Systematic calibration of epitranscriptomic maps using a synthetic modification-free RNA library.

Recent years have witnessed rapid progress in the field of epitranscriptomics. Functional interpretation of the epitranscriptome relies on sequencing technologies that determine the location and stoichiometry of various RNA modifications. However, contradictory results have been reported among studies, bringing the biological impacts of certain RNA modifications into doubt. Here, we develop a synthetic RNA library resembling the endogenous transcriptome but without any RNA modification. By incorporating this modification-free RNA library into established mapping techniques as a negative control, we reveal abundant false positives resulting from sequence bias or RNA structure. After calibration, precise and quantitative mapping expands the understanding of two representative modification types, N6-methyladenosine (m6A) and 5-methylcytosine (m5C). We propose that this approach provides a systematic solution for the calibration of various RNA-modification mappings and holds great promise in epitranscriptomic studies.

Computational pathology in cancer diagnosis, prognosis, and prediction - present day and prospects.

Computational pathology refers to applying deep learning techniques and algorithms to analyse and interpret histopathology images. Advances in artificial intelligence (AI) have led to an explosion in innovation in computational pathology, ranging from the prospect of automation of routine diagnostic tasks to the discovery of new prognostic and predictive biomarkers from tissue morphology. Despite the promising potential of computational pathology, its integration in clinical settings has been limited by a range of obstacles including operational, technical, regulatory, ethical, financial, and cultural challenges. Here, we focus on the pathologists' perspective of computational pathology: we map its current translational research landscape, evaluate its clinical utility, and address the more common challenges slowing clinical adoption and implementation. We conclude by describing contemporary approaches to drive forward these techniques. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

The association between atopic dermatitis and linear growth in children- a systematic review.

To evaluate the association between atopic dermatitis (AD) and linear growth in children, and determine factors associated with compromised linear growth in children with AD. A PRISMA-compliant systematic review was conducted. Databases (PubMed, Embase, Scopus and Cochrane) were searched from inception to June 2024 for articles that reported a quantitative relationship between AD and linear growth in children (< 18 years old). Quality of included articles was assessed using the Joanna Briggs Institute Critical Appraisal Tools while quality of evidence in these studies was evaluated using Grading of Recommendations Assessment, Development and Evaluation (GRADE) criteria. Fourteen studies (comprising 50,146 patients with AD) were included. Seven studies reported a strong positive or positive association between AD and reduced height standard deviation score (SDS) in children; the others reported no association. Only 3 studies had moderate quality of evidence, all of which reported an association between AD and poorer height SDS; the remaining 11 studies scored low in quality of evidence. Three studies reported the impact of AD on height to be transient. Secondary analysis showed AD severity, earlier AD onset, sleep disruption and, food restriction, to be risk factors for linear growth impairment in patients with AD. Topical steroid use was not associated with shorter stature in patients with AD. Conclusion: Current evidence on the association between childhood AD and poor linear growth is weak and inconsistent. However, patients with more severe AD, earlier disease onset, poorer sleep quality and higher nutritional restrictions appear more susceptible to linear growth impairment. What is known? • There is inconsistent evidence of the association between atopic dermatitis (AD) and linear growth in children in current literature, with some studies suggesting that AD may negatively impact linear height while other studies do not report similar associations. What is new? • There is no strong association between AD in childhood and poorer linear growth. • There may be a transient slowing of linear growth in children with AD, mimicking constitutional growth delay. • Children with severe AD, earlier disease onset, poorer sleep quality and nutritional restrictions may be at risk of more significant linear growth impairment. • Topical steroid use does not appear to contribute to shorter height in children with AD.

A survey exploring ophthalmologists' attitudes and beliefs in performing immediate sequential bilateral cataract surgery in Singapore.

