Cloning, Heterologous Expression, and Characterization of a βκ-Carrageenase From Marine Bacterium Wenyingzhuangia funcanilytica: A Specific Enzyme for the Hybrid Carrageenan–Furcellaran

Carrageenan is a group of important food polysaccharides with high structural heterogeneity. Furcellaran is a typical hybrid carrageenan, which contains the structure consisted of alternative beta-carrageenan and kappa-carrageenan motifs. Although several furcellaran-hydrolyzing enzymes have been characterized, their specificity for the glycosidic linkage was still unclear. In this study, we cloned, expressed, and characterized a novel GH16_13 furcellaran-hydrolyzing enzyme Cgbk16A_Wf from the marine bacterium Wenyingzhuangia fucanilytica CZ1127. Cgbk16A_Wf exhibited its maximum activity at 50 degrees C and pH 6.0 and showed high thermal stability. The oligosaccharides in enzymatic products were identified by liquid chromatography coupled with high-resolution mass spectrometry (LC-HRMS) and nuclear magnetic resonance (NMR) spectroscopy. It was confirmed that Cgbk16A_Wf specifically cleaves the beta-1,4 linkages between beta-carrageenan and kappa-carrageenan motifs from non-reducing end to reducing end. Considering the structural heterogeneity of carrageenan and for the unambiguous indication of the specificity, we recommended to name the furcellaran-hydrolyzing activity represented by Cgbk16A as beta kappa-carrageenase instead of furcellaranase

Assessing equity of healthcare utilization in rural China: results from nationally representative surveys from 1993 to 2008

Background: The phenomenon of inequitable healthcare utilization in rural China interests policymakers and researchers; however, the inequity has not been actually measured to present the magnitude and trend using nationally representative data. Methods: Based on the National Health Service Survey (NHSS) in 1993, 1998, 2003, and 2008, the Probit model with the probability of outpatient visit and the probability of inpatient visit as the dependent variables is applied to estimate need-predicted healthcare utilization. Furthermore, need-standardized healthcare utilization is assessed through indirect standardization method. Concentration index is measured to reflect income-related inequity of healthcare utilization. Results: The concentration index of need-standardized outpatient utilization is 0.0486[95% confidence interval (0.0399, 0.0574)], 0.0310[95% confidence interval (0.0229, 0.0390)], 0.0167[95% confidence interval (0.0069, 0.0264)] and −0.0108[95% confidence interval (−0.0213, -0.0004)] in 1993, 1998, 2003 and 2008, respectively. For inpatient service, the concentration index is 0.0529[95% confidence interval (0.0349, 0.0709)], 0.1543[95% confidence interval (0.1356, 0.1730)], 0.2325[95% confidence interval (0.2132, 0.2518)] and 0.1313[95% confidence interval (0.1174, 0.1451)] in 1993, 1998, 2003 and 2008, respectively. Conclusions: Utilization of both outpatient and inpatient services was pro-rich in rural China with the exception of outpatient service in 2008. With the same needs for healthcare, rich rural residents utilized more healthcare service than poor rural residents. Compared to utilization of outpatient service, utilization of inpatient service was more inequitable. Inequity of utilization of outpatient service reduced gradually from 1993 to 2008; meanwhile, inequity of inpatient service utilization increased dramatically from 1993 to 2003 and decreased significantly from 2003 to 2008. Recent attempts in China to increase coverage of insurance and primary healthcare could be a contributing factor to counteract the inequity of outpatient utilization, but better benefit packages and delivery strategies still need to be tested and scaled up to reduce future inequity in inpatient utilization in rural China

Clusterin confers gmcitabine resistance in pancreatic cancer

<p>Abstract</p> <p>Objective</p> <p>To measure clusterin expression in pancreatic cancer tissues and cell lines and to evaluate whether clusterin confers resistance to gmcitabine in pancreatic cancer cells.</p> <p>Methods</p> <p>Immunohistochemistry for clusterin was performed on 50 primary pancreatic cancer tissues and 25 matched backgrounds, and clusterin expression in 5 pancreatic cancer cell lines was quantified by Western blot and PT-PCR. The correlation between clusterin expression level and gmcitabine IC50 in pancreatic cancer cell lines was evaluated. The effect of an antisense oligonucleotide (ASO) against clusterin(OGX-011) on gmcitabine resistance was evaluated by MTT assays. Xenograft model was used to demonstrate tumor growth.</p> <p>Results</p> <p>Pancreatic cancer tissues expressed significantly higher levels of clusterin than did normal pancreatic tissues (<it>P </it>< 0.01). Clusterin expression levels were correlated with gmcitabine resistance in pancreatic cancer cell lines, and OGX-011 significantly decreased BxPc-3 cells resistance to gmcitabine (<it>P </it>< 0.01). <it>In vivo </it>systemic administration of AS clusterin and gmcitabine significantly decreased the s.c. BxPC-3 tumor volume compared with mismatch control ODN plus gmcitabine.</p> <p>Conclusion</p> <p>Our finding that clusterin expression was significantly higher in pancreatic cancer than in normal pancreatic tissues suggests that clusterin may confer gmcitabine resistance in pancreatic cancer cells.</p

