-, CD45-, and CD117-). Subsequently, all BM-MSCs were able to differentiate into adipocytes and osteocytes in vitro. Finally, 30-day-old BM-MSCs of Townes-SS showed higher expression of genes related to the maintenance of HSCs (Cxcl12, Vegfa, and Angpt1) and lower expression of pro-inflammatory genes (Tnfa and Il-6). However, 60-day-old BM-MSCs of Townes-SS started to show expression of genes related to reduced HSC maintenance and increased expression of pro-inflammatory genes. CONCLUSION: These results indicates age as a modifying factor of gene expression of BM-MSCs in the context of SCD.

Epidemiology of the SARS-CoV-2 Omicron Variant Emergence in the Southeast Brazilian Population.

The aim of this study was to describe epidemiological characteristics and perform SARS-CoV-2 genomic surveillance in the southeastern region of São Paulo State. During the first months of 2022, we compared weekly SARS-CoV-2 infection prevalence considering age, Ct value, and variants' lineages. An increase in the number of SARS-CoV-2-positive cases until the fourth epidemiological week of 2022 was observed. From the fourth epidemiological week onwards, the number of tests for SARS-CoV-2 diagnosis began to decrease, but the number of positive samples for SARS-CoV-2 remained high, reaching its most expressive level with a rate of 60% of infected individual cases. In this period, we observed a progressive increase in SARS-CoV-2 infection within the 0-10 age group throughout the epidemiological weeks, from 2.8% in the first epidemiological week to 9.2% in the eighth epidemiological week of 2022. We further observed significantly higher Ct values within younger patient samples compared to other older age groups. According to lineage assignment, SARS-CoV-2 (BA.1) was the most prevalent (74.5%) in the younger group, followed by BA.1.1 (23%), BA.2 (1.7%), and Delta (1%). Phylogenetic analysis showed that BA.2 sequences clustered together, indicating sustained transmission of this Omicron VOC sub-lineage by that time. Our results suggest the initial dissemination steps of the Omicron's sub-linage BA.2 into the younger group, due to specific genomic features of the detected sequences. These data provide interesting results related to the spread, emergence, and evolution of the Omicron variant in the southeast Brazilian population.

Dynamic clade transitions and the influence of vaccine rollout on the spatiotemporal circulation of SARS-CoV-2 variants in São Paulo, Brazil.

Since 2021, the emergence of variants of concern (VOC) has led Brazil to experience record numbers of in COVID-19 cases and deaths. The expanded spread of the SARS-CoV-2 combined with a low vaccination rate has contributed to the emergence of new mutations that may enhance viral fitness, leading to the persistence of the disease. Due to limitations in the real-time genomic monitoring of new variants in some Brazilian states, we aimed to investigate whether genomic surveillance, coupled with epidemiological data and SARS-CoV-2 variants spatiotemporal spread in a smaller region, can reflect the pandemic progression at a national level. Our findings revealed three SARS-CoV-2 variant replacements from 2021 to early 2022, corresponding to the introduction and increase in the frequency of Gamma, Delta, and Omicron variants, as indicated by peaks of the Effective Reproductive Number (Reff). These distinct clade replacements triggered two waves of COVID-19 cases, influenced by the increasing vaccine uptake over time. Our results indicated that the effectiveness of vaccination in preventing new cases during the Delta and Omicron circulations was six and eleven times higher, respectively, than during the period when Gamma was predominant, and it was highly efficient in reducing the number of deaths. Furthermore, we demonstrated that genomic monitoring at a local level can reflect the national trends in the spread and evolution of SARS-CoV-2.

Transitioning from static to suspension culture system for large-scale production of xeno-free extracellular vesicles derived from mesenchymal stromal cells.

