La maladie à coronavirus (COVID-19) est une infection virale hautement contagieuse causée par le virus SRAS-CoV-2. La maladie s’est rapidement propagée entrainant une épidémie mondiale, causant la mort des personnes à risque. De nombreuses mesures sanitaires ont été prises durant ces deux dernières années, cependant la maladie n’est pas encore éradiquée. Considérant l’importance du système immunitaire dans le contrôle des maladies infectieuses et dans l’induction d’une mémoire immunitaire de longue durée, le SRAS-CoV-2 pourrait être une cible du système immunitaire, en particulier du système adaptatif représenté par les lymphocytes B et T. La réponse humorale induite chez les personnes convalescentes est largement perçue comme étant efficace pour combattre le virus. Par conséquent, toutes les plateformes de vaccination actuelles se sont basées sur la réponse humorale induite, particulièrement par la protéine SPIKE du virus. Compte tenu de la variabilité des réponses humorales ainsi que leur déclin rapide observé chez certains patients, et l’émergence de variants, il est nécessaire d’inclure d’autres stratégies permettant de renforcer la réponse immunitaire. Nous avons émis l’hypothèse qu’une réponse cellulaire pouvait être induite par les lymphocytes T contre les différentes protéines virales du SRAS-CoV-2. Dans cette étude nous avons détecté la réactivité des lymphocytes T contre les protéines virales (SPIKE, membrane, nucléocapside) par ELISPOT chez les patients convalescents. Nous avons ensuite identifié plus précisément les séquences riches en épitopes au niveau de la protéine membranaire et de la nucléocapside en préparant des délétants successifs (approche « mRNA PCR-based epitope chase technique” ou mPec). La validation de ces séquences a été confirmée par l’utilisation de banques de peptides dans des zones définies, se chevauchant de 12 acides aminés. En vue d’inclure ces régions dans une stratégie vaccinale future, une protéine de fusion MN contenant les séquences riches en épitopes a été créée et la réactivité des lymphocytes T contre la protéine MN a été évaluée de nouveau par ELISPOT. Nos données démontrent une réponse T dirigée principalement contre la nucléocapside et la protéine membranaire. Trois zones immunogènes sont identifiées au niveau de la nucléocapside et une zone au niveau de la membrane. La réactivité observée contre la protéine de fusion MN souligne le potentiel pouvoir immunogène des protéines (membrane et nucléocapside) et l’importance d’inclure ces protéines dans une stratégie vaccinale future afin de solliciter une réponse immunitaire cellulaire protectrice. / Coronavirus disease (COVID-19) is a highly contagious viral infection caused by the SARS-CoV-2 virus. The disease has spread rapidly resulting in a worldwide epidemic, causing death in those at risk. Numerous health measures have been taken over the past two years, but the disease has not yet been eradicated. Considering the importance of the immune system in the control of infectious diseases and in the induction of a long-lasting immune memory, SARS-CoV-2 could be a target of the immune system, in particular of the adaptive system represented by B and T lymphocytes. The humoral response induced in convalescent individuals is widely used as an effective therapy to combat the virus. Therefore, all current vaccination platforms have relied on the humoral response induced particularly by the virus' SPIKE protein. Given the variability of humoral responses and their rapid decline in some patients, and the emergence of variants, it is necessary to include other strategies to enhance the immune response. We hypothesized that a cellular response could be induced by T cells against the different viral proteins of SARS-CoV-2. In this study, we detected the reactivity of T cells against viral proteins (SPIKE, membrane, nucleocapsid) by ELISPOT in convalescent patients. We then identified more precisely the immunogenic sequences at the level of the membrane protein and the nucleocapsid by successive deletants using the mRNA PCR-based epitope chase technique (mPec). The validation of these sequences was confirmed by the use of small peptide libraries (overlapping by 12 amino acids), covering defined sequences rich in epitopes. In order to include these regions in a future vaccine strategy, a MN fusion protein containing the selected sequences was created and T cell reactivity against the MN protein was further confirmed by ELISPOT. Our data show a T-cell response primarily against the nucleocapsid and membrane protein. Three immunogenic zones were identified at the nucleocapsid level and one zone at the membrane level. The reactivity observed against the MN fusion protein cassette rich in epitopes, highlights the potential immunogenicity of the proteins (membrane and nucleocapsid) and the importance of including these proteins in a future vaccine strategy.