Changes in caesarean section rates in China during the period of transition from the one-child to two-child policy era: cross-sectional National Household Health Services Surveys

Objectives: Since 2009, China has introduced policies, principally targeting health professionals, to reduce caesarean section (CS) overuse. In 2016, China endorsed a universal two-child policy. Advanced maternal age and previous CS may indicate changes in obstetric risks, which raise concerns on the need for and safety of CS. This study investigated changes in CS rates in 2008–2018, and factors associated with CS use during the period of transition from the one-child to two-child policy era. Design: We used births data from the cross-sectional National Household Health Services Surveys in 2013 and 2018. Setting: Population-based national survey. Participants: Women who had the last live birth within 5 years before the survey. Primary outcome measure: CS rate. Results: Overall CS use increased from 40.9% in 2008 to 47.2% in 2014 with significant increase in rural areas and the western region, and slightly decreased to 45.2% in 2018 with the greatest decrease among nulliparous women. Maternal request for CS by urban nulliparous women decreased from 36.8% in 2008–2009 to 22.2% in 2016–2018, but this change was not statistically significant in rural areas. Maternal age over 35 years old (OR 2.40, 95% CI 1.72 to 3.35) and births that occurred at a private hospital (OR 1.52, 95% CI 1.25 to 1.86) were associated with CS use among nulliparous women in 2016–2018. The CS rate among multiparous women increased over time. Individual socioeconomic factors associated with CS use among multiparous women. Conclusions: The CS rate rise in China in 2008–2018 is attributable to increased use in rural areas and the less developed western region. The population policy shift, alongside facility policies for unnecessary CS reduction, are likely factors in CS reduction in urban areas. The challenge remains to reduce unnecessary CS, at the same time as providing safe, universal access to CS for women in need

Tracking progress towards universal health coverage for essential health services in China, 2008-2018.

INTRODUCTION: We comprehensively evaluate whether the Chinese Government's goal of ensuring Universal Health Coverage for essential health services has been achieved. METHODS: We used data from the 2008, 2013 and 2018 National Health Services Survey to report on the coverage of a range of Sustainable Development Goals (SDG) indicator 3.8.1. We created per capita household income deciles for urban and rural samples separately. We report time trends in coverage and the slope index (SII) and relative index (RII). RESULTS: Despite much lower levels of income and education, rural populations made as much progress as their urban counterparts for most interventions. Coverage of maternal and child health interventions increased substantially in urban and rural areas, with decreasing rich-poor inequalities except for antenatal care. In rural China, one-fifth women could not access 5 or more antenatal visits. Coverage of 8 or more visits were 34% and 68%, respectively in decile D1 (the poorest) and decile D10 (the richest) (SII 35% (95% CI 22% to 48%)). More than 90% households had access to clean water, but basic sanitation was poor for rural households and the urban poorest, presenting bottom inequality. Effective coverage for non-communicable diseases was low. Medication for hypertension and diabetes were relatively high (>70%). But adequate management, counting in preventive interventions, were much lower and decreased overtime, although inequalities were small in size. Screening of cervical and breast cancer was low in both urban and rural areas, seeing no progress overtime. Cervical cancer screening was only 29% (urban) and 24% (rural) in 2018, presenting persisted top inequalities (SII 25% urban, 14% rural). CONCLUSION: China has made commendable progress in protecting the poorest for basic care. However, the 'leaving no one behind' agenda needs a strategy targeting the entire population rather than only the poorest. Blunt investing in primary healthcare facilities seems neither effective nor efficient

Development of a real-time loop-mediated isothermal amplification method for monitoring Pseudomonas lurida in raw milk throughout the year of pasture

