Association of smoking cessation with airflow obstruction in workers with silicosis: A cohort study.

BACKGROUND: Studies in general population reported a positive association between tobacco smoking and airflow obstruction (AFO), a hallmark of chronic obstructive pulmonary disease (COPD). However, this attempt was less addressed in silica dust-exposed workers. METHODS: This retrospective cohort study consisted of 4481 silicotic workers attending the Pneumoconiosis Clinic during 1981-2019. The lifelong work history and smoking habits of these workers were extracted from medical records. Spirometry was carried out at the diagnosis of silicosis (n = 4177) and reperformed after an average of 9.4 years of follow-up (n = 2648). AFO was defined as forced expiratory volume in one second (FEV1)/force vital capacity (FVC) less than lower limit of normal (LLN). The association of AFO with smoking status was determined using multivariate logistics regression, and the effect of smoking cessation on the development of AFO was evaluated Cox regression. RESULTS: Smoking was significantly associated with AFO (current smokers: OR = 1.92, 95% CI 1.51-2.44; former smokers: OR = 2.09, 95% CI 1.65-2.66). The risk of AFO significantly increased in the first 3 years of quitting smoking (OR = 1.23, 95% CI 1.02-1.47) but decreased afterwards with increasing years of cessation. Smoking cessation reduced the risk of developing AFO no matter before or after the confirmation of silicosis (pre-silicosis cessation: HR = 0.58, 95% CI 0.46-0.74; post-silicosis cessation: HR = 0.62, 95% CI 0.48-0.79). CONCLUSIONS: Smoking cessation significantly reduced the risk of AFO in the workers with silicosis, although the health benefit was not observed until 3 years of abstinence. These findings highlight the importance of early and long-term smoking cessation among silicotic or silica dust-exposed workers.

Predictive evolutionary modelling for influenza virus by site-based dynamics of mutations.

Influenza virus continuously evolves to escape human adaptive immunity and generates seasonal epidemics. Therefore, influenza vaccine strains need to be updated annually for the upcoming flu season to ensure vaccine effectiveness. We develop a computational approach, beth-1, to forecast virus evolution and select representative virus for influenza vaccine. The method involves modelling site-wise mutation fitness. Informed by virus genome and population sero-positivity, we calibrate transition time of mutations and project the fitness landscape to future time, based on which beth-1 selects the optimal vaccine strain. In season-to-season prediction in historical data for the influenza A pH1N1 and H3N2 viruses, beth-1 demonstrates superior genetic matching compared to existing approaches. In prospective validations, the model shows superior or non-inferior genetic matching and neutralization against circulating virus in mice immunization experiments compared to the current vaccine. The method offers a promising and ready-to-use tool to facilitate vaccine strain selection for the influenza virus through capturing heterogeneous evolutionary dynamics over genome space-time and linking molecular variants to population immune response.

Quantitative and qualitative subgenomic RNA profiles of SARS-CoV-2 in respiratory samples: A comparison between Omicron BA.2 and non-VOC-D614G.

The SARS-CoV-2 Omicron variants are notorious for their transmissibility, but little is known about their subgenomic RNA (sgRNA) expression. This study applied RNA-seq to delineate the quantitative and qualitative profiles of canonical sgRNA of 118 respiratory samples collected from patients infected with Omicron BA.2 and compared with 338 patients infected with non-variant of concern (non-VOC)-D614G. A unique characteristic profile depicted by the relative abundance of 9 canonical sgRNAs was reproduced by both BA.2 and non-VOC-D614G regardless of host gender, age and presence of pneumonia. Remarkably, such profile was lost in samples with low viral load, suggesting a potential application of sgRNA pattern to indicate viral activity of individual patient at a specific time point. A characteristic qualitative profile of canonical sgRNAs was also reproduced by both BA.2 and non-VOC-D614G. The presence of a full set of canonical sgRNAs carried a coherent correlation with crude viral load (AUC â€‹= â€‹0.91, 95% CI 0.88-0.94), and sgRNA ORF7b was identified to be the best surrogate marker allowing feasible routine application in characterizing the infection status of individual patient. Further potentials in using sgRNA as a target for vaccine and antiviral development are worth pursuing.

Immune modulation by rural exposures and allergy protection.

