Gene expression profiles in the testis associated with testis-ova in adult Japanese medaka (Oryziaslatipes) exposed to 17α-ethinylestradiol

The occurrence of oocytes in the testis (testis-ova) of several fish species is often associated with exposure of estrogenic chemicals. However, induction mechanisms of the testis-ova remain to be elucidated. To develop marker genes for detecting testis-ova in the testis, adult male medaka were exposed to nominal concentration of 100 ng L⁻¹ of 17α-ethinylestradiol (EE2) for 3-5 weeks, and 800 ng estradiol benzoate (EB) for 3 weeks (experiment I), and a measured concentration of 20 ng L⁻¹ EE2 for 1-6 weeks (experiment II). Histological analysis was performed for the testis, and microarray analyses were performed for the testis, liver and brain. Microarray analysis in the estrogen-exposed medaka liver showed vitellogenin and choriogenin as estrogen responsive genes. Testis-ova were induced in the testis after 4 weeks of exposure to 100 ng L⁻¹ EE2, 3 weeks of exposure to 800 ng EB, and 6 weeks of exposure to 20 ng L⁻¹ EE2. Microarray analysis of estrogen-exposed testes revealed up-regulation of genes related to zona pellucida (ZP) and the oocytes marker gene, 42Sp50. Using quantitative RT-PCR we confirmed that Zpc5 gene can be used as a marker for the detection of testis-ova in male medaka.     

Detection of early effects of a single herbicide (diuron) and a mix of herbicides and pharmaceuticals (diuron, isoproturon, ibuprofen) on immunological parameters of Pacific oyster (Crassostrea gigas) spat

In the context of massive summer mortality events of the Pacific oyster Crassostrea gigas, the aim of this study was to investigate the early effects on genes, enzymes and haemocyte parameters implicated in immune defence mechanisms in C. gigas oysters exposed to a potentially hostile environment, i.e. to an herbicide alone or within a mixture. Following 2 h of exposure to the herbicide diuron at 1 μg L⁻¹, the repression of different genes implicated in immune defence mechanisms in the haemocytes and the inhibition of enzyme activities, such as laccase-type phenoloxidase (PO) in the plasma, were observed. The inhibition of superoxide dismutase (SOD) activity in the plasma was also observed after 6 and 24 h of exposure. In the mixture with the herbicides diuron and isoproturon, and the pharmaceutical ibuprofen, catecholase-type PO activity in the plasma and the percentage of phagocytosis in the haemocytes were reduced after 6 h of exposure. Our results showed that early effects on molecular, biochemical and cellular parameters can be detected in the presence of diuron alone or within a mixture, giving an insight of its potential effect in situations that can be found in natural environments, i.e. relatively high concentrations for short periods of time.     

Algicidal activity of Salvia miltiorrhiza Bung on Microcystis aeruginosa—Towards identification of algicidal substance and determination of inhibition mechanism

The present study was to isolate and identify a potent algicidal compound from extract of Salvia miltiorrhiza and study the potential inhibition mechanism on Microcystis aeruginosa. Column chromatography and bioassay-guided fractionation methods were carried out to yield neo-przewaquinone A, which was identified by spectral analysis. The EC50 of neo-przewaquinone A on M. aeruginosa were 4.68 mg L⁻¹. In addition, neo-przewaquinone A showed relatively higher security on Chlorella pyrenoidosa and Scenedesmus obliquus, with the EC50 values of 14.78 and 10.37 mg L⁻¹, respectively. For the potential inhibition mechanisms, neo-przewaquinone A caused M. aeruginosa cells morphologic damage or lysis, increased malondialdehyde content and decreased the soluble protein content, total antioxidant and superoxide dismutase activity, and significantly inhibited three photosynthesis-related genes (psaB, psbD, and rbcL). The results demonstrated the algicidal effect of neo-przewaquinone A on M. aeruginosa and provided the possible inhibition mechanisms.     

Zebrafish embryos/larvae for rapid determination of effects on hypothalamic-pituitary-thyroid (HPT) and hypothalamic-pituitary-interrenal (HPI) axis: mRNA expression

To identify and prioritize chemicals that may affect thyroid and adrenal/interregnal endocrine system and to reduce cost and animal use by conventional toxicity assay, an in vivo screening assay was developed using zebrafish embryos/larvae based on measurement of expression of genes that were suggested to play important roles in hypothalamic-pituitary-thyroid (HPT) and hypothalamic-pituitary-interrenal (HPI) axis. Model chemicals that could modulate HPT and HPI axis in adult fish were selected in assay validation, including anti-thyroid agent 6-Propyl-2-thiouracil (PTU) and cytochrome P450 11B (Cyp11b) enzyme inhibitor metyrapone (MET). Zebrafish embryos were exposed to different concentrations of model chemical from 4 h post-fertilization (hpf) to 5 d post-fertilization (dpf). Exposure to PTU increased mRNA expression of sodium iodide symporter (nis) and thyroglobulin (tg) involved in HPT axis, and MET treatment up-regulated all the mRNA expression tested involved in HPI axis by a compensatory mechanism. These results suggested that HPT and HPI axis were active upon chemical exposure at least at 5 dpf zebrafish. Furthermore, we studied the effects of PTU or MET on the cross-talk between HPT and HPI axis. The results demonstrated that PTU and MET could affect cross-talk responses in zebrafish embryos/larvae.     

