Antigen-Specific Antibody Signature Is Associated with COVID-19 Outcome.

Numerous studies have focused on inflammation-related markers to understand COVID-19. In this study, we performed a comparative analysis of spike (S) and nucleocapsid (N) protein-specific IgA, total IgG and IgG subclass response in COVID-19 patients and compared this to their disease outcome. We observed that the SARS-CoV-2 infection elicits a robust IgA and IgG response against the N-terminal (N1) and C-terminal (N3) region of the N protein, whereas we failed to detect IgA antibodies and observed a weak IgG response against the disordered linker region (N2) in COVID-19 patients. N and S protein-specific IgG1, IgG2 and IgG3 response was significantly elevated in hospitalized patients with severe disease compared to outpatients with non-severe disease. IgA and total IgG antibody reactivity gradually increased after the first week of symptoms. Magnitude of RBD-ACE2 blocking antibodies identified in a competitive assay and neutralizing antibodies detected by PRNT assay correlated with disease severity. Generally, the IgA and total IgG response between the discharged and deceased COVID-19 patients was similar. However, significant differences in the ratio of IgG subclass antibodies were observed between discharged and deceased patients, especially towards the disordered linker region of the N protein. Overall, SARS-CoV-2 infection is linked to an elevated blood antibody response in severe patients compared to non-severe patients. Monitoring of antigen-specific serological response could be an important tool to accompany disease progression and improve outcomes.

Heterogeneity of humoral response patterns in mildly symptomatic, non-hospitalized COVID-19 patients: A one-year longitudinal study.

The duration and protectiveness of antibodies against SARS-CoV-2 in infected subjects are still uncertain; nonetheless, anti-S-specific antibodies can contribute to protective immunity against new infections. It has been described that the level of antibodies produced in COVID-19 is related to the severity of symptoms, and the majority of the humoral response studies have been conducted in hospitalized patients who have been, then, followed over time. However, about 80% of SARS-CoV-2 infections in unvaccinated people are mild to asymptomatic, and this percentage reaches more than 95% in vaccinated individuals. Therefore, understanding the long-term dynamics of the antibody responses in this predominant part of the COVID-19-affected population is essential. In this study, we followed a cohort of individuals with mild COVID-19 who did not require hospitalization. We collected blood samples at sequential times after the SARS-CoV-2-positive qRT-PCR result. From 65 recruited patients, 50 had detectable antibodies at screening. Anti-SARS-CoV-2 IgM levels peaked around two weeks post-COVID-19 diagnostics, becoming undetectable after 65 days. IgG levels reached a peak in approximately one month and remained detectable for more than one year. In contrast to the levels of anti-SARS-CoV-2, antibody neutralization potency indexes persisted over time. In this study, humoral responses in mild COVID-19 patients persisted for more than one year. This is an important long-term follow-up study that includes responses from COVID-19 patients before and after vaccination, a scenery that has become increasingly difficult to evaluate due to the growing vaccination of the world human population.

Increased mTOR Signaling and Impaired Autophagic Flux Are Hallmarks of SARS-CoV-2 Infection.

The COVID-19 (Coronavirus Disease 2019), caused by the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), severely affects mainly individuals with pre-existing comorbidities. Here our aim was to correlate the mTOR (mammalian/mechanistic Target of Rapamycin) and autophagy pathways with the disease severity. Through western blotting and RNA analysis, we found increased mTOR signaling and suppression of genes related to autophagy, lysosome, and vesicle fusion in Vero E6 cells infected with SARS-CoV-2 as well as in transcriptomic data mining of bronchoalveolar epithelial cells from severe COVID-19 patients. Immunofluorescence co-localization assays also indicated that SARS-CoV-2 colocalizes within autophagosomes but not with a lysosomal marker. Our findings indicate that SARS-CoV-2 can benefit from compromised autophagic flux and inhibited exocytosis in individuals with chronic hyperactivation of mTOR signaling.

SARS-CoV-2 infected children form early immune memory responses dominated by nucleocapsid-specific CD8+ T cells and antibodies.

