Differential patterns of antibody response against SARS-CoV-2 nucleocapsid epitopes detected in sera from patients in acute phase of COVID-19, convalescents and pre-pandemic individuals.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have already infected more than 0.7 billion people and caused over 7 million deaths worldwide. At the same time, our knowledge about this virus is still incipient. In some cases, there is a pre-pandemic immunity, however its source is unknown. The analysis of patients' humoral responses might shed a light on this puzzle. In this paper, we evaluated the antibody recognition of nucleocapsid protein, one of the structural proteins of SARS-CoV-2. For this purpose, we used pre-pandemic, acute COVID-19 and convalescent patients' sera to identify and map nucleocapsid protein epitopes. We identified a common epitope KKSAAEASKKPRQKRTATKA recognized by sera antibodies from all three groups. Some motifs of this sequence are widespread among various coronaviruses, plant or human proteins indicating that there might be more sources of nucleocapsid-reactive antibodies than previous infection with seasonal coronavirus. The two sequences MSDNGPQNQRNAPRITFGGP and KADETQALPQRQKKQQTVTL were detected as specific for sera from patients in acute phase of infection and convalescents making them suitable for future development of vaccine against SARS-CoV-2. Knowledge of the humoral response to SARS-CoV-2 infection is essential for the design of appropriate diagnostic tools and vaccine antigens.

Angiotensin-Converting Enzyme-2 (ACE2) Downregulation During Coronavirus Infection.

Angiotensin-converting enzyme-2 (ACE2) downregulation represents a detrimental factor in people with a baseline ACE2 deficiency associated with older age, hypertension, diabetes, and cardiovascular diseases. Human coronaviruses, including HCoV-NL63, SARS-CoV-1, and SARS CoV-2 infect target cells via binding of viral spike (S) glycoprotein to the ACE2, resulting in ACE2 downregulation through yet unidentified mechanisms. This downregulation disrupts the enzymatic activity of ACE2, essential in protecting against organ injury by cleaving and disposing of Angiotensin-II (Ang II), leading to the formation of Ang 1-7, thereby exacerbating the accumulation of Ang II. This accumulation activates the Angiotensin II type 1 receptor (AT1R) receptor, leading to leukocyte recruitment and increased proinflammatory cytokines, contributing to organ injury. The biological impacts and underlying mechanisms of ACE2 downregulation during SARS-CoV-2 infection have not been well defined. Therefore, there is an urgent need to establish a solid theoretical and experimental understanding of the mechanisms of ACE2 downregulation during SARS-CoV-2 entry and replication in the host cells. This review aims to discuss the physiological impact of ACE2 downregulation during coronavirus infection, the relationship between ACE2 decline and virus pathogenicity, and the possible mechanisms of ACE2 degradation, along with the therapeutic approaches.

A Gaussia luciferase reporter assay for the evaluation of coronavirus Nsp5/3CLpro activity.

Human coronaviruses (hCoVs) infect millions of people every year. Among these, MERS, SARS-CoV-1, and SARS-CoV-2 caused significant morbidity and mortality and their emergence highlights the risk of possible future coronavirus outbreaks. Therefore, broadly-active anti-coronavirus drugs are needed. Pharmacological inhibition of the hCoV protease Nsp5 (3CLpro) is clinically beneficial as shown by the wide and effective use of Paxlovid (nirmatrelvir, ritonavir). However, further treatment options are required due to the risk of drug resistance. To facilitate the assessment of coronavirus protease function and its pharmacological inhibition, we developed an assay allowing rapid and reliable quantification of Nsp5 activity under biosafety level 1 conditions. It is based on an ACE2-Gal4 transcription factor fusion protein separated by a Nsp5 recognition site. Cleavage by Nsp5 releases the Gal4 transcription factor, which then induces the expression of Gaussia luciferase. Our assay is compatible with Nsp5 proteases from all hCoVs and allows simultaneous measurement of inhibitory and cytotoxic effects of the tested compounds. Proof-of-concept measurements confirmed that nirmatrelvir, GC376 and lopinavir inhibit SARS-CoV-2 Nsp5 function. Furthermore, the assay accurately predicted the impact of Nsp5 mutations on catalytic activity and inhibitor sensitivity. Overall, the reporter assay is suitable for evaluating viral protease activity.

[Vaccinations for emerging and re-emerging viral diseases]. / Impfungen für neu- und wiederauftretende Viruserkrankungen.

