Breakthrough infections by SARS-CoV-2 variants boost cross-reactive hybrid immune responses in mRNA-vaccinated Golden Syrian hamsters.

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.

The impact of S2 mutations on Omicron SARS-CoV-2 cell surface expression and fusogenicity.

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.

Breakthrough infections by SARS-CoV-2 variants boost cross-reactive hybrid immune responses in mRNA-vaccinated Golden Syrian Hamsters.

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.

Antiviral strategies for epidemic and pandemic preparedness.

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.

Development of a robust and convenient dual-reporter high-throughput screening assay for SARS-CoV-2 antiviral drug discovery.

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.

Impact of SARS-CoV-2 Spike Mutations on Its Activation by TMPRSS2 and the Alternative TMPRSS13 Protease.

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.

The oral drug nitazoxanide restricts SARS-CoV-2 infection and attenuates disease pathogenesis in Syrian hamsters.

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.

Mutations in SARS-CoV-2 variants of concern link to increased spike cleavage and virus transmission.

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.

The SARS-CoV-2 and other human coronavirus spike proteins are fine-tuned towards temperature and proteases of the human airways.

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.

Evaluation of Nutrition Status Using the Subjective Global Assessment: Malnutrition, Cachexia, and Sarcopenia.

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.

Trends and Novel Research in Hospital Nutrition Care: A Narrative Review of Leading Clinical Nutrition Journals.

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.

N-benzyl 4,4-disubstituted piperidines as a potent class of influenza H1N1 virus inhibitors showing a novel mechanism of hemagglutinin fusion peptide interaction.

The influenza virus hemagglutinin (HA) is an attractive target for antiviral therapy due to its essential role in mediating virus entry into the host cell. We here report the identification of a class of N-benzyl-4,4,-disubstituted piperidines as influenza A virus fusion inhibitors with specific activity against the H1N1 subtype. Using the highly efficient one-step Ugi four-component reaction, diverse library of piperidine-based analogues was synthesized and evaluated to explore the structure-activity relationships (SAR). Mechanistic studies, including resistance selection with the most active compound (2) demonstrated that it acts as an inhibitor of the low pH-induced HA-mediated membrane fusion process. Computational studies identified an as yet unrecognized fusion inhibitor binding site, which is located at the bottom of the HA2 stem in close proximity to the fusion peptide. A direct π-stacking interaction between the N-benzylpiperidine moiety of 2 and F9HA2 of the fusion peptide, reinforced with an additional π-stacking interaction with Y119HA2, and a salt bridge of the protonated piperidine nitrogen with E120HA2, were identified as important interactions to mediate ligand binding. This site rationalized the observed SAR and provided a structural explanation for the H1N1-specific activity of our inhibitors. Furthermore, the HA1-S326V mutation resulting in resistance to 2 is close to the proposed new binding pocket. Our findings point to the N-benzyl-4,4,-disubstituted piperidines as an interesting class of influenza virus inhibitors, representing the first example of fusion peptide binders with great potential for anti-influenza drug development.

GLIM criteria has fair sensitivity and specificity for diagnosing malnutrition when using SGA as comparator.