This is the third year of the SARS-CoV-2 pandemic, and yet most children remain unvaccinated. COVID-19 in children manifests as mostly mild or asymptomatic, however high viral titers and strong cellular and humoral responses are observed upon acute infection. It is still unclear how long these responses persist, and if they can protect from re-infection and/or disease severity. Here, we analyzed immune memory responses in a cohort of children and adults with COVID-19. Important differences between children and adults are evident in kinetics and profile of memory responses. Children develop early N-specific cytotoxic T cell responses, that rapidly expand and dominate their immune memory to the virus. Children's anti-N, but not anti-S, antibody titers increase over time. Neutralization titers correlate with N-specific antibodies and CD8+T cells. However, antibodies generated by infection do not efficiently cross-neutralize variants Gamma or Delta. Our results indicate that mechanisms that protect from disease severity are possibly different from those that protect from reinfection, bringing novel insights for pediatric vaccine design. They also underline the importance of vaccination in children, who remain at risk for COVID-19 despite having been previously infected.

Selection of plasma donors for the production of anti-SARS-CoV-2 immunoglobulin-based therapies: Strategies for quantitative antibody measurements.

Even after two years of the pandemic, a completely effective treatment against SARS-CoV-2 has not yet been established. Considering this fact and the emergence of successive new viral variants, the development of therapies based on natural polyclonal antibodies recovered from convalescent plasma remains relevant. This study presents a comparison between different methods of screening antibodies in samples of 41 individuals previously diagnosed with COVID-19. We found a significant correlation between Abbot Architect anti-SARS-CoV-2 IgG and Abbott Allinity SARS-CoV-2 IgG II Quantitative assay intensity of reactivity and neutralizing antibody (nAb) titers. Thus, we propose an initial antibody screening with IgG anti-N Abbott Architect test, with an index of, for example, > 3.25 or SARS-CoV-2 IgG II Quantitative Abbott Allinity assay > 137.65 AU/mL as good predictors of Nab ≥ 1:80. For the quantitative method, this threshold demonstrated a 100 % sensitivity and 80 % specificity, with 97.3 % accuracy. An interesting observation was the increase in the neutralizing activity of the anti-SARS-CoV-2 antibodies with the longest interval between the end of the symptoms and the collection, demonstrating that the delay in plasma collection does not affect the achievement of adequate nAbs levels. These results demonstrate the possibility of using faster and more widely available commercial serological tests with a good correlation with viral neutralization tests in culture, allowing for optimized large-scale donor selection, which will be of utmost importance for the development of therapies such as hyperimmune immunoglobulin.

UV 254 nm is more efficient than UV 222 nm in inactivating SARS-CoV-2 present in human saliva.

Ultraviolet (UV) light can inactivate SARS-CoV-2. However, the practicality of UV light is limited by the carcinogenic potential of mercury vapor-based UV lamps. Recent advances in the development of krypton chlorine (KrCl) excimer lamps hold promise, as these emit a shorter peak wavelength (222 nm), which is highly absorbed by the skin's stratum corneum and can filter out higher wavelengths. In this sense, UV 222 nm irradiation for the inactivation of virus particles in the air and surfaces is a potentially safer option as a germicidal technology. However, these same physical properties make it harder to reach microbes present in complex solutions, such as saliva, a critical source of SARS-CoV-2 transmission. We provide the first evaluation for using a commercial filtered KrCl excimer light source to inactivate SARS-CoV-2 in saliva spread on a surface. A conventional germicidal lamp (UV 254 nm) was also evaluated under the same condition. Using plaque-forming units (PFU) and Median Tissue Culture Infectious Dose (TCID50) per milliliter we found that 99.99% viral clearance (LD99.99) was obtained with 106.3 mJ/cm2 of UV 222 nm for virus in DMEM and 2417 mJ/cm2 for virus in saliva. Additionally, our results showed that the UV 254 nm had a greater capacity to inactivate the virus in both vehicles. Effective (after discounting light absorption) LD99.99 of UV 222 nm on the virus in saliva was ∼30 times higher than the value obtained with virus in saline solution (PBS), we speculated that saliva might be protecting the virus from surface irradiation in ways other than just by intensity attenuation of UV 222 nm. Due to differences between UV 222/254 nm capacities to interact and be absorbed by molecules in complex solutions, a higher dose of 222 nm will be necessary to reduce viral load in surfaces with contaminated saliva.