BACKGROUND: Growing up on traditional farms protects children from the development of asthma and allergies. However, we have identified distinct asthma-protective factors, such as poultry exposure. This study aims to examine the biological effect of rural exposure in China. METHODS: We recruited 67 rural children (7.4 ± 0.9 years) and 79 urban children (6.8 ± 0.6 years). Depending on the personal history of exposure to domestic poultry (DP), rural children were further divided into those with DP exposure (DP+ , n = 30) and those without (DP- , n = 37). Blood samples were collected to assess differential cell counts and expression of immune-related genes. Dust samples were collected from poultry stables inside rural households. In vivo activities of nasal administration of DP dust extracts were tested in an ovalbumin-induced asthma model. RESULTS: There was a stepwise increase in the percentage of eosinophils (%) from rural DP+ children (median = 1.65, IQR = [1.28, 3.75]) to rural DP- children (3.40, [1.70, 6.50]; DP+ vs. DP- , p = .087) and to the highest of their urban counterparts (4.00, [2.00, 7.25]; urban vs. DP+ , p = .017). Similarly, rural children exhibited reduced mRNA expression of immune markers, both at baseline and following lipopolysaccharide (LPS) stimulation. Whereas LPS stimulation induced increased secretion of Th1 and proinflammatory cytokines in rural DP+ children compared to rural DP- children and urban children. Bronchoalveolar lavage of mice with intranasal instillation of dust extracts from DP household showed a significant decrease in eosinophils as compared to those of control mice (p < .05). Furthermore, DP dust strongly inhibited gene expression of Th2 signature cytokines and induced IL-17 expression in the murine asthma model. CONCLUSIONS: Immune responses of rural children were dampened compared to urban children and those exposed to DP had further downregulated immune responsiveness. DP dust extracts ameliorated Th2-driven allergic airway inflammation in mice. Determining active protective components in the rural environment may provide directions for the development of primary prevention of asthma.

Association of spirometric restriction with mortality in the silicotics: a cohort study.

BACKGROUND: Restrictive spirometry pattern (RSP), defined as reduced forced vital capacity (FVC) in absence of airflow obstruction (AFO), is associated with increased risk of mortality in general population. However, evidence in the patients with silicosis is limited. This study was aimed to investigate the relationship between RSP and the risk of death in a silicotic cohort. METHOD: This retrospective cohort study used data from the Pneumoconiosis Clinic, Hong Kong Department of Health that containing 4315 patients aged 18-80 years and diagnosed with silicosis during 1981-2019, with a follow-up till 31 December 2019. Spirometry was carried out at the diagnostic examination of silicosis. Lung function categories were classified as normal spirometry (FEV1/FVC ≥ 0.7, FVC ≥ 80% predicted), RSP only (FEV1/FVC ≥ 0.7, FVC < 80% predicted), AFO only (FEV1/FVC < 0.7, FVC ≥ 80% predicted), and RSP&AFO mixed (FEV1/FVC < 0.7, FVC < 80% predicted). The hazard ratio (HR) and 95% confidence intervals (95% CI) were computed using a Cox proportional hazards model adjusting for age, body mass index, history of tuberculosis, smoking status, pack-years, and radiographic characteristics of silicotic nodules. RESULTS: Among the 4315 patients enrolled in the study, the prevalence of RSP was 24.1% (n = 1038), including 11.0% (n = 473) with RSP only and 13.1% (n = 565) with mixed RSP and AFO. During the follow-up period, a total of 2399 (55.6%) deaths were observed. Compared with the silicotics with normal spirometry, those with RSP only had significantly increased risk of all-cause mortality (HR = 1.63, 95% CI 1.44-1.85) and respiratory-related mortality (HR = 1.56, 95% CI 1.31-1.85). Notably, a higher risk of mortality was observed in silicotics with mixed ventilatory defects of both RSP and AFO (all-cause mortality: HR = 2.22, 95% CI 1.95-2.52; respiratory-related mortality: HR = 2.59, 95% CI 2.18-3.07) than in those with RSP only. CONCLUSION: RSP is significantly associated with increased risk of all-cause and respiratory-related mortality in the silicotics, and patients with mixed restrictive and obstructive ventilatory defect have higher risk of mortality than those with single RSP or AFO. These findings emphasize the importance of recognizing RSP in the occupational settings, especially for the silicotic patients with mixed ventilatory defect.