Extracellular vesicles (EVs) derived from mesenchymal stromal cells (MSCs) have shown increasing therapeutic potential in the last years. However, large production of EV is required for therapeutic purposes. Thereby, scaling up MSC cultivation in bioreactors is essential to allow culture parameters monitoring. In this study, we reported the establishment of a scalable bioprocess to produce MSC-EV in suspension cultures using spinner flasks and human collagen-coated microcarriers (3D culture system). We compared the EV production in this 3D culture system with the standard static culture using T-flasks (2D culture system). The EV produced in both systems were characterized and quantify by western blotting and nanoparticle tracking analysis. The presence of the typical protein markers CD9, CD63, and CD81 was confirmed by western blotting analyses for EV produced in both culture systems. The cell fold-increase was 5.7-fold for the 3D culture system and 4.6-fold for the 2D culture system, signifying a fold-change of 1.2 (calculated as the ratio of fold-increase 3D to fold-increase 2D). Furthermore, it should be noted that the total cell production in the spinner flask cultures was 4.8 times higher than that in T-flask cultures. The total cell production in the spinner flask cultures was 5.2-fold higher than that in T-flask cultures. While the EV specific production (particles/cell) in T-flask cultures (4.40 ± 1.21 × 108 particles/mL, p < 0.05) was higher compared to spinner flask cultures (2.10 ± 0.04 × 108 particles/mL, p < 0.05), the spinner flask culture system offers scalability, making it capable of producing enough MSC-EV at a large scale for clinical applications. Therefore, we concluded that 3D culture system evaluated here serves as an efficient transitional platform that enables the scaling up of MSC-EV production for therapeutic purposes by utilizing stirred tank bioreactors and maintaining xeno-free conditions.

Live, Attenuated, Tetravalent Butantan-Dengue Vaccine in Children and Adults.

BACKGROUND: Butantan-Dengue Vaccine (Butantan-DV) is an investigational, single-dose, live, attenuated, tetravalent vaccine against dengue disease, but data on its overall efficacy are needed. METHODS: In an ongoing phase 3, double-blind trial in Brazil, we randomly assigned participants to receive Butantan-DV or placebo, with stratification according to age (2 to 6 years, 7 to 17 years, and 18 to 59 years); 5 years of follow-up is planned. The objectives of the trial were to evaluate overall vaccine efficacy against symptomatic, virologically confirmed dengue of any serotype occurring more than 28 days after vaccination (the primary efficacy end point), regardless of serostatus at baseline, and to describe safety up to day 21 (the primary safety end point). Here, vaccine efficacy was assessed on the basis of 2 years of follow-up for each participant, and safety as solicited vaccine-related adverse events reported up to day 21 after injection. Key secondary objectives were to assess vaccine efficacy among participants according to dengue serostatus at baseline and according to the dengue viral serotype; efficacy according to age was also assessed. RESULTS: Over a 3-year enrollment period, 16,235 participants received either Butantan-DV (10,259 participants) or placebo (5976 participants). The overall 2-year vaccine efficacy was 79.6% (95% confidence interval [CI], 70.0 to 86.3) - 73.6% (95% CI, 57.6 to 83.7) among participants with no evidence of previous dengue exposure and 89.2% (95% CI, 77.6 to 95.6) among those with a history of exposure. Vaccine efficacy was 80.1% (95% CI, 66.0 to 88.4) among participants 2 to 6 years of age, 77.8% (95% CI, 55.6 to 89.6) among those 7 to 17 years of age, and 90.0% (95% CI, 68.2 to 97.5) among those 18 to 59 years of age. Efficacy against DENV-1 was 89.5% (95% CI, 78.7 to 95.0) and against DENV-2 was 69.6% (95% CI, 50.8 to 81.5). DENV-3 and DENV-4 were not detected during the follow-up period. Solicited systemic vaccine- or placebo-related adverse events within 21 days after injection were more common with Butantan-DV than with placebo (58.3% of participants, vs. 45.6%). CONCLUSIONS: A single dose of Butantan-DV prevented symptomatic DENV-1 and DENV-2, regardless of dengue serostatus at baseline, through 2 years of follow-up. (Funded by Instituto Butantan and others; DEN-03-IB ClinicalTrials.gov number, NCT02406729, and WHO ICTRP number, U1111-1168-8679.).

