Synergism of colistin and globular endolysins against multidrug-resistant gram-negative bacteria.

Endolysins (lysins), a novel class of antibacterial agents derived from bacteriophages, efficiently lyse bacteria by degrading the peptidoglycan layer within the bacterial wall. Colistin, a classic peptide antibiotic with the ability to permeabilize the outer membrane, has recently shown great promise in synergizing with lysins against gram-negative bacteria. However, the exact mechanism responsible for their synergy remains unclear. Here, we first demonstrated the synergistic bacterial killing of various lysin and colistin combinations. With a model lysin, LysAB2, we then confirmed that there is a threshold concentration of colistin causing sufficient permeabilization of the outer membrane for lysin to access the peptidoglycan layer and subsequently exert its lytic ability. The threshold colistin concentrations were found to range 0.2-0.8 µM for the tested bacteria, with the exact value largely depending on the density of lipopolysaccharides on the outer membrane. Beyond the threshold colistin level, LysAB2 could synergize with colistin at a concentration as low as 0.31 µM. Next, we proved for the first time that lysin-induced degradation of the peptidoglycan layer facilitated the disruption of cytoplasmic membrane by colistin, elevated the level of reactive oxygen species in bacterial cells, and boosted the killing effect of colistin. Additionally, the colistin-lysin combination could effectively eliminate established biofilms due to the biofilm dispersal ability of lysin. The in-vivo efficacy was preliminary confirmed in a Galleria mellonella infection model for combination with colistin doses (≥ 1.8 µg/larvae), which could reach beyond the threshold concentration, and a fixed LysAB2 dose (10 µg/larvae). In summary, our study provided the first experimental evidence unravelling the mechanisms behind the synergy of colistin and lysins. All these findings provided important insights in guiding the dosing strategy for applying this combination in future development.

Early Metabolomic and Immunologic Biomarkers as Prognostic Indicators for COVID-19.

This prospective study in Hong Kong aimed at identifying prognostic metabolomic and immunologic biomarkers for Coronavirus Disease 2019 (COVID-19). We examined 327 patients, mean age 55 (19-89) years, in whom 33.6% were infected with Omicron and 66.4% were infected with earlier variants. The effect size of disease severity on metabolome outweighed others including age, gender, peak C-reactive protein (CRP), vitamin D and peak viral levels. Sixty-five metabolites demonstrated strong associations and the majority (54, 83.1%) were downregulated in severe disease (z score: -3.30 to -8.61). Ten cytokines/chemokines demonstrated strong associations (p < 0.001), and all were upregulated in severe disease. Multiple pairs of metabolomic/immunologic biomarkers showed significant correlations. Fourteen metabolites had the area under the receiver operating characteristic curve (AUC) > 0.8, suggesting a high predictive value. Three metabolites carried high sensitivity for severe disease: triglycerides in medium high-density lipoprotein (MHDL) (sensitivity: 0.94), free cholesterol-to-total lipids ratio in very small very-low-density lipoprotein (VLDL) (0.93), cholesteryl esters-to-total lipids ratio in chylomicrons and extremely large VLDL (0.92);whereas metabolites with the highest specificity were creatinine (specificity: 0.94), phospholipids in large VLDL (0.94) and triglycerides-to-total lipids ratio in large VLDL (0.93). Five cytokines/chemokines, namely, interleukin (IL)-6, IL-18, IL-10, macrophage inflammatory protein (MIP)-1b and tumour necrosis factor (TNF)-a, had AUC > 0.8. In conclusion, we demonstrated a tight interaction and prognostic potential of metabolomic and immunologic biomarkers enabling an outcome-based patient stratification.

Chronic post-COVID neuropsychiatric symptoms persisting beyond one year from infection: a case-control study and network analysis.

Our study aims to delineate the phenotypes of chronic neuropsychiatric symptoms among adult subjects recovering from their first COVID that occurred more than one year ago. We also aim to explore the clinical and socioeconomic risk factors of having a high loading of chronic neuropsychiatric symptoms. We recruited a post-COVID group who suffered from their first pre-Omicron COVID more than a year ago, and a control group who had never had COVID. The subjects completed app-based questionnaires on demographic, socioeconomic and health status, a COVID symptoms checklist, mental and sleep health measures, and neurocognitive tests. The post-COVID group has a statistically significantly higher level of fatigue compared to the control group (p < 0.001). Among the post-COVID group, the lack of any COVID vaccination before the first COVID and a higher level of material deprivation before the COVID pandemic predicts a higher load of chronic post-COVID neuropsychiatric symptoms. Partial correlation network analysis suggests that the chronic post-COVID neuropsychiatric symptoms can be clustered into two major (cognitive complaints -fatigue and anxiety-depression) and one minor (headache-dizziness) cluster. A higher level of material deprivation predicts a higher number of symptoms in both major clusters, but the lack of any COVID vaccination before the first COVID only predicts a higher number of symptoms in the cognitive complaints-fatigue cluster. Our result suggests heterogeneity among chronic post-COVID neuropsychiatric symptoms, which are associated with the complex interplay of biological and socioeconomic factors.