BACKGROUND: Emerging or re-emerging viral diseases have a pandemic potential and threaten global health. Vaccination is of crucial importance in the prevention of emerging and re-emerging viral diseases. OBJECTIVE: Description of the current status of vaccine development against Filoviridae, highly pathogenic coronaviruses, smallpox viruses, influenza viruses and arboviruses. MATERIAL AND METHODS: Focused literature search. RESULTS: The World Health Organization (WHO) regularly publishes a list of infectious diseases that are expected to pose a major threat to humanity as they are could potentially trigger new pandemics; however, in addition to these human-to-human transmissible diseases, some arboviruses also have pandemic potential. In recent years numerous new vaccines, some of which are highly effective, have been licensed against new and re-emerging viral diseases and other promising vaccine candidates are currently in development. CONCLUSION: There are still gaps in the development of vaccines in the area of Filoviridae and highly pathogenic coronaviruses. Vaccinations against smallpox viruses have been available for a long time. Developing influenza vaccines against novel strains in a timely manner is a challenge and universal influenza vaccines could be a possible solution. Modern vaccines are available against the arboviruses dengue and Chikungunya fever.

Optimization of Cellular Transduction by the HIV-Based Pseudovirus Platform with Pan-Coronavirus Spike Proteins.

Over the past three years, new SARS-CoV-2 variants have continuously emerged, evolving to a point where an immune response against the original vaccine no longer provided optimal protection against these new strains. During this time, high-throughput neutralization assays based on pseudoviruses have become a valuable tool for assessing the efficacy of new vaccines, screening updated vaccine candidates against emerging variants, and testing the efficacy of new therapeutics such as monoclonal antibodies. Lentiviral vectors derived from HIV-1 are popular for developing pseudo and chimeric viruses due to their ease of use, stability, and long-term transgene expression. However, the HIV-based platform has lower transduction rates for pseudotyping coronavirus spike proteins than other pseudovirus platforms, necessitating more optimized methods. As the SARS-CoV-2 virus evolved, we produced over 18 variants of the spike protein for pseudotyping with an HIV-based vector, optimizing experimental parameters for their production and transduction. In this article, we present key parameters that were assessed to improve such technology, including (a) the timing and method of collection of pseudovirus supernatant; (b) the timing of host cell transduction; (c) cell culture media replenishment after pseudovirus adsorption; and (d) the centrifugation (spinoculation) parameters of the host cell+ pseudovirus mix, towards improved transduction. Additionally, we found that, for some pseudoviruses, the addition of a cationic polymer (polybrene) to the culture medium improved the transduction process. These findings were applicable across variant spike pseudoviruses that include not only SARS-CoV-2 variants, but also SARS, MERS, Alpha Coronavirus (NL-63), and bat-like coronaviruses. In summary, we present improvements in transduction efficiency, which can broaden the dynamic range of the pseudovirus titration and neutralization assays.

Combining Short- and Long-Read Sequencing Technologies to Identify SARS-CoV-2 Variants in Wastewater.

During the COVID-19 pandemic, the monitoring of SARS-CoV-2 RNA in wastewater was used to track the evolution and emergence of variant lineages and gauge infection levels in the community, informing appropriate public health responses without relying solely on clinical testing. As more sublineages were discovered, it increased the difficulty in identifying distinct variants in a mixed population sample, particularly those without a known lineage. Here, we compare the sequencing technology from Illumina and from Oxford Nanopore Technologies, in order to determine their efficacy at detecting variants of differing abundance, using 248 wastewater samples from various Quebec and Ontario cities. Our study used two analytical approaches to identify the main variants in the samples: the presence of signature and marker mutations and the co-occurrence of signature mutations within the same amplicon. We observed that each sequencing method detected certain variants at different frequencies as each method preferentially detects mutations of distinct variants. Illumina sequencing detected more mutations with a predominant lineage that is in low abundance across the population or unknown for that time period, while Nanopore sequencing had a higher detection rate of mutations that are predominantly found in the high abundance B.1.1.7 (Alpha) lineage as well as a higher sequencing rate of co-occurring mutations in the same amplicon. We present a workflow that integrates short-read and long-read sequencing to improve the detection of SARS-CoV-2 variant lineages in mixed population samples, such as wastewater.

Coronavirus Anatomy and Its Analytical Approaches for Targeting COVID-19.

