RESUMO
Hybrid immunity (vaccination + natural infection) to SARS-CoV-2 provides superior protection to re-infection. We performed immune profiling studies during breakthrough infections in mRNA-vaccinated hamsters to evaluate hybrid immunity induction. The mRNA vaccine, BNT162b2, was dosed to induce binding antibody titers against ancestral spike, but inefficient serum virus neutralization of ancestral SARS-CoV-2 or variants of concern (VoCs). Vaccination reduced morbidity and controlled lung virus titers for ancestral virus and Alpha but allowed breakthrough infections in Beta, Delta and Mu-challenged hamsters. Vaccination primed for T cell responses that were boosted by infection. Infection back-boosted neutralizing antibody responses against ancestral virus and VoCs. Hybrid immunity resulted in more cross-reactive sera, reflected by smaller antigenic cartography distances. Transcriptomics post-infection reflects both vaccination status and disease course and suggests a role for interstitial macrophages in vaccine-mediated protection. Therefore, protection by vaccination, even in the absence of high titers of neutralizing antibodies in the serum, correlates with recall of broadly reactive B- and T-cell responses.
Assuntos
COVID-19 , SARS-CoV-2 , Animais , Cricetinae , Humanos , Vacina BNT162 , Infecções Irruptivas , COVID-19/prevenção & controle , Mesocricetus , Anticorpos Neutralizantes , Complicações Pós-Operatórias , RNA Mensageiro/genética , Imunidade , Anticorpos Antivirais , VacinaçãoRESUMO
The high transmissibility of SARS-CoV-2 is related to abundant replication in the upper airways, which is not observed for the other highly pathogenic coronaviruses SARS-CoV and MERS-CoV. We here reveal features of the coronavirus spike (S) protein, which optimize the virus towards the human respiratory tract. First, the S proteins exhibit an intrinsic temperature preference, corresponding with the temperature of the upper or lower airways. Pseudoviruses bearing the SARS-CoV-2 spike (SARS-2-S) were more infectious when produced at 33°C instead of 37°C, a property shared with the S protein of HCoV-229E, a common cold coronavirus. In contrast, the S proteins of SARS-CoV and MERS-CoV favored 37°C, in accordance with virus preference for the lower airways. Next, SARS-2-S-driven entry was efficiently activated by not only TMPRSS2, but also the TMPRSS13 protease, thus broadening the cell tropism of SARS-CoV-2. Both proteases proved relevant in the context of authentic virus replication. TMPRSS13 appeared an effective spike activator for the virulent coronaviruses but not the low pathogenic HCoV-229E virus. Activation of SARS-2-S by these surface proteases requires processing of the S1/S2 cleavage loop, in which both the furin recognition motif and extended loop length proved critical. Conversely, entry of loop deletion mutants is significantly increased in cathepsin-rich cells. Finally, we demonstrate that the D614G mutation increases SARS-CoV-2 stability, particularly at 37°C, and, enhances its use of the cathepsin L pathway. This indicates a link between S protein stability and usage of this alternative route for virus entry. Since these spike properties may promote virus spread, they potentially explain why the spike-G614 variant has replaced the early D614 variant to become globally predominant. Collectively, our findings reveal adaptive mechanisms whereby the coronavirus spike protein is adjusted to match the temperature and protease conditions of the airways, to enhance virus transmission and pathology.