Clearance of Persistent SARS-CoV-2 RNA Detection in a NFκB-Deficient Patient in Association with the Ingestion of Human Breast Milk: A Case Report.

Currently, there are no evidence-based treatment options for long COVID-19, and it is known that SARS-CoV-2 can persist in part of the infected patients, especially those with immunosuppression. Since there is a robust secretion of SARS-CoV-2-specific highly-neutralizing IgA antibodies in breast milk, and because this immunoglobulin plays an essential role against respiratory virus infection in mucosa cells, being, in addition, more potent in neutralizing SARS-CoV-2 than IgG, here we report the clinical course of an NFκB-deficient patient chronically infected with the SARS-CoV-2 Gamma variant, who, after a non-full effective treatment with plasma infusion, received breast milk from a vaccinated mother by oral route as treatment for COVID-19. After such treatment, the symptoms improved, and the patient was systematically tested negative for SARS-CoV-2. Thus, we hypothesize that IgA and IgG secreted antibodies present in breast milk could be useful to treat persistent SARS-CoV-2 infection in immunodeficient patients.

Identification and characterization of the anti-SARS-CoV-2 activity of cationic amphiphilic steroidal compounds.

The ongoing COVID-19 pandemic caused a significant loss of human lives and a worldwide decline in quality of life. Treatment of COVID-19 patients is challenging, and specific treatments to reduce COVID-19 aggravation and mortality are still necessary. Here, we describe the discovery of a novel class of epiandrosterone steroidal compounds with cationic amphiphilic properties that present antiviral activity against SARS-CoV-2 in the low micromolar range. Compounds were identified in screening campaigns using a cytopathic effect-based assay in Vero CCL81 cells, followed by hit compound validation and characterization. Compounds LNB167 and LNB169 were selected due to their ability to reduce the levels of infectious viral progeny and viral RNA levels in Vero CCL81, HEK293, and HuH7.5 cell lines. Mechanistic studies in Vero CCL81 cells indicated that LNB167 and LNB169 inhibited the initial phase of viral replication through mechanisms involving modulation of membrane lipids and cholesterol in host cells. Selection of viral variants resistant to steroidal compound treatment revealed single mutations on transmembrane, lipid membrane-interacting Spike and Envelope proteins. Finally, in vivo testing using the hACE2 transgenic mouse model indicated that SARS-CoV-2 infection could not be ameliorated by LNB167 treatment. We conclude that anti-SARS-CoV-2 activities of steroidal compounds LNB167 and LNB169 are likely host-targeted, consistent with the properties of cationic amphiphilic compounds that modulate host cell lipid biology. Although effective in vitro, protective effects were cell-type specific and did not translate to protection in vivo, indicating that subversion of lipid membrane physiology is an important, yet complex mechanism involved in SARS-CoV-2 replication and pathogenesis.

Previous Infection with SARS-CoV-2 Correlates with Increased Protective Humoral Responses after a Single Dose of an Inactivated COVID-19 Vaccine.

Previous studies have indicated that antibody responses can be robustly induced after the vaccination in individuals previously infected by SARS-CoV-2. To evaluate anti-SARS-CoV-2 humoral responses in vaccinated individuals with or without a previous history of COVID-19, we compared levels of anti-SARS-CoV-2 antibodies in the sera from 21 vaccinees, including COVID-19-recovered or -naïve individuals in different times, before and after immunization with an inactivated COVID-19 vaccine. Anti-SARS-CoV-2-specific antibodies elicited after COVID-19 and/or immunization with an inactivated vaccine were measured by ELISA and Plaque Reduction Neutralizing assays. Antibody kinetics were consistently different between the two vaccine doses for naïve individuals, contrasting with the SARS-CoV-2-recovered subjects in which we observed no additional increase in antibody levels following the second dose. Sera from SARS-CoV2-naïve individuals had no detectable neutralizing activity against lineage B.1 SARS-CoV-2 or Gamma variant five months after the second vaccine dose. Contrarily, SARS-CoV-2-recovered subjects retained considerable neutralizing activity against both viruses. We conclude that a single inactivated SARS-CoV-2 vaccine dose may be sufficient to induce protective antibody responses in individuals with previous history of SARS-CoV-2 infection.

