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Schisandrin B (Sch B) is a predominant bioactive lignan from the fruit of a Chinese medicine food homology plant, Schisandra chinensis. Previously, we observed potent anti-tumor effect of Sch-B in colorectal cancer (CRC) and enhanced chemotherapy efficacy with fluorouracil (5-FU). However, their bioavailability and reciprocal interactions under CRC conditions are unclear. In this study, we first compared the bioavailability, metabolism and tissue distribution of Sch-B between non-tumor-bearing and xenograft CRC tumor-bearing mice. Next, we examined SchB-5-FU interactions via investigating alterations in drug metabolism and multidrug resistance. Using a validated targeted metabolomics approach, five active metabolites, including Sch-B and fluorodeoxyuridine triphosphate, were found tumor-accumulative. Co-treatment resulted in higher levels of Sch-B and 5-FU metabolites, showing improved phytochemical and drug bioavailability. Multidrug resistance gene (MDR1) was significantly downregulated upon co-treatment. Overall, we demonstrated the potential of Sch-B to serve as a promising chemotherapy adjuvant via improving drug bioavailability and metabolism, and attenuating MDR., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Ltd.)

Introduction: This study investigates the impact of different feeding methods (direct breastfeeding, expressed milk feeding, formula feeding) on the infant microbiota at 6 weeks of age., Methods: A total of 217 healthy infants stool samples were collected from Hong Kong between August 2018 and December 2019., Results: Various microbial taxa, including the genera Enterobacter and Raoultella were identified in the expressed breast milk feeding group. The richness and composition of the major bacterial phyla showed similar abundance between direct breastfeeding and expressed breast milk., Discussion: These findings suggests that these bacteria may have colonized the milk during expression or could be introduced from other external sources. The mode of breastfeeding did not significantly alter microbiota parameters in the infant gut at 6 weeks., Competing Interests: Conflicts of interest None to report., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Tissue engineering for penile corpora cavernosa defects requires microvascular system reconstruction.GelMA hydrogels show promise for tissue regeneration. However, using stem cells faces challenges such as immune rejection, limited proliferation and differentiation, and biosafety concerns. Therefore, acellular tissue regeneration may avoid these issues. Exosomes are used from muscle-derived stem cells (MDSCs) to modify 3D-printed hydrogel scaffolds for acellular tissue regeneration. Hypoxia-preconditioned MDSC-derived exosomes are obtained to enhance the therapeutic effect. In contrast to normoxic exosomes (N-Exos), hypoxic exosomes (H-Exos) are found to markedly enhance the proliferation, migration, and capillary-like tube formation of human umbilical vein endothelial cells (HUVECs). High-throughput sequencing analysis of miRNAs isolated from both N-Exos and H-Exos revealed a significant upregulation of miR-21-5p in H-Exos following hypoxic preconditioning. Further validation demonstrated that the miR-21-5p/PDCD4 pathway promoted the proliferation of HUVECs. Epigallocatechin gallate (EGCG) is introduced to improve the mechanical properties and biocompatibility of GelMA hydrogels. EGCG-GelMA scaffolds loaded with different types of Exos are transplanted to repair rabbit penile corpora cavernosa defects, observed the blood flow and repair status of the defect site through color Doppler ultrasound and magnetic resonance imaging, and ultimately restored the rabbit penile erection function and successfully bred offspring. Thus, acellular hydrogel scaffolds offer an effective treatment for penile corpora cavernosa defects., (© 2024 Wiley‐VCH GmbH.)

Background and Aim: We aimed to investigate the effect of metabolic dysfunction-associated steatotic liver disease (MASLD) on three-dose BNT162b2 immunogenicity to the omicron variant., Methods: Adult recipients of three doses of BNT162b2 were prospectively recruited between May and December 2021. The serology of the neutralizing antibody by live virus microneutralization (vMN) to the omicron variant was measured at baseline, day 180, and day 360 after the first dose. The primary outcome was seroconversion (vMN titer ≥ 10) at day 360. Exposure of interest was MASLD, defined as hepatic steatosis (controlled attenuation parameter ≥ 248 dB/m on transient elastography) plus at least one of five cardiometabolic risk factors. Subjects with prior COVID-19 were excluded. A multivariable logistic regression model was used to derive the adjusted odds ratio of seroconversion with MASLD by adjusting for age, sex, antibiotic use, and proton pump inhibitor use., Results: One hundred forty-eight BNT162b2 recipients (male: 48 [32.4%]; median age: 51.0 years [interquartile range, IQR: 44.5-57.3]) were recruited. The median time from the first dose to the third dose was 8.5 months (IQR: 7.9-8.9). MASLD subjects had a lower seroconversion rate than non-MASLD ones (89.6% vs 99.0%; P = 0.007). MASLD was the only independent risk factor for seroconversion (adjusted odds ratio: 0.051, 95% confidence interval: 0.002-0.440). Subgroup analysis of immunogenicity at 4 months after the third dose shows significantly lower vMN titer (13.06 [IQR: 7.69-22.20] vs 33.49 [IQR: 24.05-46.53]; P = 0.004) and seroconversion rate (76.9% vs 97.4%; P = 0.016) in MASLD than non-MASLD subjects, but not within 4 months from the third dose (vMN titer: 46.87 [IQR: 33.12-66.02] vs 41.86 [IQR: 34.47-50.91], P = 0.240; seroconversion rate: 94.3% vs 100%, P = 0.131)., Conclusion: Metabolic dysfunction-associated steatotic liver disease was a risk factor for poorer immunogenicity to the omicron variant, with a more pronounced waning effect compared among three-dose BNT162b2 recipients., (© 2024 The Author(s). Journal of Gastroenterology and Hepatology published by Journal of Gastroenterology and Hepatology Foundation and John Wiley & Sons Australia, Ltd.)

Pseudomonas aeruginosa infections are difficult to treat due to rapid development of antibiotic drug resistance. The synergistic combination of already-in-use drugs is an alternative to developing new antibiotics to combat antibiotic-resistant bacteria. Here we demonstrate that bismuth-based drugs (bismuth subsalicylate, colloidal bismuth subcitrate) in combination with different classes of antibiotics (tetracyclines, macrolides, quinolones, rifamycins and so on) can eliminate multidrug-resistant P. aeruginosa and do not induce development of antibiotic resistance. Bismuth disrupts iron homeostasis by binding to P. aeruginosa siderophores. Inside cells, bismuth inhibits the electron transport chain, dissipates the proton motive force and impairs efflux pump activity by disrupting iron-sulfur cluster-containing enzymes, including respiration complexes. As a result, bismuth facilitates antibiotic accumulation inside bacteria, enhancing their efficacy. The combination therapy shows potent antibacterial efficacy and low toxicity in an ex vivo bacteraemia model and increases the survival rate of mice in in vivo mouse lung-infection models. Our findings highlight the potential of bismuth-based drugs to be repurposed to combat P. aeruginosa infections in combination with clinically used antibiotics., (© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)

SARS-CoV-2 entry into host cells is facilitated by the interaction between the receptor-binding domain of its spike protein (CoV2-RBD) and host cell receptor, ACE2, promoting viral membrane fusion. The virus also uses endocytic pathways for entry, but the mediating host factors remain largely unknown. It is also unknown whether mutations in the RBD of SARS-CoV-2 variants promote interactions with additional host factors to promote viral entry. Here, we used the GST pull-down approach to identify novel surface-located host factors that bind to CoV2-RBD. One of these factors, SH3BP4, regulates internalization of CoV2-RBD in an ACE2-independent but integrin- and clathrin-dependent manner and mediates SARS-CoV-2 pseudovirus entry, suggesting that SH3BP4 promotes viral entry via the endocytic route. Many of the identified factors, including SH3BP4, ADAM9, and TMEM2, show stronger affinity to CoV2-RBD than to RBD of the less infective SARS-CoV, suggesting SARS-CoV-2-specific utilization. We also found factors preferentially binding to the RBD of the SARS-CoV-2 Delta variant, potentially enhancing its entry. These data identify the repertoire of host cell surface factors that function in the events leading to the entry of SARS-CoV-2., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