BACKGROUND: To evaluate current practice patterns of Immediate Sequential Bilateral Cataract Surgery (ISBCS) by ophthalmologists in Singapore and assess their attitudes towards performing ISBCS in future cataract care. METHODS: An anonymised electronic survey, modified to local context from a similar study conducted in the United Kingdom, was distributed to members of the College of Ophthalmologists, Academy of Medicine, Singapore, from 20 June to 1 September 2023. An initial screening question on prior experience with ISBCS directed the rest of the survey. Questions explored ophthalmologists' current ISBCS practice patterns and the importance of factors affecting their willingness to perform ISBCS. Results were descriptively analysed. RESULTS: Results collated 2 months upon survey dissemination saw a total of 58 respondents from 235 eligible members (24.7% response rate). Of these, 16 (27.6%) were currently performing ISBCS, 37 (63.8%) had never performed, and 5 (8.6%) had stopped performing. In considering ISBCS, patient convenience (n = 11, 68.8%) and reduced hospital visits (n = 8, 50.0%) were the most important factors nominated. The most important barriers to performing ISBCS were medico-legal issues (n = 31, 83.8%) and risk of endophthalmitis (n = 27, 73.0%), followed by perceived lack of evidence for its effectiveness (n = 19, 51.4%). CONCLUSION: This is one of the first studies evaluating ophthalmologists' sentiments towards performing ISBCS in an Asian country. It highlights some of the most pertinent barriers and concerns that ophthalmologists face in performing and offering ISBCS. This study provides a better understanding of the potential role and prospects of ISBCS in future cataract care in Singapore.

Prevalence and risk factors for alcohol use disorders, substance use disorders, and depression anxiety and stress among users of sexual health services in Singapore: a cross-sectional survey study.

Syndemics of poor mental health also drive poorer sexual health outcomes. This study used three scales, the Alcohol Use Disorders Identification Test (AUDIT), the Drug Abuse Screening Test (DAST-10), and the Depression Anxiety Stress Scale (DASS-21) among beneficiaries of sexual health services in Singapore (n =975), respectively. We found that a prevalence of 20.4% and 18.6% of hazardous and moderate-severe alcohol use disorders and substance use risks, respectively. About 13.7%, 18.1% and 10.5% of participants reported severe to extremely severe symptoms of depression, anxiety, and stress, respectively. Further investigation and integrated interventions for mental health in sexual health settings are warranted.

Botanical Origin Differentiation of Malaysian Stingless Bee Honey Produced by Heterotrigona itama and Geniotrigona thoracica Using Chemometrics.

Stingless bee honey, specifically honeydew honey, is generally valued for its better health benefits than those of most blossom types. However, scientific studies about the differentiation of stingless bee honey based on honeydew and blossom origins are very limited. In this study, 13C NMR spectroscopy was employed to quantify the seven major sugar tautomers in stingless bee honey samples, and the major sugar compositions of both honeydew and blossom types were found not significantly different. However, several physicochemical properties of honeydew honey including moisture content, free acidity, electrical conductivity, ash content, acetic acid, diastase, hydrogen peroxide, and mineral elements levels were significantly higher; while total soluble solid, proline, and hydroxymethylfurfural were significantly lower than blossom honey. Greater antioxidant capacity in honeydew honey was proven with higher total phenolic compounds, ABTS, DPPH, superoxide radical scavenging activities, peroxyl radical inhibition, iron chelation, and ferric reducing power. Using principal component analysis (PCA), two clusters of stingless bee honey from the honeydew and blossom origin were observed. PCA also revealed that the differentiation between honeydew and blossom origin of stingless bee honey is possible with certain physicochemical and antioxidant parameters. The combination of NMR spectroscopy and chemometrics are suggested to be useful to determine the authenticity and botanical origin of stingless bee honey.

Creation of a mobile rural workforce following undergraduate longitudinal rural immersion.