The whole world has lived in fear of the Coronavirus 2019 (COVID-19) for more than one-fourth of a year in 2020. Since then, numerous studies have been done and others are still being conducted to identify the coronavirus causal agent and determine the ideal antiviral therapies for treating this illness. Despite the fact of significant global efforts, which are being made to develop vaccines and identify therapeutic medicines, the emergence of multiple variants has rumbled the research for developing an ideal diagnostic and therapeutic tool for targeting COVID-19. A thorough anatomy of the coronavirus virus and its global dissemination is essential for healthcare initiatives, as well as for predicting and preventing new epidemics. Testing for active COVID-19 infections is a critical public health strategy for tracking the epidemic. The widespread adoption of polymerase chain reaction (PCR) and antigen rapid test assays has increased the supply of test kits. In this chapter, the significance of molecular technique to develop prognostic analytical techniques for targeting COVID-19 along with the advantages and limitations of available techniques is discussed.

Discovery and characterization of a pan-betacoronavirus S2-binding antibody.

The continued emergence of deadly human coronaviruses from animal reservoirs highlights the need for pan-coronavirus interventions for effective pandemic preparedness. Here, using linking B cell receptor to antigen specificity through sequencing (LIBRA-seq), we report a panel of 50 coronavirus antibodies isolated from human B cells. Of these, 54043-5 was shown to bind the S2 subunit of spike proteins from alpha-, beta-, and deltacoronaviruses. A cryoelectron microscopy (cryo-EM) structure of 54043-5 bound to the prefusion S2 subunit of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike defined an epitope at the apex of S2 that is highly conserved among betacoronaviruses. Although non-neutralizing, 54043-5 induced Fc-dependent antiviral responses in vitro, including antibody-dependent cellular cytotoxicity (ADCC) and antibody-dependent cellular phagocytosis (ADCP). In murine SARS-CoV-2 challenge studies, protection against disease was observed after introduction of Leu234Ala, Leu235Ala, and Pro329Gly (LALA-PG) substitutions in the Fc region of 54043-5. Together, these data provide new insights into the protective mechanisms of non-neutralizing antibodies and define a broadly conserved epitope within the S2 subunit.

Large-scale deep learning identifies the antiviral potential of PKI-179 and MTI-31 against coronaviruses.

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has led to the global pandemic of Coronavirus Disease (2019) (COVID-19), underscoring the urgency for effective antiviral drugs. Despite the development of different vaccination strategies, the search for specific antiviral compounds remains crucial. Here, we combine machine learning (ML) techniques with in vitro validation to efficiently identify potential antiviral compounds. We overcome the limited amount of SARS-CoV-2 data available for ML using various techniques, supplemented with data from diverse biomedical assays, which enables end-to-end training of a deep neural network architecture. We use its predictions to identify and prioritize compounds for in vitro testing. Two top-hit compounds, PKI-179 and MTI-31, originally identified as Pi3K-mTORC1/2 pathway inhibitors, exhibit significant antiviral activity against SARS-CoV-2 at low micromolar doses. Notably, both compounds outperform the well-known mTOR inhibitor rapamycin. Furthermore, PKI-179 and MTI-31 demonstrate broad-spectrum antiviral activity against SARS-CoV-2 variants of concern and other coronaviruses. In a physiologically relevant model, both compounds show antiviral effects in primary human airway epithelial (HAE) cultures derived from healthy donors cultured in an air-liquid interface (ALI). This study highlights the potential of ML combined with in vitro testing to expedite drug discovery, emphasizing the adaptability of AI-driven approaches across different viruses, thereby contributing to pandemic preparedness.

High transmission of endemic human coronaviruses before and during the COVID-19 pandemic in adolescents in Cebu, Philippines.