SARS-CoV-2, the causative agent of COVID-19, has been intensely studied in search of effective antiviral treatments. The immunosuppressant cyclosporine A (CsA) has been suggested to be a pan-coronavirus inhibitor, yet its underlying mechanism remained largely unknown. Here, we found that non-structural protein 1 (Nsp1) of SARS-CoV-2 usurped CsA-suppressed nuclear factor of activated T cells (NFAT) signaling to drive the expression of cellular DEAD-box helicase 5 (DDX5), which facilitates viral replication. Nsp1 interacted with calcineurin A (CnA) to displace the regulatory protein regulator of calcineurin 3 (RCAN3) of CnA for NFAT activation. The influence of NFAT activation on SARS-CoV-2 replication was also validated by using the Nsp1-deficient mutant virus. Calcineurin inhibitors, such as CsA and VIVIT, inhibited SARS-CoV-2 replication and exhibited synergistic antiviral effects when used in combination with nirmatrelvir. Our study delineated the molecular mechanism of CsA-mediated inhibition of SARS-CoV-2 replication and the anti-SARS-CoV-2 action of calcineurin inhibitors., Importance: Cyclosporine A (CsA), commonly used to inhibit immune responses, is also known to have anti-SARS-CoV-2 activity, but its mode of action remains elusive. Here, we provide a model to explain how CsA antagonizes SARS-CoV-2 through three critical proteins: DDX5, NFAT1, and Nsp1. DDX5 is a cellular facilitator of SARS-CoV-2 replication, and NFAT1 controls the production of DDX5. Nsp1 is a viral protein absent from the mature viral particle and capable of activating the function of NFAT1 and DDX5. CsA and similar agents suppress Nsp1, NFAT1, and DDX5 to exert their anti-SARS-CoV-2 activity either alone or in combination with Paxlovid., Competing Interests: The authors declare no conflict of interest.

SARS-CoV-2 nsp14 functions both as an exoribonuclease (ExoN) together with its critical cofactor nsp10 and as an S -adenosyl methionine-dependent (guanine-N7) methyltransferase (MTase), which makes it an attractive target for the development of pan-anti-SARS-CoV-2 drugs. Herein, we screened a panel of compounds (and drugs) and found that certain compounds, especially Bi(III)-based compounds, could allosterically inhibit both MTase and ExoN activities of nsp14 potently. We further demonstrated that Bi(III) binds to both nsp14 and nsp10, resulting in the release of Zn(II) ions from the enzymes as well as alternation of protein quaternary structures. The in vitro activities of the compounds were also validated in SARS-CoV-2-infected mammalian cells. Importantly, we showed that nsp14 serves as an authentic target of Bi(III)-based antivirals in SARS-CoV-2-infected mammalian cells by quantification of both the protein and inhibitor. This study highlights the importance of nsp14/nsp10 as a potential target for the development of pan-antivirals against SARS-CoV-2 infection.

Mapping mutations and discovering cellular determinants that cause the spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to induce infected cells to form syncytia would facilitate the development of strategies for blocking the formation of such cell-cell fusion. Here we describe high-throughput screening methods based on droplet microfluidics and the size-exclusion selection of syncytia, coupled with large-scale mutagenesis and genome-wide knockout screening via clustered regularly interspaced short palindromic repeats (CRISPR), for the large-scale identification of determinants of cell-cell fusion. We used the methods to perform deep mutational scans in spike-presenting cells to pinpoint mutable syncytium-enhancing substitutions in two regions of the spike protein (the fusion peptide proximal region and the furin-cleavage site). We also used a genome-wide CRISPR screen in cells expressing the receptor angiotensin-converting enzyme 2 to identify inhibitors of clathrin-mediated endocytosis that impede syncytium formation, which we validated in hamsters infected with SARS-CoV-2. Finding genetic and cellular determinants of the formation of syncytia may reveal insights into the physiological and pathological consequences of cell-cell fusion., (© 2023. The Author(s).)

Background: Much attention has been paid to advocate proper oral care/hygiene provision by healthcare providers in long-term care institutions (LTCIs). This study aimed to evaluate the effects of an oral health education (OHE) programme (intervention) on knowledge, attitudes, and practice (KAP) of healthcare providers in providing oral care/hygiene to older residents in LTCIs., Methods: A case control study was conducted at two LTCIs, with one assigned as the intervention group and the other as the control group. A KAP survey was administered before and after the intervention, and oral status was assessed by standardized clinical photographs taken before and after oral hygiene provision on three older residents., Results: A total of 40 healthcare providers (20 in intervention and 20 in control groups) participated, with the attitudes and overall KAP significantly improved in the intervention group after the OHE programme. Interestingly, the knowledge of those in the control LTCI was significantly declined at re-evaluation (mean scores were from 17.25 to 14.30), indicating inadequate oral health and care training despite having more experience in taking care of older people. Significant differences in practice were observed between the two groups after the OHE programme ( p = 0.006). The three older residents exhibited poor oral health and multiple oral problems., Conclusions: This study revealed that the OHE programme effectively improved attitudes of the healthcare providers and provided a sustaining effect on attitude towards oral health and oral care. However, there were still inadequacies in oral hygiene provision by some healthcare providers, possibly due to unattended oral diseases and hygiene needs, as well as personal and environmental barriers that merit further investigation. Regular evaluation and enforcement of oral care/hygiene provision in LTCIs are necessary to maintain oral health and prevent dental and gum diseases in older residents. Immediate referral for dental treatment is recommended for older people with signs of dental/oral disease(s).

The rapid emergence of antimicrobial resistance (AMR) pathogens highlights the urgent need to approach this global burden with alternative strategies. Cefiderocol (Fetroja®) is a clinically-used sideromycin, that is utilized for the treatment of severe drug-resistant infections, caused by Gram-negative bacteria; there is evidence of cefiderocol-resistance occurring in bacterial strains however. To increase the efficacy and extend the life-span of sideromycins, we demonstrate strong synergisms between cefiderocol and metallodrugs (e.g., colloidal bismuth citrate (CBS)), against Pseudomonas aeruginosa and Burkholderia cepacia. Moreover, CBS enhances cefiderocol efficacy against biofilm formation, suppresses the resistance development in P. aeruginosa and resensitizes clinically isolated resistant P. aeruginosa to cefiderocol. Notably, the co-therapy of CBS and cefiderocol significantly increases the survival rate of mice and decreases bacterial loads in the lung in a murine acute pneumonia model. The observed phenomena are partially attributable to the competitive binding of Bi 3+ to cefiderocol with Fe 3+ , leading to enhanced uptake of Bi 3+ and reduced levels of Fe 3+ in cells. Our studies provide insight into the antimicrobial potential of metallo-sideromycins., (© 2023. Springer Nature Limited.)

