Expression and characterization of a bifunctional alginate lyase named Al163 from the Antarctic bacterium Pseudoalteromonas sp. NJ-21

In this study, an endolytic alginate lyase, named Al163, was identified, cloned, and characterized from the Antarctic bacterium Pseudoalteromonas sp. NJ-21. Comparative sequence analysis showed that the predicted amino acid sequence encoded by al163 belongs to the polysaccharide lyase 6(PL-6) family and has a molecular mass of about 80 kDa. Recombinant enzyme was purified by Ni-Sepharose affinity chromatography. Recombinant Al163 exhibited maximum activity(258 U/mg) at pH 7.0 and 40℃, and thermal stability assays showed retention of almost 90% activity after incubation at 30℃ for 30 min. Al163 activity was stimulated by Cd^(2+), Ca^(2+), Fe^(3+), and Mn^(2+), but inhibited by Cu^(2+), Si^(2+), Fe^(2+), and Ni^(2+). Thin-layer chromatographic analysis indicated that Al163 degraded sodium alginate, poly M, and poly G, generating disaccharides and trisaccharides as the final products. Only a few bacterial strains that produce a bifunctional alginate lyase have been reported. Our results indicate that recombinant Al163 exhibits broad substrate specificity and its products exhibit low degrees of polymerization. Both properties imply high potential for use of the enzyme in several industrial fields, including cosmetics and pharmaceuticals, based on the high demand for biologically active oligosaccharides.

Key genes expression of reductive tricarboxylic acid cycle from deep-sea hydrothermal chemolithoautotrophic Caminibacter profundus in response to salinity, pH and O_2

CO2 fixation pathway of Caminibacter profundus, a chemolithoautotrophic e-Proteobacteria from deep-sea hydrothermal vent, was determined and characterized by genetic and enzymatic analyses. Gene expression of key enzymes for CO2 fixation in response to salinity, pH and O2 in Medium 829 were also investigated. The results demonstrate that C. profundus contained aclB, porA and oorA, the genes encoding key enzymes of reductive tricarboxylic acid （rTCA） cycle. However, genes fragments of cbbL and cbbMencoding key enzyme of Calvin cycle were not recovered. Key enzymatic activities of ATP citrate lyase （ACL）, pyruvate： ferredoxin oxidoreductase （POR） and 2-oxoglutarate： ferredoxin oxidoreductase （OOR） were also present in C. profun- dus. The combination of genetic and enzymatic analyses confirm that C. profundus adopted rTCA cycle for carbon assimilation. The results of aclB and oorA relative expressions of C. profundus demonstrate that the ranges of environmental factors for high genes expression were sea salt 3.0%-5.0% （optimum 3.0%）, pH 5.0-6.5（optimum pH 6.5）, anaerobic to microaerobic conditions （optimum 1.0% 02）. Gene expression pat- terns under different conditions show similar patterns with bacterial growth, revealing that key rTCA cycle genes provided molecular basis for bacterial growth and propagation. Our results suggest that C. profun- dus could regulate key genes of rTCA cycle for carbon assimilation and energy metabolism in response to environmental fluctuations in hydrothermal vent.

Extraction and physicochemical characteristics of a red pigment produced by marine bacterium strain S - 9801

A red pigment that has better biological properties is produced by marine bacterium strainS- 9801. The extraction methods, physicochemical and toxicity of the pigment have been studied. Dissolubility of pigment in the five organic solvent has been tested, and ethanol is optimally chosen for extraction. Physicochemical characteristics of this pigment was stable. The absorbance of the pigment solution was no losing when put under natural light for 10 days or treated by UV for 30 minutes, color of the pigment unchanged after 100℃ hythere for 1 h or 80 ℃ xerother for 2 h. The median lethal dose (LD_50) of the rat by celiac injection was 670.04 mg/kg and minimum lethal dose of oral was greater than 2 000 mg/kg.