BACKGROUND: This study followed the workforce choices of 10-years of graduates from a longitudinal rural immersion programme, which involved living for one academic year in a rural location as a medical student. The Rural Clinical School of Western Australia is a whole-of-state Rural Clinical School partnership involving two medical schools and fourteen rural/remote towns. METHOD: For this longitudinal cohort study, all consenting graduates were contacted annually after graduation, with the outcome measure being rural work location (defined by the Australian Standard Geographical Classification -Remoteness Area) of any duration. RESULTS: There were 417 consenting graduates. Between 16 and 50% of contacted alumni worked rurally for a period of each post-graduate year. Aggregated over time, the majority took up to 30% of their postgraduate training rurally. There was considerable movement in and out of rural work. About 17% of contacted and practicing graduates were working full time rurally at the 2013 contact point. The majority remained in their state of training. The majority identified with GP and other rural-related colleges, and College-affiliation predicted amount of rural training time. Entry into rural work was equivalent for urban-origin and rural origin alumni, suggesting one year of RCS is sufficient to convert commitment to rural work. CONCLUSION: Undergraduate rural immersion is sufficient to create a graduate rural workforce that is far more mobile that was previously appreciated.

ANTI-STAPHYLOCOCCAL ACTIVITY OF MELALEUCA HONEY.

Honey is well-known for its antioxidant properties due to the presence of phytochemical compounds, which are also involved in antibacterial activities. In this study, properties (total phenolic compounds, flavonoids, free radical scavenging activity, ferric reducing-antioxidant power, reducing sugar content, and pH) of Malaysian Melaleuca honey were investigated for their antistaphylococcal activity, against both methicillin-sensitive (MSSA) and resistant (MRSA) Staphylococcus aureus strains using a hole-in-plate diffusion method. The outcome revealed there is a significant positive correlation between the antistaphylococcal activity and increase in honey concentration [20%-80% (v/v) in distilled water], which also correlated with each of the above mentioned parameter in honey, except for pH value that shows negative correlation. Furthermore, there was no difference in susceptibility to Melaleuca honey between MSSA and MRSA strains. Thus, in addition to being an antioxidant product, Melaleuca honey has a potential as a natural antistaphylococcal agent.

Tiger Milk Mushroom: A Comprehensive Review of Nutritional Composition, Phytochemicals, Health Benefits, and Scientific Advancements with Emphasis on Chemometrics and Multi-Omics.

This article presents a comprehensive overview of tiger milk mushroom (TMM), covering its nutritional composition, phytochemicals, health benefits, and related scientific advancements. It describes various potential positive health benefits of TMM, including anticancer, anti-inflammatory, respiratory function enhancement, antioxidant, anti-aging, neuroprotective, photoprotective, antidiabetic, wound-healing, and anti-HIV, among others. This article also underlines the importance of further research into the phytochemicals present in TMM for additional discoveries. It underscores the importance of further research into phytochemicals content of TMM for additional discoveries and emphasizes the potential applications of TMM in nutrition, health, and well-being. Sophisticated techniques, such as chemometrics and multi-omics technologies revealed latest scientific advancements of TMM. This comprehensive overview provides a foundation for future research and development in harnessing TMM's potential for human health.

Antioxidant Peptides and Protein Hydrolysates from Tilapia: Cellular and In Vivo Evidences for Human Health Benefits.

Antioxidant peptides derived from aquatic organisms have attracted tremendous research interest due to their potential applications in human health. Tilapia is one of the most widely farmed aquaculture species globally. The current understanding of tilapia-derived antioxidant peptides is gradually expanding. This review discusses the current knowledge of peptides and protein hydrolysates derived from tilapia muscle, skin, and scales, whose antioxidant capacity has been validated in various cellular and in vivo models. To date, at least 16 peptides and several hydrolysates have been identified from tilapia that protect human and non-human cell models against oxidative injury. Tilapia hydrolysates and peptide mixtures have also shown protective effects in animal models of oxidative stress-associated diseases and exercise-induced oxidative injury and fatigue. The key mechanisms of tilapia hydrolysates and peptide mixtures involve enhancing antioxidant enzyme activities and suppressing radical production. Notably, such hydrolysates also exerted additional in vivo functions, such as anti-inflammatory, anti-diabetic, wound healing, and antiaging properties. Taken together, tilapia-derived antioxidant peptides and hydrolysates represent a valuable source of functional ingredients for applications in functional food, dietary supplements, and therapeutic applications. Continued research into their health benefits is warranted in the future.