Assuntos
COVID-19/metabolismo , Sistema Respiratório/metabolismo , Sistema Respiratório/virologia , SARS-CoV-2/metabolismo , Glicoproteína da Espícula de Coronavírus/metabolismo , COVID-19/transmissão , Coronavirus Humano 229E/metabolismo , Furina/metabolismo , Humanos , Proteínas de Membrana/metabolismo , Coronavírus da Síndrome Respiratória do Oriente Médio/metabolismo , Peptídeo Hidrolases/metabolismo , Serina Endopeptidases/metabolismo , Glicoproteína da Espícula de Coronavírus/genética , Temperatura , Internalização do Vírus , Replicação Viral/fisiologiaRESUMO
Influenza A virus (IAV) and influenza B virus (IBV) cause yearly epidemics with significant morbidity and mortality. When zoonotic IAVs enter the human population, the viral hemagglutinin (HA) requires adaptation to achieve sustained virus transmission. In contrast, IBV has been circulating in humans, its only host, for a long period of time. Whether this entailed adaptation of IBV HA to the human airways is unknown. To address this question, we compared two seasonal IAVs (A/H1N1 and A/H3N2) and two IBVs (B/Victoria and B/Yamagata lineages) with regard to host-dependent activity of HA as the mediator of membrane fusion during viral entry. We first investigated proteolytic activation of HA by covering all type II transmembrane serine protease (TTSP) and kallikrein enzymes, many of which proved to be present in human respiratory epithelium. The IBV HA0 precursor is cleaved by a broader panel of TTSPs and activated with much higher efficiency than IAV HA0. Accordingly, knockdown of a single protease, TMPRSS2, abrogated spread of IAV but not IBV in human respiratory epithelial cells. Second, the HA fusion pH values proved similar for IBV and human-adapted IAVs (with one exception being the HA of 1918 IAV). Third, IBV HA exhibited higher expression at 33°C, a temperature required for membrane fusion by B/Victoria HA. This indicates pronounced adaptation of IBV HA to the mildly acidic pH and cooler temperature of human upper airways. These distinct and intrinsic features of IBV HA are compatible with extensive host adaptation during prolonged circulation of this respiratory virus in the human population.IMPORTANCE Influenza epidemics are caused by influenza A and influenza B viruses (IAV and IBV, respectively). IBV causes substantial disease; however, it is far less studied than IAV. While IAV originates from animal reservoirs, IBV circulates in humans only. Virus spread requires that the viral hemagglutinin (HA) is active and sufficiently stable in human airways. We resolve here how these mechanisms differ between IBV and IAV. Whereas human IAVs rely on one particular protease for HA activation, this is not the case for IBV. Superior activation of IBV by several proteases should enhance shedding of infectious particles. IBV HA exhibits acid stability and a preference for 33°C, indicating pronounced adaptation to the human upper airways, where the pH is mildly acidic and a cooler temperature exists. These adaptive features are rationalized by the long existence of IBV in humans and may have broader relevance for understanding the biology and evolution of respiratory viruses.
Assuntos
Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H3N2/genética , Vírus da Influenza B/genética , Influenza Humana/virologia , Pulmão/virologia , Replicação Viral/genética , Linhagem Celular , Células Epiteliais/patologia , Células Epiteliais/virologia , Regulação da Expressão Gênica , Glicoproteínas de Hemaglutininação de Vírus da Influenza/química , Glicoproteínas de Hemaglutininação de Vírus da Influenza/metabolismo , Interações Hospedeiro-Patógeno/genética , Humanos , Concentração de Íons de Hidrogênio , Vírus da Influenza A Subtipo H1N1/metabolismo , Vírus da Influenza A Subtipo H1N1/patogenicidade , Vírus da Influenza A Subtipo H3N2/metabolismo , Vírus da Influenza A Subtipo H3N2/patogenicidade , Vírus da Influenza B/metabolismo , Vírus da Influenza B/patogenicidade , Influenza Humana/patologia , Calicreínas/classificação , Calicreínas/genética , Calicreínas/metabolismo , Pulmão/patologia , Fusão de Membrana , Proteínas de Membrana/classificação , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Proteólise , Mucosa Respiratória/patologia , Mucosa Respiratória/virologia , Serina Endopeptidases/deficiência , Serina Endopeptidases/genética , Serina Proteases/classificação , Serina Proteases/genética , Serina Proteases/metabolismo , Especificidade da Espécie , Temperatura , Internalização do VírusRESUMO
The possible resistance of influenza virus against existing antiviral drugs calls for new therapeutic concepts. One appealing strategy is to inhibit virus entry, in particular at the stage of internalization. This requires a better understanding of virus-host interactions during the entry process, including the role of receptor tyrosine kinases (RTKs). To search for cellular targets, we evaluated a panel of 276 protein kinase inhibitors in a multicycle antiviral assay in Madin-Darby canine kidney cells. The RTK inhibitor Ki8751 displayed robust anti-influenza A and B virus activity and was selected for mechanistic investigations. Ki8751 efficiently disrupted the endocytic process of influenza virus in different cell lines carrying platelet-derived growth factor receptor ß (PDGFRß), an RTK that is known to act at GM3 ganglioside-positive lipid rafts. The more efficient virus entry in CHO-K1 cells compared to the wild-type ancestor (CHO-wt) cells indicated a positive effect of GM3, which is abundant in CHO-K1 but not in CHO-wt cells. Entering virus localized to GM3-positive lipid rafts and the PDGFRß-containing endosomal compartment. PDGFRß/GM3-dependent virus internalization involved PDGFRß phosphorylation, which was potently inhibited by Ki8751, and desialylation of activated PDGFRß by the viral neuraminidase. Virus uptake coincided with strong activation of the Raf/MEK/Erk cascade, but not of PI3K/Akt or phospholipase C-γ. We conclude that influenza virus efficiently hijacks the GM3-enhanced PDGFRß signalling pathway for cell penetration, providing an opportunity for host cell-targeting antiviral intervention.
