Best Practices and Recommendations for Research Using Virtual Real-Time Data Collection: Protocol for Virtual Data Collection Studies.

BACKGROUND: The COVID-19 pandemic and the subsequent need for social distancing required the immediate pivoting of research modalities. Research that had previously been conducted in person had to pivot to remote data collection. Researchers had to develop data collection protocols that could be conducted remotely with limited or no evidence to guide the process. Therefore, the use of web-based platforms to conduct real-time research visits surged despite the lack of evidence backing these novel approaches. OBJECTIVE: This paper aims to review the remote or virtual research protocols that have been used in the past 10 years, gather existing best practices, and propose recommendations for continuing to use virtual real-time methods when appropriate. METHODS: Articles (n=22) published from 2013 to June 2023 were reviewed and analyzed to understand how researchers conducted virtual research that implemented real-time protocols. "Real-time" was defined as data collection with a participant through a live medium where a participant and research staff could talk to each other back and forth in the moment. We excluded studies for the following reasons: (1) studies that collected participant or patient measures for the sole purpose of engaging in a clinical encounter; (2) studies that solely conducted qualitative interview data collection; (3) studies that conducted virtual data collection such as surveys or self-report measures that had no interaction with research staff; (4) studies that described research interventions but did not involve the collection of data through a web-based platform; (5) studies that were reviews or not original research; (6) studies that described research protocols and did not include actual data collection; and (7) studies that did not collect data in real time, focused on telehealth or telemedicine, and were exclusively intended for medical and not research purposes. RESULTS: Findings from studies conducted both before and during the COVID-19 pandemic suggest that many types of data can be collected virtually in real time. Results and best practice recommendations from the current protocol review will be used in the design and implementation of a substudy to provide more evidence for virtual real-time data collection over the next year. CONCLUSIONS: Our findings suggest that virtual real-time visits are doable across a range of participant populations and can answer a range of research questions. Recommended best practices for virtual real-time data collection include (1) providing adequate equipment for real-time data collection, (2) creating protocols and materials for research staff to facilitate or guide participants through data collection, (3) piloting data collection, (4) iteratively accepting feedback, and (5) providing instructions in multiple forms. The implementation of these best practices and recommendations for future research are further discussed in the paper. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/53790.

Seasonal human coronavirus NL63 epidemics in children in Guilin, China, reveal the emergence of a new subgenotype of HCoV-NL63.

Introduction: Seasonal human coronavirus NL63 (HCoV-NL63) is a frequently encountered virus linked to mild upper respiratory infections. However, its potential to cause more severe or widespread disease remains an area of concern. This study aimed to investigate a rare localized epidemic of HCoV-NL63-induced respiratory infections among pediatric patients in Guilin, China, and to understand the viral subtype distribution and genetic characteristics. Methods: In this study, 83 pediatric patients hospitalized with acute respiratory infections and positive for HCoV-NL63 were enrolled. Molecular analysis was conducted to identify the viral subgenotypes and to assess genetic variations in the receptor-binding domain of the spiking protein. Results: Among the 83 HCoV-NL63-positive children, three subgenotypes were identified: C4, C3, and B. Notably, 21 cases exhibited a previously unreported subtype, C4. Analysis of the C4 subtype revealed a unique amino acid mutation (I507L) in the receptor-binding domain of the spiking protein, which was also observed in the previously reported C3 genotype. This mutation may suggest potential increases in viral transmissibility and pathogenicity. Discussion: The findings of this study highlight the rapid mutation dynamics of HCoV-NL63 and its potential for increased virulence and epidemic transmission. The presence of a unique mutation in the C4 subtype, shared with the C3 genotype, raises concerns about the virus's evolving nature and its potential public health implications. This research contributes valuable insights into the understanding of HCoV-NL63's epidemiology and pathogenesis, which is crucial for effective disease prevention and control strategies. Future studies are needed to further investigate the biological significance of the observed mutation and its potential impact on the virus's transmissibility and pathogenicity.