Live, attenuated, tetravalent Butantan- Dengue vaccine in children and adults

BACKGROUND Butantan–Dengue Vaccine (Butantan-DV) is an investigational, single-dose, live, attenuated, tetravalent vaccine against dengue disease, but data on its overall efficacy are needed. METHODS In an ongoing phase 3, double-blind trial in Brazil, we randomly assigned participants to receive Butantan-DV or placebo, with stratification according to age (2 to 6 years, 7 to 17 years, and 18 to 59 years); 5 years of follow-up is planned. The objectives of the trial were to evaluate overall vaccine efficacy against symptomatic, virologically con firmed dengue of any serotype occurring more than 28 days after vaccination (the primary efficacy end point), regardless of serostatus at baseline, and to describe safety up to day 21 (the primary safety end point). Here, vaccine efficacy was assessed on the basis of 2 years of follow-up for each participant, and safety as solicited vaccine-related adverse events reported up to day 21 after injection. Key secondary objectives were to assess vaccine efficacy among participants according to dengue serostatus at baseline and according to the dengue viral serotype; efficacy according to age was also assessed. RESULTS Over a 3-year enrollment period, 16,235 participants received either Butantan-DV (10,259 participants) or placebo (5976 participants). The overall 2-year vaccine effi cacy was 79.6% (95% confidence interval [CI], 70.0 to 86.3) — 73.6% (95% CI, 57.6 to 83.7) among participants with no evidence of previous dengue exposure and 89.2% (95% CI, 77.6 to 95.6) among those with a history of exposure. Vaccine effi cacy was 80.1% (95% CI, 66.0 to 88.4) among participants 2 to 6 years of age, 77.8% (95% CI, 55.6 to 89.6) among those 7 to 17 years of age, and 90.0% (95% CI, 68.2 to 97.5) among those 18 to 59 years of age. Efficacy against DENV-1 was 89.5% (95% CI, 78.7 to 95.0) and against DENV-2 was 69.6% (95% CI, 50.8 to 81.5). DENV-3 and DENV-4 were not detected during the follow-up period. Solicited systemic vaccine- or placebo-related adverse events within 21 days after injection were more common with Butantan-DV than with placebo (58.3% of participants, vs. 45.6%). CONCLUSIONS A single dose of Butantan-DV prevented symptomatic DENV-1 and DENV-2, regard less of dengue serostatus at baseline, through 2 years of follow-up. (Funded by Instituto Butantan and others; DEN-03-IB ClinicalTrials.gov number, NCT02406729, and WHO ICTRP number, U1111-1168-8679.)

PARP1 Characterization as a Potential Biomarker for BCR::ABL1 p190+ Acute Lymphoblastic Leukemia.

Detection of t(9;22), and consequent BCR::ABL1 fusion, is still a marker of worse prognosis for acute lymphoblastic leukemia (ALL), with resistance to tyrosine-kinase inhibitor therapy being a major obstacle in the clinical practice for this subset of patients. In this study, we investigated the effectiveness of targeting poly-ADP-ribose polymerase (PARP) in a model of BCR::ABL1 p190+ ALL, the most common isoform to afflict ALL patients, and demonstrated the use of experimental PARP inhibitor (PARPi), AZD2461, as a therapeutic option with cytotoxic capabilities similar to that of imatinib, the current gold standard in medical care. We characterized cytostatic profiles, induced cell death, and biomarker expression modulation utilizing cell models, also providing a comprehensive genome-wide analysis through an aCGH of the model used, and further validated PARP1 differential expression in samples of ALL p190+ patients from local healthcare institutions, as well as in larger cohorts of online and readily available datasets. Overall, we demonstrate the effectiveness of PARPi in the treatment of BCR::ABL1 p190+ ALL cell models and that PARP1 is differentially expressed in patient samples. We hope our findings help expand the characterization of molecular profiles in ALL settings and guide future investigations into novel biomarker detection and pharmacological choices in clinical practice.

Exploring Viral Metagenomics in Pediatric Patients with Acute Respiratory Infections: Unveiling Pathogens beyond SARS-CoV-2.

The emergence of SARS-CoV-2 and the subsequent pandemic have prompted extensive diagnostic and clinical efforts to mitigate viral spread. However, these strategies have largely overlooked the presence of other respiratory viruses. Acute respiratory diseases in pediatric patients can be caused by a diverse range of viral agents, and metagenomics represents a powerful tool for their characterization. This study aimed to investigate the viral abundance in pediatric patients with acute respiratory symptoms who tested negative for SARS-CoV-2 during the Omicron pandemic wave. To achieve this, viral metagenomics and next-generation sequencing were employed on 96 nasopharyngeal swab samples, which were organized into 12 pools, with each pool consisting of eight individual samples. Metagenomic analysis revealed that the most prevalent viruses associated with acute disease in pediatric patients were respiratory syncytial virus (detected in all pools) and enteroviruses, which are known to cause significant morbidity and mortality in children. Additionally, clinically significant viruses such as mumps orthorubulavirus, human metapneumovirus, influenza A, and a wide array of human herpesviruses (1, 3-7) were identified. These findings highlight the extensive potential of viral metagenomics in identifying viruses other than SARS-CoV-2 that contribute to acute infections in children. Consequently, this methodology should garner clinical attention in terms of differential diagnosis and the development of public policies to address such conditions in the global pediatric population.