BACKGROUND: SARS-CoV-2, the causative agent of COVID-19, is a betacoronavirus belonging to the same genus as endemic human coronaviruses (hCoVs) OC43 and HKU1 and is distinct from alpha hCoVs 229E and NL63. In a study of adolescents in the Philippines, we evaluated seroprevalence to the hCoVs, whether pre-pandemic hCoV immunity modulated subsequent risk of SARS-CoV-2 infection, and if SARS-CoV-2 infection affected the transmission of the hCoVs. METHODS: From 499 individuals screened in 2021 for SARS-CoV-2 receptor binding domain (RBD) antibodies by enzyme-linked immunosorbent assay (ELISA), we randomly selected 59 SARS-CoV-2 negative and 61 positive individuals for further serological evaluation. We measured RBD and spike antibodies to the four hCoVs and SARS-CoV-2 by ELISA in samples from the same participants collected pre-pandemic (2018-2019) and mid-pandemic (2021), before COVID-19 vaccination. RESULTS: We observed over 72% seropositivity to the four hCoVs pre-pandemic. Binding antibodies increased with age to 229E and OC43, suggesting endemic circulation, while antibody levels was flat across ages for HKU1 and NL63. During the COVID-19 pandemic, antibodies increased significantly to the RBDs of OC43, NL63, and 229E and spikes of all four hCoVs in both SARS-CoV-2 negative and positive adolescents. Those aged 12-15 years old in 2021 had higher antibodies to RBD and spike of OC43, NL63, and 229E than adolescents the same age in 2019, further demonstrating intense transmission of the hCoVs during the pandemic. CONCLUSIONS: We observe a limited impact of the COVID-19 pandemic on endemic hCoV transmission. This study provides insight into co-circulation of hCoVs and SARS-CoV-2.

GeneRaMeN enables integration, comparison, and meta-analysis of multiple ranked gene lists to identify consensus, unique, and correlated genes.

High-throughput experiments often produce ranked gene outputs, with forward genetic screening being a notable example. While there are various tools for analyzing individual datasets, those that perform comparative and meta-analytical examination of such ranked gene lists remain scarce. Here, we introduce Gene Rank Meta Analyzer (GeneRaMeN), an R Shiny tool utilizing rank statistics to facilitate the identification of consensus, unique, and correlated genes across multiple hit lists. We focused on two key topics to showcase GeneRaMeN: virus host factors and cancer dependencies. Using GeneRaMeN 'Rank Aggregation', we integrated 24 published and new flavivirus genetic screening datasets, including dengue, Japanese encephalitis, and Zika viruses. This meta-analysis yielded a consensus list of flavivirus host factors, elucidating the significant influence of cell line selection on screening outcomes. Similar analysis on 13 SARS-CoV-2 CRISPR screening datasets highlighted the pivotal role of meta-analysis in revealing redundant biological pathways exploited by the virus to enter human cells. Such redundancy was further underscored using GeneRaMeN's 'Rank Correlation', where a strong negative correlation was observed for host factors implicated in one entry pathway versus the alternate route. Utilizing GeneRaMeN's 'Rank Uniqueness', we analyzed human coronaviruses 229E, OC43, and SARS-CoV-2 datasets, identifying host factors uniquely associated with a defined subset of the screening datasets. Similar analyses were performed on over 1000 Cancer Dependency Map (DepMap) datasets spanning 19 human cancer types to reveal unique cancer vulnerabilities for each organ/tissue. GeneRaMeN, an efficient tool to integrate and maximize the usability of genetic screening datasets, is freely accessible via https://ysolab.shinyapps.io/GeneRaMeN.

Immune imprinting: The persisting influence of the first antigenic encounter with rapidly evolving viruses.

Immune imprinting is a phenomenon that stems from the fundamentals of immunological memory. Upon recurrent exposures to an evolving pathogen, the immune system must weigh the benefits of rapidly recalling established antibody repertoires with greater affinity to the initial variant or invest additional time and energy in producing de novo responses specific to the emerging variant. In this review, we delve into the mechanistic complexities of immune imprinting and its role in shaping subsequent immune responses, both de novo and recall, against rapidly evolving respiratory viruses such as influenza and coronaviruses. By exploring the duality of immune imprinting, we examine its potential to both enhance or hinder immune protection against disease, while emphasizing the role of host and viral factors. Finally, we explore how different vaccine platforms may affect immune imprinting and comment on vaccine strategies that can favor de novo variant-specific antibody responses.

Profiling of coronaviral Mpro and enteroviral 3Cpro specificity provides a framework for the development of broad-spectrum antiviral compounds.

The main protease from coronaviruses and the 3C protease from enteroviruses play a crucial role in processing viral polyproteins, making them attractive targets for the development of antiviral agents. In this study, we employed a combinatorial chemistry approach-HyCoSuL-to compare the substrate specificity profiles of the main and 3C proteases from alphacoronaviruses, betacoronaviruses, and enteroviruses. The obtained data demonstrate that coronavirus Mpros exhibit overlapping substrate specificity in all binding pockets, whereas the 3Cpro from enterovirus displays slightly different preferences toward natural and unnatural amino acids at the P4-P2 positions. However, chemical tools such as substrates, inhibitors, and activity-based probes developed for SARS-CoV-2 Mpro can be successfully applied to investigate the activity of the Mpro from other coronaviruses as well as the 3Cpro from enteroviruses. Our study provides a structural framework for the development of broad-spectrum antiviral compounds.