Chemical structures, extraction and analysis technologies, and bioactivities of edible fungal polysaccharides from Poria cocos: An updated review.

Poria cocos is a popular medicinal food. Polysaccharides are the key component of Poria cocos, forming 70-90 % of the dry sclerotia mass. Recent studies indicate that Poria cocos polysaccharides (PCP-Cs) have multiple beneficial functions and applications. A literature search was conducted using the Web of Science Core Collection and PubMed databases. For this review, we provided an updated research progress in chemical structures, various extraction and analysis technologies, bioactivities of PCP-Cs, and insights into the directions for future research. The main polysaccharides identified in Poria cocos are water-soluble polysaccharides and acidic polysaccharides. Hot water, alkali, supercritical fluid, ultrasonic, enzyme, and deep eutectic solvent-based methods are the most common methods for PCP-Cs extraction. Technologies such as near-infrared spectroscopy, high-performance liquid chromatography, and ultraviolet-visible spectrophotometry, are commonly used to evaluate the qualities of PCP-Cs. In addition, PCP-Cs have antioxidant, immunomodulatory, neuroregulatory, anticancer, hepatoprotective, and gut microbiota regulatory properties. Future research is needed to focus on scaling up extraction, enhancing quality control, elucidating mechanisms of bioactivities, and the utilisation of PCP-Cs in food industries. Overall, Poria cocos is a good source of edible fungi polysaccharides, which can be developed into functional foods with potential health benefits.

Chondroitin Sulfate Proteoglycan 4 (CSPG4) as an Emerging Target for Immunotherapy to Treat Melanoma.

Immunotherapies, including checkpoint inhibitor antibodies, have precipitated significant improvements in clinical outcomes for melanoma. However, approximately half of patients do not benefit from approved treatments. Additionally, apart from Tebentafusp, which is approved for the treatment of uveal melanoma, there is a lack of immunotherapies directly focused on melanoma cells. This is partly due to few available targets, especially those expressed on the cancer cell surface. Chondroitin sulfate proteoglycan 4 (CSPG4) is a cell surface molecule overexpressed in human melanoma, with restricted distribution and low expression in non-malignant tissues and involved in several cancer-promoting and dissemination pathways. Here, we summarize the current understanding of the expression and functional significance of CSPG4 in health and melanoma, and we outline immunotherapeutic strategies. These include monoclonal antibodies, antibody-drug conjugates (ADCs), chimeric-antigen receptor (CAR) T cells, and other strategies such as anti-idiotypic and mimotope vaccines to raise immune responses against CSPG4-expressing melanomas. Several showed promising functions in preclinical models of melanoma, yet few have reached clinical testing, and none are approved for therapeutic use. Obstacles preventing that progress include limited knowledge of CSPG4 function in human cancer and a lack of in vivo models that adequately represent patient immune responses and human melanoma biology. Despite several challenges, immunotherapy directed to CSPG4-expressing melanoma harbors significant potential to transform the treatment landscape.

Is the National Registration website (Australian Health Practitioner Regulation Agency) effective in tracking Australian medical graduates' rural work?