IntroductionThe psychrophilic bacterium Pseudomonas lurida (P. lurida) and its thermostable alkaline proteases can seriously damage raw milk quality.MethodsIn this study, specific primers were designed for P. lurida’s gyrB and aprX genes, and a real-time loop-mediated isothermal amplification (RealAmp) rapid detection method was developed for the early monitoring of P. lurida and its proteases in raw milk. A phylogenetic tree of the gyrB and aprX genes of P. lurida was constructed to analyze the homology of the design sequence of the RealAmp primer. The DNA of 2 strains of P. lurida and 44 strains of non-P. lurida were detected via RealAmp to analyze the specificity of the primer.ResultsIt was found that aprX-positive proteases were produced by P. lurida-positive strains only when Pseudomonas fluorescens was negative. The dissociation temperatures of gyrB and aprX in the RealAmp-amplified products were approximately 85.0°C and 90.0°C, respectively. Moreover, DNA was detected through a 10-fold dilution of P. lurida in a pure bacterial solution and artificially contaminated skimmed milk. The limit of detection of P. lurida DNA copy number in the pure bacterial solution was 8.6 copies/μL and that in the 10% skimmed milk was 5.5 copies/μL. Further, 144 raw milk samples throughout the year from three farms in Hebei province were analyzed using RealAmp. The highest detection rate of P. lurida was 56% in the first and third quarters, and that of proteases was 36% in the second quarter. The detection rates of P. lurida and its proteases were the highest in samples collected from pasture 2 (52 and 46%, respectively), and the ability of P. lurida to produce proteases reached 88%.DiscussionIn conclusion, RealAmp established an early and rapid method for the detection of P. lurida and its proteases in raw milk samples, allowing the identification and control of contamination sources in a timely manner to ensure the quality of milk and dairy products

Selection and environmental adaptation along a path to speciation in the Tibetan frog Nanorana parkeri.

Tibetan frogs, Nanorana parkeri, are differentiated genetically but not morphologically along geographical and elevational gradients in a challenging environment, presenting a unique opportunity to investigate processes leading to speciation. Analyses of whole genomes of 63 frogs reveal population structuring and historical demography, characterized by highly restricted gene flow in a narrow geographic zone lying between matrilines West (W) and East (E). A population found only along a single tributary of the Yalu Zangbu River has the mitogenome only of E, whereas nuclear genes of W comprise 89-95% of the nuclear genome. Selection accounts for 579 broadly scattered, highly divergent regions (HDRs) of the genome, which involve 365 genes. These genes fall into 51 gene ontology (GO) functional classes, 14 of which are likely to be important in driving reproductive isolation. GO enrichment analyses of E reveal many overrepresented functional categories associated with adaptation to high elevations, including blood circulation, response to hypoxia, and UV radiation. Four genes, including DNAJC8 in the brain, TNNC1 and ADORA1 in the heart, and LAMB3 in the lung, differ in levels of expression between low- and high-elevation populations. High-altitude adaptation plays an important role in maintaining and driving continuing divergence and reproductive isolation. Use of total genomes enabled recognition of selection and adaptation in and between populations, as well as documentation of evolution along a stepped cline toward speciation

A simplified frailty index and nomogram to predict the postoperative complications and survival in older patients with upper urinary tract urothelial carcinoma

PurposeThis study was designed to investigate the clinical value of a simplified five-item frailty index (sFI) for predicting short- and long-term outcomes in older patients with upper urinary tract urothelial carcinoma (UTUC) patients after radical nephroureterectomy (RNU).MethodThis retrospective study included 333 patients (aged ≥65 years) with UTUC. Patients were classified into five groups: 0, 1, 2, 3, and 3+, according to sFI score. The variable importance and minimum depth methods were used to screen for significant variables, and univariable and multivariable logistic regression models applied to investigated the relationships between significant variables and postoperative complications. Survival differences between groups were analyzed using Kaplan-Meier plots and log-rank tests. Cox proportional hazards regression was used to evaluate risk factors associated with overall survival (OS) and cancer-specific survival (CSS). Further, we developed a nomogram based on clinicopathological features and the sFI. The area under the curve (AUC), Harrel’s concordance index (C-index), calibration curve, and decision curve analysis (DCA) were used to evaluate the nomogram.ResultOf 333 cases identified, 31.2% experienced a Clavien-Dindo grade of 2 or greater complication. Random forest–logistic regression modeling showed that sFI significantly influenced the incidence of postoperative complications in older patients (AUC= 0.756). Compared with patients with low sFI score, those with high sFI scores had significantly lower OS and CSS (p &lt; 0.001). Across all patients, the random survival forest–Cox regression model revealed that sFI score was an independent prognostic factor for OS and CSS, with AUC values of 0.815 and 0.823 for predicting 3-year OS and CSS, respectively. The nomogram developed was clinically valuable and had good ability to discriminate abilities for high-risk patients. Further, we developed a survival risk classification system that divided all patients into high-, moderate-, and low-risk groups based on total nomogram points for each patient.ConclusionA simple five-item frailty index may be considered a prognostic factor for the prognosis and postoperative complications of UTUC following RNU. By using this predictive model, clinicians may increase their accuracy in predicting complications and prognosis and improve preoperative decision-making