A filter pad design-based multiplexed lateral flow immunoassay for rapid simultaneous detection of PDCoV, TGEV, and PEDV in swine feces.

Swine Enteric Coronaviruses (SECoVs), with high lethality and infectiousness, are the main pathogens causing fatal and watery diarrhea in piglets and spreading globally. Moreover, these SECoVs can cause similar clinical manifestations and are often co-infected, requiring an accurate assay suitable for rapid, in situ, and differential detection. Here, we developed a multiplexed fluorescent-based lateral flow immunoassay (mFB-LFIA) for the detection of three SECoVs, including porcine delta coronaviruses (PDCoV), transmissible gastroenteritis virus (TGEV), and porcine epidemic diarrhea virus (PEDV), in swine fecal samples. Thanks to the filter pad design and reasonable optimization, the mFB-LFIA was achieved within 15 min for three SECoVs detection simultaneously and improved the tolerance of the strips for feces samples. The limit of detection (LoD) of detecting PDCoV, TGEV, and PEDV were 2.1 × 104 TCID50 mL-1, 3.4 × 102 TCID50 mL-1, and 3.6 × 102 TCID50 mL-1, respectively. Additionally, the proposed assay was successfully applied to the detection of PDCoV, TGEV, and PEDV in swine feces with high accuracy. Compared with the gold standard nucleic acid testing, the total coincidence rate of the proposed assay was more than 90 %. Moreover, the mFB-LFIA performed excellent stability and repeatability. The proposed mFB-LFIA allows for rapid, in situ, more cost-effective and simultaneous detection of PDCoV, TGEV, and PEDV compared with nucleic acid testing. To the best of our knowledge, this is the first report to describe a multiplexed point-of-care assay capable of detecting PDCoV, TGEV, and PEDV in swine fecal samples. We believe our approach has a great potential for application to pig farm.

Hyperforin, the major metabolite of St. John's wort, exhibits pan-coronavirus antiviral activity.

Introduction: The COVID-19 pandemic caused by the SARS-CoV-2 virus has underscored the urgent necessity for the development of antiviral compounds that can effectively target coronaviruses. In this study, we present the first evidence of the antiviral efficacy of hyperforin, a major metabolite of St. John's wort, for which safety and bioavailability in humans have already been established. Methods: Antiviral assays were conducted in cell culture with four human coronaviruses: three of high virulence, SARS-CoV-2, SARS-CoV, and MERS-CoV, and one causing mild symptoms, HCoV-229E. The antiviral activity was also evaluated in human primary airway epithelial cells. To ascertain the viral step inhibited by hyperforin, time-of-addition assays were conducted. Subsequently, a combination assay of hyperforin with remdesivir was performed. Results: The results demonstrated that hyperforin exhibited notable antiviral activity against the four tested human coronaviruses, with IC50 values spanning from 0.24 to 2.55 µM. Kinetic studies indicated that the observed activity occur at a post-entry step, potentially during replication. The antiviral efficacy of hyperforin was additionally corroborated in human primary airway epithelial cells. The results demonstrated a reduction in both intracellular and extracellular SARS-CoV-2 viral RNA, confirming that hyperforin targeted the replication step. Finally, an additive antiviral effect on SARS-CoV-2 was observed when hyperforin was combined with remdesivir. Discussion: In conclusion, hyperforin has been identified as a novel pan-coronavirus inhibitor with activity in human primary airway epithelial cells, a preclinical model for coronaviruses. These findings collectively suggest that hyperforin has potential as a candidate antiviral agent against current and future human coronaviruses.

Recent evolutionary origin and localized diversity hotspots of mammalian coronaviruses.