Transcriptome analysis provides insights for understanding the adverse effects of endosulfan in Drosophila melanogaster

Indiscriminate use of agrochemicals worldwide, particularly, persistent organic pollutants (POPs), is of concern. Endosulfan, a POP, is used by various developing/developed nations and is known to adversely affect the development and the hormonal profiles of humans and animals. However, little is known about the molecular players/pathways underlying the adverse effects of endosulfan. We therefore analyzed the global gene expression changes and subsequent adverse effects of endosulfan using Drosophila. We used Drosophila melanogaster keeping in view of its well annotated genome and the wealth of genetic/molecular reagents available for this model organism. We exposed third instar larvae of D. melanogaster to endosulfan (2.0 μg mL⁻¹) for 24 h and using microarray, we identified differential expression of 256 genes in exposed organisms compared to controls. These genes are associated with cellular processes such as development, stress and immune response and metabolism. Microarray results were validated through quantitative PCR and biochemical assay on a subset of genes/proteins. Taking cues from microarray data, we analyzed the effect of endosulfan on development, emergence and survival of the organism. In exposed organisms, we observed deformities in hind-legs, reminiscent of those observed in higher organisms exposed to endosulfan. In addition, we observed delayed and/or reduced emergence in exposed organisms when compared to their respective controls. Together, our studies not only highlight the adverse effects of endosulfan on the organism but also provide an insight into the possible genetic perturbations underlying these effects, which might have potential implications to higher organisms.     

Hydrocarbon degradation, plant colonization and gene expression of alkane degradation genes by endophytic Enterobacter ludwigii strains

The genus Enterobacter comprises a range of beneficial plant-associated bacteria showing plant growth promotion. Enterobacter ludwigii belongs to the Enterobacter cloacae complex and has been reported to include human pathogens but also plant-associated strains with plant beneficial capacities. To assess the role of Enterobacter endophytes in hydrocarbon degradation, plant colonization, abundance and expression of CYP153 genes in different plant compartments, three plant species (Italian ryegrass, birdsfoot trefoil and alfalfa) were grown in sterile soil spiked with 1% diesel and inoculated with three endophytic E. ludwigii strains. Results showed that all strains were capable of hydrocarbon degradation and efficiently colonized the rhizosphere and plant interior. Two strains, ISI10-3 and BRI10-9, showed highest degradation rates of diesel fuel up to 68% and performed best in combination with Italian ryegrass and alfalfa. All strains expressed the CYP153 gene in all plant compartments, indicating an active role in degradation of diesel in association with plants.     

Expression of alkane monooxygenase (alkB) genes by plant-associated bacteria in the rhizosphere and endosphere of Italian ryegrass (Lolium multiflorum L.) grown in diesel contaminated soil

For phytoremediation of organic contaminants, plants have to host an efficiently degrading microflora. To assess the role of endophytes in alkane degradation, Italian ryegrass was grown in sterile soil with 0, 1 or 2% diesel and inoculated either with an alkane degrading bacterial strain originally derived from the rhizosphere of Italian ryegrass or with an endophyte. We studied plant colonization of these strains as well as the abundance and expression of alkane monooxygenase (alkB) genes in the rhizosphere, shoot and root interior. Results showed that the endophyte strain better colonized the plant, particularly the plant interior, and also showed higher expression of alkB genes suggesting a more efficient degradation of the pollutant. Furthermore, plants inoculated with the endophyte were better able to grow in the presence of diesel. The rhizosphere strain colonized primarily the rhizosphere and showed low alkB gene expression in the plant interior.     

基于模糊神经网络的机动车驾驶教练员职业适宜性研究

提出应用模糊神经网络系统,建构教练员职业适宜性的3个要素(即心理素质、驾驶技能和知识的表达阐述能力)的学习样本,分析"三要素"的8项特征参数指标——场依存性、速度估计能力、交通安全意识、简单反应、选择反应、跟踪能力、行车注意力、表达阐述能力等,使用K-均值法对实验样本进行初始分类,形成标准学习样本,使用该样本对所构建系统进行训练和调试。利用经调试训练后的系统,依据所测教练员的心理、心理参数对其职业适宜性进行评价。试验表明:建立的教练员职业适宜性仿真模型能取得很好的评价效果。     

甲基营养型酵母表达载体系统研究进展

酵母作为生产真核异源蛋白的宿主菌 ,具备了易于分子遗传操作 ,原核微生物的生长性质及真核蛋白的翻译后加工修饰等特点 .其中Ｓａｃｃｈａｒｏｍｙｃｅｓｃｅｒｅｖｉｓｉａｅ (酿酒酵母 )是第一个作为异源蛋白基因表达系统并加以应用的酵母 .人们认为Ｓ .ｃｅｒｅｖｉｓｉａｅ并非表达外源基因的最理想的酵母宿主菌 .在一群优于Ｓ .ｃｅｒｅｖｉｓｉａｅ的表达菌中 ,甲基营养型酵母Ｐｉｃｈｉａｐａｓｔｏｒｉｓ研究得最多 ,最具应用潜力 .在 2 0世纪 70年代期间 ,Ｐ .ｐａｓｔｏｒｉｓ是单细胞蛋白 (Ｓｉｎｇｌｅ ｃｅｌｌｐｒｏｔｅｉｎ…