胚胎移植不同结局人群的游离DNA核小体印迹差异表达分析

目的分析体外受精-胚胎移植(IVF-ET)治疗后不同结局患者的血浆游离DNA(cfDNA)核小体印迹和差异基因表达谱,探讨影响妊娠结局及胚胎植入的差异基因的生物进程及细胞组分。方法收集2021年5月至2021年10月在广东医科大学顺德妇女儿童医院生殖中心的IVF-ET病例,根据胚胎植入后的结局分为失败组(n=5)和成功组(n=9),按胚胎植入前1天、植入后14天和植入后28天三个时间点分别检测患者血浆游离DNA,通过高通量测序对胚胎植入后不同结局和不同时间点的cfDNA核小体印迹和差异基因表达谱进行分析。结果血浆cfDNA核小体印迹中高表达基因的覆盖深度低于低表达基因,同一结局不同时间和同一时间不同结局的cfDNA核小体印迹的基因表达差异有统计学意义(P<0.05)。基于基因本体(GO)进行功能富集和信号通路分析发现,失败组在不同时间点的差异基因主要与免疫系统变化、发育调节和有机化合物的代谢相关,而成功组的与细胞发育和有机化合物的代谢相关。失败组差异表达基因在植入后14天的有机化合物代谢过程中表达下调,在植入后28天的免疫相关的生物过程表达上调。结论基于血浆游离DNA核小体印迹的差异基因表达分析可有效筛选和富集到胚胎植入后不同结局在不同时间点之间的差异表达基因和相关生物信号通路,为改善IVF-ET结局提供理论依据,利于研究和临床参考。

老年住院患者跌倒风险评估量表及工具的研究进展

跌倒是全球老年人关注的重要问题[1],是我国65岁以上老年人伤害死亡的首位原因[2]。中国老年人跌倒的年发生率为14.7%~34.0%[3]。而医疗机构中的跌倒发生率更高[4],患者住院期间跌倒的发生率是医院护理管理质量重要评价指标之一。因此,在临床护理中跌倒风险的评估是非常重要的工作内容。

Production and characterization of an extracellular polysaccharide of antarctic marine bacteria Pseudoalteromonas sp. S-15-13

Twenty-seven antarctic bacteria producing extracellular polysaccharide （EPS） were selected from 57 strains by staining technology. The effects of major environmental factors on the growth and EPS production of Pseudoalteromonas sp. S - 15 - 13 were investigated, and the EPS was separated and purified for characterization analysis. The results showed that the optimal conditions for the EPS production were culture period, 56 h; growth temperature, 8 ℃ ; carbon source, 1.0% glucose; NaCI concentration, 3.0% ; pH 6.0 - 7.0. The EPS was purified by cold ethanol precipitation, proteins removal, ion exchange chromatography and gel chromatography technology. The molecular mass of EPS - H was 62 kDa as determined by the high performance gel permeation chromatography. Its sugar composition was a homopolymer of marmose analyzed by gas chromatograph spectroscopy. After repeated freezing and thawing of the bacteria hiomass in the presence of EPS, the bacterial growth was much higher than that observed after freezing in the absence of EPS and the difference augmented with the increase of freeze-thaw cycles. It is hypothesized that the adaptation of Pseudoalteromonas sp. S- 15 - 13 to the antarctic marine conditions, characterized by low temperature, high NaCl concentration and repeated freeze-thaw cycles, might be related to the EPS production ability.

Biological characteristics of marine bacterium S-9801 strain and its culture conditions of pigment production

Strain of Flavobacterium sp.(S-9801),was screened from 207 strains of marine bacteria isolated from the Bohai Sea continental shelf and the Zhujiang Estuary,for its red pigment production.The biological characteristics of strain S-9801 and culture conditions of pigment production have been checked out in this study. The color of the bacterial colony on 2216E medium was from coccineus to rose bengal. Optimum culture conditions were sodium chloride concentration(g/dm3),10～30;pH,3～8;temperature,25～28℃;tryptone and yeast extract as nitrogen sources and glucose as carbon source. Under optimum conditions,pigment accumulation started after 12 h,reaching a maximum rate of synthesis at 36 h.