Clusters of SARS-CoV-2 Lineage B.1.1.7 Infection after Vaccination with Adenovirus-Vectored and Inactivated Vaccines.

A SARS-CoV-2 B.1.1.7 variant of concern (VOC) has been associated with increased transmissibility, hospitalization, and mortality. This study aimed to explore the factors associated with B.1.1.7 VOC infection in the context of vaccination. On March 2021, we detected SARS-CoV-2 RNA in nasopharyngeal samples from 14 of 22 individuals vaccinated with a single-dose of ChAdOx1 (outbreak A, n = 26), and 22 of 42 of individuals with two doses of the CoronaVac vaccine (outbreak B, n = 52) for breakthrough infection rates for ChAdOx1 of 63.6% and 52.4% for CoronaVac. The outbreaks were caused by two independent clusters of the B.1.1.7 VOC. The serum of PCR-positive symptomatic SARS-CoV-2-infected individuals had ~1.8-3.4-fold more neutralizing capacity against B.1.1.7 compared to the serum of asymptomatic individuals. These data based on exploratory analysis suggest that the B.1.1.7 variant can infect individuals partially immunized with a single dose of an adenovirus-vectored vaccine or fully immunized with two doses of an inactivated vaccine, although the vaccines were able to reduce the risk of severe disease and death caused by this VOC, even in the elderly.

Ultraviolet germicidal irradiation is effective against SARS-CoV-2 in contaminated makeup powder and lipstick.

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is mainly transmitted by airborne droplets generated by infected individuals. Since this and many other pathogens are able to remain viable on inert surfaces for extended periods of time, contaminated surfaces play an important role in SARS-CoV-2 fomite transmission. Cosmetic products are destined to be applied on infection-sensitive sites, such as the lips and eyelids. Therefore, special biosafety precautions should be incorporated into the routine procedures of beauty parlors and shops. Indeed, innovative cosmetics companies are currently searching for disinfection protocols that ensure the customers' safety in makeup testing. Here, we propose an ultraviolet germicidal irradiation (UVGI) strategy that can be used to reduce the odds of COVID-19 fomite transmission by makeup testers. It is well-known that UVGI effectively inactivates pathogens on flat surfaces and clear fluids. However, ultraviolet-C (UVC) radiation at 254 nm penetrates poorly in turbid and porous materials, such as makeup and lipstick formulations. Thus, we investigated the virucidal effect of UVGI against SARS-CoV-2 deposited on such substrates and compared their performance to that of flat polystyrene surfaces, used as controls. Concentrated infectious SARS-CoV-2 inoculum (106 PFU/mL) deposited on lipstick and makeup powder was completely inactivated (>5log10 reduction) following UVC exposures at 1,260 mJ/cm2, while flat plastic surfaces required 10 times less exposure (126 mJ/cm2) to reach the same microbicidal performance. We conclude that UVGI comprises an effective disinfection strategy to promote biosafety for cosmetics testers. However, appropriate UVC dosimetry must be implemented to overcome inefficiencies caused by the optical properties of turbid materials in lipsticks and makeup powders.

Identification of SARS-CoV-2 on the ocular surface in a cohort of COVID-19 patients from Brazil.

