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To map the spread of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) and evaluate immune response variations against this virus, it is essential to set up efficient serological tests locally. The SARS-CoV-2 immunogenic proteins were very expensive and not affordable for lower- middle-income countries (LMICs). For this purpose, the commonly used antigen, receptor- binding domain (RBD) of spike S1 protein (S1RBD), was produced using the baculovirus expression vector system (BEVS). In the current study, the expression of S1RBD was monitored using Western blot under different culture conditions. Different parameters were studied: the multiplicity of infection (MOI), cell density at infection, and harvest time. Hence, optimal conditions for efficient S1RBD production were identified: MOI 3; cell density at infection 2-3 × 106 cells/mL; and time post-infection (tPI or harvest time) of 72 h and 72-96 h, successively, for expression in shake flasks and a 7L bioreactor. A high production yield of S1RBD varying between 4 mg and 70 mg per liter of crude cell culture supernatant was achieved, respectively, in the shake flasks and 7L bioreactor. Moreover, the produced S1RBD showed an excellent antigenicity potential against COVID-19 (Wuhan strain) patient sera evaluated by Western blot. Thus, additional serological assays, such as in-house ELISA and seroprevalence studies based on the purified S1RDB, were developed.

Background: Vaccination constitutes the best strategy against COVID-19. In Tunisia, seven vaccines standing for the three main platforms, namely RNA, viral vector, and inactivated vaccines, have been used to vaccinate the population at a large scale. This study aimed to assess, in our setting, the kinetics of vaccine-induced anti-RBD IgG and IgA antibody responses. Methods: Using in-house developed and validated ELISA assays, we measured anti-RBD IgG and IgA serum antibodies in 186 vaccinated workers at the Institut Pasteur de Tunis over 12 months. Results: We showed that RNA vaccines were the most immunogenic vaccines, as compared to alum-adjuvanted inactivated and viral-vector vaccines, either in SARS-CoV-2-naive or in SARS-CoV-2-experienced individuals. In addition to the IgG antibodies, the vaccination elicited RBD-specific IgAs. Vaccinated individuals with prior SARS-CoV-2 infection exhibited more robust IgG and IgA antibody responses, as compared to SARS-CoV-2-naive individuals. Conclusions: After following up for 12 months post-immunization, we concluded that the hierarchy between the platforms for anti-RBD antibody-titer dynamics was RNA vaccines, followed by viral-vector and alum-adjuvanted inactivated vaccines.

Seroprevalence studies are essential to get an accurate estimate of the actual SARS-CoV-2 diffusion within populations. We report on the findings of the first serosurvey conducted in Tunis prior to the implementation of mass vaccination and analyzed factors associated with seropositivity. A household cross sectional survey was conducted (March–April 2021) in Tunis, spanning the end of the second wave and the beginning of the third wave of COVID-19. SARS-CoV-2 specific immunoglobulin G (IgG) antibodies to the spike (S-RBD) or the nucleocapsid (N) proteins were detected by in-house ELISA tests. The survey included 1676 individuals from 431 households. The mean age and sex ratio were 43.3 ± 20.9 years and 0.6, respectively. The weighted seroprevalence of anti-N and/or anti-S-RBD IgG antibodies was equal to 38.0% (34.6–41.5). In multivariate analysis, age under 10, no tobacco use, previous diagnosis of COVID-19, a history of COVID-19 related symptoms and contact with a COVID-19 case within the household, were independently associated with higher SARS-CoV-2 seroprevalence. More than one third of people living in Tunis obtained antibodies to SARS-CoV-2. Further studies are needed to monitor changes in these figures as Tunisian population is confronted to the subsequent epidemic waves and to guide the vaccine strategy.

The aim of this study was to measure the extent of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection among workers at the Institut Pasteur de Tunis (IPT), a public health laboratory involved in the management of the COVID-19 pandemic in Tunisia, and to identify risk factors for infection in this occupational setting. A cross-sectional survey was conducted on IPT workers not vaccinated against coronavirus disease 2019 (COVID-19). Participants completed a questionnaire that included a history of reverse transcription-polymerase chain reaction (RT-PCR)-confirmed SARS-CoV-2 infection. Immunoglobulin G antibodies against the receptor-binding domain of the spike antigen (anti-S-RBD IgG) and the nucleocapsid protein (anti-N IgG) of the SARS-CoV-2 virus were detected by enzyme-linked immunoassay (ELISA). A multivariate analysis was used to identify factors significantly associated with SARS-CoV-2 infection. A total of 428 workers were enrolled in the study. The prevalence of anti-S-RBD and/or anti-N IgG antibodies was 32.9% [28.7–37.4]. The cumulative incidence of SARS-CoV-2 infection (positive serology and/or previous positive RT-PCR test) was 40.0% [35.5–44.9], while the proportion with asymptomatic infection was 32.9%. One-third of the participants with RT-PCR-confirmed infection tested seronegative more than 90 days postinfection. Participants aged over 40 and laborers were more susceptible to infection (adjusted OR [AOR] = 1.65 [1.08–2.51] and AOR = 2.67 [1.45–4.89], respectively), while tobacco smokers had a lower risk of infection (AOR = 0.54 [0.29–0.97]). The SARS-CoV-2 infection rate among IPT workers was not significantly different from that detected concurrently in the general population. Hence, the professional activities conducted in this public health laboratory did not generate additional risk to that incurred outside the institute in day-to-day activities.