Prevalence of oral human papillomavirus infection among the general adult population in Hong Kong.

A cross-sectional study in 2021-23 collected oral rinse gargle samples from an human papillomaviruses (HPV) vaccine-naïve general adult population in Hong Kong. HPV was detected by a PCR using SPF10 primers, and genotyped by a linear array covering 25 genotypes. Epidemiologic information including sociodemographics, medical history, oral health, and sexual behavior were collected by a self-administered questionnaire. Altogether, 2323 subjects aged 18-75 (median 47) years with 50.1% male were recruited. The prevalence for oral HPV infection with all genotypes combined, high-risk, and low-risk genotypes was 1.5%, 0.7%, and 0.7%, respectively; and with no statistically significant difference between participant gender. The prevalence increased with age and was highest in women at 45-54 years (2.7% for all genotypes combined), and highest in men aged >64 years (4.1% for all genotypes combined). HPV52 was the most common genotype among all participants. Univariate analysis suggested more lifetime sexual or oral sexual partners as risk factors, but they did not reach statistical significance upon multivariate analysis; whereas higher educational level had an independent protective effect. To conclude, oral HPV prevalence increased with age in Hong Kong. Strategies to prevent oral HPV infection and the associated cancers are urgently needed.

Changes in the Incidence and Human Papillomavirus-Positive Portion of Oropharyngeal Squamous Cell Carcinoma in Hong Kong.

The incidence of human papillomavirus (HPV)-associated oropharyngeal squamous cell carcinoma (OPSCC) is rising in the West, but little is known in Asia. This study elucidated changes in the incidence and HPV-positive portion of OPSCC in Hong Kong. Data from population-based cancer registry were used to analyze the incidence of OPSCC in association with other head and neck cancers. Archived tumor tissues were tested for HPV. From 1986 to 2020, there was a marked decrease in the incidence of nasopharyngeal and laryngeal cancers, but a persistent increase in OPSCC from 36 cases in 1986 to 116 cases in 2020. The average positive rate for high-risk HPV was 36.1% (112/310) among OPSCC diagnosed in 2010-2020. The HPV-positive rate in recent years was significantly higher than earlier cases (tonsil SCC: 64.7% (55/85) in 2016-2020 vs. 40.4% (19/47) in 2010-2015, p = 0.007). Patients with HPV-positive tonsil cancers were significantly younger than those negative (mean [SD]: 58.9 [9.9] vs. 64.3 [13.3] years, p = 0.006), but no significant difference was observed between genders. A persistent increase in the incidence of oropharyngeal cancer over the last few decades was observed in Hong Kong, which can be explained by the remarkable increase in HPV-positive tonsil cancers.

Longitudinal monitoring reveals the emergence and spread of blaGES-5-harboring carbapenem-resistant Klebsiella quasipneumoniae in a Hong Kong hospital wastewater discharge line.

Testing hospital wastewater (HWW) is potentially an effective, long-term approach for monitoring trends in antimicrobial resistance (AMR) patterns in health care institutions. Over a year, we collected wastewater samples from the clinical and non-clinical sites of a tertiary hospital and from a downstream wastewater treatment plant (WWTP). We focused on the extent of carbapenem resistance among Enterobacteriaceae isolates given their clinical importance. Escherichia coli and Klebsiella spp. were the most frequently isolated Enterobacteriaceae species at all sampling sites. Additionally, a small number of isolates belonging to ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species), except K. pneumoniae, were detected. Of the 232 Klebsiella spp. isolates, 100 (43.1 %) were multi-drug resistant (MDR), with 46 being carbapenem-resistant. Most of these carbapenem-resistant isolates were K. quasipneumoniae (CRKQ) (n = 44). All CRKQ isolates were isolated from the wastewater of a clinical site that includes intensive care units, which also yielded significantly more multi-drug resistant isolates compared to all other sampling sites. Among the CRKQ isolates, blaGES-5 genes (n = 42) were the primary genetic determinant of carbapenem resistance. Notably, three different CRKQ isolates, collected within the same month in HWW and the influent and effluent flow of the WWTP, shared >99 % sequence similarity between their blaGES-5 genes and between their flanking regions and upstream integron-integrase region. The influent isolate was phylogenetically close to K. quasipnuemoniae isolates from wastewater collected in Japan. Its blaGES-5 gene and surrounding sequences were > 99 % identical to blaGES-24 genes found in the Japanese isolates. Our results suggest that testing samples from sites located closer to hospitals could support antibiotic stewardship programs compared to samples collected further downstream. Moreover, testing samples collected regularly from WWTPs may reflect the local and global spread of pathogens and their resistances.