Seroprevalence of the novel swine acute diarrhea syndrome coronavirus in China assessed by enzyme-linked immunosorbent assay.

The endemic outbreak of SADS-CoV has resulted in economic losses and potentially threatened the safety of China's pig industry. The molecular epidemiology of SADS-CoV in pig herds has been investigated in many provinces in China. However, there are no data over a long-time span, and there is a lack of extensive serological surveys to assess the prevalence of SADS-CoV in Chinese swine herds since the discovery of SADS-CoV. In this study, an indirect anti-SADS-CoV IgG enzyme-linked immunosorbent assay (ELISA) based on the SADS-CoV S1 protein was established to investigate the seroprevalence of SADS-CoV in Chinese swine herds. Cross-reactivity assays, indirect immunofluorescence, and western blotting assays showed that the developed ELISA had excellent SADS-CoV specificity. In total, 12,978 pig serum samples from 29 provinces/municipalities/autonomous regions in China were tested from 2022 to 2023. The results showed that the general seroprevalence of SADS-CoV in China was 59.97%, with seroprevalence ranging from 16.7% to 77.12% in different provinces and from 42.61% to 68.45% in different months. SADS-CoV is widely prevalent in China, and its seroprevalence was higher in Northeast China, North China, and Central China than in other regions. Among the four seasons, the prevalence of SADS-CoV was the highest in spring and the lowest in autumn. The results of this study provide the general seroprevalence profile of SADS-CoV in China, facilitating the understanding of the prevalence of SADS-CoV in pigs. More importantly, this study is beneficial in formulating preventive and control measures for SADS-CoV and may provide directions for vaccine development.

Lessons learned from the COVID-19 pandemic: The importance of physician leadership in responding to rural community ecosystem disruptions.

INTRODUCTION: The COVID-19 pandemic presented an unprecedented challenge for rural family physicians. The lessons learned over the course of 2 years have potential to help guide responses to future ecosystem disruption. This qualitative study aims to explore the leadership experiences of rural Canadian family physicians during the COVID-19 pandemic as both local care providers and community health leaders and to identify potential supports and barriers to physician leadership. METHODS: Semi-structured, virtual, qualitative interviews were completed with participants from rural communities in Canada from December 2021 to February 2022 inclusive. Participant recruitment involved identifying seed contacts and conducting snowball sampling. Participants were asked about their experiences during the COVID-19 pandemic, including the role of physician leadership in building community resilience. Data collection was completed on theoretical saturation. Data were thematically analysed using NVivo 12. RESULTS: Sixty-four participants took part from 22 rural communities in 4 provinces. Four key factors were identified that supported physician leadership towards rural resilience during ecosystem disruption: (1) continuity of care, (2) team-based care models, (3) physician well-being and (4) openness to innovative care models. CONCLUSION: Healthcare policy and practice transformation should prioritise developing opportunities to strengthen physician leadership, particularly in rural areas that will be adversely affected by ecosystem disruption. INTRODUCTION: La pandémie de COVID-19 a représenté un défi sans précédent pour les médecins de famille en milieu rural. Les leçons tirées au cours des deux années écoulées peuvent aider à orienter les réponses aux futures perturbations de l'écosystème. Cette étude qualitative vise à explorer les expériences de leadership des médecins de famille ruraux canadiens pendant la pandémie de COVID-19, en tant que prestataires de soins locaux et chefs de file de la santé communautaire, et à identifier les soutiens et les obstacles potentiels au leadership des médecins. MTHODES: Des entretiens qualitatifs virtuels semi-structurés ont été réalisés avec des participants issus de communautés rurales du Canada entre décembre 2021 et février 2022 inclus. Le recrutement des participants a consisté à identifier des contacts de base et à procéder à un échantillonnage boule de neige. Les participants ont été interrogés sur leurs expériences durant la pandémie de COVID-19, notamment sur le rôle du leadership des médecins dans le renforcement de la résilience des communautés. La collecte des données s'est achevée après saturation théorique. Les données ont été analysées thématiquement à l'aide de NVivo 12. RSULTATS: Soixante-quatre participants provenant de 22 communautés rurales de quatre provinces ont pris part à l'étude. Quatre facteurs clés ont été identifiés pour soutenir le leadership des médecins en faveur de la résilience rurale en cas de perturbation de l'écosystème: (1) la continuité des soins, (2) les modèles de soins en équipe, (3) le bien-être des médecins et (4) l'ouverture à des modèles de soins novateurs. CONCLUSION: La politique de santé et la transformation des pratiques devraient donner la priorité au développement d'opportunités pour renforcer le leadership des médecins, en particulier dans les zones rurales qui seront négativement affectées par la perturbation de l'écosystème.