To identify ancestry-linked genetic risk variants associated with COVID-19 hospitalization, we performed an integrative analysis of two genome-wide association studies and resolved four single nucleotide polymorphisms more frequent in COVID-19-hospitalized patients with non-European ancestry. Among them, the COVID-19 risk SNP rs16831827 shows the largest difference in minor allele frequency (MAF) between populations with African and European ancestry and also shows higher MAF in hospitalized COVID-19 patients among cohorts of mixed ancestry (odds ratio [OR] = 1.20, 95% CI: 1.10-1.30) and entirely African ancestry (OR = 1.30, 95% CI: 1.02-1.67). rs16831827 is an expression quantitative trait locus of MAP3K19 . MAP3K19 expression is induced during ciliogenesis and most abundant in ciliated tissues including lungs. Single-cell RNA sequencing analyses revealed that MAP3K19 is highly expressed in multiple ciliated cell types. As rs16831827∗T is associated with reduced MAP3K19 expression, it may increase the risk of severe COVID-19 by reducing MAP3K19 expression., Competing Interests: The authors declare no competing interests., (© 2023 The Author(s).)

Objectives: A partnership model in interprofessional education (IPE) is important in promoting a sense of global citizenship while preparing students for cross-sector problem-solving. However, the literature remains scant in providing useful guidance for the development of an IPE programme co-implemented by external partners. In this pioneering study, we describe the processes of forging global partnerships in co-implementing IPE and evaluate the programme in light of the preliminary data available., Methods: This study is generally quantitative. We collected data from a total of 747 health and social care students from four higher education institutions. We utilized a descriptive narrative format and a quantitative design to present our experiences of running IPE with external partners and performed independent t-tests and analysis of variance to examine pretest and posttest mean differences in students' data., Results: We identified factors in establishing a cross-institutional IPE programme. These factors include complementarity of expertise, mutual benefits, internet connectivity, interactivity of design, and time difference. We found significant pretest-posttest differences in students' readiness for interprofessional learning (teamwork and collaboration, positive professional identity, roles, and responsibilities). We also found a significant decrease in students' social interaction anxiety after the IPE simulation., Conclusions: The narrative of our experiences described in this manuscript could be considered by higher education institutions seeking to forge meaningful external partnerships in their effort to establish interprofessional global health education., (© 2023. The Author(s).)

Objectives: The aim of this study was to estimate carbapenem resistance in Pseudomonas aeruginosa and Enterobacterales isolated from infected patients in intensive care unit (ICU) and non-ICU hospital wards in Hong Kong., Methods: Isolates of Pseudomonas aeruginosa (ICU, n = 35; non-ICU, n = 264) and Enterobacterales (ICU, n = 129; non-ICU, n = 1390) were collected in four Hong Kong hospitals in 2017-2020. Clinical and Laboratory Standards Institute broth microdilution minimum inhibitory concentrations (MICs) were interpreted according to Clinical and Laboratory Standards Institute 2021 M100 breakpoints. β-lactamase genes were identified in imipenem-, imipenem/relebactam-, and ceftolozane/tazobactam-nonsusceptible isolates., Results: Ceftolozane/tazobactam demonstrated potent in vitro activity against both P. aeruginosa (ICU, 88.6%; non-ICU, 98.5%) and Enterobacterales (96.1%; 97.1%). Percent susceptible values for P. aeruginosa isolates from ICU and non-ICU patients, respectively, were as follows: meropenem (ICU, 74.3%; non-ICU, 84.1%) and imipenem (68.6%; 73.1%). Only 1 of 77 isolates tested for β-lactamase genes carried a carbapenemase (VIM-2). Percent susceptible values for Enterobacterales isolates from ICU and non-ICU patients were as follows: meropenem (100%; 99.4%), ertapenem (100%; 98.0%), and imipenem (88.4%; 88.6%). A total of 62 Enterobacterales isolates were tested for β-lactamase genes. Only three isolates carried a carbapenemase gene; two (both Escherichia coli) were metallo-β-lactamase-positive (both NDM-5), and one (Klebsiella pneumoniae) was OXA-48-like-positive., Conclusions: Carbapenem-nonsusceptible isolates of P. aeruginosa were common (>15% of isolates). P. aeruginosa percent susceptible values for ceftolozane/tazobactam (97.3% susceptible overall) were ≥14% higher than those for carbapenems in both ICU and non-ICU isolates. Carbapenemases were rare among both P. aeruginosa (one isolate) and Enterobacterales (three isolates). Most Enterobacterales isolates tested from ICU and non-ICU patients in Hong Kong hospitals in 2017-2020 were susceptible to meropenem and ertapenem (≥98%); imipenem was less active (89% susceptible)., (Copyright © 2023 Merck Sharp & Dohme LLC., a subsidiary Merck & Co., Inc, The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Purpose: We evaluated the evolution of thyroid function and autoimmunity among COVID-19 survivors over 6 months in relation to interferon beta-1b treatment and long COVID., Methods: We included COVID-19 survivors managed in a major COVID-19 centre between July 2020 and May 2021 who were reassessed three and/or six months after acute COVID-19. Thyroid function tests (TFTs) and anti-thyroid antibody titres were measured at acute COVID-19, 3-month and 6-month., Results: 250 COVID-19 survivors were included (mean age 52.7 years, 50.4% men). Persistent thyroid function abnormalities were more likely in those with abnormal TFTs in acute COVID-19 (P < 0.001). Among 51 patients with abnormal TFTs in acute COVID-19, 82.4% resolved upon follow-up. Of 199 patients with normal TFTs in acute COVID-19, only 4.5% had incident abnormal TFTs, more likely in interferon-treated patients (P = 0.044) and none clinically overt. Among 129 patients with complete 6-month follow-up for anti-thyroid antibody titres, there was no significant change overall, except for modest increase in anti-thyroid antibody titres among the 84 interferon-treated patients (P < 0.05 at both 3 months and 6 months). Long COVID occurred in 19.5% and 10.4% at 3 and 6 months respectively, where TFTs and anti-thyroid antibody titres were not predictive of its occurrence., Conclusion: Over 6 months, most abnormal TFTs in acute COVID-19 resolved, with no significant incident thyroid dysfunction. SARS-CoV-2 infection did not lead to change in thyroid autoimmunity, while interferon treatment was associated with modest increase in anti-thyroid antibody titres. Thyroid function and anti-thyroid antibodies did not play a significant role in long COVID., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Fungal infections usually occur in immunocompromised patients. Intravenous immunoglobulin (IVIG) has been used as therapeutic interventions for many infectious diseases, but seldom applied in mycosis due to unknown antifungal specificity. This study aims to determine the presence of antifungal antibodies in IVIG. Binding reactivity of IVIG with crude and recombinant antigens of Candida albicans, Aspergillus fumigatus, Cryptococcus neoformans and Talaromyces marneffei were observed in a dose-dependent manner, similar with mixed normal human sera. The antifungal specificity was further confirmed by competitive enzyme-linked immunosorbent assays (ELISA) inhibited by rabbit specific antifungal polyclonal antibodies (PAbs) and homogenous crude antigens with inhibitions of 65.5-87.2% and 73.1-94.2%, respectively. Moreover, IVIG also reacted with fungal glycoproteins (Csa2, Cpl1 and Mp1p) in a dose-dependent way, which was inhibited by specific rabbit PAbs and homogenous antigens with different inhibitions and pulled down 72.8-83.8% of specific antibodies if preabsorption IVIG with Dynabeads® coupled with homogenous glycoproteins. These results furthermore verified the antifungal specificity of IVIG. Among four brands of IVIG, there was different antifungal IgG against C. albicans (P < 0.05) and C. neoformans (P < 0.05), while no difference for A. fumigatus (P = 0.086) and T. marneffei (P = 0.057). IVIG contained a significantly higher level of specific IgG for C. albicans than other three fungi (P <0.001). In conclusion, we proved antifungal IgG against C. albicans, A. fumigatus, C. neoformans and T. marneffei present in IVIG, which might be expected to provide a possible immunoregulation choice for mycosis and an evaluation to humoral immunity against fungi., (© 2023. The Author(s).)