In this cross-sectional study, we investigate the presence of Severe Acute Respiratory Syndrome Coronavirus 2 Ribonucleic Acid (SARS-CoV-2 RNA) in the tears of hospitalized COVID-19 patients. After laboratory confirmation of SARS-CoV-2 infection by reverse transcription polymerase chain reaction (RT-PCR) analysis, tear samples from both eyes of each patient were collected using conjunctival swab for RT-PCR. Detailed demographic profile, systemic and ocular symptoms, comorbidities, clinical, ancillary, and ocular manifestations were evaluated. Of the 83 patients enrolled in the study, 7 (8.43%) had SARS-CoV-2 RNA detected in the tear samples. Neutrophils' count, C-reactive protein, and D-dimer were higher in patients with SARS-CoV-2 detected in tears than in patients without virus in ocular surface samples. One patient with SARS-CoV-2 in tears showed mild ocular eyelid edema, hyperemia, and chemosis. No relevant ocular manifestations were detected in the other patients. Although the levels of viral RNA on ocular surface samples were low for most patients (5/7), with positivity only for gene N and CT higher than 30, two patients were positive for all viral targets tested (N, E, and RpRd), with viral load near 1 × 105 ePFU/mL, indicating that the ocular transmission of SARS-CoV-2 is a possibility that needs to be considered, especially in the hospital environment. Further studies need to be conducted to demonstrate whether infective viral particles could be isolated from tears.

Respiratory Viral Shedding in Healthcare Workers Reinfected with SARS-CoV-2, Brazil, 2020.

We documented 4 cases of severe acute respiratory syndrome coronavirus 2 reinfection by non-variant of concern strains among healthcare workers in Campinas, Brazil. We isolated infectious particles from nasopharyngeal secretions during both infection episodes. Improved and continued protection measures are necessary to mitigate the risk for reinfection among healthcare workers.

Rapid clinical recovery of a SARS-CoV-2 infected common variable immunodeficiency patient following the infusion of COVID-19 convalescent plasma.

BACKGROUND: Common variable immunodeficiency is the most prevalent symptomatic primary immunodeficiency in adults. Affected patients fail to mount an appropriate humoral response against community acquired infectious diseases and recent reports have provided data supporting the increased susceptibility of these patients to severe SARS-CoV-2 infections. In this context, the infusion of COVID-19 convalescent plasma could represent an effective therapeutic strategy. CASE PRESENTATION: 25-year old woman diagnosed with common variable immunodeficiency in 2013, developed severe COVID-19 that rapidly progressed to pneumonia presenting with multiple bilateral lung opacities that were both central and peripheral and presented as ground-glass and consolidation types involving all lobes, bilaterally. As blood oxygen saturation decayed and lung abnormalities were not responsive to large spectrum antibiotics and corticosteroids, patient was placed on mechanical ventilation and compassionate-use of approved COVID-19 convalescent donor plasma was introduced. The patient presented a rapid response to the approach and mechanical ventilation could be interrupted 24 h after first dose of COVID-19 convalescent donor plasma. As a whole, the patient received four doses of 200 mL convalescent plasma during a period of 6 days. There was rapid improvement of clinical status, with interruption of supplemental oxygen therapy after 6 days and reduction of lung abnormalities as evidence by sequential computed tomography scans. CONCLUSIONS: This is a single patient report that adds to other few reports on common variable immunodeficiency and agammaglobulinemia, suggesting that COVID-19 convalescent donor plasma could be a valuable therapeutic approach to treat patients affected by dysgammaglobulinemias and presenting severe COVID-19.

Evolution and epidemic spread of SARS-CoV-2 in Brazil.

Brazil currently has one of the fastest-growing severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) epidemics in the world. Because of limited available data, assessments of the impact of nonpharmaceutical interventions (NPIs) on this virus spread remain challenging. Using a mobility-driven transmission model, we show that NPIs reduced the reproduction number from >3 to 1 to 1.6 in São Paulo and Rio de Janeiro. Sequencing of 427 new genomes and analysis of a geographically representative genomic dataset identified >100 international virus introductions in Brazil. We estimate that most (76%) of the Brazilian strains fell in three clades that were introduced from Europe between 22 February and 11 March 2020. During the early epidemic phase, we found that SARS-CoV-2 spread mostly locally and within state borders. After this period, despite sharp decreases in air travel, we estimated multiple exportations from large urban centers that coincided with a 25% increase in average traveled distances in national flights. This study sheds new light on the epidemic transmission and evolutionary trajectories of SARS-CoV-2 lineages in Brazil and provides evidence that current interventions remain insufficient to keep virus transmission under control in this country.