Assuntos
Gangliosídeo G(M3)/metabolismo , Influenza Humana/metabolismo , Influenza Humana/virologia , Infecções por Orthomyxoviridae/metabolismo , Orthomyxoviridae/patogenicidade , Receptor beta de Fator de Crescimento Derivado de Plaquetas/metabolismo , Transdução de Sinais/fisiologia , Animais , Células CHO , Linhagem Celular , Cricetulus , Cães , Células HEK293 , Humanos , Influenza Humana/tratamento farmacológico , Células Madin Darby de Rim Canino , Orthomyxoviridae/efeitos dos fármacos , Infecções por Orthomyxoviridae/tratamento farmacológico , Compostos de Fenilureia/farmacologia , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação/fisiologia , Inibidores de Proteínas Quinases/farmacologia , Quinolinas/farmacologia , Receptores Proteína Tirosina Quinases/metabolismo , Transdução de Sinais/efeitos dos fármacos , Internalização do Vírus/efeitos dos fármacosRESUMO
The CCL20/CCR6 chemokine/receptor axis has previously been shown to contribute to the initiation and progression of hepatocellular carcinoma (HCC) through the recruitment of CCR6-positive leukocytes to the tumor microenvironment. In particular, high serum levels of CCL20 are reported in patients with HCC induced by the hepatitis C virus (HCV). A potential non-immune role for the CCL20/CCR6 axis in HCC development has not yet been investigated. Microarray analysis (Benkheil et al., paper submitted for publication), revealed that CCL20 is highly upregulated in hepatoma cells infected with HCV compared with non-infected hepatoma cells. To determine the role of the CCL20/CCR6 axis in HCV-related HCC, we first explored which cell populations express CCR6 in human liver tissue with chronic disease or HCC. Immunohistochemical (IHC) analysis revealed that CCR6 is present on endothelial cells (ECs) of portal blood vessels in livers with chronic HCV infection and in HCV- and alcoholic-HCC tissue. In addition, we found CCR6 to be expressed on primary macrovascular (HUVECs) and microvascular ECs (HMVEC-ds) where it co-expressed with the endothelial marker CD31. In vitro angiogenesis experiments revealed that CCL20 is a direct pro-angiogenic molecule that induces EC invasion, sprouting and migration through CCR6. Moreover, using the angiogenesis matrigel plug assay in immunodeficient NMRI-nu mice, we clearly showed that CCL20 induces blood vessel formation, by attracting CCR6-positive ECs. Finally, we demonstrated that HCV-induced CCL20 protein expression and secretion in hepatoma cells could be abolished by antiviral treatment, indicating that CCL20 expression is dependent on HCV replication. In contrast to HCV, HBV-infection resulted in a decreased expression of CCL20, implying a virus-specific effect. Taken together, we identified HCV-induced CCL20 as a direct pro-angiogenic factor that acts on endothelial CCR6. These results suggest that the CCL20/CCR6 axis contributes to hepatic angiogenesis, promoting the hypervascular state of HCV-HCC.