Emergency Department Use Across Income Groups Following an Increase in Cost-Sharing.

Importance: To encourage the appropriate utilization of emergency care, cost-sharing for emergency care was increased from HK$100 (US $12.8) to HK$180 (US $23.1) per visit in June 2017 in all public hospitals in Hong Kong. However, there are concerns that this increase could deter appropriate emergency department (ED) visits and be associated with income-related disparities. Objective: To examine changes in ED visits after the fee increase. Design, Setting, and Participants: This retrospective cohort study used administrative data from June 2015 to May 2019 from all public hospitals in Hong Kong. Participants included all Hong Kong residents aged 64 years and younger, categorized into low-income, middle-income, and high-income groups according to the median household income in their district of residence. Data analysis was performed from May to June 2023. Main Outcomes and Measures: The primary outcome was the ED visit rate per 100 000 people per month, categorized into 3 severity levels (emergency, urgent, and nonurgent). Secondary outcomes include general outpatient (GOP) visit rate, emergency admission rate, and in-hospital mortality rate per month at public hospitals. Segmented regression analyses were used to estimate changes in the level and slope of outcome variables before and after the fee increase. Results: This study included a total of 5 441 679 ED patients (2 606 332 male patients [47.9%]; 2 108 933 patients [38.5%] aged 45-64 years), with 2 930 662 patients (1 407 885 male patients [48.0%]; 1 111 804 patients [37.9%] aged 45-64 years) from the period before the fee increase. The fee increase was associated with an 8.0% (95% CI, 7.1%-9.0%) immediate reduction in ED visits after June 2017, including a 5.9% (95% CI, 3.3%-8.5%) reduction in urgent visits and an 8.9% (95% CI, 8.0%-9.8%) reduction in nonurgent visits. In addition, a 5.7% (95% CI, 4.7%-6.8%) reduction of emergency admissions was found, whereas no significant changes were observed in in-hospital mortality. Specifically, a statistically significant increase in GOP visits (4.1%; 95% CI, 0.9%-7.2%) was found within the low-income group, but this association became insignificant after controlling for the social security group, who were exempted from payment, as a control. Conclusions and Relevance: In this cohort study, the fee increase was not associated with changes in ED visits for emergency conditions, but there was a negative and significant association with both urgent and nonurgent conditions across all income groups. Considering the marginal increase in public GOP services, further study is warranted to examine strategies to protect low-income people from avoiding necessary care.

CAGI6 ID-Challenge: Assessment of phenotype and variant predictions in 415 children with Neurodevelopmental Disorders (NDDs).

In the context of the Critical Assessment of the Genome Interpretation, 6th edition (CAGI6), the Genetics of Neurodevelopmental Disorders Lab in Padua proposed a new ID-challenge to give the opportunity of developing computational methods for predicting patient's phenotype and the causal variants. Eight research teams and 30 models had access to the phenotype details and real genetic data, based on the sequences of 74 genes (VCF format) in 415 pediatric patients affected by Neurodevelopmental Disorders (NDDs). NDDs are clinically and genetically heterogeneous conditions, with onset in infant age. In this study we evaluate the ability and accuracy of computational methods to predict comorbid phenotypes based on clinical features described in each patient and causal variants. Finally, we asked to develop a method to find new possible genetic causes for patients without a genetic diagnosis. As already done for the CAGI5, seven clinical features (ID, ASD, ataxia, epilepsy, microcephaly, macrocephaly, hypotonia), and variants (causative, putative pathogenic and contributing factors) were provided. Considering the overall clinical manifestation of our cohort, we give out the variant data and phenotypic traits of the 150 patients from CAGI5 ID-Challenge as training and validation for the prediction methods development.

Comparison of COVID-19 with influenza A in the ICU: a territory-wide, retrospective, propensity matched cohort on mortality and length of stay.