A portable transistor immunosensor for fast identification of porcine epidemic diarrhea virus.

Widespread distribution of porcine epidemic diarrhea virus (PEDV) has led to catastrophic losses to the global pig farming industry. As a result, there is an urgent need for rapid, sensitive and accurate tests for PEDV to enable timely and effective interventions. In the present study, we develop and validate a floating gate carbon nanotubes field-effect transistor (FG CNT-FET)-based portable immunosensor for rapid identification of PEDV in a sensitive and accurate manner. To improve the affinity, a unique PEDV spike protein-specific monoclonal antibody is prepared by purification, and subsequently modified on FG CNT-FET sensor to recognize PEDV. The developed FET biosensor enables highly sensitive detection (LoD: 8.1 fg/mL and 100.14 TCID50/mL for recombinant spike proteins and PEDV, respectively), as well as satisfactory specificity. Notably, an integrated portable platform consisting of a pluggable FG CNT-FET chip and a portable device can discriminate PEDV positive from negative samples and even identify PEDV and porcine deltacoronavirus within 1 min with 100% accuracy. The portable sensing platform offers the capability to quickly, sensitively and accurately identify PEDV, which further points to a possibility of point of care (POC) applications of large-scale surveillance in pig breeding facilities.

Combination of S1-N-Terminal and S1-C-Terminal Domain Antigens Targeting Double Receptor-Binding Domains Bolsters Protective Immunity of a Nanoparticle Vaccine against Porcine Epidemic Diarrhea Virus.

Variants of coronavirus porcine epidemic diarrhea virus (PEDV) frequently emerge, causing an incomplete match between the vaccine and variant strains, which affects vaccine efficacy. Designing vaccines with rapidly replaceable antigens and high efficacy is a promising strategy for the prevention of infection with PEDV variant strains. In our study, three different types of self-assembled nanoparticles (nps) targeting receptor-binding N-terminal domain (NTD) and C-terminal domain (CTD) of S1 protein, named NTDnps, CTDnps, and NTD/CTDnps, were constructed and evaluated as vaccine candidates against PEDV. NTDnps and CTDnps vaccines mediated significantly higher neutralizing antibody (NAb) titers than NTD and CTD recombinant proteins in mice. The NTD/CTDnps in varying ratios elicited significantly higher NAb titers when compared with NTDnps and CTDnps alone. The NTD/CTDnps (3:1) elicited NAb with titers up to 92.92% of those induced by the commercial vaccine. Piglets immunized with NTD/CTDnps (3:1) achieved a passive immune protection rate of 83.33% of that induced by the commercial vaccine. NTD/CTDnps (3:1) enhanced the capacity of mononuclear macrophages and dendritic cells to take up and present antigens by activating major histocompatibility complex I and II molecules to stimulate humoral and cellular immunity. These data reveal that a combination of S1-NTD and S1-CTD antigens targeting double receptor-binding domains strengthens the protective immunity of nanoparticle vaccines against PEDV. Our findings will provide a promising vaccine candidate against PEDV.

Desorption Separation Ionization Mass Spectrometry (DSI-MS) for Rapid Analysis of COVID-19.