Assuntos
Carcinoma Hepatocelular/genética , Quimiocina CCL20/genética , Neoplasias Hepáticas/genética , Neovascularização Patológica/genética , Receptores CCR6/genética , Animais , Carcinoma Hepatocelular/patologia , Carcinoma Hepatocelular/virologia , Quimiotaxia/genética , Células Endoteliais/fisiologia , Células Endoteliais/virologia , Regulação Neoplásica da Expressão Gênica , Hepacivirus/genética , Hepacivirus/patogenicidade , Células Endoteliais da Veia Umbilical Humana , Humanos , Fígado/metabolismo , Fígado/patologia , Fígado/virologia , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/virologia , Camundongos , Neovascularização Patológica/patologia , Neovascularização Patológica/virologia , Microambiente Tumoral/genéticaRESUMO
The Integrated Nutrition Pathway for Acute Care (INPAC) is an evidence and consensus based pathway developed to guide health care professionals in the prevention, detection, and treatment of malnutrition in medical and surgical patients. From 2015 to 2017, the More-2-Eat implementation project (M2E) used a participatory action research approach to determine the feasibility, and evaluate the implementation of INPAC in 5 hospital units across Canada. Based on the findings of M2E and consensus with M2E stakeholders, updates have been made to INPAC to enhance feasibility in Canadian hospitals. The learnings from M2E have been converted into an online toolkit that outlines how to implement the key steps within INPAC. The aim of this short report is to highlight the updated version of INPAC, and introduce the implementation toolkit that was used to support practice improvements towards this standard.
Assuntos
Pacientes Internados , Desnutrição/diagnóstico , Desnutrição/terapia , Avaliação Nutricional , Terapia Nutricional/métodos , Canadá , Estudos de Viabilidade , Humanos , Desnutrição/prevenção & controle , Avaliação de Programas e Projetos de SaúdeRESUMO
This prospective cohort study was conducted in eighteen Canadian hospitals with the aim of examining factors associated with nutritional decline in medical and surgical patients. Nutritional decline was defined based on subjective global assessment (SGA) performed at admission and discharge. Data were collected on demographics, medical information, food intake and patients' satisfaction with nutrition care and meals during hospitalisation; 424 long-stay (≥7 d) patients were included; 38% of them had surgery; 51% were malnourished at admission (SGA B or C); 37% had in-hospital changes in SGA; 19·6% deteriorated (14·6% from SGA A to B/C and 5% from SGA B to C); 17·4% improved (10·6% from SGA B to A, 6·8% from SGA C to B/A); and 63·0 % patients were stable (34·4% were SGA A, 21·3% SGA B, 7·3% SGA C). One SGA C patient had weight loss ≥5%, likely due to fluid loss and was designated as stable. A subset of 364 patients with admission SGA A and B was included in the multiple logistic regression models to determine factors associated with nutritional decline. After controlling for SGA at admission and the presence of a surgical procedure, lower admission BMI, cancer, two or more diagnostic categories, new in-hospital infection, reduced food intake, dissatisfaction with food quality and illness affecting food intake were factors significantly associated with nutritional decline in medical patients. For surgical patients, only male sex was associated with nutritional decline. Factors associated with nutritional decline are different in medical and surgical patients. Identifying these factors may assist nutritional care.
Assuntos
Hospitalização , Desnutrição/epidemiologia , Estado Nutricional , Idoso , Canadá/epidemiologia , Estudos de Coortes , Ingestão de Alimentos , Feminino , Humanos , Tempo de Internação , Masculino , Refeições , Avaliação Nutricional , Terapia Nutricional , Satisfação do Paciente , Cuidados Pós-Operatórios , Estudos Prospectivos , Fatores Sexuais , Redução de PesoRESUMO
BACKGROUND: Malnutrition is commonly underdiagnosed and undertreated in acute care patients. Implementation of current pathways of care is limited, potentially as a result of the perception that they are not feasible with current resources. There is a need for a pathway based on expert consensus, best practice and evidence that addresses this crisis in acute care, while still being feasible for implementation. METHODS: A modified Delphi was used to develop consensus on a new pathway. Extant literature and other resources were reviewed to develop an evidence-informed background document and draft pathway, which were considered at a stakeholder meeting of 24 experts. Two rounds of an on-line Delphi survey were completed (n = 28 and 26 participants respectively). Diverse clinicians from four hospitals participated in focus groups to face validate the draft pathway and a final stakeholder meeting confirmed format changes to make the pathway conceptually clear and easy to follow for end-users. Experts involved in this process were researchers and clinicians from dietetics, medicine and nursing, including management and frontline personnel. RESULTS: 80% of stakeholders who were invited, participated in the first Delphi survey. The two rounds of the Delphi resulted in consensus for all but two minor components of the Integrated Nutrition Pathway for Acute Care (INPAC). The format of the INPAC was revised based on the input of focus group participants, stakeholders and investigators. CONCLUSIONS: This evidence-informed, consensus based pathway for nutrition care has greater depth and breadth than prior guidelines that were commonly based on systematic reviews. As extant evidence for many best practices is absent, the modified Delphi process has allowed for consensus to be developed based on better practices. Attention to feasibility during development has created a pathway that has greater implementation potential. External validation specifically with practitioner groups promoted a conceptually easy to use format. Test site implementation and evaluation is needed to identify resource requirements and demonstrate process and patient reported outcomes resulting from embedding INPAC into clinical practice.