OBJECTIVES: Direct comparisons between COVID-19 and influenza A in the critical care setting are limited. The objective of this study was to compare their outcomes and identify risk factors for hospital mortality. DESIGN AND SETTING: This was a territory-wide, retrospective study on all adult (≥18 years old) patients admitted to public hospital intensive care units in Hong Kong. We compared COVID-19 patients admitted between 27 January 2020 and 26 January 2021 with a propensity-matched historical cohort of influenza A patients admitted between 27 January 2015 and 26 January 2020. We reported outcomes of hospital mortality and time to death or discharge. Multivariate analysis using Poisson regression and relative risk (RR) was used to identify risk factors for hospital mortality. RESULTS: After propensity matching, 373 COVID-19 and 373 influenza A patients were evenly matched for baseline characteristics. COVID-19 patients had higher unadjusted hospital mortality than influenza A patients (17.5% vs 7.5%, p<0.001). The Acute Physiology and Chronic Health Evaluation IV (APACHE IV) adjusted standardised mortality ratio was also higher for COVID-19 than influenza A patients ((0.79 (95% CI 0.61 to 1.00) vs 0.42 (95% CI 0.28 to 0.60)), p<0.001). Adjusting for age, PaO2/FiO2, Charlson Comorbidity Index and APACHE IV, COVID-19 (adjusted RR 2.26 (95% CI 1.52 to 3.36)) and early bacterial-viral coinfection (adjusted RR 1.66 (95% CI 1.17 to 2.37)) were directly associated with hospital mortality. CONCLUSIONS: Critically ill patients with COVID-19 had substantially higher hospital mortality when compared with propensity-matched patients with influenza A.

Statistical Methods for Disease Risk Prediction with Genotype Data.

Single-nucleotide polymorphism (SNP) is the basic unit to understand the heritability of complex traits. One attractive application of the susceptible SNPs is to construct prediction models for assessing disease risk. Here, we introduce prediction methods for human traits using SNPs data, including the polygenic risk score (PRS), linear mixed models (LMMs), penalized regressions, and methods for controlling population stratification.

Comparing the incubation period, serial interval, and infectiousness profile between SARS-CoV-2 Omicron and Delta variants.

In January 2022, the SARS-CoV-2 Omicron variants initiated major outbreaks and dominated the transmissions in Hong Kong, displacing an earlier outbreak seeded by the Delta variants. To provide insight into the transmission potential of the emerging variants, we aimed to compare the epidemiological characteristics of the Omicron and Delta variants. We analyzed the line-list clinical and contact tracing data of the SARS-CoV-2 confirmed cases in Hong Kong. Transmission pairs were constructed based on the individual contact history. We fitted bias-controlled models to the data to estimate the serial interval, incubation period and infectiousness profile of the two variants. Viral load data were extracted and fitted to the random effect models to investigate the potential risk modifiers for the clinical viral shedding course. Totally 14 401 confirmed cases were reported between January 1 and February 15, 2022. The estimated mean serial interval (4.4 days vs. 5.8 days) and incubation period (3.4 days vs. 3.8 days) were shorter for the Omicron than the Delta variants. A larger proportion of presymptomatic transmission was observed for the Omicron (62%) compared to the Delta variants (48%). The Omicron cases had higher mean viral load over an infection course than the Delta cases, with the elder cases appearing more infectious than the younger cases for both variants. The epidemiological features of Omicron variants were likely an obstacle to contact tracing measures, imposed as a major intervention in settings like Hong Kong. Continuously monitoring the epidemiological feature for any emerging SARS-CoV-2 variants in the future is needed to assist officials in planning measures for COVID-19 control.

Estimation of Vaccine Effectiveness of CoronaVac and BNT162b2 Against Severe Outcomes Over Time Among Patients With SARS-CoV-2 Omicron.