During the coronavirus disease 2019 (COVID-19) pandemic, which has witnessed over 772 million confirmed cases and over 6 million deaths globally, the outbreak of COVID-19 has emerged as a significant medical challenge affecting both affluent and impoverished nations. Therefore, there is an urgent need to explore the disease mechanism and to implement rapid detection methods. To address this, we employed the desorption separation ionization (DSI) device in conjunction with a mass spectrometer for the efficient detection and screening of COVID-19 urine samples. The study encompassed patients with COVID-19, healthy controls (HC), and patients with other types of pneumonia (OP) to evaluate their urine metabolomic profiles. Subsequently, we identified the differentially expressed metabolites in the COVID-19 patients and recognized amino acid metabolism as the predominant metabolic pathway involved. Furthermore, multiple established machine learning algorithms validated the exceptional performance of the metabolites in discriminating the COVID-19 group from healthy subjects, with an area under the curve of 0.932 in the blind test set. This study collectively suggests that the small-molecule metabolites detected from urine using the DSI device allow for rapid screening of COVID-19, taking just three minutes per sample. This approach has the potential to expand our understanding of the pathophysiological mechanisms of COVID-19 and offers a way to rapidly screen patients with COVID-19 through the utilization of machine learning algorithms.

Infectious bronchitis virus: an overview of the "chicken coronavirus".

Infectious bronchitis virus (IBV) is a highly contagious avian Gammacoronavirus that affects mainly chickens (Gallus gallus) but can circulate in other avian species. IBV constitutes a significant threat to the poultry industry, causing reduced egg yield, growth and mortality levels that can vary in impact. The virus can be transmitted horizontally by inhalation or direct/indirect contact with infected birds or contaminated fomites, vehicles, farm personnel and litter (Figure 1). The error-prone viral polymerase and recombination mechanisms mean diverse viral population results, with multiple genotypes, serotypes, pathotypes and protectotypes. This significantly complicates control and mitigation strategies based on vigilance in biosecurity and the deployment of vaccination.

SP2509, a specific antagonist of LSD1, exhibits antiviral properties against Porcine epidemic diarrhea virus.

BACKGROUND: Porcine epidemic diarrhea virus (PEDV), a type of coronavirus, is one of the main pathogens that can infect pigs of all ages. It causes diarrhea and acute death of newborn piglets, resulting in massive economic losses to the worldwide swine industry. While vaccination remains the primary approach in combating PEDV, it often fails to address all the challenges posed by the infection, particularly in light of the emergence of evolving mutant strains. Therefore, there is a critical need to identify potent antiviral drugs that can effectively safeguard pigs against PEDV infection. RESULTS: In this study, the antiviral efficacy of SP2509, a specific antagonist of Lysine-specific demethylase 1(LSD1), was evaluated in vitro. The RT-qPCR, Western blot, TCID50, and IFA showed that at a concentration of 1µmol/L, SP2509 significantly inhibited PEDV infection. Additionally, viral life cycle assays showed that SP2509 operates by impeding PEDV internalization and replication rather than attachment and release. Regarding mechanism, in Huh-7 cells, knockdowns LSD1 can suppress PEDV replication. This indicated that the inhibition effect of SP2509 on PEDV largely depends on the activity of its target protein, LSD1. CONCLUSION: Our results in vitro show that SP2509 can inhibit PEDV infection during the internalization and replication stage and revealed a role of LSD1 as a restriction factor for PEDV. These imply that LSD1 might be a target for interfering with the viral infection, and SP2509 could be developed as an effective anti-PEDV agent.

Quantitative assay to analyze neutralization and inhibition of authentic Middle East respiratory syndrome coronavirus.