Assuntos
Consenso , Cuidados Críticos/métodos , Desnutrição/dietoterapia , Terapia Nutricional/métodos , Adulto , Técnica Delphi , Feminino , Grupos Focais , Humanos , Masculino , Pessoa de Meia-Idade , Inquéritos Nutricionais , Estado NutricionalRESUMO
SARS-CoV-2 Omicron subvariants are still emerging and spreading worldwide. These variants contain a high number of polymorphisms in the spike (S) glycoprotein that could potentially impact their pathogenicity and transmission. We have previously shown that the S:655Y and P681H mutations enhance S protein cleavage and syncytia formation. Interestingly, these polymorphisms are present in Omicron S protein. Here, we characterized the cleavage efficiency and fusogenicity of the S protein of different Omicron sublineages. Our results showed that Omicron BA.1 subvariant is efficiently cleaved but it is poorly fusogenic compared to previous SARS-CoV-2 strains. To understand the basis of this phenotype, we generated chimeric S protein using combinations of the S1 and S2 domains from WA1, Delta and Omicron BA.1 variants. We found that the S2 domain of Omicron BA.1 hindered efficient cell-cell fusion. Interestingly, this domain only contains six unique polymorphisms never detected before in ancestral SARS-CoV-2 variants. WA1614G S proteins containing the six individuals S2 Omicron mutations were assessed for their fusogenicity and S surface expression after transfection in cells. Results showed that the S:N856K and N969K substitutions decreased syncytia formation and impacted S protein cell surface levels. However, we observed that "first-generation" Omicron sublineages that emerged subsequently, had convergently evolved to an enhanced fusogenic activity and S expression on the surface of infected cells while "second-generation" Omicron variants have highly diverged and showed lineage-specific fusogenic properties. Importantly, our findings could have potential implications in the improvement and redesign of COVID-19 vaccines.
Assuntos
Vacinas contra COVID-19 , COVID-19 , Humanos , SARS-CoV-2/genética , Mutação , Glicoproteína da Espícula de Coronavírus/genéticaRESUMO
Broad(er)-acting antiviral drugs, active against entire genera or families of viruses, should be developed and stockpiled in epidemic/pandemic peacetime. They can be used to counter outbreaks as soon as the new virus has been identified and will also remain important pharmacological tools after the introduction of vaccines and monoclonal antibodies.
Assuntos
Virus da Influenza A Subtipo H5N1 , Vacinas contra Influenza , Influenza Humana , Humanos , Influenza Humana/prevenção & controle , Pandemias/prevenção & controle , Antivirais/farmacologia , Antivirais/uso terapêutico , Surtos de Doenças/prevenção & controleRESUMO
Massive efforts on both vaccine development and antiviral research were launched to combat the new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We contributed, amongst others, by the development of a high-throughput screening (HTS) antiviral assay against SARS-CoV-2 using a fully automated, high-containment robot system. Here, we describe the development of this novel, convenient and phenotypic dual-reporter virus-cell-based high-content imaging assay using the A549+hACE2+TMPRSS2_mCherry reporter lung carcinoma cell line and an ancestral SARS-CoV-2_Wuhan_mNeonGreen reporter virus. Briefly, by means of clonal selection, a host cell subclone was selected that (i) efficiently supports replication of the reporter virus with high expression, upon infection, of the NeonGreen fluorescent reporter protein, (ii) that is not affected by virus-induced cytopathogenic effects and, (iii) that expresses a strong fluorescent mCherry signal in the nucleus. The selected clone matched these criteria with an infection rate on average of 75% with limited cell death. The average (R)Z'-factors of the assay plates were all >0.8, which indicates a robust assay suitable for HTS purposes. A selection of reference compounds that inhibits SARS-CoV-2 replication in vitro were used to validate this novel dual-reporter assay and confirms the data reported in the literature. This assay is a convenient and powerful tool for HTS of large compound libraries against SARS-CoV-2.