Characterization of upper airway microbiome across severity of COVID-19 during hospitalization and treatment.

Longitudinal studies on upper respiratory tract microbiome in coronavirus disease 2019 (COVID-19) without potential confounders such as antimicrobial therapy are limited. The objective of this study is to assess for longitudinal changes in the upper respiratory microbiome, its association with disease severity, and potential confounders in adult hospitalized patients with COVID-19. Serial nasopharyngeal and throat swabs (NPSTSs) were taken for 16S rRNA gene amplicon sequencing from adults hospitalized for COVID-19. Alpha and beta diversity was assessed between different groups. Principal coordinate analysis was used to assess beta diversity between groups. Linear discriminant analysis was used to identify discriminative bacterial taxa in NPSTS taken early during hospitalization on need for intensive care unit (ICU) admission. A total of 314 NPSTS samples from 197 subjects (asymptomatic = 14, mild/moderate = 106, and severe/critical = 51 patients with COVID-19; non-COVID-19 mechanically ventilated ICU patients = 11; and healthy volunteers = 15) were sequenced. Among all covariates, antibiotic treatment had the largest effect on upper airway microbiota. When samples taken after antibiotics were excluded, alpha diversity (Shannon, Simpson, richness, and evenness) was similar across severity of COVID-19, whereas beta diversity (weighted GUniFrac and Bray-Curtis distance) remained different. Thirteen bacterial genera from NPSTS taken within the first week of hospitalization were associated with a need for ICU admission (area under the receiver operating characteristic curve, 0.96; 95% CI, 0.91-0.99). Longitudinal analysis showed that the upper respiratory microbiota alpha and beta diversity was unchanged during hospitalization in the absence of antimicrobial therapy.

Alterations in faecal microbiome and resistome in Chinese international travellers: a metagenomic analysis.

BACKGROUND: International travel increases the risk of acquisition of antibiotic-resistant bacteria and antibiotic resistance genes (ARGs). Previous studies have characterized the changes in the gut microbiome and resistome of Western travellers; however, information on non-Western populations and the effects of travel-related risk factors on the gut microbiome and resistome remains limited. METHODS: We conducted a prospective observational study on a cohort of 90 healthy Chinese adult residents of Hong Kong. We characterized the microbiome and resistome in stools collected from the subjects before and after travelling to diverse international locations using shotgun metagenomic sequencing and examined their associations with travel-related variables. RESULTS: Our results showed that travel neither significantly changed the taxonomic composition of the faecal microbiota nor altered the alpha (Shannon) or beta diversity of the faecal microbiome or resistome. However, travel significantly increased the number of ARGs. Ten ARGs, including aadA, TEM, mgrB, mphA, qnrS9 and tetR, were significantly enriched in relative abundance after travel, eight of which were detected in metagenomic bins belonging to Escherichia/Shigella flexneri in the post-trip samples. In sum, 30 ARGs significantly increased in prevalence after travel, with the largest changes observed in tetD and a few qnrS variants (qnrS9, qnrS and qnrS8). We found that travel to low- or middle-income countries, or Africa or Southeast Asia, increased the number of ARG subtypes, whereas travel to low- or middle-income countries and the use of alcohol-based hand sanitizer (ABHS) or doxycycline as antimalarial prophylaxis during travel resulted in increased changes in the beta diversity of the faecal resistome. CONCLUSIONS: Our study highlights travel to low- or middle-income countries, Africa or Southeast Asia, a long travel duration, or the use of ABHS or doxycycline as antimalarial prophylaxis as important risk factors for the acquisition/enrichment of ARGs during international travel.