The emergence and rapid spread of the mobile colistin resistance gene mcr-1 among bacterial species and hosts significantly challenge the efficacy of "last-line" antibiotic colistin. Previously, we reported silver nitrate and auranofin serve as colistin adjuvants for combating mcr-1-positive bacteria. Herein, we uncovered more gold-based drugs and nanoparticles, and found that they exhibited varying degree of synergisms with colistin on killing mcr-1-positive bacteria. However, pre-activation of the drugs by either glutathione or N-acetyl cysteine, thus releasing and accumulating gold ions, is perquisite for their abilities to substitute zinc cofactor from MCR-1 enzyme. X-ray crystallography and biophysical studies further supported the proposed mechanism. This study not only provides basis for combining gold-based drugs and colistin for combating mcr-1-positive bacterial infections, but also undoubtedly opens a new horizon for metabolism details of gold-based drugs in overcoming antimicrobial resistance., (© 2023. The Author(s), under exclusive licence to Society for Biological Inorganic Chemistry (SBIC).)

Background: We aimed to investigate the effect of non-alcoholic fatty liver disease (NAFLD) on BNT162b2 immunogenicity against wild-type SARS-CoV-2 and variants and infection outcome, as data are lacking., Methods: Recipients of two doses of BNT162b2 were prospectively recruited. Outcomes of interest were seroconversion of neutralizing antibody by live virus microneutralization (vMN) to SARS-CoV-2 strains (wild-type, delta and omicron variants) at day 21, 56 and 180 after first dose. Exposure of interest was moderate-to-severe NAFLD (controlled attenuation parameter ≥ 268 dB/M on transient elastography). We calculated adjusted odds ratio (aOR) of infection with NAFLD by adjusting for age, sex, overweight/obesity, diabetes and antibiotic use., Results: Of 259 BNT162b2 recipients (90 (34.7%) male; median age: 50.8 years (IQR: 43.6-57.8)), 68 (26.3%) had NAFLD. For wild type, there was no difference in seroconversion rate between NAFLD and control groups at day 21 (72.1% vs. 77.0%; p = 0.42), day 56 (100% vs. 100%) and day 180 (100% and 97.2%; p = 0.22), respectively. For the delta variant, there was no difference also at day 21 (25.0% vs. 29.5%; p = 0.70), day 56 (100% vs. 98.4%; p = 0.57) and day 180 (89.5% vs. 93.3%; p = 0.58), respectively. For the omicron variant, none achieved seroconversion at day 21 and 180. At day 56, there was no difference in seroconversion rate (15.0% vs. 18.0%; p = 0.76). NAFLD was not an independent risk factor of infection (aOR: 1.50; 95% CI: 0.68-3.24)., Conclusions: NAFLD patients receiving two doses of BNT162b2 had good immunogenicity to wild-type SARS-CoV-2 and the delta variant but not the omicron variant, and they were not at higher risk of infection compared with controls.

Background: Early antiviral therapy was effective in the treatment of coronavirus disease 2019 (COVID-19). We assessed the efficacy and safety of combined interferon beta-1b and remdesivir treatment in hospitalized COVID-19 patients., Methods: We conducted a multicentre, prospective open-label, randomized-controlled trial involving high-risk adults hospitalized for COVID-19. Patients were randomly assigned to a 5-day interferon beta-1b 16 million units daily and remdesivir 200 mg loading on day 1 followed by 100 mg daily on day 2 to 5 (combination group), or to remdesivir only of similar regimen (control group) (1:1). The primary endpoint was the time to complete alleviation of symptoms (NEWS2 = 0)., Results: Two-hundred and twelve patients were enrolled. The median days of starting treatment from symptom onset was 3 days. The median age was 65 years, and 159 patients (75%) had chronic disease. The baseline demographics were similar. There was no mortality. For the primary endpoint, the combination group was significantly quicker to NEWS2 = 0 (4 vs 6.5 days; hazard ratio [HR], 6.59; 95% confidence interval [CI], 6.1-7.09; P < .0001) when compared to the control group. For the secondary endpoints, the combination group was quicker to negative nasopharyngeal swab (NPS) viral load (VL) (6 vs 8 days; HR, 8.16; 95% CI, 7.79-8.52; P < .0001) and to develop seropositive immunoglobulin G (IgG) (8 vs 10 days; HR, 10.78; 95% CI, 9.98-11.58; P < .0001). All adverse events resolved upon follow-up. Combination group (HR, 4.1 95% CI, 1.9-8.6, P < .0001) was the most significant independent factor associated with NEWS2 = 0 on day 4., Conclusions: Early treatment with interferon beta-1b and remdesivir was safe and better than remdesivir only in alleviating symptoms, and in shortening viral shedding and hospitalization with earlier seropositivity in high-risk COVID-19 patients., Clinical Trials Registration: NCT04647695., Competing Interests: Potential conflicts of interest. I. F. H received honoraria as speaker from MSD for COVID-19 Regional Expert Input Forum 2021 and Herpes Zoster lecture 2021 and was a member of the Advisory Board for Pfizer on COVID-19 Management 2022 and Gilead on Evolving Treatment Landscape in COVID-19 2021. All other authors report no potential conflicts. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed., (© The Author(s) 2022. Published by Oxford University Press on behalf of Infectious Diseases Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Natural products (NPs) especially the secondary metabolites originated from microbes exhibit great importance in biomedical, industrial and agricultural applications. However, mining biosynthetic gene clusters (BGCs) to produce novel NPs has been hindered owing that a large population of environmental microbes are unculturable. In the past decade, strategies to explore BGCs directly from (meta)genomes have been established along with the fast development of high-throughput sequencing technologies and the powerful bioinformatics data-processing tools, which greatly expedited the exploitations of novel BGCs from unculturable microbes including the extremophilic microbes. In this review, we firstly summarized the popular bioinformatics tools and databases available to mine novel BGCs from (meta)genomes based on either pure cultures or pristine environmental samples. Noticeably, approaches rooted from machine learning and deep learning with focuses on the prediction of ribosomally synthesized and post-translationally modified peptides (RiPPs) were dramatically increased in recent years. Moreover, synthetic biology techniques to express the novel BGCs in culturable native microbes or heterologous hosts were introduced. This working pipeline including the discovery and biosynthesis of novel NPs will greatly advance the exploitations of the abundant but unexplored microbial BGCs.

Tailam paramyxovirus (TlmPV) was identified in Sikkim Rats in Hong Kong, China in 2011. Its negative sense RNA genome is similar to J paramyxovirus (JPV) and Beilong paramyxovirus (BeiPV), the prototypes of the recently established genus Jeilongvirus. TlmPV genome is predicted to have eight genes in the order 3'-N-P/V/C-M-F-SH-TM-G/X-L-5'. The predicted size of the TlmPV G protein is 1,052 amino acid (aa) residues and much larger than G proteins of typical paramyxoviruses, which are often less than 800 aa. In addition to G open reading frame (ORF) in the G gene, another ORF, termed ORF-X exists in the G gene transcript. Similar ORF-X exists in JPV and BeiPV G gene, but their expression in virus-infected cells has not been confirmed. In this study, we generated infectious TlmPV using a newly developed reverse genetics system. We have found that the G protein of TlmPV is truncated in cultured cells: stop codons emerged in the G open reading frame, resulting in deletions of amino acid residues beyond residue 732. We have obtained infectious TlmPV lacking the C-terminal 307 aa (rTlmPV-G745) and TlmPV lacking the C-terminal 306 aa and the ORF-X (rTlmPV-GΔ746-X). The recombinant TlmPVs lacking the C-terminal 300 aa reach a higher peak viral titer and have improved genome stability in tissue cultured cells. The work indicates that the C-terminal of the G protein of TlmPV and ORF-X are not required for replication in tissue culture cells, and the deletion of the C-terminal confers a growth advantage in tissue culture cells. IMPORTANCE TlmPV is a member of the recently established genus Jeilongvirus . TlmPV encodes a large G protein and its G gene contains ORF-X. In this work, infectious TlmPV was recovered using reverse genetics. Using this system, we have demonstrated that 300 aa of C-terminal of G and the ORF-X are not required for viral replication in tissue culture cells.