Engineering NK-CAR.19 cells with the IL-15/IL-15Rα complex improved proliferation and anti-tumor effect in vivo.

Introduction: Natural killer 92 (NK-92) cells are an attractive therapeutic approach as alternative chimeric antigen receptor (CAR) carriers, different from T cells, once they can be used in the allogeneic setting. The modest in vivo outcomes observed with NK-92 cells continue to present hurdles in successfully translating NK-92 cell therapies into clinical applications. Adoptive transfer of CAR-NK-92 cells holds out the promise of therapeutic benefit at a lower rate of adverse events due to the absence of GvHD and cytokine release syndrome. However, it has not achieved breakthrough clinical results yet, and further improvement of CAR-NK-92 cells is necessary. Methods: In this study, we conducted a comparative analysis between CD19-targeted CAR (CAR.19) co-expressing IL-15 (CAR.19-IL15) with IL-15/IL-15Rα (CAR.19-IL15/IL15Rα) to promote NK cell proliferation, activation, and cytotoxic activity against B-cell leukemia. CAR constructs were cloned into lentiviral vector and transduced into NK-92 cell line. Potency of CAR-NK cells were assessed against CD19-expressing cell lines NALM-6 or Raji in vitro and in vivo in a murine model. Tumor burden was measured by bioluminescence. Results: We demonstrated that a fourth- generation CD19-targeted CAR (CAR.19) co-expressing IL-15 linked to its receptor IL-15/IL-15Rα (CAR.19-IL-15/IL-15Rα) significantly enhanced NK-92 cell proliferation, proinflammatory cytokine secretion, and cytotoxic activity against B-cell cancer cell lines in vitro and in a xenograft mouse model. Conclusion: Together with the results of the systematic analysis of the transcriptome of activated NK-92 CAR variants, this supports the notion that IL-15/IL-15Rα comprising fourth-generation CARs may overcome the limitations of NK-92 cell-based targeted tumor therapies in vivo by providing the necessary growth and activation signals.

Human Pegivirus-1 Detection and Genotyping in Brazilian Patients with Fulminant Hepatitis.

Fulminant hepatitis is a severe clinical disease characterized by a marked decline in liver function and encephalopathy. In a previous survey, using metagenomics in a group of 27 patients with this clinical condition, we observed an expressive quantity of reads of the Human pegivirus-1 (HPgV-1). Therefore, the objective of this study was to evaluate the frequency, molecular features, and HPgV-1 circulating genotypes in patients with fulminant hepatitis. After testing the collected plasma samples, we discovered twelve samples (44.4%) that were positive for HPgV-1 RNA (using both real-time and nested PCR). The positive samples presented a mean cycle threshold (Ct) of 28.5 (±7.3). Genotyping assignments revealed that all HPgV-1 positive samples belonged to the HPgV-1 genotype 2 (both subgenotypes 2A and 2B were identified). Although HPgV-1 is considered a commensal virus, little is known regarding its prevalence and genotypes in cases of fulminant hepatitis. More research is needed to understand whether HPgV-1 can be implicated in clinical disorders and infectious diseases.

Genome Editing for Engineering the Next Generation of Advanced Immune Cell Therapies.

Our current genetic engineering capacity through synthetic biology and genome editing is the foundation of a revolution in biomedical science: the use of genetically programmed cells as therapeutics. The prime example of this paradigm is the adoptive transfer of genetically engineered T cells to express tumor-specific receptors, such as chimeric antigen receptors (CARs) or engineered T-cell receptors (TCR). This approach has led to unprecedented complete remission rates in patients with otherwise incurable hematological malignancies. However, this approach is still largely ineffective against solid tumors, which comprise the vast majority of neoplasms. Also, limitations associated with the autologous nature of this therapy and shared markers between cancer cells and T cells further restrict the access to these therapies. Here, we described how cutting-edge genome editing approaches have been applied to unlock the full potential of these revolutionary therapies, thereby increasing therapeutic efficacy and patient accessibility.

Time-dependent vaccine efficacy estimation quantified by a mathematical model.

In this paper we calculate the variation of the estimated vaccine efficacy (VE) due to the time-dependent force of infection resulting from the difference between the moment the Clinical Trial (CT) begins and the peak in the outbreak intensity. Using a simple mathematical model we tested the hypothesis that the time difference between the moment the CT begins and the peak in the outbreak intensity determines substantially different values for VE. We exemplify the method with the case of the VE efficacy estimation for one of the vaccines against the new coronavirus SARS-CoV-2.