Several coronaviruses infect humans, with three, including the SARS-CoV2, causing diseases. While coronaviruses are especially prone to induce pandemics, we know little about their evolutionary history, host-to-host transmissions, and biogeography. One of the difficulties lies in dating the origination of the family, a particularly challenging task for RNA viruses in general. Previous cophylogenetic tests of virus-host associations, including in the Coronaviridae family, have suggested a virus-host codiversification history stretching many millions of years. Here, we establish a framework for robustly testing scenarios of ancient origination and codiversification versus recent origination and diversification by host switches. Applied to coronaviruses and their mammalian hosts, our results support a scenario of recent origination of coronaviruses in bats and diversification by host switches, with preferential host switches within mammalian orders. Hotspots of coronavirus diversity, concentrated in East Asia and Europe, are consistent with this scenario of relatively recent origination and localized host switches. Spillovers from bats to other species are rare, but have the highest probability to be towards humans than to any other mammal species, implicating humans as the evolutionary intermediate host. The high host-switching rates within orders, as well as between humans, domesticated mammals, and non-flying wild mammals, indicates the potential for rapid additional spreading of coronaviruses across the world. Our results suggest that the evolutionary history of extant mammalian coronaviruses is recent, and that cases of long-term virus-host codiversification have been largely over-estimated.

The SARS-CoV-2 virus, which caused the recent global coronavirus pandemic, is the latest in a string of coronaviruses that have caused serious outbreaks. This group of coronaviruses can also infect other mammals and likely jumped between species ­ including from non-humans to humans ­ over the course of evolution. Determining when and how viruses evolved to infect humans can help scientists predict and prevent outbreaks. However, tracking the evolutionary trajectory of coronaviruses is challenging, and there are conflicting views on how often coronaviruses crossed between species and when these transitions likely occurred. Some studies suggest that coronaviruses originated early on in evolution and evolved together with their mammalian hosts, only occasionally jumping to and from different species. While others suggest they appeared more recently, and rapidly diversified by regularly transferring between species. To determine which is the most likely scenario, Maestri, Perez-Lamarque et al. developed a computational approach using already available data on the genetics and evolutionary history of mammals and coronaviruses. This revealed that coronaviruses originated recently in bats from East Asia and Europe, and primarily evolved by rapidly transferring between different mammalian species. This has led to geographical hotspots of diverse coronaviruses in East Asia and Europe. Maestri, Perez-Lamarque et al. found that it was rare for coronaviruses to spill over from bats to other types of mammals. Most of these spillovers resulted from coronaviruses jumping from bats to humans or domesticated animals. Humans appeared to be the main intermediary host that coronaviruses temporarily infected as they transferred from bats to other mammals. These findings ­ that coronaviruses emerged recently in evolution, jumped relatively frequently between species, and are geographically restricted ­ suggest that future transmissions are likely. Gathering more coronavirus samples from across the world and using even more powerful analysis tools could help scientists understand more about how these viruses recently evolved. These insights may lead to strategies for preventing new coronaviruses from emerging and spreading among humans.

Respiratory Viral Infections from 2015 to 2022 in the HIVE Cohort of American Households: Incidence, illness characteristics, and seasonality.

BACKGROUND: Viral respiratory illnesses are the most common acute illnesses experienced and generally follow a predicted pattern over time. The SARS-CoV-2 pandemic interrupted that pattern. METHODS: The HIVE (Household Influenza Vaccine Evaluation) study was established in 2010 to follow a cohort of Southeast Michigan households over time. Initially focused on influenza, surveillance was expanded to include other major respiratory pathogens, and, starting in 2015, the population was followed year-round. Symptoms of acute illness were reported, and respiratory specimens were collected and tested to identify viral infections. Based on the known population being followed, virus-specific incidence was calculated. RESULTS: From 2015 to 2022, 1755 participants were followed in HIVE for 7785 person-years with 7833 illnesses documented. Before the pandemic, rhinovirus (RV) and common cold human coronaviruses (HCoVs) were the viruses most frequently identified, and incidence decreased with increasing age. Type A influenza was next but with comparable incidence by age. Parainfluenza and respiratory syncytial viruses were less frequent overall, followed by human metapneumoviruses. Incidence was highest in young children, but infections were frequently documented in all age groups. Seasonality followed patterns established decades ago. The SARS-CoV-2 pandemic disrupted these patterns, except for RV and, to a lesser extent, HCoVs. In the first two years of the pandemic, RV incidence far exceeded that of SARS-CoV-2. CONCLUSION: Longitudinal cohort studies are important in comparing the incidence, seasonality, and characteristics of different respiratory viral infections. Studies documented the differential effect of the pandemic on the incidence of respiratory viruses in addition to SARS-CoV-2.

COVID-19: did the masks work?

