623. DACTYLICAPNOS MACROCAPNOS

Summary Dactylicapnos macrocapnos (Prain) Hutch., a tuberous rooted perennial climber from China, India and Nepal, is described and illustrated. Characters distinguishing it from D. scandens are provided, although the two species in cultivation are easily recognized.     

625. DACTYLICAPNOS TORULOSA

Summary Dactylicapnos torulosa (Hook.f. & T. Thomson) Hutch., an annual or short‐lived perennial, is described and illustrated and its fruit dehiscence is discussed.     

应用正交实验法优化大肠杆菌表达CTB-PSMA624-632蛋白的条件

通过基因工程方法,将含霍乱毒素B亚单位(CTB)和前列腺特异性膜抗原(PSMA)表位肽的重组质粒pET28a-CTB-PSMA624-632,转化表达宿主菌BL21 (DE3)。CTB蛋白的表达有利于开发粘膜佐剂和霍乱弧菌基因工程疫苗,同时抗原肽疫苗也是肿瘤免疫治疗的理想候选。本研究采用正交实验法,优化条件以增加大肠杆菌中CTB-PSMA624-632蛋白的表达量。选取了5个单因素作为考察因子,分别为:预诱导时间、诱导时间、诱导温度、IPTG浓度、诱导转速。先做了4水平的实验L16(45)。根据极差分析的数据对影响较大的3个因素又做了细调型的3水平实验L9(33)。结果表明,在早期指数生长期的诱导能得到相对高水平的包涵体形式的CTB-PSMA624-632蛋白,并且此蛋白的表达随着诱导温度升高到37℃而逐渐增加;相比于使用正常浓度的IPTG诱导得到的蛋白表达量,用100%μmol/L诱导足以诱导蛋白的表达,这比通常使用的IPTG浓度小10倍;此外,诱导时间5 h与7 h对蛋白的表达量差距甚微,因此选定较短时间的5 h,240 r/min的诱导转速也被认为是能诱导目的蛋白CTB-PSMA624-632具有高表达量的最优条件。     

Bleaching of Hardwood Kraft Pulp with Manganese Peroxidase Secreted from Phanerochaete sordida YK-624

In vitro bleaching of an unbleached hardwood kraft pulp was performed with manganese peroxidase (MnP) from the fungus Phanerochaete sordida YK-624. When the kraft pulp was treated with partially purified MnP in the presence of MnSO₄, Tween 80, and sodium malonate with continuous addition of H₂O₂ at 37°C for 24 h, the pulp brightness increased by about 10 points and the kappa number decreased by about 6 points compared with untreated pulp. The pulp brightness was also increased by 43 points to 75.5% by multiple (six) treatments with MnP combined with alkaline extraction. Our results indicate that in vitro degradation of residual lignin in hardwood kraft pulp with MnP is possible.     

624例宁波市基层地区女性阴道感染调查分析

目的调查宁波市基层地区女性阴道感染的分布情况,为基层卫生医疗机构制订规范的阴道感染诊治方案提供参考。方法采用生理盐水涂片法和预成酶谱生化检测对624例基层女性患者的阴道分泌物进行联合检测,并记录患者的年龄、职业等情况。结果检出阴道感染患者425例,随着清洁度数的升高,各种阴道感染的检出率呈明显递增趋势,清洁度I~II患者BV、VVC、BV＋VVC的检出率明显低于清洁度III~IV的患者;单纯感染患者295例（69.41%）,其中VVC患者105例（24.70%）,BV患者88例（20.70%）,厌氧菌增殖52例（12.24%）,TV患者24例（5.65%）,其他感染患者26例（6.12%）。混合感染患者130例（30.59%）,其中BV＋VVC患者109例（25.65%）,BV＋TV患者21例（4.94%）;不同年龄组阴道感染发病率比较差异有统计学意义,20岁以下年龄组检出率最低,21~40岁年龄段检出率最高,尤其是BV、VVC、BV＋VVC患病率明显高于其他年龄组。结论基层地区各种阴道感染的发病率均处于较高水平,应加强基层地区女性阴道感染防治的健康教育,规范基层医疗机构的诊治方案,常规涂片镜检联合预成酶谱生化检测技术操作简单、快速、准确,可给临床医生提供科学明确的诊断依据,特别适宜在基层地区广泛推广。     

Dimerization of Bisphenol A by Hyper Lignin-Degrading Fungus Phanerochaete sordida YK-624 Under Ligninolytic Condition

Bisphenol A (BPA) was treated with hyper lignin-degrading fungus Phanerochaete sordida YK-624 under ligninolytic condition. After preculturing P. sordida YK-624 for 4 days, BPA (final concentration, 1 and 0.1 mM) was added to cultures. Both 1- and 0.1-mM BPA were effectively decreased within a 24-h treatment and two metabolites were detected. Two metabolites (5,5′-bis-[1-(4-hydroxy-phenyl)1-methyl-ethyl]-biphenyl-2,2′-diol and 4-(2-(4-hydroxy-phenyl) propan-2-yl)-2-(4-(2-(4-hydroxyphenyl) propan-2-yl) phenoxy)phenol) were identified by ESI–MS and NMR analysis. These results indicated that BPA was oxidized to BPA phenoxy radicals by ligninolytic enzymes and then dimerized at extracellular region.     