Assuntos
Antivirais , COVID-19 , Humanos , Antivirais/farmacologia , Antivirais/metabolismo , Ensaios de Triagem em Larga Escala/métodos , SARS-CoV-2 , Descoberta de Drogas , Replicação ViralRESUMO
Hybrid immunity to SARS-CoV-2 provides superior protection to re-infection. We performed immune profiling studies during breakthrough infections in mRNA-vaccinated hamsters to evaluate hybrid immunity induction. mRNA vaccine, BNT162b2, was dosed to induce binding antibody titers against ancestral spike, but inefficient serum virus neutralization of ancestral SARS-CoV-2 or variants of concern (VoCs). Vaccination reduced morbidity and controlled lung virus titers for ancestral virus and Alpha but allowed breakthrough infections in Beta, Delta and Mu-challenged hamsters. Vaccination primed T cell responses that were boosted by infection. Infection back-boosted neutralizing antibody responses against ancestral virus and VoCs. Hybrid immunity resulted in more cross-reactive sera. Transcriptomics post-infection reflects both vaccination status and disease course and suggests a role for interstitial macrophages in vaccine-mediated protection. Therefore, protection by vaccination, even in the absence of high titers of neutralizing antibodies in the serum, correlates with recall of broadly reactive B- and T-cell responses.
RESUMO
We explored the availability of parameters for a nutrition screening system among elderly people in New Brunswick (NB) health care facilities. Patients aged 65 or older were asked to participate in the study; each participant had been admitted to one of four hospitals or lived in one of six nursing homes. Availability of nutrition screening parameters (weight, height, weight change, serum albumin level, appetite, and food intake record) was assessed by auditing the participants' medical charts. When data were not available, the feasibility of obtaining them was determined. Additional data related to nutrition screening were also obtained. In total, 421 participants were recruited for the study: 140 (33.2%) who lived in nursing homes and 281 (66.8%) who were in hospitals. Parameters needed to conduct nutrition screening, such as weight upon admission, were available for 83.6% of participants; usual weight was available for 43.0%, height for 86.0%, and serum albumin level for 47.5%. Our findings show that basic parameters for nutrition screening are available, and that implementation of a nutrition screening system is feasible for patients in NB health care facilities.
