Tropane alkaloids (TAs) are a class of anticholinergic drugs widely used in clinical practice and mainly extracted from plant, among which Atopa belladonna is the main commercial drug source. It is of great industrial value to obtain TAs in large quantities by plant metabolic engineering. In TAs pathway, cytochrome oxidase CYP82M3 catalyze the synthesis of tropinone and then tropinone reductase I (TRI) compete with TRII for tropinone to form tropine leading to the TAs synthesis (drainage). In this study, based on the "increasing flow and drainage" metabolic engineering strategy, two genes, namely HnCYP82M3 and DsTRI from Hyoscyamus niger and Datura stramonium, respectively, were overexpressed in the hair roots of A. belladonna, with a view to promote the TAs accumulation. The HnCYP82M3 gene was cloned from the root of H. niger, and it encoded amino acid with 91.7% sequence identity with AbCYP82M3 from A. belladonna. Overexpression of HnCYP82M3 alone did not affect the content of TAs in hair roots of A. belladonna, indicating that CYP82M3 was not a key enzyme in TAs biosynthesis. Simultaneous overexpression of HnCYP82M3 and DsTRI greatly promoted the accumulation of the three TAs, and the contents of hyoscyamine, anisodamine and scopolamine were 4.97 times, 2.83 times and 2.19 times that of the control, respectively, and the increase amplitude was greater than that of single overexpression of DsTRI. This study showed that the "increasing flow and drainage" strategy of enzyme genes co-expression at branch points was a promising metabolic engineering method to effectively improve the biosynthesis of TAs in A. belladonna, and laid a theoretical and technical foundation for the large-scale industrial acquisition of TAs.

Objective: To construct a prognostic model for unsuccessful removal of nasogastric tube (NGT) was the aim of our study. Methods: This study examined patients with swallowing disorders receiving NGT feeding due to stroke or traumatic brain injury in a regional hospital. Clinical data was collected, such as age, sex, body mass index (BMI), level of activities of daily living (ADLs) dependence. Additionally, gather information regarding the enhancement in Functional Oral Intake Scale (FOIS) levels and the increase in food types according to the International Dysphagia Diet Standardization Initiative (IDDSI) after one month of swallowing training. A stepwise logistic regression analysis model was employed to predict NGT removal failure using these parameters. Results: Out of 203 patients, 53 patients (26.1%) had experienced a failed removal of NGT after six months of follow-up. The strongest predictors for failed removal were age over 60 years, underweight BMI, total dependence in ADLs, and ischemic stroke. The admission prediction model categorized patients into high, moderate, and low-risk groups for removal failure. The failure rate of NGT removal was high not only in the high-risk group but also in the moderate-risk groups when there was no improvement in FOIS levels and IDDSI food types. Conclusion Our predictive model categorizes patients with brain insults into risk groups for swallowing disorders, enabling advanced interventions such as percutaneous endoscopic gastrostomy for high-risk patients struggling with NGT removal, while follow-up assessments using FOIS and IDDSI aid in guiding rehabilitation decisions for those at moderate risk.