Importance: Few studies have evaluated the waning of vaccine effectiveness against severe outcomes caused by SARS-CoV-2 Omicron infection. Hong Kong is providing inactivated and mRNA vaccines, but the population had limited protection from natural infections before the Omicron variant emerged. Objective: To examine the change in vaccine effectiveness against hospitalization and mortality due to the Omicron variant over time. Design, Setting, and Participants: This case-control study included adults with SARS-CoV-2 Omicron variant infection who died or were hospitalized in Hong Kong from January 1 to June 5, 2022 (ie, case participants), and adults with SARS-CoV-2 Omicron, sampled from the public health registry during the study period (ie, control participants), who were matched to case participants by propensity score. Exposures: Vaccination status of the individuals. Main Outcomes and Measures: Estimated vaccine effectiveness against death, death or hospitalization, and death among hospitalized patients. Vaccine effectiveness was calculated as 1 - adjusted odds ratio obtained by conditional logistic regression adjusted with covariates for each period following vaccination. Results: There were 32 823 case participants (25 546 [77.8%] ≥65 years; 16 930 [47.4%] female) and 131 328 control participants (100 041 [76.2%] ≥65 years; 66 625 [46.6%] female) in the sample analyzed for the death or hospitalization outcome. Vaccine effectiveness against death or hospitalization was maintained for at least 6 months after the second dose of both CoronaVac (74.0%; 95% CI, 71.8%-75.8%) and BNT162b2 (77.4%; 95% CI, 75.5%-79.0%) vaccines. Vaccine effectiveness against death in those aged 18 to 49 years was 86.4% (95% CI, 85.8%-87.0%) and 92.9% (95% CI, 92.6%-93.2%) for those receiving 2 doses of CoronaVac and BNT162b2, respectively, while for patients aged 80 years or older, it dropped to 61.4% (95% CI, 59.8%-63.2%) and 52.7% (95% CI, 50.2%-55.6%) for CoronaVac and BNT162b2, respectively. Nevertheless, overall vaccine effectiveness against death at 4 to 6 months after the third dose was greater than 90% for CoronaVac, BNT162b2, and the mixed vaccine schedule (eg, mixed vaccines: vaccine effectiveness, 92.2%; 95% CI, 89.2%-95.1%). Conclusions and Relevance: While vaccines were generally estimated to be effective against severe outcomes caused by SARS-CoV-2 Omicron infection, this analysis found that protection in older patients was more likely to wane 6 months after the second dose. Hence, a booster dose is recommended for older patients to restore immunity. This is especially critical in a setting like Hong Kong, where third-dose coverage is still insufficient among older residents.

Extra cup of tea intake associated with increased risk of Alzheimer's disease: Genetic insights from Mendelian randomization.

Background: Observational studies report inconclusive effects of tea consumption on the risk of Alzheimer's disease (AD), and the mechanisms are unclear. This study aims to investigate the effects of genetically predicted tea intake (cups of tea consumed per day) on AD, brain volume, and cerebral small vessel disease (CSVD) using the two-sample Mendelian randomization (MR) method. Methods: Summary statistics of tea intake were obtained from UK Biobank (N = 447,485), and AD was from the International Genomics of Alzheimer's Project (N = 54,162). Genetic instruments were retrieved from UK Biobank using brain imaging-derived phenotypes for brain volume outcomes (N > 33,224) and genome-wide association studies for CSVD (N: 17,663-48,454). Results: In the primary MR analysis, tea intake significantly increased the risk of AD using two different methods (ORIVW = 1.48, 95% CI: [1.14, 1.93]; ORWM = 2.00, 95% CI: [1.26, 3.18]) and reached a weak significant level using MR-Egger regression (p < 0.1). The result passed all the sensitivity analyses, including heterogeneity, pleiotropy, and outlier tests. In the secondary MR analysis, per extra cup of tea significantly decreased gray matter (ßWM = -1.63, 95% CI: [-2.41, -0.85]) and right hippocampus volume (ßWM = -1.78, 95% CI: [-2.76, -0.79]). We found a nonlinear association between tea intake and AD in association analysis, which suggested that over-drinking with more than 13 cups per day might be a risk factor for AD. Association analysis results were consistent with MR results. Conclusion: This study revealed a potential causal association between per extra cup of tea and an increased risk of AD. Genetically predicted tea intake was associated with a decreased brain volume of gray matter and the right hippocampus, which indicates that over-drinking tea might lead to a decline in language and memory functions. Our results shed light on a novel possible mechanism of tea intake to increase the risk of AD by reducing brain volume.

A statistical framework for tracking the time-varying superspreading potential of COVID-19 epidemic.