Induction of Volatile Organic Compounds in Leaves of Lycopersicon Esculentum by Nd^(3+)

The effects of Nd^3＋ on the quality and quantity of volatile organic compounds （VOCs） in the leaves of Lycopersicon esculentum were studied. The results demonstrate that Nd^3＋ can increase the total amount of VOC by 75% after treatment for 120 h, as compared with the control. Phyto-oxylipins, terpenoids and aromatic compounds were increased by 73%, 38% and 21%, respectively. （E）-2-hexenal, the most abundant constituent is increased by 74%, β- phellandrene and α-caryophyllene in terpenoids,

基于单细胞RNA测序的人类宫腔中度粘连组织中独特的细胞分子学特征研究

目的构建人类宫腔粘连(IUA)组织的细胞景观及其分子生物学特征,以便更好地了解其免疫微环境,为临床治疗提供新的思路。方法选取2022年2月至2022年4月在东莞市妇幼保健院接受宫腔镜治疗的4例IUA患者为研究对象。利用宫腔镜采集IUA组织,并根据患者的病史、月经史及IUA状态等评分和分级,样本采集后立即送检。严格按照单细胞RNA测序流程进行文库构建、测序、数据比对和基因表达矩阵构建,之后进行细胞分群UMAP降维分析和基于细胞类型的差异基因分析。结果收集到4例分级为中度的IUA组织样本,共获得27511个细胞转录组,分配到6个细胞谱系,分别为T细胞、单核吞噬细胞、上皮细胞、成纤维细胞、内皮细胞和红细胞。与文献报道的正常子宫组织细胞相比,本研究的4例样本具有不同的细胞分布,IUA0202204号样本中单核吞噬细胞与T细胞的占比明显偏高,提示存在较强的细胞免疫反应。结论描述了中度IUA组织的细胞多样性和异质性,以及各细胞亚群的分子学特征,为进一步研究IUA的发病机制与患者的异质性提供了线索。

原发性卵巢功能不全患者血清miRNA-146、OX-LDL和ROS的表达及临床意义

目的探讨原发性卵巢功能不全(primary ovarian insufficiency,POI)患者miRNA-146、氧化低密度脂蛋白(oxidized low density lipoprotein,OX-LDL)、活性氧(reactive oxygen species,ROS)的表达及临床意义。方法100例POI患者作为观察组,随机匹配100例卵巢功能正常的妇女作为对照组。通过荧光定量PCR(real-time PCR,qRT-PCR)检测血清miRNA-146,酶联免疫吸附测定(enzyme-linked immunosorbent assay,ELISA)检测OX-LDL和ROS,超声评估卵巢体积和卵巢动脉收缩期峰值血流速度。miRNA-146模拟物或抑制剂转染小鼠卵巢颗粒细胞,与OX-LDL共培养,评估其细胞活力、集落形成能力、细胞凋亡率和Toll样受体4(toll like receptor 4,TLR4)/核因子-κB(nuclear factor-κB,NF-κB)通路蛋白的表达。结果与对照组相比,POI组miRNA-146表达水平显著下降(P<0.05),OX-LDL和ROS表达水平显著升高(P<0.05),卵巢体积显著缩小(P<0.05),卵巢动脉收缩期峰值血流速度显著下降(P<0.05)。OX-LDL呈剂量依赖性抑制颗粒细胞活性,上调miRNA-146表达可增加卵巢颗粒细胞活力和集落数,降低其凋亡率和TLR4/NF-κB表达水平。结论miRNA-146可能靶向调控下游TLR4/NF-κB信号通路,影响OX-LDL和ROS诱导的POI氧化应激和炎症反应。miRNA-146有望成为早期预测POI的生物标志物和治疗新靶点。

胎儿游离DNA在单基因遗传病检测的研究进展

单基因遗传病种类多而复杂,且有不断增长的趋势,总体发病率较高,临床表现各异,通过产前筛查和诊断,及早阻止致死或严重致残的单基因遗传病患儿的出生,是行之有效的防控策略。基于胎儿游离DNA的无创产前单基因遗传病检测技术已应用于临床,检测的疾病种类不断增加,技术也在不断取得突破,本文就该领域的研究进展做一综述。