To date, there is no licensed vaccine for Middle East respiratory syndrome coronavirus (MERS-CoV). Therefore, MERS-CoV is one of the diseases targeted by the Coalition for Epidemic Preparedness Innovations (CEPI) vaccine development programs and has been classified as a priority disease by the World Health Organization (WHO). An important measure of vaccine immunogenicity and antibody functionality is the detection of virus-neutralizing antibodies. We have developed and optimized a microneutralization assay (MNA) using authentic MERS-CoV and standardized automatic counting of virus foci. Compared to our standard virus neutralization assay, the MNA showed improved sensitivity when analyzing 30 human sera with good correlation of results (Spearman's correlation coefficient r = 0.8917, p value < 0.0001). It is important to use standardized materials, such as the WHO international standard (IS) for anti-MERS-CoV immunoglobulin G, to compare the results from clinical trials worldwide. Therefore, in addition to the neutralizing titers (NT50 = 1384, NT80 = 384), we determined the IC50 and IC80 of WHO IS in our MNA to be 0.67 IU/ml and 2.6 IU/ml, respectively. Overall, the established MNA is well suited to reliably quantify vaccine-induced neutralizing antibodies with high sensitivity.

COVID­19 detection from chest X-ray images using transfer learning.

COVID-19 is a kind of coronavirus that appeared in China in the Province of Wuhan in December 2019. The most significant influence of this virus is its very highly contagious characteristic which may lead to death. The standard diagnosis of COVID-19 is based on swabs from the throat and nose, their sensitivity is not high enough and so they are prone to errors. Early diagnosis of COVID-19 disease is important to provide the chance of quick isolation of the suspected cases and to decrease the opportunity of infection in healthy people. In this research, a framework for chest X-ray image classification tasks based on deep learning is proposed to help in early diagnosis of COVID-19. The proposed framework contains two phases which are the pre-processing phase and classification phase which uses pre-trained convolution neural network models based on transfer learning. In the pre-processing phase, different image enhancements have been applied to full and segmented X-ray images to improve the classification performance of the CNN models. Two CNN pre-trained models have been used for classification which are VGG19 and EfficientNetB0. From experimental results, the best model achieved a sensitivity of 0.96, specificity of 0.94, precision of 0.9412, F1 score of 0.9505 and accuracy of 0.95 using enhanced full X-ray images for binary classification of chest X-ray images into COVID-19 or normal with VGG19. The proposed framework is promising and achieved a classification accuracy of 0.935 for 4-class classification.

Characterization and immunogenicity assessment of MERS-CoV pre-fusion spike trimeric oligomers as vaccine immunogen.

Middle East respiratory syndrome coronavirus (MERS-CoV) is a lethal beta-coronavirus that emerged in 2012. The virus is part of the WHO blueprint priority list with a concerning fatality rate of 35%. Scientific efforts are ongoing for the development of vaccines, anti-viral and biotherapeutics, which are majorly directed toward the structural spike protein. However, the ongoing effort is challenging due to conformational instability of the spike protein and the evasion strategy posed by the MERS-CoV. In this study, we have expressed and purified the MERS-CoV pre-fusion spike protein in the Expi293F mammalian expression system. The purified protein was extensively characterized for its biochemical and biophysical properties. Thermal stability analysis showed a melting temperature of 58°C and the protein resisted major structural changes at elevated temperature as revealed by fluorescence spectroscopy and circular dichroism. Immunological assessment of the MERS-CoV spike immunogen in BALB/c mice with AddaVaxTM and Imject alum adjuvants showed elicitation of high titer antibody responses but a more balanced Th1/Th2 response with AddaVaxTM squalene like adjuvant. Together, our results suggest the formation of higher-order trimeric pre-fusion MERS-CoV spike proteins, which were able to induce robust immune responses. The comprehensive characterization of MERS-CoV spike protein warrants a better understanding of MERS spike protein and future vaccine development efforts.

Genetic characterization of bovine coronavirus strain isolated in Inner Mongolia of China.