Background: Because the regular vaccine campaign started in Guinea one year after the COVID-19 index case, the profile of naturally acquired immunity following primary SARS-CoV-2 infection needs to be deepened. Methods: Blood samples were collected once from 200 patients (90% of African extraction) who were recovered from COVID-19 for at least ~2.4 months (72 days), and their sera were tested for IgG antibodies to SARS-CoV-2 using an in-house ELISA assay against the Receptor Binding Domain (RBD) of the SARS-CoV-2 spike1 protein (RBD/S1-IH kit). Results: Results revealed that 73% of sera (146/200) were positive for IgG to SARS-CoV-2 with an Optical Density (OD) ranging from 0.13 to 1.19 and a median value of 0.56 (IC95: 0.51–0.61). The median OD value at 3 months (1.040) suddenly decreased thereafter and remained stable around OD 0.5 until 15 months post-infection. The OD median value was slightly higher in males compared to females (0.62 vs. 0.49), but the difference was not statistically significant (p-value: 0.073). In contrast, the OD median value was significantly higher among the 60–100 age group (0.87) compared to other groups, with a noteworthy odds ratio compared to the 0–20 age group (OR: 9.69, p-value: 0.044*). Results from the RBD/S1-IH ELISA kit demonstrated superior concordance with the whole spike1 protein ELISA commercial kit compared to a nucleoprotein ELISA commercial kit. Furthermore, anti-spike1 protein ELISAs (whole spike1 and RBD/S1) revealed higher seropositivity rates. Conclusions: These findings underscore the necessity for additional insights into naturally acquired immunity against COVID-19 and emphasize the relevance of specific ELISA kits for accurate seropositivity rates

Understanding mechanisms of pathogenesis and protection in human tuberculosis (TB) remain major global health challenges. While organized granulomas have long been the focus of TB research, growing evidence for asymptomatic transmission highlights the need to study earlier disease stages, particularly TB pneumonia, which remains underexplored. Defining the alveolar immune niche that governs bacillary expansion before granuloma formation is essential for interrupting transmission. Here, we integrate spatial transcriptomics, single cell RNA sequencing, and high resolution imaging of human lung biopsies to map early TB pneumonia and compare with adjacent granulomas within the same tissues. Pneumonic alveolar spaces were dominated by TREM2-associated macrophages, characterized by sparse T cell infiltration, minimal antimicrobial gene expression, and abundant antigens and transcripts. In contrast, granuloma cores were enriched for antimicrobial pathways, were surrounded by multiple cell types that walled off infection, and contained comparatively fewer bacterial markers. Our findings identify TREM2 positive 'foamy' macrophages as a key permissive alveolar niche for survival and growth. These cells represent an attractive target for early intervention to restrict infection and limit transmission. Early TB pneumonia defines an alveolar niche that fosters bacterial persistence and transmission before granuloma formation.

Tuberculosis (TB) remains a major global health challenge. While organized granulomas have long been the focus of TB pathogenesis research, the early development of TB pneumonia typically preceding granuloma formation has been underexplored. Using spatial transcriptomics, high-resolution proteomics, and scRNA-seq on human pulmonary TB lesions, we reveal a striking compartmentalization of immune responses between early pneumonia and mature granulomas. The immunologic composition of granulomas was distinct from the pneumonia; granulomas are enriched for antimicrobial gene expression in both macrophages and T cells and show reduced bacterial antigen burden. In contrast, TREM2-expressing foamy macrophages are the predominant cell type occupying alveolar spaces in TB pneumonia with T cells infrequent. These TREM2⁺ macrophages exhibit a lipid-associated gene program, accumulate lipid droplets, and harbor Mycobacterium tuberculosis antigens and mRNA corresponding to increased bacterial viability in vitro. We further show that the M. tuberculosis virulence lipids, PDIM and mycolic acids, potently induce and activate TREM2 signaling in TREM2-expressing macrophages, promoting an intracellular environment permissive for bacterial growth. These findings establish TREM2⁺ macrophages as an early niche for M. tuberculosis survival and implicate TB pneumonia as a critical stage in disease transmission. Targeting this foamy macrophage population may offer opportunities to interrupt early TB progression and transmission.Tuberculosis (TB) remains a major global health challenge. While organized granulomas have long been the focus of TB pathogenesis research, the early development of TB pneumonia typically preceding granuloma formation has been underexplored. Using spatial transcriptomics, high-resolution proteomics, and scRNA-seq on human pulmonary TB lesions, we reveal a striking compartmentalization of immune responses between early pneumonia and mature granulomas. The immunologic composition of granulomas was distinct from the pneumonia; granulomas are enriched for antimicrobial gene expression in both macrophages and T cells and show reduced bacterial antigen burden. In contrast, TREM2-expressing foamy macrophages are the predominant cell type occupying alveolar spaces in TB pneumonia with T cells infrequent. These TREM2⁺ macrophages exhibit a lipid-associated gene program, accumulate lipid droplets, and harbor Mycobacterium tuberculosis antigens and mRNA corresponding to increased bacterial viability in vitro. We further show that the M. tuberculosis virulence lipids, PDIM and mycolic acids, potently induce and activate TREM2 signaling in TREM2-expressing macrophages, promoting an intracellular environment permissive for bacterial growth. These findings establish TREM2⁺ macrophages as an early niche for M. tuberculosis survival and implicate TB pneumonia as a critical stage in disease transmission. Targeting this foamy macrophage population may offer opportunities to interrupt early TB progression and transmission.