Assuntos
Avaliação Geriátrica , Serviços de Saúde para Idosos , Programas de Rastreamento , Avaliação Nutricional , Admissão do Paciente , Idoso , Idoso de 80 Anos ou mais , Registros Eletrônicos de Saúde , Estudos de Viabilidade , Feminino , Instituição de Longa Permanência para Idosos , Hospitais Comunitários , Humanos , Masculino , Novo Brunswick , Casas de SaúdeRESUMO
The continuous emergence of new variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) urges better understanding of the functional motifs in the spike (S) protein and their tolerance to mutations. Here, we focused on the S2' motif, which, during virus entry, requires cleavage by a host cell protease to release the fusion peptide. Though belonging to an immunogenic region, the SARS-CoV-2 S2' motif (811-KPSKR-815) has shown hardly any variation, with its three basic (K/R) residues being >99.99% conserved thus far. By creating a series of mutant pseudoviruses bearing the spikes of Wuhan-Hu-1, its G614 mutant or the Delta and Omicron variants, we show that residue K814 (preceding the scissile R815) is dispensable for TMPRSS2 yet favored by the alternative TMPRSS13 protease. Activation by TMPRSS13 was drastically reduced when the SARS-CoV-2 S2' motif was swapped with that of the low pathogenic 229E coronavirus (685-RVAGR-689), and also, the reverse effect was seen. This swap had no impact on recognition by TMPRSS2. In the Middle East respiratory syndrome coronavirus (MERS-CoV) spike, introducing a dibasic scissile motif was easily accepted by TMPRSS13 but less so by TMPRSS2, confirming that TMPRSS13 favors a sequence rich in K/R residues. Pseudovirus entry experiments in Calu-3 cells confirmed that the S2' mutations have minor impact on TMPRSS2. Our findings are the first to demonstrate which S2' residues are important for SARS-CoV-2 spike activation by these two airway proteases, with TMPRSS2 being more tolerant to variation than TMPRSS13. This preemptive insight will help to estimate the impact of S2' motif changes as they appear in new SARS-CoV-2 variants. IMPORTANCE Since its introduction in humans, SARS-CoV-2 is evolving with frequent appearance of new variants. The surveillance would benefit from proactive characterization of the functional motifs in the spike (S) protein, the most variable viral factor. This is linked to immune evasion but also influences spike functioning. Remarkably, though located in a strongly immunogenic region, the S2' cleavage motif has, thus far, remained highly conserved. This suggests that its sequence is critical for spike activation by airway proteases. To investigate this, we assessed how pseudovirus entry is affected by changes in the S2' motif. We demonstrate that TMPRSS2 readily accepts variations in this motif, whereas the alternative TMPRSS13 protease is more fastidious. The Wuhan-Hu-1, G614, Delta and Omicron spikes showed no difference in this regard. Being the first in its kind, our study will help to assess the impact of S2' variations as soon as they are detected during variant surveillance.
Assuntos
COVID-19 , SARS-CoV-2 , Humanos , Proteínas de Membrana/genética , Mutação , Peptídeo Hidrolases/genética , SARS-CoV-2/genética , Serina Endopeptidases/genética , Glicoproteína da Espícula de Coronavírus/metabolismo , Internalização do VírusRESUMO
A well-tolerated and cost-effective oral drug that blocks SARS-CoV-2 growth and dissemination would be a major advance in the global effort to reduce COVID-19 morbidity and mortality. Here, we show that the oral FDA-approved drug nitazoxanide (NTZ) significantly inhibits SARS-CoV-2 viral replication and infection in different primate and human cell models including stem cell-derived human alveolar epithelial type 2 cells. Furthermore, NTZ synergizes with remdesivir, and it broadly inhibits growth of SARS-CoV-2 variants B.1.351 (beta), P.1 (gamma), and B.1617.2 (delta) and viral syncytia formation driven by their spike proteins. Strikingly, oral NTZ treatment of Syrian hamsters significantly inhibits SARS-CoV-2-driven weight loss, inflammation, and viral dissemination and syncytia formation in the lungs. These studies show that NTZ is a novel host-directed therapeutic that broadly inhibits SARS-CoV-2 dissemination and pathogenesis in human and hamster physiological models, which supports further testing and optimization of NTZ-based therapy for SARS-CoV-2 infection alone and in combination with antiviral drugs.
RESUMO
SARS-CoV-2 lineages have diverged into highly prevalent variants termed "variants of concern" (VOCs). Here, we characterized emerging SARS-CoV-2 spike polymorphisms in vitro and in vivo to understand their impact on transmissibility and virus pathogenicity and fitness. We demonstrate that the substitution S:655Y, represented in the gamma and omicron VOCs, enhances viral replication and spike protein cleavage. The S:655Y substitution was transmitted more efficiently than its ancestor S:655H in the hamster infection model and was able to outcompete S:655H in the hamster model and in a human primary airway system. Finally, we analyzed a set of emerging SARS-CoV-2 variants to investigate how different sets of mutations may impact spike processing. All VOCs tested exhibited increased spike cleavage and fusogenic capacity. Taken together, our study demonstrates that the spike mutations present in VOCs that become epidemiologically prevalent in humans are linked to an increase in spike processing and virus transmission.