BACKGROUND & AIMS: The Global Leadership Initiative on Malnutrition (GLIM) proposed a new framework for diagnosing malnutrition based on combinations of phenotypic and etiologic criteria. The aim of this study was to compare GLIM criteria to Subjective Global Assessment (SGA) judged to be the most validated standardized assessment of malnutrition. METHODS: This is a retrospective analysis of variables extracted from a prospective cohort study assessing malnutrition at admission, in 18 Canadian hospitals. Based on the available parameters, GLIM was compared to SGA using the following combinations of one phenotypic and one etiologic criteria: A. weight loss and low intake; B. weight loss and high C-reactive protein (CRP); C. low body mass index (BMI) and low intake; D. low BMI, high CRP. Data were not available for fat-free mass. Since all patients had acute or chronic active disease as per GLIM etiologic criterion, CRP was used as a more specific measure to define inflammation. Sensitivity, specificity, positive (PPV) and negative (NPV) predictive values were calculated. Data are expressed as mean and Clopper-Pearson exact 95% confidence interval (CI). RESULTS: From 1022 patients in the original dataset, 784 had all considered parameters with a prevalence of malnutrition (SGA B or C) of 45.15% (CI 41.60, 48.70), where severe malnutrition (SGA C) was 11.73% (CI 9.57, 14.20). Using the available GLIM parameters with the above combinations of two-criteria, the prevalence of malnutrition was 33.29% (CI 30.00, 36.71) and severe malnutrition was 19.77% (CI 17.00, 22.70). For all criteria combinations of GLIM together versus SGA, sensitivity was 61.30% (CI 56.0, 66.4), specificity was 89.77% (CI 86.5, 92.5) and PPV was 83.14% (CI 78.0, 87.5) while NPV was 73.80 (CI 69.8, 77.5). Sensitivity was improved when only SGA C for severe malnutrition was used as the criterion (82.61%; CI 73.3, 89.7) but PPV was greatly reduced (29.12%; CI 23.7, 35.0). Similarly, when using GLIM criteria for severe malnutrition only, sensitivity improved (76.09%; CI 66.1, 84.4). Any two criteria combinations of GLIM had much poorer sensitivity with the highest being weight loss + high CRP (46.33%) with a specificity of 93.02% (PPV: 84.54%; NPV: 67.80%), while the combination of low BMI + low intake had the highest specificity (98.84%) but with a sensitivity of 15.54% (PPV 91.67%; NPV: 58.70%). CONCLUSIONS: Based on the CMTF dataset and using SGA as the most validated tool for diagnosing malnutrition, the two criteria combinations used for GLIM in the present study had fair criterion validity for the diagnosis of malnutrition, regardless of severity status. The best combinations were weight loss and high CRP or weight loss and low intake, both having high specificity at diagnosing malnutrition but unacceptably low sensitivity, and thus were considered poor. There may be potential for the full framework to be used to diagnose malnutrition, but individual combinations of two criteria when used exclusively will miss malnourished patients, as defined by SGA.

Handgrip strength predicts length of stay and quality of life in and out of hospital.

BACKGROUND: Handgrip strength (HGS) is a practical measure of strength and physical function that can be used to identify frailty among hospitalized patients, but its utility in this setting is unclear. To be considered useful, any functional measure needs to provide pertinent information on the patient and predict relevant outcomes such as health-care utilization (e.g., length of stay (LOS)) and patient-reported quality of life (QOL). The purpose of this study was to determine if HGS predicted LOS and QOL. A second aim was to examine the best sensitivity (SE) and specificity (SP) for predicting length of stay (>7 or >13 days) using previously published cut-points for HGS. METHODOLOGY: HGS was measured on 1136 medical patients shortly after admission with a Lafayette dynamometer. QOL was assessed with the self-reported SF-12 completed with an interviewer during hospitalization and 30- days after discharge via telephone. Physical (PCS) and mental (MCS) component scores of SF-12 were calculated. A variety of covariates were assessed (e.g., nutritional status). Multivariate analyses stratified by sex were completed. RESULTS: The mean LOS was 12.71 days (median = 8.00; SD = 13.20), 12.88 days (SD = 13.82) for males, and 12.58 days (SD = 12.68) for females. Lower admission HGS scores were associated with longer LOS (male X2 = 7.85, p < 0.05; female X2 = 14.9, p < 0.0001). The average quality of life scores were as follows: in hospital PCS: 34.66, MCS: 46.49; post discharge PCS: 36.17; MCS: 51.22. HGS predicted PCS during hospitalization (male X2 = 36.22, p < 0.0001; female X2 = 19.87, p < 0.0001) and post hospitalization (male X2 = 6.98, p < 0.01; female X2 = 10.99, p < 0.01). Various reference cut-points for HGS were tested against LOS, with none being considered appropriate (e.g., SE and SP both < 70) when adjusting for age and sex. CONCLUSION: Admission HGS adds predictive value for both LOS and physical components of QOL and is worth pursuing in practice to identify potential frailty and the need for proactive steps to mitigate further functional decline during hospitalization. However, HGS cut-points for LOS specific to acute care patients need to be defined and tested.