BACKGROUND: Bovine coronavirus (BCoV) is implicated in severe diarrhea in calves and contributes to the bovine respiratory disease complex; it shares a close relationship with human coronavirus. Similar to other coronaviruses, remarkable variability was found in the genome and biology of the BCoV. In 2022, samples of feces were collected from a cattle farm. A virus was isolated from 7-day-old newborn calves. In this study, we present the genetic characteristics of a new BCoV isolate. The complete genomic, spike protein, and nucleocapsid protein gene sequences of the BCoV strain, along with those of other coronaviruses, were obtained from the GenBank database. Genetic analysis was conducted using MEGA7.0 and the Neighbor-Joining (NJ) method. The reference strains' related genes were retrieved from GenBank for comparison and analysis using DNAMAN. RESULTS: The phylogenetic tree and whole genome consistency analysis showed that it belonged to the GIIb subgroup, which is epidemic in Asia and America, and was quite similar to the Chinese strains in the same cluster. Significantly, the S gene was highly consistent with QH1 (MH810151.1) isolated from yak. This suggests that the strain may have originated from interspecies transmission involving mutations of wild strains. The N gene was conserved and showed high sequence identity with the epidemic strains in China and the USA. CONCLUSIONS: Genetic characterization suggests that the isolated strain could be a new mutant from a wild-type lineage, which is in the same cluster as most Chinese epidemic strains but on a new branch.

SCARLET (Supplemental Citicoline Administration to Reduce Lung injury Efficacy Trial): study protocol for a single-site, double-blinded, placebo-controlled, and randomized Phase 1/2 trial of i.v. citicoline (CDP-choline) in hospitalized SARS CoV-2-infected patients with hypoxemic acute respiratory failure.

BACKGROUND: The SARS CoV-2 pandemic has resulted in more than 1.1 million deaths in the USA alone. Therapeutic options for critically ill patients with COVID-19 are limited. Prior studies showed that post-infection treatment of influenza A virus-infected mice with the liponucleotide CDP-choline, which is an essential precursor for de novo phosphatidylcholine synthesis, improved gas exchange and reduced pulmonary inflammation without altering viral replication. In unpublished studies, we found that treatment of SARS CoV-2-infected K18-hACE2-transgenic mice with CDP-choline prevented development of hypoxemia. We hypothesize that administration of citicoline (the pharmaceutical form of CDP-choline) will be safe in hospitalized SARS CoV-2-infected patients with hypoxemic acute respiratory failure (HARF) and that we will obtain preliminary evidence of clinical benefit to support a larger Phase 3 trial using one or more citicoline doses. METHODS: We will conduct a single-site, double-blinded, placebo-controlled, and randomized Phase 1/2 dose-ranging and safety study of Somazina® citicoline solution for injection in consented adults of any sex, gender, age, or ethnicity hospitalized for SARS CoV-2-associated HARF. The trial is named "SCARLET" (Supplemental Citicoline Administration to Reduce Lung injury Efficacy Trial). We hypothesize that SCARLET will show that i.v. citicoline is safe at one or more of three doses (0.5, 2.5, or 5 mg/kg, every 12 h for 5 days) in hospitalized SARS CoV-2-infected patients with HARF (20 per dose) and provide preliminary evidence that i.v. citicoline improves pulmonary outcomes in this population. The primary efficacy outcome will be the SpO2:FiO2 ratio on study day 3. Exploratory outcomes include Sequential Organ Failure Assessment (SOFA) scores, dead space ventilation index, and lung compliance. Citicoline effects on a panel of COVID-relevant lung and blood biomarkers will also be determined. DISCUSSION: Citicoline has many characteristics that would be advantageous to any candidate COVID-19 therapeutic, including safety, low-cost, favorable chemical characteristics, and potentially pathogen-agnostic efficacy. Successful demonstration that citicoline is beneficial in severely ill patients with SARS CoV-2-induced HARF could transform management of severely ill COVID patients. TRIAL REGISTRATION: The trial was registered at www. CLINICALTRIALS: gov on 5/31/2023 (NCT05881135). TRIAL STATUS: Currently enrolling.

Nucleic acid vaccine candidates encapsulated with mesoporous silica nanoparticles against MERS-CoV.