There has been a rapid surge of hospitalization due to the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variants globally. The severity of Omicron BA.2 in unexposed, unvaccinated, hospitalized children is unknown. We investigated the severity and clinical outcomes of COVID-19 infection during the Omicron wave in uninfected, unvaccinated hospitalized children and in comparison with influenza and parainfluenza viral infections. This population-based study retrieved data from the HK territory-wide CDARS database of hospitalisations in all public hospitals and compared severe outcomes for the Omicron BA.2-dominant fifth wave (5-28 February 2022, n = 1144), and influenza and parainfluenza viruses (1 January 2015-31 December 2019, n = 32212 and n = 16423, respectively) in children 0-11 years old. Two deaths (0.2%) out of 1144 cases during the initial Omicron wave were recorded. Twenty-one (1.8%) required PICU admission, and the relative risk was higher for Omicron than influenza virus (n = 254, 0.8%, adjusted RR = 2.1, 95%CI 1.3-3.3, p  = 0.001). The proportion with neurological complications was 15.0% (n = 171) for Omicron, which was higher than influenza and parainfluenza viruses (n = 2707, 8.4%, adjusted RR = 1.6, 95%CI 1.4-1.9 and n = 1258, 7.7%, adjusted RR = 1.9, 95%CI 1.6-2.2, p  < 0.001 for both, respectively). Croup occurred for Omicron (n = 61, 5.3%) more than influenza virus (n = 601, 1.9%, adjusted RR = 2.0, 95%CI 1.5-2.6, p  < 0.001) but not parainfluenza virus (n = 889, 5.4%). Our findings showed that for hospitalized children who had no past COVID-19 or vaccination, Omicron BA.2 was not mild. Omicron BA.2 appeared to be more neuropathogenic than influenza and parainfluenza viruses. It targeted the upper airways more than influenza virus.

Purpose: Thyroid dysfunction in COVID-19 carries clinical and prognostic implications. In this study, we developed a prediction score (ThyroCOVID) for abnormal thyroid function (TFT) on admission amongst COVID-19 patients., Methods: Consecutive COVID-19 patients admitted to Queen Mary Hospital were prospectively recruited during July 2020-May 2021. Thyroid-stimulating hormone (TSH), free thyroxine (fT4) and free triiodothyronine (fT3) were measured on admission. Multivariable logistic regression analysis was performed to identify independent determinants of abnormal TFTs. ThyroCOVID was developed based on a clinical model with the lowest Akaike information criteria., Results: Five hundred and forty six COVID-19 patients were recruited (median age 50 years, 45.4% men, 72.9% mild disease on admission). 84 patients (15.4%) had abnormal TFTs on admission. Patients with abnormal TFTs were more likely to be older, have more comorbidities, symptomatic, have worse COVID-19 severity, higher SARS-CoV-2 viral loads and more adverse profile of acute-phase reactants, haematological and biochemical parameters. ThyroCOVID consisted of five parameters: symptoms (malaise), comorbidities (ischaemic heart disease/congestive heart failure) and laboratory parameters (lymphocyte count, C-reactive protein, and SARS-CoV-2 cycle threshold values). It was able to identify abnormal TFT on admission with an AUROC of 0.73 (95% CI 0.67-0.79). The optimal cut-off of 0.15 had a sensitivity of 75.0%, specificity of 65.2%, negative predictive value of 93.5% and positive predictive value of 28.1% in identifying abnormal TFTs on admission amongst COVID-19 patients., Conclusion: ThyroCOVID, a prediction score to identify COVID-19 patients at risk of having abnormal TFT on admission, was developed based on a cohort of predominantly non-severe COVID-19 patients., (© 2022. The Author(s), under exclusive licence to Italian Society of Endocrinology (SIE).)

Background: Patients with COVID-19 display a broad spectrum of manifestations from asymptomatic to life-threatening disease with dysregulated immune responses. Mechanisms underlying the detrimental immune responses and disease severity remain elusive., Methods: We investigated a total of 137 APs infected with SARS-CoV-2. Patients were divided into mild and severe patient groups based on their requirement of oxygen supplementation. All blood samples from APs were collected within three weeks after symptom onset. Freshly isolated PBMCs were investigated for B cell subsets, their homing potential, activation state, mitochondrial functionality and proliferative response. Plasma samples were tested for cytokine concentration, and titer of Nabs, RBD-, S1-, SSA/Ro- and dsDNA-specific IgG., Results: While critically ill patients displayed predominantly extrafollicular B cell activation with elevated inflammation, mild patients counteracted the disease through the timely induction of mitochondrial dysfunction in B cells within the first week post symptom onset. Rapidly increased mitochondrial dysfunction, which was caused by infection-induced excessive intracellular calcium accumulation, suppressed excessive extrafollicular responses, leading to increased neutralizing potency index and decreased inflammatory cytokine production. Patients who received prior COVID-19 vaccines before infection displayed significantly decreased extrafollicular B cell responses and mild disease., Conclusion: Our results reveal an immune mechanism that controls SARS-CoV-2-induced detrimental B cell responses and COVID-19 severity, which may have implications for viral pathogenesis, therapeutic interventions and vaccine development., (© 2022 The Authors. Clinical and Translational Medicine published by John Wiley & Sons Australia, Ltd on behalf of Shanghai Institute of Clinical Bioinformatics.)