Background: Vaccination against SARS-CoV-2 significantly reduces the transmissibility of the virus and the likelihood of a severe course of COVID-19, and is thus a critical component in overcoming the current pandemic. The factors associated with adverse reactions after vaccination against SARS-CoV-2 have not yet been sufficiently evaluated. Methods: We used the Disease Analyzer database (IQVIA) to identify 531,468 individuals who received a total of 908,869 SARS-CoV-2 vaccinations in 827 general practices in Germany between April and September 2021. Cox regression models were used to analyze the frequency of vaccination-related side effects reported within 14 days after SARS-CoV-2 vaccination, as well as subjects’ demographic characteristics and comorbidities. Results: The total number of side effects documented was 28,287 (3.1% of all vaccinations). Pain in the limb (24.3%), fatigue (21.0%), dizziness (17.9%), joint pain (15.7%), fever (9.5%), nausea (7.5%), and myalgia (6.4%) were the most common side effects documented among the 12,575 vaccinations with definite side effects. In the multivariate regression analysis, young age was associated with much higher odds of reported side effects (OR18–30 years: 4.45, OR31–40 years: 3.50, OR41–50 years: 2.89). In addition, pre-existing comorbidities such as dementia (OR: 1.54), somatoform disorder (OR: 1.53), anxiety disorder (OR: 1.43), depression (OR: 1.37), chronic respiratory tract disease (OR: 1.27), hypertension (OR: 1.20), and obesity (1.14) significantly increased the odds of side effects. Finally, the male sex was associated with increased odds of reported side effects (OR: 1.17). Conclusion: Our study, based on a large outpatient database from Germany, identified young age, male sex, and pre-existing comorbidities such as dementia, somatoform disorders, anxiety disorders, and depression as factors associated with vaccine-related adverse events diagnosed in GP practices. These data could help to identify subgroups needing particular advice and care in the context of SARS-CoV-2 vaccinations.

Introduction: Community mitigation strategies (CMS) have demonstrated to be effective in the reduction of transmission and incidence of COVID-19, especially in the population with symptoms associated with the disease. The aim of this study was to evaluate the association between the presence of COVID-19 symptoms and adherence to CMS in Latin American adults. Methods: We carried out a secondary analysis of a database developed by the University of Maryland and Facebook social network during the COVID-19 pandemic. We included Latin American adults that used the Facebook platform and participated in a survey conducted from April 23 to May 23, 2020. The principal outcome variable was reported compliance with the three main CMS (physical distancing, use of face masks, and hand washing). The exposure variable included symptoms suspicious for COVID-19 defined as the presence of three or more symptoms of an acute clinical case of COVID-19. We performed generalized linear models of the Poisson family with a logarithmic link function to evaluate the association between the presence of COVID-19 symptoms and reported compliance with CMS. We calculated crude and adjusted prevalence ratios (PR) with their 95% confidence intervals (95%CI). Results: We analyzed 1,310,690 adults from Latin America; 48.1% were male and 42.9% were under 35 years of age. The prevalence of suspicious symptoms of COVID-19 was 18.5% and reported compliance with the three CMS was 45.3%. The countries with the highest proportion of reported compliance with the three CMS were Peru, Bolivia and Panama, while those with the lowest reported compliance were Costa Rica, Nicaragua and Honduras. In the adjusted model, people with suspicious symptoms for COVID-19 had a 14% lower compliance with the three CMS (aPR = 0.86; 95%CI: 0.85–0.87; p < 0.001). Conclusions: Less than half of the participants complied with the CMS, and those presenting suspicious symptoms for COVID-19 had lower reported compliance with the three CMS. / University of Maryland / Revisión por pares

PURPOSE OF REVIEW: This review article aims to explore the GI changes induced by SARS-CoV-2 and how gut microbial homeostasis can influence these changes and affect the lung-gut axis and its relationship with the induction of the cytokine release syndrome in severe COVID-19 patients. RECENT FINDINGS: Coronavirus disease 2019 (COVID-19) affects not only the respiratory system but can produce multi-systemic damage. The expression of angiotensin-converting enzyme 2 (ACE-2) receptors in the gastrointestinal (GI) tract, the high prevalence of GI symptoms in severely ill COVID-19 patients, and the abnormalities described in the gut microbiome in these patients have raised concerns about the influence of GI tract as a risk factor or as a potential modulator to reduce the severity of COVID-19. Understanding the mechanisms by which gut dysbiosis may influence viral transmission and disease progression in COVID-19 may help in shaping how accessible therapies, like diet modulation, can potentially help beat the devastating consequences of COVID-19.