The rol genes on pRiA4 plasmid of Agrobacterium rhizogenes are potent genes that promote secondary metabolism. Molecular breeding of Atropa belladonna can be conducted by introducing rol genes to increase tropane alkaloids (TAs) content in A. belladonna. In this study, the rolB gene was overexpressed in A. belladonna plants to study the effect of rolB gene on the biosynthesis of TAs. The phenotype, TAs content and expression levels of key enzyme genes in the pathway of TAs biosynthesis of transgenic A. belladonna were analyzed. The results showed that transgenic A. belladonna had developed root system, enlarged leaves, increased leaf fresh weight, deepened leaf color, enlarged flowers, changed flower shape, reduced pistil height and decreased pollen vitality. The content of TAs in the stems of transgenic A. belladonna was significantly higher than that of the control, and the contents of scopolamine, anisodamine, hyoscyamine can reach 2.11-2.91, 1.23-2.37 and 4.88-5.20 times of the control, respectively. Compared with the control group, the expressions of key enzymes putrescine N-methyltransferase (PMT), type III polyketide synthase (PYKS), tropinone reductase I (TRI), aromatic amino acid aminotransferase 4 (ArAT4), UDP-glycosyltransferase 1 (UGT1) and hyoscyamine 6-β-hydroxylase (H6H) in the TAs biosynthesis pathway were up-regulated, and the expression of tropinone reductase II (TRII) as a metabolic shunting gene was down-regulated. The results indicated that rolB gene enhanced TAs synthesis ability in roots and accumulation in stems of A. belladonna by enhancing metabolic flow of TAs synthesis pathway and weakening the metabolic shunt of competing pathway. This study laid a foundation for molecular breeding of A. belladonna with high-yield TAs content using rolB gene.

Mycena subpiligera, a new taxon in sect. Fragilipedes that can strongly enhance the germination efficiency of Gastrodia elata seeds, was discovered in subtropical areas of China. As revealed by a morphological comparison with related Mycena species as well as maximum likelihood (ML) and Bayesian phylogenetic analyses based on sequences of the internal transcribed spacer (ITS) and the large subunit (LSU) regions of nuclear ribosomal RNA, the new taxon can be distinguished from phenotypically similar and phylogenetically related species. Optimal cultural conditions for M. subpiligera basidiomata are reported, and the germination rate of the new species is compared with that of M. citrinomarginata.

Hair roots induced by Agrobacterium rhizogene produce higher levels of secondary metabolites than non-induced plants, and the enhanced metabolic capacity is driven by the rol gene. We hypothesized that rol genes can be utilized to improve the biosynthesis of tropane alkaloids (TAs) in Atropa belladonna. In this study, the rolC gene from Agrobacterium rhizogene pRiA4 plasmid, driven by a CaMV35S promoter, was overexpressed in A. belladonna. The phenotypes, TAs content and transcriptional expression of key genes in TAs biosynthesis were analyzed in transgenic A. belladonna plants. Results show that transgenic A. belladonna exhibited a well-developed root system, male sterility, higher stamen column length than pistil, early flowering, internode shortening, smaller but more flowers, increased axillary buds and lateral buds, decreased apical dominance, and long and narrow leaves as compared to wild-type plants. Transgenic A. belladonna produced more TAs than wild-type plants, with the content of hyoscyamine, anisodamine and scopolamine reaching 2.58, 3.59 and 15.77-fold that of the control group, respectively. The gene expression of putrescine N-methyltransferase (PMT), tropinone reductase I (TRⅠ) and hyoscyamine 6-β-hydroxylase (H6H), key enzymes in TAs biosynthesis, were up-regulated compared with the control group. The above results indicate that the rolC gene enhances TAs biosynthesis in A. belladonna by up-regulating the expression of key enzymes in the TAs biosynthesis pathway, laying a foundation for genetic manipulation of A. belladonna to increase TAs content by increasing rolC gene expression.

Objective: To analyze the course of disease and epidemiological parameters of COVID-19 and provide evidence for making prevention and control strategies. Methods: To display the distribution of course of disease of the infectors who had close contacts with COVID-19 cases from January 1 to March 15, 2020 in Guangdong Provincial, the models of Lognormal, Weibull and gamma distribution were applied. A descriptive analysis was conducted on the basic characteristics and epidemiological parameters of course of disease. Results: In total, 515 of 11 580 close contacts were infected, with an attack rate about 4.4%, including 449 confirmed cases and 66 asymptomatic cases. Lognormal distribution was fitting best for latent period, incubation period, pre-symptomatic infection period of confirmed cases and infection period of asymptomatic cases; Gamma distribution was fitting best for infectious period and clinical symptom period of confirmed cases; Weibull distribution was fitting best for latent period of asymptomatic cases. The latent period, incubation period, pre-symptomatic infection period, infectious period and clinical symptoms period of confirmed cases were 4.50 (95%CI:3.86-5.13) days, 5.12 (95%CI:4.63-5.62) days, 0.87 (95%CI:0.67-1.07) days, 11.89 (95%CI:9.81-13.98) days and 22.00 (95%CI:21.24-22.77) days, respectively. The latent period and infectious period of asymptomatic cases were 8.88 (95%CI:6.89-10.86) days and 6.18 (95%CI:1.89-10.47) days, respectively. Conclusion: The estimated course of COVID-19 and related epidemiological parameters are similar to the existing data.
COVID-19 ; Cohort Studies ; Contact Tracing ; Humans ; Incidence ; Prospective Studies