Background: CRISPR-Cas12a has been integrated with nanomaterial-based optical techniques, such as surface-enhanced Raman scattering (SERS), to formulate a powerful amplification-free nucleic acid detection system. However, nanomaterials impose steric hindrance to limit the accessibility of CRISPR-Cas12a to the narrow gaps (SERS hot spots) among nanoparticles (NPs) for producing a significant change in signals after nucleic acid detection. Methods: To overcome this restriction, we specifically design chimeric DNA/RNA hairpins (displacers) that can be destabilized by activated CRISPR-Cas12a in the presence of target DNA, liberating excessive RNA that can disintegrate a core-satellite nanocluster via toehold-mediated strand displacement for orchestrating a promising "on-off" nucleic acid biosensor. The core-satellite nanocluster comprises a large gold nanoparticle (AuNP) core surrounded by small AuNPs with Raman tags via DNA hybridization as an ultrabright Raman reporter, and its disassembly leads to a drastic decrease of SERS intensity as signal readouts. We further introduce a magnetic core to the large AuNPs that can facilitate their separation from the disassembled nanostructures to suppress the background for improving detection sensitivity. Results: As a proof-of-concept study, our findings showed that the application of displacers was more effective in decreasing the SERS intensity of the system and attained a better limit of detection (LOD, 10 aM) than that by directly using activated CRISPR-Cas12a, with high selectivity and stability for nucleic acid detection. Introducing magnetic-responsive functionality to our system further improves the LOD to 1 aM. Conclusion: Our work not only offers a platform to sensitively and selectively probe nucleic acids without pre-amplification but also provides new insights into the design of the CRISPR-Cas12a/SERS integrated system to resolve the steric hindrance of nanomaterials for constructing biosensors., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Background: Expression of genes of interest from plasmids or lentiviral vectors is one of the most common tools in molecular and gene therapy. Aberrant splicing between the inserted gene of interest and downstream vector sequence has not been systematically analyzed. Methods: Formation of aberrant fusion transcripts and proteins was detected by RT-PCR, sequencing, Western blotting and mass spectrometry. Bioinformatic analysis was performed to identify all human and mouse genes prone to vector-dependent aberrant splicing. Selected genes were experimentally validated. Results: When we expressed human FACI in cultured cells, an aberrant splicing event was found to occur between FACI transcript and downstream plasmid sequence through one exon-exon junction in FACI that accidentally contributes a splice donor site. To explore whether this could be a general phenomenon, we searched the whole human and mouse genomes for protein-coding genes that harbor an exon-exon junction resembling a splice donor site. Almost all genes prone to this type of aberrant splicing were identified. A total of 17 genes among the hits were randomly selected for experimental validation. RT-PCR and sequencing results verified that 13 genes were aberrantly spliced on the identified exon-exon junctions. In addition, all 17 genes were aberrantly spliced on their V5 tag sequence. Aberrant fusion protein expression from all 17 genes was validated by immunoblotting. Aberrant splicing was prevented by recoding the V5 tag or the splice sites. Conclusions: Our study revealed an unexpectedly high frequency of vector-dependent aberrant splicing events. Aberrant formation of the resulting fusion proteins could undermine the accuracy of gain-of-function studies and might cause potential side effects when the therapeutic gene is expressed in vivo . Our work has implications in improving vector construction and epitope tagging for gene expression and therapy., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Background/aims: Studies of hepatic steatosis (HS) effect on COVID-19 vaccine immunogenicity are lacking. We aimed to compare immunogenicity of BNT162b2 and CoronaVac among moderate/severe HS and control subjects., Methods: Two hundred ninety-five subjects who received BNT162b2 or CoronaVac vaccines from five vaccination centers were categorized into moderate/severe HS (controlled attenuation parameter ≥268 dB/m on transient elastography) (n=74) or control (n=221) groups. Primary outcomes were seroconversion rates of neutralising antibody by live virus Microneutralization (vMN) assay (titer ≥10) at day21 (BNT162b2) or day28 (CoronaVac) and day56 (both). Secondary outcome was highest-tier titer response (top 25% of vMN titer; cutoff: 160 [BNT162b2] and 20 [CoronaVac]) at day 56., Results: For BNT162b2 (n=228, 77.3%), there was no statistical differences in seroconversion rates (day21: 71.7% vs. 76.6%; day56: 100% vs. 100%) or vMN geometric mean titer (GMT) (day21: 13.2 vs. 13.3; day56: 91.9 vs. 101.4) among moderate/severe HS and control groups respectively. However, lower proportion of moderate/severe HS patients had highest-tier response (day56: 5.0% vs. 15.5%; P=0.037). For CoronaVac (n=67, 22.7%), there was no statistical differences in seroconversion rates (day21: 7.1% vs. 15.1%; day56: 64.3% vs. 83.0%) or vMN GMT (5.3 vs. 5.8,) at day28. However, moderate/severe HS patients had lower vMN GMT (9.1 vs. 14.8, P=0.021) at day 56 with lower proportion having highest-tier response (21.4% vs. 52.8%, P=0.036)., Conclusion: While there was no difference in seroconversion rate between moderate/severe HS and control groups after two doses of vaccine, a lower proportion of moderate/severe HS patients achieved highest-tier response for either BNT162b2 or CoronaVac.

Vertical transmission of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) along a vertical column of flats has been documented in several outbreaks of coronavirus disease 2019 (COVID-19) in Guangdong and Hong Kong. We describe an outbreak in Luk Chuen House, involving two vertical columns of flats associated with an unusually connected two-stack drainage system, in which nine individuals from seven households were infected. The index case resided in Flat 812 (8th floor, Unit 12), two flats (813, 817) on its opposite side reported one case each (i.e., a horizontal sub-cluster). All other flats with infected residents were vertically associated, forming a vertical sub-cluster. We injected tracer gas (SF 6 ) into drainage stacks via toilet or balcony of Flat 812, monitored gas concentrations in roof vent, toilet, façade, and living room in four of the seven flats with infected residents and four flats with no infected residents. The measured gas concentration distributions agreed with the observed distribution of affected flats. Aerosols leaking into drainage stacks may generate the vertical sub-cluster, whereas airflow across the corridor probably caused the horizontal sub-cluster. Sequencing and phylogenetic analyses also revealed a common point-source. The findings provided additional evidence of probable roles of drainage systems in SARS-CoV-2 transmission., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Human adenoviruses (HAdVs) within species B, C, and E are responsible for highly contagious and potentially severe respiratory disease infections. The traditional method to type these pathogens was based on virus neutralization and hemagglutination assays, which are both time-consuming and difficult, particularly due to the nonavailability of reagents. Subsequent molecular typing based on the partial characterization of the hexon gene and/or the restriction enzyme analysis (REA) of the genomes is inadequate, particularly in identifying recombinants. Here, a rapid, simple, and cost-effective method for molecular typing HAdV respiratory pathogens is presented. This incorporates three pairs of universal PCR primers that target the variable regions of the three major capsid genes, i.e., hexon, penton base, and fiber genes, that span the genome. The protocol enables typing and characterization of genotypes within species B, C, and E, as well as of some genotypes within species D and F. To validate this method, we surveyed 100 children with HAdV-associated acute respiratory infections identified by direct immunofluorescence (Hong Kong; July through October, 2014). Throat swab specimens were collected and analyzed by PCR amplification and sequencing; these sequences were characterized by BLAST. HAdVs were detected in 98 out of 100 (98%) samples, distributing as follows: 74 HAdV-B3 (74%); 10 HAdV-E4 (10%); 7 HAdV-C2 (7%); 2 HAdV-C6 (2%); 1 HAdV-B7 (1%); 1 HAdV-C1 (1%); 2 co-infection (2%); and 1 novel recombinant (1%). This study is the first detailed molecular epidemiological survey of HAdVs in Hong Kong. The developed method allows for the rapid identification of HAdV respiratory pathogens, including recombinants, and bypasses the need for whole genome sequencing for real-time surveillance of circulating adenovirus strains in outbreaks and populations by clinical virologists, public health officials, and epidemiologists., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Wu, Zhang, Lan, Quan, Ou, Zhao, Wu, Woo, Seto and Zhang.)

Olfactory dysfunction (OD) is a common symptom in coronavirus disease 2019 (COVID-19) patients. Moreover, many neurological manifestations have been reported in these patients, suggesting central nervous system involvement. The default mode network (DMN) is closely associated with olfactory processing. In this study, we investigated the internetwork and intranetwork connectivity of the DMN and the olfactory network (ON) in 13 healthy controls and 22 patients presenting with COVID-19-related OD using independent component analysis and region of interest functional magnetic resonance imaging (fMRI) analysis. There was a significant correlation between the butanol threshold test (BTT) and the intranetwork connectivity in ON. Meanwhile, the COVID-19 patients with OD showed significantly higher intranetwork connectivity in the DMN, as well as higher internetwork connectivity between ON and DMN. However, no significant difference was found between groups in the intranetwork connectivity within ON. We postulate that higher intranetwork functional connectivities compensate for the deficits in olfactory processing and general well-being in COVID-19 patients. Nevertheless, the compensation process in the ON may not be obvious at this stage. Our results suggest that resting-state fMRI is a potentially valuable tool to evaluate neurosensory dysfunction in COVID-19 patients.