Transformation by complementation of a uracil auxotroph of the hyper lignin-degrading basidiomycete Phanerochaete sordida YK-624

Phanerochaete sordida YK-624 is a hyper lignin-degrading basidiomycete possessing greater ligninolytic selectivity than either P. chrysosporium or Trametes versicolor. To construct a gene transformation system for P. sordida YK-624, uracil auxotrophic mutants were generated using a combination of ultraviolet (UV) radiation and 5-fluoroorotate resistance as a selection scheme. An uracil auxotrophic strain (UV-64) was transformed into a uracil prototroph using the marker plasmid pPsURA5 containing the orotate phosphoribosyltransferase gene from P. sordida YK-624. This system generated approximately 50 stable transformants using 2 × 10⁷ protoplasts. Southern blot analysis demonstrated that the transformed pPsURA5 was ectopically integrated into the chromosomal DNA of all transformants. The enhanced green fluorescent protein (EGFP) gene was also introduced into UV-64. The transformed EGFP was expressed in the co-transformants driven by P. sordida glyceraldehyde-3-phosphate dehydrogenase gene promoter and terminator regions.     

Detoxification of aflatoxin B1 by manganese peroxidase from the white-rot fungus Phanerochaete sordida YK-624

Aflatoxin B(1) (AFB(1) ) is a potent mycotoxin with mutagenic, carcinogenic, teratogenic, hepatotoxic, and immunosuppressive properties. In order to develop a bioremediation system for AFB(1) -contaminated foods by white-rot fungi or ligninolytic enzymes, AFB(1) was treated with manganese peroxidase (MnP) from the white-rot fungus Phanerochaete sordida YK-624. AFB(1) was eliminated by MnP. The maximum elimination (86.0%) of AFB(1) was observed after 48 h in a reaction mixture containing 5 nkat of MnP. The addition of Tween 80 enhanced AFB(1) elimination. The elimination of AFB(1) by MnP considerably reduced its mutagenic activity in an umu test, and the treatment of AFB(1) by 20 nkat MnP reduced the mutagenic activity by 69.2%. (1) H-NMR and HR-ESI-MS analysis suggested that AFB(1) is first oxidized to AFB(1) -8,9-epoxide by MnP and then hydrolyzed to AFB(1) -8,9-dihydrodiol. This is the first report that MnP can effectively remove the mutagenic activity of AFB(1) by converting it into AFB(1) -8,9-dihydrodiol.     

Platelets in patients with premature coronary artery disease exhibit upregulation of miRNA340* and miRNA624*

Background

Coronary artery disease (CAD) is the leading cause of human morbidity and mortality worldwide, underscoring the need to improve diagnostic strategies. Platelets play a major role, not only in the process of acute thrombosis during plaque rupture, but also in the formation of atherosclerosis itself. MicroRNAs are endogenous small non-coding RNAs that control gene expression and are expressed in a tissue and disease-specific manner. Therefore they have been proposed to be useful biomarkers. It remains unknown whether differences in miRNA expression levels in platelets can be found between patients with premature CAD and healthy controls.

Methodology/Principal Findings

In this case-control study we measured relative expression levels of platelet miRNAs using microarrays from 12 patients with premature CAD and 12 age- and sex-matched healthy controls. Six platelet microRNAs were significantly upregulated (miR340*, miR451, miR454*, miR545:9.1. miR615-5p and miR624*) and one miRNA (miR1280) was significantly downregulated in patients with CAD as compared to healthy controls. To validate these results, we measured the expression levels of these candidate miRNAs by qRT-PCR in platelets of individuals from two independent cohorts; validation cohort I consisted of 40 patients with premature CAD and 40 healthy controls and validation cohort II consisted of 27 patients with artery disease and 40 healthy relatives. MiR340* and miR624* were confirmed to be upregulated in patients with CAD as compared to healthy controls in both validation cohorts.

Conclusion/Significance

Two miRNAs in platelets are significantly upregulated in patients with CAD as compared to healthy controls. Whether the two identified miRNAs can be used as biomarkers and whether they are cause or consequence of the disease remains to be elucidated in a larger prospective study.     

Improvement of ligninolytic properties by recombinant expression of glyoxal oxidase gene in hyper lignin-degrading fungus Phanerochaete sordida YK-624

Glyoxal oxidase (GLOX) is a source of the extracellular H₂O₂ required for the oxidation reactions catalyzed by the ligninolytic peroxidases. In the present study, the GLOX-encoding gene (glx) of Phanerochaete chrysosporium was cloned, and bee2 promoter of P. sordida YK-624 was used to drive the expression of glx. The expression plasmid was transformed into a P. sordida YK-624 uracil auxotrophic mutant (strain UV-64), and 16 clones were obtained as GLOX-introducing transformants. These transformants showed higher GLOX activities than wild-type P. sordida YK-624 and control transformants harboring marker plasmid. RT-PCR analysis indicated that the increased GLOX activity was associated with elevated recombinant glx expression. Moreover, these transformants showed higher ligninolytic activity than control transformants. These results suggest that the ligninolytic properties of white-rot fungi can be improved by recombinant expression of glx.     

Cloning and transcriptional analysis of the gene encoding 5-aminolevulinic acid synthase of the white-rot fungus Phanerochaete sordida YK-624

In this study, we cloned the gene encoding 5-aminolevulinic acid synthase (ALAS) from the hyper-lignin-degrading fungus Phanerochaete sordida YK-624. The deduced amino acid sequence showed highest identity (93.0%) to ALAS of P. chrysosporium. Expression of the gene encoding ALAS, which we named aas, corresponded temporally with the expression and activity of manganese peroxidase.