Determining meteorologically-favorable zones for seasonal influenza activity in Hong Kong.

Investigations of simple and accurate meteorology classification systems for influenza epidemics, particularly in subtropical regions, are limited. To assist in preparing for potential upsurges in the demand on healthcare facilities during influenza seasons, our study aims to develop a set of meteorologically-favorable zones for epidemics of influenza A and B, defined as the intervals of meteorological variables with prediction performance optimized. We collected weekly detection rates of laboratory-confirmed influenza cases from four local major hospitals in Hong Kong between 2004 and 2019. Meteorological and air quality records for hospitals were collected from their closest monitoring stations. We employed classification and regression trees to identify zones that optimize the prediction performance of meteorological data in influenza epidemics, defined as a weekly rate > 50th percentile over a year. According to the results, a combination of temperature > 25.1℃ and relative humidity > 79% was favorable to epidemics in hot seasons, whereas either temperature < 16.4℃ or a combination of < 20.4℃ and relative humidity > 76% was favorable to epidemics in cold seasons. The area under the receiver operating characteristic curve (AUC) in model training achieved 0.80 (95% confidence interval [CI], 0.76-0.83) and was kept at 0.71 (95%CI, 0.65-0.77) in validation. The meteorologically-favorable zones for predicting influenza A or A and B epidemics together were similar, but the AUC for predicting influenza B epidemics was comparatively lower. In conclusion, we established meteorologically-favorable zones for influenza A and B epidemics with a satisfactory prediction performance, even though the influenza seasonality in this subtropical setting was weak and type-specific.

Extent of Oral-Gut Transmission of Bacterial and Fungal Microbiota in Healthy Chinese Adults.

Recent studies have provided evidence on the presence of an oral-gut microbiota axis in gastrointestinal diseases; however, whether a similar axis exists in healthy individuals is still in debate. Here, we characterized the bacterial and fungal microbiomes in paired oral rinse and stool samples collected from 470 healthy Chinese adults by sequencing the 16S rRNA V3-V4 and ITS1 regions, respectively. We hypothesized that there is limited oral-gut transmission of both the bacterial and fungal microbiota in healthy Chinese adults. Our results showed that the oral and gut microbiota in healthy individuals differed in taxonomic composition, alpha and beta diversity, metabolic potential, and network properties. Bayesian analysis showed that the vast majority of subjects had negligible or low bacterial and fungal oral-to-stool contribution. Detailed examination of the prevalent amplicon sequence variants (ASVs) also revealed limited cases of sharing between the oral and stool samples within the same individuals, except a few bacterial and fungal ASVs. Association analysis showed that sharing of the potentially transmissible fungal ASVs was associated with host factors, including an older age and a higher body mass index. Our findings indicate that oral-gut transmission of both bacterial and fungal microbiota in healthy adults is limited. Detection of a large amount of shared bacterial or fungal members between the oral and gut microbiome of an individual may indicate medical conditions that warrant detailed checkup. IMPORTANCE The oral-gut microbiota axis in health is a fundamentally important and clinically relevant topic; however, our current understanding of it remains biased and incomplete. By characterizing the bacterial and fungal microbiomes in paired oral rinse and stool samples from a large cohort of healthy Chinese adults, here we provided new evidence that oral-gut microbiota transmission is limited in non-Western population and across biological domains. Our study has established an important baseline of a healthy oral-gut microbiota axis, with which other disease conditions can be compared. Besides, our findings have practical implications that detection of a large amount of shared bacterial or fungal members between the oral cavity and gut within the same individual as an indicator of potential medical conditions.

Limited Impact of SARS-CoV-2 on the Human Naso-Oropharyngeal Microbiota in Hospitalized Patients.