During the early stages of the COVID-19 pandemic, we called for mandatory public masking to 'flatten the curve'. We helped formulate a national standard (SWiFT 19) for barrier facemasks, and, using a novel laser-based approach, we determined that mask efficacy is dependent on both fabric and fit; with both variables being inversely related. Herein, we take a retrospective view of the role of masks during the pandemic and surmise that, on the balance of evidence to date, masks were effective at stemming the spread of SARS-CoV-2 and may well be an effective early control strategy for potential future respiratory pandemics.

Face coverings, which cover the nose and mouth, are a means of preventing infections that travel in the air. These include viruses such as SARS-CoV-2, which causes COVID-19. Face coverings, or masks, played a key role during the COVID-19 pandemic by reducing person-to-person spread of the virus. The key features of a mask that make it effective are the material from which it is made and how closely the mask fits the face. A loosely fitting mask, for example, will lead to gaps around the nose and cheeks through which droplets can escape. A better fitting mask will have less leakage. Masks made from light single-layer material is less able to prevent droplet penetration than thicker, multi-layered fabric. Properly fashioned and fitted face masks are an effective means of slowing the spread of infections that travel in the air.

Prevalence of Respiratory Pathogens in Nasopharyngeal Swabs of Febrile Patients with or without Respiratory Symptoms in the Niakhar Area of Rural Senegal.

Acute respiratory tract infections are one of the leading causes of morbidity and mortality worldwide. More data are needed on circulating respiratory microorganisms in different geographical areas and ecosystems. We analyzed nasopharyngeal swabs from 500 febrile patients living in the Niakhar area (Senegal), using FTDTM multiplex qPCR and simplex qPCR to target a panel of 25 microorganisms. We detected at least one microorganism for 366/500 patients (73.2%), at least one virus for 193/500 (38.6%), and at least one bacterium for 324/500 (64.8%). The most frequently detected microorganisms were Streptococcus pneumoniae (36.8%), Haemophilus influenzae (35.8%), adenovirus (11.8%), influenza viruses (6.4%), rhinovirus (5.0%), SARS-CoV-2 (4.0%), and RSV (4.0%). The main microorganisms significantly associated with respiratory symptoms, with a p-value ≤ 0.05, were influenza virus (11.9% in patients with respiratory symptoms versus 2.9% in patients without), RSV (6.5% versus 2.6%), metapneumovirus (5.4% versus 1.3%), HPIVs (7.6% versus 1.0%), S. pneumoniae (51.9% versus 28.0%), and H. influenzae (54.6% versus 24.5%). Co-infections were significantly associated with respiratory symptoms (65.4% versus 32.9%). All the epidemiological data show a high level of circulation of respiratory pathogens among febrile patients, including those preventable by vaccination such as S. pneumoniae, raising the question of the serotypes currently circulating. Furthermore, the availability of affordable real-time etiological diagnostic tools would enable management to be adapted as effectively as possible.

Identification of novel and potent inhibitors of SARS-CoV-2 main protease from DNA-encoded chemical libraries.

In vitro screening of large compound libraries with automated high-throughput screening is expensive and time-consuming and requires dedicated infrastructures. Conversely, the selection of DNA-encoded chemical libraries (DECLs) can be rapidly performed with routine equipment available in most laboratories. In this study, we identified novel inhibitors of SARS-CoV-2 main protease (Mpro) through the affinity-based selection of the DELopen library (open access for academics), containing 4.2 billion compounds. The identified inhibitors were peptide-like compounds containing an N-terminal electrophilic group able to form a covalent bond with the nucleophilic Cys145 of Mpro, as confirmed by x-ray crystallography. This DECL selection campaign enabled the discovery of the unoptimized compound SLL11 (IC50 = 30 nM), proving that the rapid exploration of large chemical spaces enabled by DECL technology allows for the direct identification of potent inhibitors avoiding several rounds of iterative medicinal chemistry. As demonstrated further by x-ray crystallography, SLL11 was found to adopt a highly unique U-shaped binding conformation, which allows the N-terminal electrophilic group to loop back to the S1' subsite while the C-terminal amino acid sits in the S1 subsite. MP1, a close analog of SLL11, showed antiviral activity against SARS-CoV-2 in the low micromolar range when tested in Caco-2 and Calu-3 (EC50 = 2.3 µM) cell lines. As peptide-like compounds can suffer from low cell permeability and metabolic stability, the cyclization of the compounds will be explored in the future to improve their antiviral activity.