Background: SARS-CoV-2 is the causative agent of COVID-19. Overproduction and release of proinflammatory cytokines are the underlying cause of severe COVID-19. Treatment of this condition with JAK inhibitors is a double-edged sword, which might result in the suppression of proinflammatory cytokine storm and the concurrent enhancement of viral infection, since JAK signaling is essential for host antiviral response. Improving the current JAK inhibitor therapy requires a detailed molecular analysis on how SARS-CoV-2 modulates interferon (IFN)-induced activation of JAK-STAT signaling., Results: In this study, we focused on the molecular mechanism by which SARS-CoV-2 NSP13 helicase suppresses IFN signaling. Expression of SARS-CoV-2 NSP13 alleviated transcriptional activity driven by type I and type II IFN-responsive enhancer elements. It also prevented nuclear translocation of STAT1 and STAT2. The suppression of NSP13 on IFN signaling occurred at the step of STAT1 phosphorylation. Nucleic acid binding-defective mutant K345A K347A and NTPase-deficient mutant E375A of NSP13 were found to have largely lost the ability to suppress IFN-β-induced STAT1 phosphorylation and transcriptional activation, indicating the requirement of the helicase activity for NSP13-mediated inhibition of STAT1 phosphorylation. NSP13 did not interact with JAK1 nor prevent STAT1-JAK1 complex formation. Mechanistically, NSP13 interacted with STAT1 to prevent JAK1 kinase from phosphorylating STAT1., Conclusion: SARS-CoV-2 NSP13 helicase broadly suppresses IFN signaling by targeting JAK1 phosphorylation of STAT1., (© 2022. The Author(s).)

Colistin is considered the last-line antimicrobial for the treatment of multidrug-resistant gram-negative bacterial infections. The emergence and spread of superbugs carrying the mobile colistin resistance gene ( mcr ) have become the most serious and urgent threat to healthcare. Here, we discover that silver (Ag + ), including silver nanoparticles, could restore colistin efficacy against mcr -positive bacteria. We show that Ag + inhibits the activity of the MCR-1 enzyme via substitution of Zn 2+ in the active site. Unexpectedly, a tetra-silver center was found in the active-site pocket of MCR-1 as revealed by the X-ray structure of the Ag-bound MCR-1, resulting in the prevention of substrate binding. Moreover, Ag + effectively slows down the development of higher-level resistance and reduces mutation frequency. Importantly, the combined use of Ag + at a low concentration with colistin could relieve dermonecrotic lesions and reduce the bacterial load of mice infected with mcr -1–carrying pathogens. This study depicts a mechanism of Ag + inhibition of MCR enzymes and demonstrates the potentials of Ag + as broad-spectrum inhibitors for the treatment of mcr -positive bacterial infection in combination with colistin.

Aims: We carried out this prospective study of predominantly non-severe COVID-19 patients, to evaluate the influence of glycaemic status on clinical outcomes and neutralising antibody (Nab) responses, potentially relevant to the COVID-19 vaccination programme., Methods: We included consecutive adults admitted to Queen Mary Hospital for COVID-19 from July 2020-May 2021. Glycaemic status was defined by admission HbA1c. Clinical deterioration was defined by radiological progression/new oxygen requirement/intensive care requirement/death. COVID-19 survivors had Nab measurements at 1-month, 2-month, 3-month and 6-month post-discharge., Results: Among 605 patients (96.9% non-severe COVID-19; 325 normoglycaemia, 185 prediabetes, 95 diabetes), 74 (12.2%) had clinical deterioration, more likely with worse glycaemic status and higher HbA1c (p < 0.001). Older age (p < 0.001), higher viral loads (p < 0.001), higher C-reactive protein (CRP) (p < 0.001) and symptomatic presentation (p = 0.008), but not glycaemic status/HbA1c, independently predicted clinical deterioration. Older age (p = 0.001), higher CRP (p = 0.038), elevated lactate dehydrogenase (p = 0.046) and interferon treatment (p = 0.001), but not glycaemic status/HbA1c, independently predicted Nab titres. Rate of Nab titre decline was comparable across glycaemic status., Conclusions: COVID-19 patients with worse glycaemic status were more likely to deteriorate clinically, mediated through the association of worse glycaemic status with older age, more severe inflammation and higher viral loads. Importantly, Nab responses did not differ across glycaemic status., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Defective biosynthesis or function of proteoglycans causes pathological conditions in a variety of tissue systems. Osteoarthritis (OA) is a prevalent degenerative joint disorder characterized by progressive cartilage destruction caused by imbalanced proteoglycan synthesis and degradation. Identifying agents that regulate proteoglycan metabolism may benefit the development of OA-modifying therapeutics. High-throughput screening (HTS) of chemical libraries has paved the way for achieving this goal. However, the implementation and adaptation of HTS assays based on proteoglycan measurement remain underexploited. Using primary porcine chondrocytes as a model, we report a miniaturized dimethyl-methylene blue (DMMB) assay, which is commonly used to quantitatively evaluate sulfated glycosaminoglycan (GAG) content, with an optimized detection range and reproducibility and its integration with HTS. Treatment with TGF-β1 and IL1-α, known as positive and negative proteoglycan regulators, respectively, supported the assay specificity. A pre-test of chemical screening of 960 compounds identified both stimulators (4.48%) and inhibitors (6.04%) of GAG production. Fluorophore-assisted carbohydrate electrophoresis validated the activity of selected hits on chondroitin sulfate expression in an alginate culture system. Our findings support the implementation of this simple colorimetric assay in HTS to discover modifiers of OA or other diseases related to dysregulated proteoglycan metabolism., (© 2022. The Author(s).)

Background: Both lymphopenia and thyroid dysfunction are commonly observed among COVID-19 patients. Whether thyroid function independently correlates with lymphocyte counts (LYM) remains to be elucidated., Methods: We included consecutive adults without known thyroid disorder admitted to Queen Mary Hospital for COVID-19 from July 2020 to April 2021 who had thyroid-stimulating hormone (TSH), free thyroxine (fT4), free triiodothyronine (fT3) and LYM measured on admission., Results: A total of 541 patients were included. Median LYM was 1.22 x 10 9 /L, with 36.0% of the cohort lymphopenic. 83 patients (15.4%) had abnormal thyroid function tests (TFTs), mostly non-thyroidal illness syndrome (NTIS). Patients with lymphopenia had lower TSH, fT4 and fT3 levels than those without. Multivariable stepwise linear regression analysis revealed that both TSH (standardized beta 0.160, p<0.001) and fT3 (standardized beta 0.094, p=0.023), but not fT4, remained independently correlated with LYM, in addition to age, SARS-CoV-2 viral load, C-reactive protein levels, coagulation profile, sodium levels and more severe clinical presentations. Among the 40 patients who had reassessment of TFTs and LYM after discharge, at a median of 9 days from admission, there were significant increases in TSH (p=0.031), fT3 (p<0.001) and LYM (p<0.001). Furthermore, patients who had both lymphopenia and NTIS were more likely to deteriorate compared to those who only had either one alone, and those without lymphopenia or NTIS (p for trend <0.001)., Conclusion: TSH and fT3 levels showed independent positive correlations with LYM among COVID-19 patients, supporting the interaction between the hypothalamic-pituitary-thyroid axis and immune system in COVID-19., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Lui, Lee, Chow, Lee, Tam, Pang, Ho, Cheung, Fong, Law, To, Lam, Tan, Woo, Hung and Lam.)