OBJECTIVE: This study compared rural location identified through the National Registration (AHPRA) registry with location obtained through labour-intensive personal contact. DESIGN: Longitudinal cohort study with two methods to identify the work locations of medical graduates from The Rural Clinical School of Western Australia (RCSWA). PARTICIPANTS: Consenting alumni from the University of Western Australia and the University of Notre Dame Fremantle participating in RCSWA between 2002 and 2009 inclusive and available to contact in 2011. MAIN OUTCOME MEASURE: Percentage location matches between two contact methods. RESULTS: There was 80% agreement for principal suburb, 92% agreement for principal city and 94% agreement for principal state between RCSWA personal contact and the AHPRA registry. AHPRA identified nearly two times as many graduate locations. However, there was only 31% agreement for a rural placement location (of any length). In more detail, for year-long rural placement, personal contact was 88% concordant with AHPRA; work six months or more were less concordant (44% agreement); work less than six months were not concordant (4% agreement). CONCLUSIONS: AHPRA data matched RCSWA alumni data only for graduates in full-time rural work. Since medical alumni spend up to 10 years in pre-vocational and vocational training, which includes many rural options, personal contact was able to pick up the myriad of rural choices, whereas the AHPRA database was not sensitive enough to identify them. Until graduates have stably finished training, the optimal method to identify rural work is through personal contact but statistical correction for missing data needs to be considered.

A review on matcha: Chemical composition, health benefits, with insights on its quality control by applying chemometrics and multi-omics.

This review discussed the origin, manufacturing process, chemical composition, factors affecting quality and health benefits of matcha (Camellia sinensis), and the application of chemometrics and multi-omics in the science of matcha. The discussion primarily distinguishes between matcha and regular green tea with processing and compositional factors, and demonstrates beneficial health effects of consuming matcha. Preferred Reporting Items for Systematic Reviews and Meta-Analyses was adopted to search for relevant information in this review. Boolean operators were incorporated to explore related sources in various databases. Notably, climate, cultivar, maturity of tea leaves, grinding process and brewing temperature impact on the overall quality of matcha. Besides, sufficient shading prior to harvesting significantly increases the contents of theanine and chlorophyll in the tea leaves. Furthermore, the ground whole tea leaf powder delivers matcha with the greatest benefits to the consumers. The health promoting benefits of matcha are mainly contributed by its micro-nutrients and the antioxidative phytochemicals, specifically epigallocatechin-gallate, theanine and caffeine. Collectively, the chemical composition of matcha affected its quality and health benefits significantly. To this end, more studies are required to elucidate the biological mechanisms of these compounds for human health. Chemometrics and multi-omics technologies are useful to fill up the research gaps identified in this review.

Ginger-Enriched Honey Attenuates Antibiotic Resistant Pseudomonas aeruginosa Quorum Sensing Virulence Factors and Biofilm Formation.

Quorum sensing (QS) in Pseudomonas aeruginosa plays an essential role in virulence factors, biofilm formation as well as antibiotic resistance. Approaches that target virulence factors are known to be more sustainable than antibiotics in weakening the infectivity of bacteria. Although honey has been shown to exert antipseudomonal activities, the enhancement of such activity in ginger-enriched honey is still unknown. The main objective of this study was to determine the impacts of honey and ginger-enriched honey on the QS virulence factors and biofilm formation of antibiotic resistant P. aeruginosa clinical isolates. Outcomes showed honey and/or ginger-enriched honey significantly reduced the protease activity, pyocyanin production and exotoxin A concentration of the isolates. The swarming and swimming motility together with biofilm formation in all clinical isolates were also significantly inhibited by both honey samples. Notable morphological alteration of bacterial cells was also observed using scanning electron microscopy. A principal component analysis (PCA) managed to distinguish the untreated group and treatment groups into two distinct clusters, although honey and ginger-enriched honey groups were not well differentiated. This study revealed the effectiveness of honey including ginger-enriched honey to attenuate QS virulence factors and biofilm formation of P. aeruginosa.

Self-Assessment of INTERHEART Risk Stratification among the Middle-Aged Community in Malaysia.