Introduction: Apical Hypertrophic Cardiomyopathy (Apical HCM) is an uncommon variant of hypertrophic cardiomyopathy, but it is relatively more common in Asian countries. This is a retrospective, non-randomised, single centre study of patients with Apical HCM focusing on their diastolic dysfunction grading, echocardiographic parameters and electrocardiograms (ECG). Methods: All Apical HCM patients coming for clinic visits at the Institut Jantung Negara from September 2017 to September 2018 were included. We assessed their echocardiography images, grade their diastolic function and reviewed their ECG on presentation. Results: Fifty patient were included, 82% (n=41) were males and 18% (n=9) females. The diastolic function grading of 37 (74%) patients were able to be determined using the updated 2016 American Society of Echocardiography (ASE) diastolic guidelines. Fifty percent (n=25) had the typical ace-ofspades shape left ventricle (LV) appearance in diastole and 12% (n=6) had apical pouch. All patients had T inversion in the anterior leads of their ECG, and only 52% (n=26) fulfilled the ECG left ventricular hypertrophy (LVH) criteria. Majority of our patients presented with symptoms of chest pain (52%, n=26) and dyspnoea (42%, n=21). Conclusion: The updated 2016 ASE guideline makes it easier to evaluate LV diastolic function in most patients with Apical HCM. It also helps in elucidating the aetiology of dyspnoea, based on left atrial pressure. Clinicians should have a high index of suspicion for Apical HCM when faced with deep T inversion on ECG, in addition to a thick LV apex with an aceof-spades appearance during diastole.

Cystatin-C (Cys-C) has been reported as a valuable prognostic biomarker in various malignancies. However, its effect on upper tract urothelial carcinoma (UTUC) patients has not been investigated before. Thus, to explore the impact of Cys-C on survival outcomes in patients undergoing radical nephroureterectomy (RNU), a total of 538 patients with UTUC who underwent RNU between 2005 and 2014 in our center (West China Hospital, Chengdu, China) were included in this study. Kaplan-Meier method and Cox regression analyses were performed to assess the relationship between Cys-C and survival outcomes using SPSS version 22.0. The cutoff value of Cys-C was set as 1.4 mg l^-1 using the receiver operating characteristic (ROC) curves and Youden index. The mean age of patients included was 66.1 ± 11.1 years, and the median follow-up duration was 38 (interquartile range: 19-56) months. Overall, 162 (30.1%) patients had elevated Cys-C, and they were much older and had worse renal function than those with Cys-C <1.4 mg l^-1 (both P < 0.001). Meanwhile, Kaplan-Meier analysis revealed that the group with elevated Cys-C had worse cancer-specific survival (CSS, P = 0.001), disease recurrence-free survival (RFS, P = 0.003), and overall survival (OS, P < 0.001). Multivariable Cox analysis suggested that the elevated Cys-C was identified as an independent prognostic predictor of CSS (hazard ratio [HR]: 1.997, 95% confidential interval [CI]: 1.331-2.996), RFS (HR: 1.429, 95% CI: 1.009-2.023), and OS (HR: 1.989, 95% CI: 1.366-2.896). In conclusion, our result revealed that the elevated preoperative serum Cys-C was significantly associated with worse outcomes in UTUC patients undergoing RNU.
Age Factors ; Aged ; Biomarkers, Tumor/blood* ; Carcinoma, Transitional Cell/surgery* ; China ; Cystatin C/blood* ; Female ; Humans ; Male ; Middle Aged ; Preoperative Period ; Prognosis ; Retrospective Studies ; Survival Rate ; Urologic Neoplasms/surgery*