Numerous studies have reported dysbiosis in the naso- and/or oro-pharyngeal microbiota of COVID-19 patients compared with healthy individuals; however, only a few small-scale studies have also included a disease control group. In this study, we characterized and compared the bacterial communities of pooled nasopharyngeal and throat swabs from hospitalized COVID-19 patients (n = 76), hospitalized non-COVID-19 patients with respiratory symptoms or related illnesses (n = 69), and local community controls (n = 76) using 16S rRNA gene V3-V4 amplicon sequencing. None of the subjects received antimicrobial therapy within 2 weeks prior to sample collection. Both COVID-19 and non-COVID-19 hospitalized patients differed in the composition, alpha and beta diversity, and metabolic potential of the naso-oropharyngeal microbiota compared with local controls. However, the microbial communities in the two hospitalized patient groups did not differ significantly from each other. Differential abundance analysis revealed the enrichment of nine bacterial genera in the COVID-19 patients compared with local controls; however, six of them were also enriched in the non-COVID-19 patients. Bacterial genera uniquely enriched in the COVID-19 patients included Alloprevotella and Solobacterium. In contrast, Mogibacterium and Lactococcus were dramatically decreased in COVID-19 patients only. Association analysis revealed that Alloprevotella in COVID-19 patients was positively correlated with the level of the inflammation biomarker C-reactive protein. Our findings reveal a limited impact of SARS-CoV-2 on the naso-oropharyngeal microbiota in hospitalized patients and suggest that Alloprevotella and Solobacterium are more specific biomarkers for COVID-19 detection. IMPORTANCE Our results showed that while both COVID-19 and non-COVID-19 hospitalized patients differed in the composition, alpha and beta diversity, and metabolic potential of the naso-oropharyngeal microbiota compared with local controls, the microbial communities in the two hospitalized patient groups did not differ significantly from each other, indicating a limited impact of SARS-CoV-2 on the naso-oropharyngeal microbiota in hospitalized patients. Besides, we identified Alloprevotella and Solobacterium as bacterial genera uniquely enriched in COVID-19 patients, which may serve as more specific biomarkers for COVID-19 detection.

Overcoming the rising incidence and evolving mechanisms of antibiotic resistance by novel drug delivery approaches - An overview.

Antimicrobial resistance is a normal evolutionary process for microorganisms. Antibiotics exerted accelerated selective pressure that hasten bacterial resistance through mutation, and acquisition external genes. These genes often carry multiple antibiotic resistant determinants allowing the recipient microbe an instant "super-bug" status. The extent of Antimicrobial Resistance (AMR) has reached a level of global crisis, existing antimicrobials are no long effective in treating infections caused by AMR pathogens. The great majority of clinically available antimicrobial agents are administered through oral and intra-venous routes. Overcoming antibacterial resistance by novel drug delivery approach offered new hopes, particularly in the treatment of AMR pathogens in sites less assessible through systemic circulation such as the lung and skin. In the current review, we will revisit the mechanism and incidence of important AMR pathogens. Finally, we will discuss novel drug delivery approaches including novel local antibiotic delivery systems, hybrid antibiotics, and nanoparticle-based antibiotic delivery systems.

COVID-19 Vaccine Hesitancy in a City with Free Choice and Sufficient Doses.

BACKGROUND: Vaccine hesitancy represents one of the major global health issues around the world. We examined the perception, attitude, perceived barriers and facilitation measures of receiving the COVID-19 vaccine in a Chinese population with free vaccine choices (Sinovac [Coronavac] vs. BioNTech/Fosun [Comirnaty]) and adequate doses. METHOD: We conducted a random telephone survey of the general population in 1195 subjects aged 18 years or above from 23 April 2021 to 8 May 2021 after two months of vaccine rollout. A descriptive analysis of the levels of enabling factors, obstacles and perception of COVID-19 vaccination was conducted using ANOVA and Chi-square tests for trend. RESULTS: Only 10.1% and 13.5% had received one and two COVID-19 vaccine doses, respectively. Among those who had not received any COVID-19 vaccine (75.4%), only 25.1% expressed their intention to receive in the coming 6 months. The barriers with the highest scores included "having heard of cases with serious adverse events or death after vaccination" (score: 8.17 out 10, 95% C.I. 7.99, 8.35), "lack of confidence on governmental recommendations" (7.69, 95% C.I. 7.47, 7.91), and "waiting for a better vaccine" (7.29, 95% C.I. 7.07, 7.52). The highest score for the impact of various incentives for vaccination was for "vaccine passports for overseas travel" (4.44, 95% C.I. 4.18, 4.71). CONCLUSIONS: Vaccine hesitancy is commonly observed in this Chinese population despite adequate provision of vaccine doses and choices. No single incentive is strong enough to promote vaccination, and multiple facilitation measures for different groups of population are needed to encourage vaccine uptake. Active clarification and promotion by medical professionals together with a variety of incentives are needed to drive vaccine uptake.

Seroprevalence of Unidentified SARS-CoV-2 Infection in Hong Kong During 3 Pandemic Waves.