Hemagglutinin Cleavability, Acid Stability, and Temperature Dependence Optimize Influenza B Virus for Replication in Human Airways.

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.

Influenza virus entry via the GM3 ganglioside-mediated platelet-derived growth factor receptor ß signalling pathway.

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.

Multi-site implementation of nutrition screening and diagnosis in medical care units: Success of the More-2-Eat project.

BACKGROUND: Improving the detection and treatment of malnourished patients in hospital is needed to promote recovery. AIM: To describe the change in rates of detection and triaging of care for malnourished patients in 5 hospitals that were implementing an evidence-based nutrition care algorithm. To demonstrate that following this algorithm leads to increased detection of malnutrition and increased treatment to mitigate this condition. METHODS: Sites worked towards implementing the Integrated Nutrition Pathway for Acute Care (INPAC), including screening (Canadian Nutrition Screening Tool) and triage (Subjective Global Assessment; SGA) to detect and diagnose malnourished patients. Implementation occurred over a 24-month period, including developmental (Period 1), implementation (Periods 2-5), and sustainability (Period 6) phases. Audits (n = 36) of patient health records (n = 5030) were conducted to identify nutrition care practices implemented with a variety of strategies and behaviour change techniques. RESULTS: All sites increased nutrition screening from Period 1, with three achieving the goal of 75% of admitted patients being screened by Period 3, and the remainder achieving a rate of 70% by end of implementation. No sites were conducting SGA at Period 1, and sites reached the goal of a 75% completion rate or referral for those identified to be at nutrition risk, by Period 3 or 4. By Period 2, 100% of patients identified as SGA C (severely malnourished) were receiving a comprehensive nutritional assessment. In Period 1, the nutrition diagnosis and documentation by the dietitian of 'malnutrition' was a modest 0.37%, increasing to over 5% of all audited health records. The overall use of any Advanced Nutrition Care practices increased from 31% during Period 1 to 63% during Period 6. CONCLUSION: The success of this multi-site study demonstrated that implementation of nutrition screening and diagnosis is feasible and leads to appropriate care. INPAC promotes efficiency in nutrition care while minimizing the risk of missing malnourished patients. TRIAL REGISTRATION: Retrospectively registered ClinicalTrials.gov Identifier: NCT02800304, June 7, 2016.

Handgrip Strength, but Not 5-Meter Walk, Adds Value to a Clinical Nutrition Assessment.

BACKGROUND: Decreased physical functioning is associated with malnutrition and common in acute care patients; determining loss of function is often considered part of a comprehensive nutrition assessment. Handgrip strength (HGS) and 5-meter timed walk (5m) are functional measures used in a variety of settings. This analysis sought to determine which functional measure could be added to a hospital nutrition assessment, based on its feasibility and capacity to discriminate patient subgroups. METHODS: Eligible medical patients (no delirium/dementia, admitted from community; n = 1250), recruited from 5 hospitals that participated in a previous multisite action research study, provided data on demographics, HGS, 5m, nutrition status, perceived disability, and other characteristics. RESULTS: Significantly more patients (z = 17.39, P < .00001) were able to complete HGS than 5m (92% versus 43%, respectively). Median HGS was 28.0 kg for men and 14.7 kg for women. Of patients who completed the 5m, mean completion time was 8.98 seconds (median, 6.79 seconds, SD = 6.59). 5m and HGS scores were significantly worse with patient-perceived disability (z = -9.56, t = 10.69, respectively; P < .0001; 95% confidence interval [CI], [7.33, 10.63]; [1.76, 3.18]). HGS was associated with nutrition status (t = 4.13, P < .001; 95% CI [2.02, 5.67]), although it showed poor validity as a single nutrition indicator. CONCLUSIONS: These data indicate that HGS is a more useful functional measure than 5m when added to a hospital nutrition assessment. Determination of HGS cutpoints to identify low strength in acute care patients will promote its use.

CCL20, a direct-acting pro-angiogenic chemokine induced by hepatitis C virus (HCV): Potential role in HCV-related liver cancer.

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.

Update on the Integrated Nutrition Pathway for Acute Care (INPAC): post implementation tailoring and toolkit to support practice improvements.

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.