INTRODUCTIONTinea unguium is a common nail infection. We conducted a retrospective ten-year study of the patient demographics and species distribution of dermatophytes causing tinea unguium in a tertiary hospital from Singapore.

METHODSResults of fungal nail cultures were retrieved from our hospital's microbiology department. Samples from nail scrapings and clippings were inoculated onto agar plates (Sabouraud dextrose agar with chloramphenicol and Mycosel agar). Nail specimens that grew dermatophytes were included in the study.

RESULTSOverall, 229 (male: n = 164, 71.6%; female: n = 65, 28.4%) nail specimens grew dermatophytes. Mean patient age was 58 (range 18-93) years. A majority of specimens came from patients aged over 50 years (n = 162, 70.7%) and 60-79 years (n = 100, 43.7%). Ethnically, 160 (69.9%) patients were Chinese, 36 (15.7%) Indian, 18 (7.9%) Malay and 15 (6.6%) of other ethnicities. Among dermatophytes isolated were Trichophyton rubrum (n = 93, 40.6%), Trichophyton mentagrophytes (n = 60, 26.2%), unidentified Trichophyton spp. (n = 57, 24.9%), Trichophyton tonsurans (n = 10, 4.4%), Epidermophyton floccosum (n = 5, 2.2%), Trichophyton verrucosum (n = 2, 0.9%), Trichophyton soudanense (n = 1, 0.4%) and Trichophyton violaceum (n = 1, 0.4%).

CONCLUSIONA majority of isolates were from elderly patients. Compared to Singapore's general population, patients of Indian and other ethnicities were over-represented for tinea unguium when compared to Chinese and Malay patients. Trichophyton rubrum was the most common dermatophyte isolated, while Trichophyton verrucosum, Trichophyton violaceum and Trichophyton soudanense were rare causes of tinea unguium.

Chinese herbal medicinal formulation Yinzhihuang injection is widely used in clinic for jaundice and chronic liver diseases in eastern Asian countries. However, the pharmacologically active components and the underlying mechanism are unclear. In this study, 30 male ICR mice were randomly assigned into 6 groups： normal control, model control, chlorogenic acid group, geniposide group, baicalin group and wogonoside group. The liver function, liver pathological changes and bile acid metabolism-related gene expression in mice were assayed. The serum levels of ALT, AST, ALP, TBA in chlorogenic acid group (15.89±2.53), (18.32±2.56), (26.38±9.87) U•L⁻¹, (40.63±7.67) μmol•L⁻¹, respectively and geniposide group (20.54±2.36), (24.28±5.19), (35.09±5.03) U•L⁻¹, (42.86±7.11) μmol•L⁻¹, respectively were lower than those in the model control group (59.52±10.94), (128.37±17.97),(169.52±9.62) U•L⁻¹, (132.50±33.00) μmol•L⁻¹, respectively. Hematoxylin-eosin staining showed necrosis, infiltration of inflammation cell in chlorogenic acid group and geniposide group were milder than those in the model control group. Q-PCR analysis revealed the expression of bile acid metabolism-related genes was normalized after treatment with chlorogenic acid or geniposide. Chlorogenic acid and geniposide improved the liver injury and cholestasis effectively, and reversed the mRNA expression of bile acid metabolism-related genes induced by ANIT. So, chlorogenic acid and geniposide were protective for cholestasis, suggesting their pharmacodynamic effect in Yinzhihuang injection.