Importance: Seroprevalence studies inform the extent of infection and assist evaluation of mitigation strategies for the COVID-19 pandemic. Objective: To estimate the prevalence of unidentified SARS-CoV-2 infection in the general population of Hong Kong. Design, Setting, and Participants: A prospective cross-sectional study was conducted in Hong Kong after each major wave of the COVID-19 pandemic (April 21 to July 7, 2020; September 29 to November 23, 2020; and January 15 to April 18, 2021). Adults (age ≥18 years) who had not been diagnosed with COVID-19 were recruited during each period, and their sociodemographic information, symptoms, travel, contact, quarantine, and COVID-19 testing history were collected. Main Outcomes and Measures: The main outcome was prevalence of SARS-CoV-2 infection. SARS-CoV-2 IgG antibodies were detected by an enzyme-linked immunosorbent assay based on spike (S1/S2) protein, followed by confirmation with a commercial electrochemiluminescence immunoassay based on the receptor binding domain of spike protein. Results: The study enrolled 4198 participants (2539 [60%] female; median age, 50 years [IQR, 25 years]), including 903 (22%), 1046 (25%), and 2249 (53%) during April 21 to July 7, 2020; during September 29 to November 23, 2020; and during January 15 to April 18, 2021, respectively. The numbers of participants aged 18 to 39 years, 40 to 59 years, and 60 years or older were 1328 (32%), 1645 (39%), and 1225 (29%), respectively. Among the participants, 2444 (58%) stayed in Hong Kong since November 2019 and 2094 (50%) had negative SARS-CoV-2 RNA test results. Only 170 (4%) reported ever having contact with individuals with confirmed cases, and 5% had been isolated or quarantined. Most (2803 [67%]) did not recall any illnesses, whereas 737 (18%), 212 (5%), and 385 (9%) had experienced respiratory symptoms, gastrointestinal symptoms, or both, respectively, before testing. Six participants were confirmed to be positive for anti-SARS-CoV-2 IgG; the adjusted prevalence of unidentified infection was 0.15% (95% CI, 0.06%-0.32%). Extrapolating these findings to the whole population, there were fewer than 1.9 unidentified infections for every recorded confirmed case. The overall prevalence of SARS-CoV-2 infection in Hong Kong before the roll out of vaccination was less than 0.45%. Conclusions and Relevance: In this cross-sectional study of participants from the general public in Hong Kong, the prevalence of unidentified SARS-CoV-2 infection was low after 3 major waves of the pandemic, suggesting the success of the pandemic mitigation by stringent isolation and quarantine policies even without complete city lockdown. More than 99.5% of the general population of Hong Kong remain naive to SARS-CoV-2, highlighting the urgent need to achieve high vaccine coverage.

Temporal-Geographical Dispersion of SARS-CoV-2 Spike Glycoprotein Variant Lineages and Their Functional Prediction Using in Silico Approach.

SARS-CoV-2 is a positive-sense single-stranded RNA virus with emerging mutations, especially on the Spike glycoprotein (S protein). To delineate the genomic diversity in association with geographic dispersion of SARS-CoV-2 variant lineages, we collected 939,591 complete S protein sequences deposited in the Global Initiative on Sharing All Influenza Data (GISAID) from December 2019 to April 2021. An exponential emergence of S protein variants was observed since October 2020 when the four major variants of concern (VOCs), namely, alpha (α) (B.1.1.7), beta (ß) (B.1.351), gamma (γ) (P.1), and delta (δ) (B.1.617), started to circulate in various communities. We found that residues 452, 477, 484, and 501, the 4 key amino acids located in the hACE2 binding domain of S protein, were under positive selection. Through in silico protein structure prediction and immunoinformatics tools, we discovered D614G is the key determinant to S protein conformational change, while variations of N439K, T478I, E484K, and N501Y in S1-RBD also had an impact on S protein binding affinity to hACE2 and antigenicity. Finally, we predicted that the yet-to-be-identified hypothetical N439S, T478S, and N501K mutations could confer an even greater binding affinity to hACE2 and evade host immune surveillance more efficiently than the respective native variants. This study documented the evolution of SARS-CoV-2 S protein over the first 16 months of the pandemic and identified several key amino acid changes that are predicted to confer a substantial impact on transmission and immunological recognition. These findings convey crucial information to sequence-based surveillance programs and the design of next-generation vaccines. IMPORTANCE Our study showed the global distribution of SARS-CoV-2 S protein variants from January 2020 to the end of April 2021. We highlighted the key amino acids of S protein subjected to positive selection. Using computer-aided approaches, we predicted the impact of the amino acid variations in S protein on viral infectivity and antigenicity. We also predicted the potential amino acid mutations that could arise in favor of SARS-CoV-2 virulence. These findings are vital for vaccine designing and anti-SARS-CoV-2 drug discovery in an effort to combat COVID-19.