Research background and Objectives: Age is an independent risk factor for cardiovascular disease (CVD), but CVD risk factors are preventable, and lack of awareness of its risk factors is a contributing factor to CVDs. Middle-aged people may be more likely to engage in unhealthy lifestyle behaviours which can increase the risk of CVD. Health self-assessment is crucial for early detection and management of health issues and early lifestyle intervention for better personalised health management. This study aims to determine the self-assessment of INTERHEART risk classification among the middle-aged community in Malaysia. Method: Local community members aged 40-60 years and who are currently residing in Malaysia were recruited via non-randomised sampling. Sociodemographic characteristics and dietary pattern related to salt, fibre, fat (deep fried/snacks), poultry/meat intakes, and other cardiovascular risk factors (waist-hip ratio, medical history related to diabetes/hypertension, history/exposure of tobacco use, psychosocial status, and level of physical activity) were assessed; INTERHEART risk scores were then computed and stratified into low, medium and high risks. Results: Approximately 45% (n = 273/602) of middle-aged respondents in Malaysia are at moderate-to-high risk of cardiovascular events, with men being more likely to develop CVD compared to women. The results of the survey indicated that poultry/meat intake (61%), physical inactivity (59%), and second-hand smoke (SHS) exposure (54%) are the most prevalent risk factors among the respondents. One-third of the respondents consumed excessive salty food and deep fried foods/snacks/fast food, and only one-third of them consumed vegetables/fruits at a recommended level. It is worrying that about a quarter of the respondents felt several periodical/permanent stresses and even felt sad/blue/depressed for two weeks or more in a row. Males, labour workers, and those with lower educational levels are more likely to develop CVD events. Conclusions: This study found that 45% of the middle-aged respondents were having moderate-to-high risk for cardiovascular events with multiple risk factors related to unhealthy lifestyle habits and environmental factors. In addition to non-modifiable factors such as gender and age, sociodemographic factors, i.e., educational level and occupation, are equally important factors to determine CVD risk. Overall, the findings of this study emphasize the clinical relevance of assessing multiple factors in the determination of CVD risks for early prevention and management of cardiovascular diseases.

Spatial analysis of ambulance response times related to prehospital cardiac arrests in the city-state of Singapore.

OBJECTIVES: The main objective of this study was to establish the spatial variation in ambulance response times for out-of-hospital cardiac arrests (OHCAs) in the city-state of Singapore. The secondary objective involved studying the relationships between various covariates, such as traffic condition and time and day of collapse, and ambulance response times. METHODS: The study design was observational and ecological in nature. Data on OHCAs were collected from a nationally representative database for the period October 2001 to October 2004. We used the conditional autoregressive (CAR) model to analyze the data. Within the Bayesian framework of analysis, we used a Weibull regression model that took into account spatial random effects. The regression model was used to study the independent effects of each covariate. RESULTS: Our results showed that there was spatial heterogeneity in the ambulance response times in Singapore. Generally, areas in the far outskirts (suburbs), such as Boon Lay (in the west) and Sembawang (in the north), fared badly in terms of ambulance response times. This improved when adjusted for key covariates, including distance from the nearest fire station. Ambulance response time was also associated with better traffic conditions, weekend OHCAs, distance from the nearest fire station, and OHCAs occurring during nonpeak driving hours. For instance, the hazard ratio for good ambulance response time was 2.35 (95% credible interval [CI] 1.97-2.81) when traffic conditions were light and 1.72 (95% CI 1.51-1.97) when traffic conditions were moderate, as compared with heavy traffic. CONCLUSIONS: We found a clear spatial gradient for ambulance response times, with far-outlying areas' exhibiting poorer response times. Our study highlights the utility of this novel approach, which may be helpful for planning emergency medical services and public emergency responses.

Effectiveness of antihypertensive drugs in hypertensive patients with end stage renal failure.