BACKGROUND: Telepresence robots provide real-time audio, video, and mobility features, allowing faculty and students to engage in learning experiences without being physically present. PROBLEM: With multiple students and faculty members needing to quarantine due to the COVID-19 pandemic, a flexible learning environment was essential. APPROACH: The telepresence robots were used as an innovative approach for both faculty and students to engage in learning experiences offered in a variety of settings. OUTCOME: Feedback was obtained from faculty and students about the use of and satisfaction with telepresence robots. The robots were easy to use and posed only a few technological challenges, which were easily overcome. CONCLUSIONS: Telepresence robots were effective tools in overcoming teaching and learning barriers caused by the COVID-19 pandemic. The telepresence robots have many applications, including use in clinical and community settings.

During the past three years, we assisted to the rise of a new pathogen that afflicted the world with a global pandemic. Working in an era of rapid change has posed important challenges and the focus of research has shifted more and more toward topics of greater social utility. However, this period has also brought a new role for wastewater highlighting how it can provide insight into the health of a community. This is the approach of Wastewater-based Epidemiology (WBE). The work presented here aimed to deepen this approach not only at the theoretical level but also contributing with an ongoing monitoring of about 30 months. The main objectives were (i) to collect information on the recently discovered SARS-CoV-2 virus, its biology, transmission mechanism, and role in wastewater treatment plants (WWTP); (ii) set up a surveillance system that would allow to monitor SARS-CoV-2 infections over time, obtaining early information on its spread among the population to support the Health Authority. Starting from a detailed study of the shedding mechanisms of SARS-CoV-2 in the feces of infected patients, we moved on to the evaluation of the viral concentrations in the sewage system and the wastewater entering the WWTP. The possibility of a faecal-oral transmission route of the virus was investigated by evaluating the data about viability and infectivity in wastewater. The natural processes of decay of the virus in wastewater and the reduction of its concentration in the different treatment stages of WWTPs were explored in literature and with experimental data. At the same time, we developed a SARS-CoV-2 surveillance system in wastewater by applying different detection methods. Some practical and scientific aspects of the analysis protocol have been studied in depth such as the choice of the type of sample, the storage temperatures, and the pre-heat treatments aimed at making the analysis safer for the operator. The choice of the concentration method was evaluated to comply with the low concentration of the viral titer and therefore the crucial importance of this phase of the protocol. During the monitoring campaign, we further investigated aspects related to data processing and developed normalization approaches. Samples from WWTPs in the province of Trento were analysed weekly and sampling frequencies and curve smoothing methods deriving from those data were evaluated. The trend curves thus obtained were compared with those deriving from clinical data provided by the local Health Authority and signals of early warnings of virus diffusion trends in the population were highlighted. With the alternation of the different variants of the virus and the evidence of their importance in the development of new waves of infection, a PCR based genotyping method has been devised to rapidly identify the already known variants. In conclusion, this research project addressed a broad spectrum of aspects related to the WBE approach in contrasting the COVID-19 emergency and confirmed that wastewater could be a valuable source of information and management support for this and other emerging pathogens or micropollutants.

<p dir="ltr">The recent global pandemic, COVID-19, has revealed to all the importance of understanding the complex relationship between viruses and hosts. Before COVID-19, I started my study of viral protein-host lipid interactions in the hemorrhagic fevers Ebola and Marburg viruses. These viruses contain a matrix protein that interacts with the plasma membrane to facilitate the formation of both authentic viruses and virus-like particles. My goal was to understand the limitations of their specific host lipid interactions. However, when the COVID-19 pandemic began, so to be our swift response in the development of a biosafety level 2 compatible model. This model can be used for studying severe acute respiratory distress syndrome 2 (SARS-CoV-2) assembly, egress, and entry. This model enabled exponentially greater access to more facilities to study the intricacies of SARS-CoV-2 assembly. With more access to studying the virus in a safe model, our goal is to push the understanding of viral assembly faster. I then began to take apart the individual pieces of the model and started to look at understanding the roles that they play independently. The membrane protein is the most abundant structural protein and I studied the specific lipid interactions of the soluble fraction of the protein. Physicians observed nucleocapsid protein mutations in the clinic with the increasing number of SARS-CoV-2 variants that are on the rise. The microscopy data collected can give us more insight into perhaps how the nucleocapsid protein induces the formation of filopodia structures at the plasma membrane. The envelope protein proved to be a challenge, but I determined a specific envelope and ceramide interaction in cells. The envelope protein was also causing the formation of microvesicles for an undefined function. I was able to determine the subcellular localization of the protein to the mitochondria. The localization to the mitochondria appears to induce depolarization of the mitochondria membrane action potential and induces the increase in mitochondria dysfunction signal, cytochrome c. Although the mitochondria were dysfunctional, there was no increase in apoptosis signal in the presence of the protein alone.</p>