Longitudinal Genomic Characterization of Carbapenemase-producing Enterobacteriaceae (CPE) Reveals Changing Pattern of CPE Isolated in Hong Kong Hospitals.

An active, territory-wide, CPE surveillance program implemented from 2011 showed increasing levels of carbapenemase-producing Enterobacteriaceae (CPE) isolates from patients in Hong Kong hospitals. The molecular epidemiology of 567 CPE from patients of three of seven public hospital clusters in Hong Kong are described. During a 7-year period, the incidence of CPE isolation increased from 0.05 to 9.6/100 000 patient-days. The carbapenemase genes identified were polyclonal, including blaKPC, blaNDM and blaIMP, which were mainly associated with hospitalization overseas in previous years. However, increasing CPE isolation from patients without hospitalization overseas occurred in 2015, with blaNDM (52.6%) predominant followed by blaIMP (30.0%). Escherichia coli (46.4%) and Klebsiella spp. (38.3%) were the dominant species. Whole-genome sequencing was performed on 169 representative isolates with a combination of short and long reads using Illumina and Nanopore technology. Two distinct lineages of blaKPC-2-positive Klebsiella pneumoniae (ST11 and ST258) were identified with ST11 carrying yersiniabactin gene ybt-9 on ICEKp3. ST131 E. coli producing IMP-4 was present throughout the study period. The blaNDM and blaIMP genes were mainly carried in IncX3 and IncN-ST7 plasmids, respectively. blaOXA-48-like gene was carried in the IncX3 plasmid in E. coli and in the ColKP3 plasmid in K. pneumoniae. A lineage of K. pneumoniae with blaNDM-1 plus blaOXA-232 in distinct plasmids of IncF1B/IncHI1B was identified and associated with prior hospitalization overseas. This study highlights the threat of multiple types of CPE, with the predominance of blaNDM and blaIMP among CPE in our hospitals. Enhanced containment strategies are needed to mitigate the trend of rapidly rising CPE in healthcare settings.

A global analysis of replacement of genetic variants of SARS-CoV-2 in association with containment capacity and changes in disease severity.

OBJECTIVES: To examine severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variant replacement in association with containment capacity and changes in case fatality at country level. METHODS: Altogether, 69 571 full SARS-CoV-2 genomes collected globally within the first 6 months of the pandemic were examined. The correlation between variant replacement and containment capacity was examined by logistic regression models using the WHO International Health Regulation (IHR) score, the Oxford COVID-19 Government Response Tracker (OxCGRT) and the vulnerability index INFORM as proxies, while correlation with changes in monthly crude case fatality ratios was examined by a mixed effect model. RESULTS: At the global level, variant lineage G∗, characterized by the S-D614G mutation, replaced the older lineages L and S in March 2020. European countries-including Finland, France and Italy-were the first to reach a 50% increment of G∗, whereas only Singapore and South Korea had non-G∗ persisting throughout the first 6 months. Countries with higher IHR scores (ß-coefficient -0.001, 95%CI -0.016, -0.001; p 0.034) and higher stringency indexes (OxCGRT) (ß-coefficient -0.011, 95%CI -0.020, -0.001; p 0.035) were associated with lower levels of G∗ replacement, whereas higher vulnerability indexes (INFORM) (ß-coefficient 0.049, 95%CI 0.001, 0.097; p 0.044) were associated with higher replacement levels. Crude case fatality ratio showed a positive correlation with G∗ replacement (ß-coefficient: 0.034, 95%CI 0.011, 0.058; p 0.004), even after adjusting for testing capacity and other country-specific characteristics. CONCLUSIONS: SARS-CoV-2 variant lineage G∗ (S-D614G) replaced older lineages more efficiently in countries with lower containment capacity, and its possible association with increased disease severity deserves further investigation.

Acceptance of the COVID-19 vaccine based on the health belief model: A population-based survey in Hong Kong.