Timely detection of an evolving event of an infectious disease with superspreading potential is imperative for territory-wide disease control as well as preventing future outbreaks. While the reproduction number (R) is a commonly-adopted metric for disease transmissibility, the transmission heterogeneity quantified by dispersion parameter k, a metric for superspreading potential is seldom tracked. In this study, we developed an estimation framework to track the time-varying risk of superspreading events (SSEs) and demonstrated the method using the three epidemic waves of COVID-19 in Hong Kong. Epidemiological contact tracing data of the confirmed COVID-19 cases from 23 January 2020 to 30 September 2021 were obtained. By applying branching process models, we jointly estimated the time-varying R and k. Individual-based outbreak simulations were conducted to compare the time-varying assessment of the superspreading potential with the typical non-time-varying estimate of k over a period of time. We found that the COVID-19 transmission in Hong Kong exhibited substantial superspreading during the initial phase of the epidemics, with only 1 % (95 % Credible interval [CrI]: 0.6-2 %), 5 % (95 % CrI: 3-7 %) and 10 % (95 % CrI: 8-14 %) of the most infectious cases generated 80 % of all transmission for the first, second and third epidemic waves, respectively. After implementing local public health interventions, R estimates dropped gradually and k estimates increased thereby reducing the risk of SSEs to approaching zero. Outbreak simulations indicated that the non-time-varying estimate of k may overlook the possibility of large outbreaks. Hence, an estimation of the time-varying k as a compliment of R as a monitoring of both disease transmissibility and superspreading potential, particularly when public health interventions were relaxed is crucial for minimizing the risk of future outbreaks.

A Prism Vote method for individualized risk prediction of traits in genotype data of Multi-population.

Multi-population cohorts offer unprecedented opportunities for profiling disease risk in large samples, however, heterogeneous risk effects underlying complex traits across populations make integrative prediction challenging. In this study, we propose a novel Bayesian probability framework, the Prism Vote (PV), to construct risk predictions in heterogeneous genetic data. The PV views the trait of an individual as a composite risk from subpopulations, in which stratum-specific predictors can be formed in data of more homogeneous genetic structure. Since each individual is described by a composition of subpopulation memberships, the framework enables individualized risk characterization. Simulations demonstrated that the PV framework applied with alternative prediction methods significantly improved prediction accuracy in mixed and admixed populations. The advantage of PV enlarges as genetic heterogeneity and sample size increase. In two real genome-wide association data consists of multiple populations, we showed that the framework considerably enhanced prediction accuracy of the linear mixed model in five-group cross validations. The proposed method offers a new aspect to analyze individual's disease risk and improve accuracy for predicting complex traits in genotype data.

Rapid evaluation of COVID-19 vaccine effectiveness against symptomatic infection with SARS-CoV-2 variants by analysis of genetic distance.

Timely evaluation of the protective effects of Coronavirus Disease 2019 (COVID-19) vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern is urgently needed to inform pandemic control planning. Based on 78 vaccine efficacy or effectiveness (VE) data from 49 studies and 1,984,241 SARS-CoV-2 sequences collected from 31 regions, we analyzed the relationship between genetic distance (GD) of circulating viruses against the vaccine strain and VE against symptomatic infection. We found that the GD of the receptor-binding domain of the SARS-CoV-2 spike protein is highly predictive of vaccine protection and accounted for 86.3% (P = 0.038) of the VE change in a vaccine platform-based mixed-effects model and 87.9% (P = 0.006) in a manufacturer-based model. We applied the VE-GD model to predict protection mediated by existing vaccines against new genetic variants and validated the results by published real-world and clinical trial data, finding high concordance of predicted VE with observed VE. We estimated the VE against the Delta variant to be 82.8% (95% prediction interval: 68.7-96.0) using the mRNA vaccine platform, closely matching the reported VE of 83.0% from an observational study. Among the four sublineages of Omicron, the predicted VE varied between 11.9% and 33.3%, with the highest VE predicted against BA.1 and the lowest against BA.2, using the mRNA vaccine platform. The VE-GD framework enables predictions of vaccine protection in real time and offers a rapid evaluation method against novel variants that may inform vaccine deployment and public health responses.

Single-Hit Inactivation Drove Tumor Suppressor Genes Out of the X Chromosome during Evolution.

Cancer-related genes are under intense evolutionary pressure. In this study, we conjecture that X-linked tumor suppressor genes (TSG) are not protected by the Knudson's two-hit mechanism and are therefore subject to negative selection. Accordingly, nearly all mammalian species exhibited lower TSG-to-noncancer gene ratios on their X chromosomes compared with nonmammalian species. Synteny analysis revealed that mammalian X-linked TSGs were depleted shortly after the emergence of the XY sex-determination system. A phylogeny-based model unveiled a higher X chromosome-to-autosome relocation flux for human TSGs. This was verified in other mammals by assessing the concordance/discordance of chromosomal locations of mammalian TSGs and their orthologs in Xenopus tropicalis. In humans, X-linked TSGs are younger or larger in size. Consistently, pan-cancer analysis revealed more frequent nonsynonymous somatic mutations of X-linked TSGs. These findings suggest that relocation of TSGs out of the X chromosome could confer a survival advantage by facilitating evasion of single-hit inactivation. SIGNIFICANCE: This work unveils extensive trafficking of TSGs from the X chromosome to autosomes during evolution, thus identifying X-linked TSGs as a genetic Achilles' heel in tumor suppression.