Umbilical Cord Mesenchymal Stromal Cells for Steroid-Refractory Acute Graft-versus-Host Disease.

BACKGROUND: Steroid-refractory acute graft-vs.-host disease (SR-aGVHD) is a complication of allogeneic hematopoietic stem cell transplantation with a dismal prognosis and for which there is no consensus-based second-line therapy. Ruxolitinib is not easily accessible in many countries. A possible therapy is the administration of mesenchymal stromal cells (MSCs). METHODS: In this retrospective study, 52 patients with severe SR-aGVHD were treated with MSCs from umbilical cord (UC-MSCs) in nine institutions. RESULTS: The median (range) age was 12.5 (0.3-65) years and the mean ± SD dose (×106/kg) was 4.73 ± 1.3 per infusion (median of four infusions). Overall (OR) and complete response (CR) rates on day 28 were 63.5% and 36.6%, respectively. Children (n = 35) had better OR (71.5% vs. 47.1%, p = 0.12), CR (48.6% vs. 11.8%, p = 0.03), overall survival (p = 0.0006), and relapse-free survival (p = 0.0014) than adults (n = 17). Acute adverse events (all of them mild or moderate) were detected in 32.7% of patients, with no significant difference in children and adult groups (p = 1.0). CONCLUSIONS: UC-MSCs are a feasible alternative therapy for SR-aGVHD, especially in children. The safety profile is favorable.

Virulence Profiles of Wild-Type, P.1 and Delta SARS-CoV-2 Variants in K18-hACE2 Transgenic Mice.

Since December 2019, the world has been experiencing the COVID-19 pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and we now face the emergence of several variants. We aimed to assess the differences between the wild-type (Wt) (Wuhan) strain and the P.1 (Gamma) and Delta variants using infected K18-hACE2 mice. The clinical manifestations, behavior, virus load, pulmonary capacity, and histopathological alterations were analyzed. The P.1-infected mice showed weight loss and more severe clinical manifestations of COVID-19 than the Wt and Delta-infected mice. The respiratory capacity was reduced in the P.1-infected mice compared to the other groups. Pulmonary histological findings demonstrated that a more aggressive disease was generated by the P.1 and Delta variants compared to the Wt strain of the virus. The quantification of the SARS-CoV-2 viral copies varied greatly among the infected mice although it was higher in P.1-infected mice on the day of death. Our data revealed that K18-hACE2 mice infected with the P.1 variant develop a more severe infectious disease than those infected with the other variants, despite the significant heterogeneity among the mice.

Antibody response dynamics to CoronaVac vaccine and booster immunization in adults and the elderly: A long-term, longitudinal prospective study.

Background: The long-term humoral immune response after vaccination varies between vaccines and is dependent on the accuracy of the antibody test. A better understanding of the vaccine immune response may help to define vaccination strategies against coronavirus disease 2019 (COVID-19). Objective: To investigate the long-term immunological response to CoronaVac vaccine and determinants of breakthrough COVID-19 infection. Methods: A long-term, prospective cohort study involving vaccinated adult and elderly subjects was conducted to investigate the presence of anti-RBD-specific immunoglobulin (Ig)G, anti-nucleocapsid IgG and anti-spike trimeric protein IgG. Antibody level dynamics and risk factors associated with breakthrough COVID-19 infection were investigated. Results: In total, 3902 participants were included in this study. Vaccination with two doses of CoronaVac and a booster dose increased the levels of anti-RBD-specific IgG, anti-nucleocapsid IgG and anti-spike trimeric IgG significantly. In adults, anti-nucleocapsid IgG and anti-spike trimeric IgG levels decreased significantly 7 months after the second dose. In adults and the elderly, the levels of anti-spike trimeric IgG and anti-RBD IgG decreased significantly 4 and 6 months after the booster dose, respectively. Previous exposure to severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and anti-spike trimeric IgG titres was independently associated with a lower probability of post-vaccination infection. Conclusions: A significant increase in antibody levels was found after two doses of CoronaVac and a booster dose. Antibody titres declined significantly 7 months post-vaccination in participants who did not receive a booster dose. Higher levels of antibodies and previous SARS-CoV-2 infection were associated with protection against breakthrough COVID-19.