BACKGROUND: Vaccines for COVID-19 are anticipated to be available by 2021. Vaccine uptake rate is a crucial determinant for herd immunity. We examined factors associated with acceptance of vaccine based on (1). constructs of the Health Belief Model (HBM), (2). trust in the healthcare system, new vaccine platforms and manufacturers, and (3). self-reported health outcomes. METHODS: A population-based, random telephone survey was performed during the peak of the third wave of COVID-19 outbreak (27/07/2020 to 27/08/2020) in Hong Kong. All adults aged ≥ 18 years were eligible. The survey included sociodemographic details; self-report health conditions; trust scales; and self-reported health outcomes. Multivariable regression analyses were applied to examine independent associations. The primary outcome is the acceptance of the COVID-19 vaccine. RESULTS: We conducted 1200 successful telephone interviews (response rate 55%). The overall vaccine acceptance rate after adjustment for population distribution was 37.2% (95% C.I. 34.5-39.9%). The projected acceptance rates exhibited a "J-shaped" pattern with age, with higher rates among young adults (18-24 years), then increased linearly with age. Multivariable regression analyses revealed that perceived severity, perceived benefits of the vaccine, cues to action, self-reported health outcomes, and trust in healthcare system or vaccine manufacturers were positive correlates of acceptance; whilst perceived access barriers and harm were negative correlates. Remarkably, perceived susceptibility to infection carried no significant association, whereas recommendation from Government (aOR = 10.2, 95% C.I. 6.54 to 15.9, p < 0.001) was as the strongest driving factor for acceptance. Other key obstacles of acceptance included lack of confidence on newer vaccine platforms (43.4%) and manufacturers without track record (52.2%), which are of particular relevance to the current context. CONCLUSIONS: Governmental recommendation is an important driver, whereas perceived susceptibility is not associated with acceptance of COVID-19 vaccine. These HBM constructs and independent predictors inform evidence-based formulation and implementation of vaccination strategies.

Laboratory testing for the diagnosis of COVID-19.

Rapid and accurate laboratory diagnosis of active COVID-19 infection is one of the cornerstones of pandemic control. With the myriad of tests available in the market, the use of correct specimen type and laboratory-testing technique in the right clinical scenario could be challenging for non-specialists. In this mini-review, we will discuss the difference in diagnostic performance for different upper and lower respiratory tract specimens, and the role of blood and fecal specimens. We will analyze the performance characteristics of laboratory testing techniques of nucleic acid amplification tests, antigen detection tests, antibody detection tests, and point-of-care tests. Finally, the dynamics of viral replication and antibody production, and laboratory results interpretation in conjunction with clinical scenarios will be discussed.

Stringent containment measures without complete city lockdown to achieve low incidence and mortality across two waves of COVID-19 in Hong Kong.

INTRODUCTION: An international city, Hong Kong, in proximity to the first epicentre of COVID- 19, experienced two epidemic waves with different importation pressure. We compared the epidemiological features of patients with COVID-19 in the context of containment policies between the first and second waves. METHODS: We retrieved information on the first 1038 cases detected in Hong Kong (23 January to 25 April 2020) to analyse the epidemiological characteristics including age/gender-specific incidence, clustering, reproduction number (Rt ) and containment delay; in relation to the containment measures implemented. Factors associated with containment delay were evaluated by multiple linear regression analysis with age, gender, epidemic wave and infection source as covariates. A time series of 5-day moving average was plotted to examine the changes across the two epidemic waves. RESULTS: The incidence and mortality (135.5 and 0.5 per 1 000 000 population) was among the lowest in the world. Aggressive escalation of border control correlated with reductions in Rt from 1.35 to 0.57 and 0.92 to 0.18, and aversions of 450 and 1650 local infections during the first and second waves, respectively. Implementing COVID-19 tests for overseas returners correlated with an upsurge of asymptomatic case detection, and shortened containment delay in the second wave. Medium-sized cluster events in the first wave were family gatherings, whereas those in the second wave were leisure activities among youngsters. Containment delay was associated with older age (adjusted OR (AOR)=1.01, 95% CI 1.00 to 1.02, p=0.040), male gender (AOR=1.41, 95% CI 1.02 to 1.96, p=0.039) and local cases (AOR=11.18, 95% CI 7.43 to 16.83, p<0.001), and with significant improvement in the second wave compared with the first wave (average: 6.8 vs 3.7 days). A higher incidence rate was observed for males, raising possibility of gender predilection in susceptibility of developing symptoms. CONCLUSION: Prompt and stringent all-round containment strategies represent successful measures in pandemic control. These findings could inform formulation and implementation of pandemic mitigation strategies.