Middle East respiratory coronavirus (MERS-CoV) is a newly emergent, highly pathogenic coronavirus that is associated with 34% mortality rate. MERS-CoV remains listed as priority pathogen by the WHO. Since its discovery in 2012 and despite the efforts to develop coronaviruses vaccines to fight against SARS-CoV-2, there are currently no MERS-CoV vaccine that has been approved. Therefore, there is high demand to continue on the development of prophylactic vaccines against MERS-CoV. Current advancements in vaccine developments can be adapted for the development of improved MERS-CoV vaccines candidates. Nucleic acid-based vaccines, including pDNA and mRNA, are relatively new class of vaccine platforms. In this work, we developed pDNA and mRNA vaccine candidates expressing S.FL gene of MERS-CoV. Further, we synthesized a silane functionalized hierarchical aluminosilicate to encapsulate each vaccine candidates. We tested the nucleic acid vaccine candidates in mice and evaluated humoral antibodies response. Interestingly, we determined that the non-encapsulated, codon optimized S.FL pDNA vaccine candidate elicited the highest level of antibody responses against S.FL and S1 of MERS-CoV. Encapsulation of mRNA with nanoporous aluminosilicate increased the humoral antibody responses, whereas encapsulation of pDNA did not. These findings suggests that MERS-CoV S.FL pDNA vaccine candidate induced the highest level of humoral responses. This study will enhance further optimization of nanosilica as potential carrier for mRNA vaccines. In conclusion, this study suggests MERS-CoV pDNA vaccine candidate as a suitable vaccine platform for further pivotal preclinical testings.

Inhibition Mechanism of SARS-CoV-2 Infection by a Cholesterol Derivative, Nat-20(S)-yne.

The coronavirus disease 2019 (COVID-19) is caused by the etiological agent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19, with the recurrent epidemics of new variants of SARS-CoV-2, remains a global public health problem, and new antivirals are still required. Some cholesterol derivatives, such as 25-hydroxycholesterol, are known to have antiviral activity against a wide range of enveloped and non-enveloped viruses, including SARS-CoV-2. At the entry step of SARS-CoV-2 infection, the viral envelope fuses with the host membrane dependent of viral spike (S) glycoproteins. From the screening of cholesterol derivatives, we found a new compound 26,27-dinorcholest-5-en-24-yne-3ß,20-diol (Nat-20(S)-yne) that inhibited the SARS-CoV-2 S protein-dependent membrane fusion in a syncytium formation assay. Nat-20(S)-yne exhibited the inhibitory activities of SARS-CoV-2 pseudovirus entry and intact SARS-CoV-2 infection in a dose-dependent manner. Among the variants of SARS-CoV-2, inhibition of infection by Nat-20(S)-yne was stronger in delta and Wuhan strains, which predominantly invade into cells via fusion at the plasma membrane, than in omicron strains. The interaction between receptor-binding domain of S proteins and host receptor ACE2 was not affected by Nat-20(S)-yne. Unlike 25-hydroxycholesterol, which regulates various steps of cholesterol metabolism, Nat-20(S)-yne inhibited only de novo cholesterol biosynthesis. As a result, plasma membrane cholesterol content was substantially decreased in Nat-20(S)-yne-treated cells, leading to inhibition of SARS-CoV-2 infection. Nat-20(S)-yne having a new mechanism of action may be a potential therapeutic candidate for COVID-19.

Lactobacillus rhamnosus GG powder supplementation alleviates intestinal injury in piglets challenged by porcine epidemic diarrhea virus.

Porcine epidemic diarrhea virus (PEDV) has become a challenging problem in pig industry worldwide, causing significant profit losses. Lactobacillus rhamnosus GG (LGG) has been regarded as a safe probiotic strain and has been shown to exert protective effects on the intestinal dysfunction caused by PEDV. This study evaluated the effect of LGG on the gut health of lactating piglets challenged with PEDV. Fifteen piglets at 7 days of age were equally assigned into 3 groups (5 piglets per group): 1) control group (basal diet); 2) PEDV group: (basal diet + PEDV challenged); 3) LGG + PEDV group (basal diet + 3×109 CFU/pig/day LGG + PEDV). The trial lasted 11 days including 3 days of adaptation. The treatment with LGG was from D4 to D10. PEDV challenge was carried out on D8. PEDV infection disrupted the cell structure, undermined the integrity of the intestinal tract, and induced oxidative stress, and intestinal damage of piglets. Supplementation of LGG improved intestinal morphology, enhanced intestinal antioxidant capacity, and alleviated jejunal mucosal inflammation and lipid metabolism disorders in PEDV-infected piglets, which may be regulated by LGG by altering the expression of TNF signaling pathway, PPAR signaling pathway, and fat digestion and absorption pathway.