Hypertension has been identified as one of the causes for end stage renal failure (ESRF) and is likely to worsen kidney function. This retrospective study was carried out at a tertiary hospital in Malaysia with the objective of determining the effectiveness of combination antihypertensive drugs in hypertensive patients with ESRF admitted between 2006 and 2008. Patients with incomplete data and who were on monotherapy were excluded from this study. Although six different combinations gave significant reductions in systolic blood pressure (SBP) (13.38 +/- 9.11 mmHg, p < 0.05) and diastolic blood pressure (DBP) (6.03 +/- 11.39 mmHg, p < 0.05), 69.16% patients did not achieve target blood pressure (BP) (< or = 130/80 mmHg). Combination of beta blocker (BB) with calcium channel blocker (CCB) was the most commonly used. The CCB-diuretic regimen achieved highest percentage of BP control compared to others (40%). Comparison of blood pressure reduction between different combinations of antihypertensive drugs were not significant (p > 0.05) except for CCB-diuretics and BB-CCB-alpha blocker. The findings suggested better BP control with CCB-diuretic relative to other combinations used.

Clinicopathological predictors of survival in resected primary lung adenocarcinoma.

AIMS: Primary lung adenocarcinoma consists of a spectrum of clinical and pathological subtypes that may impact on overall survival (OS). Our study aims to evaluate the impact of adenocarcinoma subtype and intra-alveolar spread on survival after anatomical lung resection and identify different prognostic factors based on stage and histological subtype. METHODS: Newly diagnosed patients undergoing anatomical lung resections without induction therapy, for pT1-3, N0-2 lung adenocarcinoma from April 2011 to March 2013, were included. The effect of clinical-pathological factors on survival was retrospectively assessed. RESULTS: Two hundred and sixty-two patients were enrolled. The 1-year, 3-year and 5-year OS were 88.8%, 64.3% and 51.1%, respectively. Univariate analysis showed lymphovascular, parietal pleural and chest wall invasion to confer a worse 1-year and 5-year prognosis (all p<0.0001). Solid predominant adenocarcinomas exhibited a significantly worse OS (p=0.014). Multivariate analysis did not identify solid subtype as an independent prognostic factor; however, identified stage >IIa, lymphovascular invasion (p=0.002) and intra-alveolar spread (p=0.009) as significant independent predictors of worse OS. Co-presence of intra-alveolar spread and solid predominance significantly reduced OS. Disease-free survival (DFS) was reduced with parietal pleural (p=0.0007) and chest wall invasion (p<0.0001), however, adenocarcinoma subtype had no significant impact on DFS. CONCLUSIONS: Our study demonstrates that solid predominant adenocarcinoma, intra-alveolar spread and lymphovascular invasion confer a worse prognosis and should be used as a prognostic tool to determine appropriate adjuvant treatment.

High-precision mapping reveals rare N6-deoxyadenosine methylation in the mammalian genome.

N6-deoxyadenosine methylation (6mA) is the most widespread type of DNA modification in prokaryotes and is also abundantly distributed in some unicellular eukaryotes. However, 6mA levels are remarkably low in mammals. The lack of a precise and comprehensive mapping method has hindered more advanced investigations of 6mA. Here, we report a new method MM-seq (modification-induced mismatch sequencing) for genome-wide 6mA mapping based on a novel detection principle. We found that modified DNA bases are prone to form a local open region that allows capture by antibody, for example, via a DNA breathing or base-flipping mechanism. Specified endonuclease or exonuclease can recognize the antibody-stabilized mismatch-like structure and mark the exact modified sites for sequencing readout. Using this method, we examined the genomic positions of 6mA in bacteria (E. coli), green algae (C. reinhardtii), and mammalian cells (HEK239T, Huh7, and HeLa cells). In contrast to bacteria and green algae, human cells possess a very limited number of 6mA sites which are sporadically distributed across the genome of different cell types. After knocking out the RNA m6A methyltransferase METTL3 in mouse ES cells, 6mA becomes mostly diminished. Our results imply that rare 6mA in the mammalian genome is introduced by RNA m6A machinery via a non-targeted mechanism.