Profiling of SARS-CoV-2 Subgenomic RNAs in Clinical Specimens.

SARS-CoV-2 transcribes a set of subgenomic RNAs (sgRNAs) essential for the translation of structural and accessory proteins to sustain its life cycle. We applied RNA-seq on 375 respiratory samples from individual COVID-19 patients and revealed that the majority of the sgRNAs were canonical transcripts with N being the most abundant (36.2%), followed by S (11.6%), open reading frame 7a (ORF7a; 10.3%), M (8.4%), ORF3a (7.9%), ORF8 (6.0%), E (4.6%), ORF6 (2.5%), and ORF7b (0.3%); but ORF10 was not detected. The profile of most sgRNAs, except N, showed an independent association with viral load, time of specimen collection after onset, age of the patient, and S-614D/G variant with ORF7b and then ORF6 being the most sensitive to changes in these characteristics. Monitoring of 124 serial samples from 10 patients using sgRNA-specific real-time RT-PCR revealed a potential of adopting sgRNA as a marker of viral activity. Respiratory samples harboring a full set of canonical sgRNAs were mainly collected early within 1 to 2 weeks from onset, and most of the stool samples (90%) were negative for sgRNAs despite testing positive by diagnostic PCR targeting genomic RNA. ORF7b was the first to become undetectable and again being the most sensitive surrogate marker for a full set of canonical sgRNAs in clinical samples. The potential of using sgRNA to monitor viral activity and progression of SARS-CoV-2 infection, and hence as one of the objective indicators to triage patients for isolation and treatment should be considered. IMPORTANCE Attempts to use subgenomic RNAs (sgRNAs) of SARS-CoV-2 to identify active infection of COVID-19 have produced diverse results. In this work, we applied next-generation sequencing and RT-PCR to profile the full spectrum of SARS-CoV-2 sgRNAs in a large cohort of respiratory and stool samples collected throughout infection. Numerous known and novel discontinuous transcription events potentially encoding full-length, deleted and frameshift proteins were observed. In particular, the expression profile of canonical sgRNAs was associated with genomic RNA level and clinical characteristics. Our study found sgRNAs as potential biomarkers for monitoring infectivity and progression of SARS-CoV-2 infection, which provides an alternative target for the management and treatment of COVID-19 patients.

SARS-CoV-2 non-structural protein 6 triggers NLRP3-dependent pyroptosis by targeting ATP6AP1.

A recent mutation analysis suggested that Non-Structural Protein 6 (NSP6) of the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is a key determinant of the viral pathogenicity. Here, by transcriptome analysis, we demonstrated that the inflammasome-related NOD-like receptor signaling was activated in SARS-CoV-2-infected lung epithelial cells and Coronavirus Disease 2019 (COVID-19) patients' lung tissues. The induction of inflammasomes/pyroptosis in patients with severe COVID-19 was confirmed by serological markers. Overexpression of NSP6 triggered NLRP3/ASC-dependent caspase-1 activation, interleukin-1ß/18 maturation, and pyroptosis of lung epithelial cells. Upstream, NSP6 impaired lysosome acidification to inhibit autophagic flux, whose restoration by 1α,25-dihydroxyvitamin D3, metformin or polydatin abrogated NSP6-induced pyroptosis. NSP6 directly interacted with ATP6AP1, a vacuolar ATPase proton pump component, and inhibited its cleavage-mediated activation. L37F NSP6 variant, which was associated with asymptomatic COVID-19, exhibited reduced binding to ATP6AP1 and weakened ability to impair lysosome acidification to induce pyroptosis. Consistently, infection of cultured lung epithelial cells with live SARS-CoV-2 resulted in autophagic flux stagnation, inflammasome activation, and pyroptosis. Overall, this work supports that NSP6 of SARS-CoV-2 could induce inflammatory cell death in lung epithelial cells, through which pharmacological rectification of autophagic flux might be therapeutically exploited.