Interferon signaling in the nasal epithelium distinguishes among lethal and common cold coronaviruses and mediates viral clearance.

All respiratory viruses establish primary infections in the nasal epithelium, where efficient innate immune induction may prevent dissemination to the lower airway and thus minimize pathogenesis. Human coronaviruses (HCoVs) cause a range of pathologies, but the host and viral determinants of disease during common cold versus lethal HCoV infections are poorly understood. We model the initial site of infection using primary nasal epithelial cells cultured at an air-liquid interface (ALI). HCoV-229E, HCoV-NL63, and human rhinovirus-16 are common cold-associated viruses that exhibit unique features in this model: early induction of antiviral interferon (IFN) signaling, IFN-mediated viral clearance, and preferential replication at nasal airway temperature (33 °C) which confers muted host IFN responses. In contrast, lethal SARS-CoV-2 and MERS-CoV encode antagonist proteins that prevent IFN-mediated clearance in nasal cultures. Our study identifies features shared among common cold-associated viruses, highlighting nasal innate immune responses as predictive of infection outcomes and nasally directed IFNs as potential therapeutics.

More than what meets the eye in COVID-19 critical illness: A case report of bilateral femoral neuropathy due to psoas hematomas.

Bilateral femoral neuropathy is rare, especially that caused by bilateral compressive iliopsoas, psoas, or iliacus muscle hematomas. We present a case of bilateral femoral neuropathy due to spontaneous psoas hematomas developed during COVID-19 critical illness. A 41-year-old patient developed COVID-19 pneumonia, and his condition deteriorated rapidly. A decrease in the hemoglobin level prompted imaging studies during his intensive care unit (ICU) stay. Bilateral psoas hematomas were identified as the source of bleeding. Thereafter, the patient complained of weakness in both upper and lower limbs and numbness in the lower limb. He was considered to have critical illness neuropathy and was referred to rehabilitation. Electrodiagnostic testing suggested bilateral femoral neuropathy because of compression due to hematomas developed during the course of his ICU stay. The consequences of iliopsoas hematomas occurring in the critically ill can be catastrophic, ranging from hemorrhagic shock to severe weakness, highlighting the importance of recognizing this entity.

Evidence for cross-species transmission of human coronavirus OC43 through bioinformatics and modeling infections in porcine intestinal organoids.

Cross-species transmission of coronaviruses has been continuously posing a major challenge to public health. Pigs, as the major animal reservoirs for many zoonotic viruses, frequently mediate viral transmission to humans. This study comprehensively mapped the relationship between human and porcine coronaviruses through in-depth bioinformatics analysis. We found that human coronavirus OC43 and porcine coronavirus PHEV share a close phylogenetic relationship, evidenced by high genomic homology, similar codon usage patterns and comparable tertiary structure in spike proteins. Inoculation of infectious OC43 viruses in organoids derived from porcine small and large intestine demonstrated that porcine intestinal organoids (pIOs) are highly susceptible to human coronavirus OC43 infection and support infectious virus production. Using transmission electron microscopy, we visualized OC43 viral particles in both intracellular and extracellular compartments, and observed abnormalities of multiple organelles in infected organoid cells. Robust OC43 infections in pIOs result in a significant reduction of organoids viability and widespread cell death. This study bears essential implications for better understanding the evolutionary origin of human coronavirus OC43, and provides a proof-of-concept for using pIOs as a model to investigate cross-species transmission of human coronavirus.