Effective immunotherapy treats cancers by eradicating tumourigenic cells by activated tumour antigen-specific and bystander CD8 + T-cells. However, T-cells can gradually lose cytotoxicity in the tumour microenvironment, known as exhaustion. Recently, DNA methylation, histone modification, and chromatin architecture have provided novel insights into epigenetic regulations of T-cell differentiation/exhaustion, thereby controlling the translational potential of the T-cells. Thus, developing strategies to govern epigenetic switches of T-cells dynamically is critical to maintaining the effector function of antigen-specific T-cells. In this mini-review, we 1) describe the correlation between epigenetic states and T cell phenotypes; 2) discuss the enzymatic factors and intracellular/extracellular microRNA imprinting T-cell epigenomes that drive T-cell exhaustion; 3) highlight recent advances in epigenetic interventions to rescue CD8 + T-cell functions from exhaustion. Finally, we express our perspective that regulating the interplay between epigenetic changes and transcriptional programs provides translational implications of current immunotherapy for cancer treatments., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Wong, Yin, Lam, Huang, Yan, Tan and Wong.)

COVID-19 is a trending topic worldwide due to its immense impact on society. Recent trends have shifted from acute effects towards the long-term morbidity of COVID-19. In this review, we hypothesize that SARS-CoV-2 contributes to age-related perturbations in endothelial and adipose tissue, which are known to characterize the early aging process. This would explain the long-lasting symptoms of SARS-CoV-2 as the result of an accelerated aging process. Connective tissues such as adipose tissue and musculoskeletal tissue are the primary sites of aging. Therefore, current literature was analyzed focusing on the musculoskeletal symptoms in COVID-19 patients. Hypovitaminosis D, increased fragility, and calcium deficiency point towards bone aging, while joint and muscle pain are typical for joint and muscle aging, respectively. These characteristics could be classified as early osteoarthritis-like phenotype. Exploration of the impact of SARS-CoV-2 and osteoarthritis on endothelial and adipose tissue, as well as neuronal function, showed similar perturbations. At a molecular level, this could be attributed to the angiotensin-converting enzyme 2 expression, renin-angiotensin system dysfunction, and inflammation. Finally, the influence of the nicotinic cholinergic system is being evaluated as a new treatment strategy. This is combined with the current knowledge of musculoskeletal aging to pave the road towards the treatment of long-term COVID-19.

Background & Aims: CREB-H is a key liver-enriched transcription factor governing lipid metabolism. Additional targets of CREB-H remain to be identified and characterized. Here, we identified a novel fasting- and CREB-H-induced (FACI) protein that inhibits intestinal lipid absorption and alleviates diet-induced obesity in mice., Methods: FACI was identified by reanalysis of existing transcriptomic data. Faci -/- mice were generated by clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9)-mediated genome engineering. RNA sequencing was performed to identify differentially expressed genes in Faci -/- mice. Lipid accumulation in the villi was assessed by triglyceride measurement and Oil red O staining. In vitro fatty acid uptake assay was performed to verify in vivo findings., Results: FACI expression was enriched in liver and intestine. FACI is a phospholipid-binding protein that localizes to plasma membrane and recycling endosomes. Hepatic transcription of Faci was regulated by not only CREB-H, but also nutrient-responsive transcription factors sterol regulatory element-binding protein 1 (SREBP1), hepatocyte nuclear factor 4α (HNF4α), peroxisome proliferator-activated receptor γ coactivator-1α (PGC1α), and CREB, as well as fasting-related cyclic adenosine monophosphate (cAMP) signaling. Genetic knockout of Faci in mice showed an increase in intestinal fat absorption. In accordance with this, Faci deficiency aggravated high-fat diet-induced obesity, hyperlipidemia, steatosis, and other obesity-related metabolic dysfunction in mice., Conclusions: FACI is a novel CREB-H-induced protein. Genetic disruption of Faci in mice showed its inhibitory effect on fat absorption and obesity. Our findings shed light on a new target of CREB-H implicated in lipid homeostasis., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

In February 2020, we highlighted the top nine important research questions on SARS-CoV-2 and COVID-19 concerning virus transmission, asymptomatic and presymptomatic virus shedding, diagnosis, treatment, vaccine development, origin of virus and viral pathogenesis. These and related questions are revisited at the end of 2021 to shed light on the roadmap of bringing an end to the pandemic., (© 2021. The Author(s).)

Antiviral short-hairpin RNAs (shRNAs) delivered by recombinant adeno-associated virus (rAAV) were investigated for their potential prophylactic and therapeutic applications related to the influenza A virus (IAV). To express shRNAs efficiently, an H1 promoter was inserted into the commercial rAAV2 system. The modified rAAV2 system could express shRNAs, and the purified rAAV was obtained at levels over 10 13 viral genomes/ml and 10 10 viral infection units/ml. The shNP-1496-n and shM2-925 delivered by rAAV could inhibit the replication of the H1N1 and H5N1 virus by targeting the conserved regions of the IAV nucleoprotein and matrix 2 genes in MDCK cells. The shNP-1496-n and shM2-925 expressed by rAAV could provide potent and long-term anti-H5N1 virus effects in rAAV-shRNA-enriched MDCK cells. Our findings provide a rational basis for developing RNA interference for the prevention and therapy of IAV infection., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Effective treatments for coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are urgently needed. Dexamethasone has been shown to confer survival benefits to certain groups of hospitalized patients, but whether glucocorticoids such as dexamethasone and methylprednisolone should be used together with antivirals to prevent a boost of SARS-CoV-2 replication remains to be determined. Here, we show the beneficial effect of methylprednisolone alone and in combination with remdesivir in the hamster model of SARS-CoV-2 infection. Treatment with methylprednisolone boosted RNA replication of SARS-CoV-2 but suppressed viral induction of proinflammatory cytokines in human monocyte-derived macrophages. Although methylprednisolone monotherapy alleviated body weight loss as well as nasal and pulmonary inflammation, viral loads increased and antibody response against the receptor-binding domain of spike protein attenuated. In contrast, a combination of methylprednisolone with remdesivir not only prevented body weight loss and inflammation, but also dampened viral protein expression and viral loads. In addition, the suppressive effect of methylprednisolone on antibody response was alleviated in the presence of remdesivir. Thus, combinational anti-inflammatory and antiviral therapy might be an effective, safer and more versatile treatment option for COVID-19. These data support testing of the efficacy of a combination of methylprednisolone and remdesivir for the treatment of COVID-19 in randomized controlled clinical trials.

Zika virus (ZIKV) is an emerging mosquito-borne flavivirus that poses significant threats to global public health. Macrophages and dendritic cells are both key sentinel cells in the host immune response and play critical roles in the pathogenesis of flavivirus infections. Recent studies showed that ZIKV could productively infect monocyte-derived dendritic cells (moDCs), but the role of macrophages in ZIKV infection remains incompletely understood. In this study, we first compared ZIKV infection in monocyte-derived macrophages (MDMs) and moDCs derived from the same donors. We demonstrated that while both MDMs and moDCs were susceptible to epidemic (Puerto Rico) and pre-epidemic (Uganda) strains of ZIKV, virus replication was largely restricted in MDMs but not in moDCs. ZIKV induced significant apoptosis in moDCs but not MDMs. The restricted virus replication in MDMs was not due to inefficient virus entry but was related to post-entry events in the viral replication cycle. In stark contrast with moDCs, ZIKV failed to inhibit STAT1 and STAT2 phosphorylation in MDMs. This resulted in the lack of efficient antagonism of the host type I interferon-mediated antiviral responses. Importantly, depletion of STAT2 but not STAT1 in MDMs significantly rescued the replication of ZIKV and the prototype flavivirus yellow fever virus. Overall, our findings revealed a differential interplay between macrophages and dendritic cells with ZIKV. While dendritic cells may be exploited by ZIKV to facilitate virus replication, macrophages restricted ZIKV infection.