Assuntos
COVID-19 , SARS-CoV-2 , Humanos , Mutação , SARS-CoV-2/genética , Glicoproteína da Espícula de Coronavírus/genéticaRESUMO
PURPOSE: Several studies show that malnutrition is prevalent in health care facilities, especially among elderly patients and nursing home residents. Although validated screening tools exist, little evidence exists on the feasibility of implementing nutrition screening in health care facilities. We examined New Brunswick health care professionals' perceptions of and practices involving nutrition screening in elderly clients, as well as barriers to screening. METHODS: A survey was conducted with questionnaires intended for physicians, nurses, and dietitians. RESULTS: Participants were 457 health care professionals (physicians, 34.6%; nurses, 50.3%; dietitians, 15.1%). Perceptions of nutrition screening varied. For example, most nurses (94.7%) and dietitians (98.5%) indicated that screening was important/very important, while only 63.5% of physicians indicated this. Screening methods also differed among professionals and few used a screening tool. Several barriers to implementing nutrition screening were reported, such as lack of time, lack of professional resources, and clients' short stays. CONCLUSIONS: These findings will help professionals address the feasibility of implementing standardized screening tools in health care facilities. A more consistent and systematic approach for detecting populations at high nutritional risk may result.
Assuntos
Pesquisas sobre Atenção à Saúde , Pessoal de Saúde , Hospitais , Avaliação Nutricional , Estado Nutricional , Idoso , Estudos Transversais , Dietética , Humanos , Desnutrição/diagnóstico , Novo Brunswick , Enfermeiras e Enfermeiros , Casas de Saúde , Médicos , Inquéritos e QuestionáriosRESUMO
The subjective global assessment (SGA) is a nutrition assessment tool that refers to an overall evaluation of a patient's history and physical examination and uses structured clinical parameters to diagnose malnutrition. The SGA is known to be a reliable and valid tool that predicts morbidity and mortality associated with malnutrition. The objective of SGA is to identify patients likely to benefit from nutrition intervention and therefore to identify persons in whom inadequate nutrition intake or absorption explain features of malnutrition, including body wasting. There are other conditions that cause weight loss, muscle wasting, and fat loss, including cachexia and sarcopenia. Acknowledging that these 2 last conditions differ in their mechanism of body wasting and consequently in the outcomes of nutrition intervention, the practitioner needs a tool to identify when malnutrition is the dominating factor to explain body wasting. The SGA form has been revised to clearly reflect the key concepts behind the diagnosis of malnutrition and help to distinguish this condition from other wasting conditions. This review presents the revised SGA form and guidance document. Using case studies, it illustrates the 3 wasting conditions, their overlap, and how the SGA identifies malnutrition as a dominating factor of body wasting and thus individuals who require nutrition intervention.
Assuntos
Desnutrição , Sarcopenia , Caquexia/diagnóstico , Caquexia/etiologia , Humanos , Desnutrição/diagnóstico , Desnutrição/epidemiologia , Avaliação Nutricional , Estado Nutricional , Sarcopenia/diagnóstico , Sarcopenia/epidemiologiaRESUMO
Hospital malnutrition is a longstanding problem that continues to be underrecognized and undertreated. The aim of this narrative review is to summarize novel, solution-focused, recent research or commentary to update providers on the prevention of iatrogenic malnutrition as well as the detection and treatment of hospital malnutrition. A narrative review was completed using the top 11 clinically relevant nutrition journals. Of the 13,850 articles and editorials published in these journals between 2013 and 2019, 511 were related to hospital malnutrition. A duplicate review was used to select (n = 108) and extract key findings from articles and editorials. Key criteria for selection were population of interest (adult hospital patients, no specific diagnostic group), solution-focused, and novel perspectives. Articles were categorized (6 classified in >1 category) as Screening and Assessment (n = 17), Standard (n = 25), Advanced (n = 12) and Specialized Nutrition Care (n = 8), Transitions (n = 15), Multicomponent (n = 21), Education and Empowerment (n = 9), Economic Impact (n = 3), and Guidelines (n = 4) for summarizing. Research advances in screening implementation, standard nutrition care, transitions, and multicomponent interventions provide new strategies to consider for malnutrition prevention (iatrogenic), detection, and care. However, several areas requiring further research were identified. Specifically, larger and more rigorous studies that examine health outcomes and economic analyses are urgently needed.