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Enveloped RNA viruses like human immunodeficiency virus type-1 (HIV-1) and SARS-CoV-2 enter host cells through fusion with the plasma membrane, a process facilitated by specific viral envelope proteins that recognize and bind to receptors expressed on the host cell surface. These receptors can diverge based on the type of cell and virus. For HIV-1, the primary receptors on myeloid cells are CD4 and CCR5 or CXCR4. For SARS-CoV-2, although the primary receptor is ACE2, other myeloid-cell specific sialic acid binding lectins can also facilitate entry. Following cellular invasion, different viral RNA species can be detected by distinct host nucleic acid sensors, resulting in type I interferons and pro-inflammatory cytokine induction. While these innate immune responses are essential for controlling viral infections, overactivation can lead to chronic inflammation, tissue damage, and disease pathogenesis. Herein, I examine the contribution of HIV-1 and SARS-CoV-2 de-novo RNA expression and the molecular mechanisms that contribute to innate immune activation in myeloid cells. Despite advancements in combination antiretroviral therapy (ART) in suppressing systemic viral replication in individuals infected with HIV, residual viral RNA expression in tissue reservoirs remains a significant hindrance to curative efforts. I hypothesized that persistent expression of viral RNAs in myeloid cells triggers dysregulated innate immune activation, and inflammasomes activation. This study centers on the long-lived tissue-resident innate immune cells - macrophages and microglia, which, owing to their self-renewing nature, operate as reservoirs of viral RNA production, and are thought to lead to chronic immune activation even in the absence of productive replication. Our previous studies suggest that de novo expression of unspliced intron-containing HIV-1 RNA (herein referred to as icRNA) triggers activation of pro-inflammatory cytokines in myeloid cells. Here, I demonstrate that cytosolic expression of HIV-1 icRNA, but not multiply-spliced viral RNAs induces inflammasome activation, LDH release and IL-1β secretion in productively infected monocyte-derived macrophages (MDM) and induced pluripotent stem cell (iPSC)-derived microglia. Interestingly, knockdown of RLRs, RIG-I and MDA5 or endosomal TLRs failed to abrogate HIV-1 icRNA-induced IL-1β secretion. Rather, knockdown of NLRP1, but not NLRP3, inflammasome resulted in a significant reduction in IL-1β secretion, underscoring NLRP1's pivotal role in the HIV-1 icRNA-induced IL-1β secretion. Furthermore, Rev-Crm1-dependent nucleocytoplasmic export of HIV-1 icRNA was required for NLRP1-mediated Caspase-1 activation, IL-1β secretion, LDH release and cell death. Similarly, SARS-CoV-2, while not establishing productive infection in macrophages, can activate these cells, contributing to a hyper-inflammatory response marked by the heightened expression of pro-inflammatory cytokines, which is understood to be a principal driver of COVID-19 pathology. SARS-CoV-2 established an abortive infection in macrophages. CD169, a macrophage-specific sialic-acid binding lectin, mediated ACE2-independent SARS-CoV-2 entry in human macrophages and establishment of restricted infection. Interestingly, CD169-mediated SARS-CoV-2 entry in macrophages led to the expression of viral genomic and subgenomic RNAs, with negligible viral protein expression and no release of infectious virus particles, implying a post-entry restriction to SARS-CoV-2 replication in macrophages that was curbed by exogenous ACE2 expression. Despite restricted viral RNA expression, cytoplasmic RLRs, RIG-I and MDA5, sensed abortive viral transcripts, and induced pro-inflammatory responses in a MAVS dependent manner. This dissertation reveals striking parallels between the role of viral RNAs in driving pro-inflammatory responses in HIV-1 and SARS-CoV-2 infections. These findings collectively underscore the central role of cytoplasmic sensing of viral RNAs and their contribution to chronic inflammation in virus-infected myeloid cells. Elucidating these molecular mechanisms further may pave the way for novel therapeutic interventions to mitigate the persistent innate immune activation and immunopathology detected in HIV-1 and SARS-CoV-2 infected individuals.

Previous studies reported on the broad-spectrum antiviral function of heparin. Here we investigated the antiviral function of magnesium-modified heparin and found that modified heparin displayed a significantly enhanced antiviral function against human adenovirus (HAdV) in immortalized and primary cells. Nuclear magnetic resonance analyses revealed a conformational change of heparin when complexed with magnesium. To broadly explore this discovery, we tested the antiviral function of modified heparin against herpes simplex virus type 1 (HSV-1) and found that the replication of HSV-1 was even further decreased compared to aciclovir. Moreover, we investigated the antiviral effect against the new severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) and measured a 55-fold decreased viral load in the supernatant of infected cells associated with a 38-fold decrease in virus growth. The advantage of our modified heparin is an increased antiviral effect compared to regular heparin.