Wetland macrophyte decomposition under different nutrient conditions: Relationships between decomposition rate, enzyme activities and microbial biomass

We used oligotrophic, P-limited herbaceous wetlands of northern Belize as a model system, on which to document and explain how changes in nutrient content along a salinity gradient affect activities of extracellular enzymes involved in macrophyte decomposition. To determine what is more important for decomposition, the initial litter quality, or site differences, we used reciprocal litter placement in a combined “site quality” and “litter quality” experiment running from August 2003 to April 2004. The experiment was set up in long-term control and nutrient addition plots (P, N, and NP) established in 2001 in 15 limestone-based inland marshes with a wide range of water conductivities (200-6000 μS) and a uniform pH (7.0-7.7) dominated by emergent macrophytes, Eleocharis spp. There were no differences among the plots in total sediment N and water NH₄-N, but total and KCl-extractable sediment P and water PO₄-P were significantly higher in P and NP plots throughout the duration of the experiment. The initial litter N content was slightly but significantly different between control and N plots versus P and NP plots (5.7 and 7.1 mg g⁻¹, respectively). The difference was much bigger for litter P content, 0.1 and 0.7 mg g⁻¹, respectively. Enzyme activities of alkaline phosphatase, leucine-aminopeptidase, arylsulfatase, and β-glucosidase were measured fluorometrically in Eleocharis litter in both the litterbag experiment and the naturally decomposing material. Total phospholipid fatty acid (PLFA) content in litter samples was used as a measure of microbial biomass present. Phosphatase always exhibited the highest activity of the enzymes studied, followed by leucine-aminopeptidase, arylsulfatase and β-glucosidase. There were no significant differences between enzyme activities from litterbags and the unconfined litter. Phosphatase activity was significantly suppressed in P-addition plots under all salinity levels while the activities of the remaining enzymes were significantly higher in P-enriched plots. There was a strong correlation between decomposition coefficient k-values and most of the enzymes as well as between the amount of PLFA and enzyme activities. PLFA, arylsulfatase, and litter C/P were the best predictors of k-values.     

miR-148a-3p靶向PPARγ基因抑制鹅颗粒细胞孕酮的合成

旨在探究miR-148a-3p是否通过靶基因PPARγ调节鹅卵泡颗粒细胞中类固醇激素的合成。本研究选用12只健康的处于产蛋高峰期的天府肉鹅母系母鹅分别用于组织样品采集和颗粒细胞培养。利用qPCR检测miR-148a-3p在鹅不同阶段卵泡颗粒层中的表达;通过MEGA 7软件分析miR-148a-3p在不同物种的保守性,预测miR-148a-3p的靶基因,采用双荧光素酶报告试验验证miR-148a-3p与靶基因的靶标关系;在鹅颗粒细胞中转染miR-148a-3p mimics和inhibitor,qPCR检测miR-148a-3p对靶基因及其下游基因表达水平的影响;同时利用ELISA技术检测激素含量的变化。结果表明,miR-148a-3p在不同物种中高度保守;在鹅卵泡发育过程中,miR-148a-3p的表达量随卵泡直径的增加呈先升高后下降的趋势。在鹅卵泡颗粒细胞体外培养过程中,miR-148a-3p mimics能显著下调3β-HSD的表达（P<0.05）,抑制孕酮的合成（P<0.05）;而miR-148a-3p inhibitor的作用则相反（P<0.05）。生物信息学分析和双荧光素酶报告试验证实,PPARγ为miR-148a-3p的靶基因。在颗粒细胞中,miR-148a-3p mimics能显著降低PPARγ的表达（P<0.05）,而miR-148a-3p inhibitor能显著上调PPARγ的表达（P<0.05）。当干扰PPARγ后,3β-HSD的表达量显著降低（P<0.05）,孕酮的含量也降低。这些结果表明,在鹅等级卵泡颗粒细胞中,miR-148a-3p可靶向结合PPARγ来抑制3β-HSD的表达,从而降低颗粒细胞孕酮的合成。     

β-内酰胺类药物对副猪嗜血杆菌流行病学临界值的建立及耐药性的测定

副猪嗜血杆菌病是副猪嗜血杆菌主要针对保育仔猪的一种全身多发性的传染病,对猪场的危害极大。β-内酰胺类抗生素是临床上使用比较多的一类抗生素,具有杀菌活性强、毒性低、适应症广及临床疗效好等优点,常用于治疗副猪嗜血杆菌病,因此有必要了解副猪嗜血杆菌对β-内酰胺类抗生素的耐药情况,从而更科学地指导临床用药和新药开发。本研究参考CLSI-VET和VETCAST中流行病学临界值的建立方法,汇总不同地区来源的菌株对14种β-内酰胺类药物的药敏结果,建立β-内酰胺类药物的流行病学临界值。结果显示,头孢克洛、头孢吡肟、头孢噻肟、头孢喹肟、头孢噻呋、头孢氨苄、阿莫西林-克拉维酸、阿莫西林、克拉维酸、氨苄西林、青霉素、苯唑西林、亚胺培南和美罗培南的流行病学折点值分别为16、0.5、0.125、0.031 25、0.5、32、0.25、1、0.5、1、2、8、0.25、0.062 5 μg·mL^-1,可以得出副猪嗜血杆菌对氨苄西林、阿莫西林以及头孢吡肟的耐药率较高,对亚胺培南和阿莫西林-克拉维酸的敏感性较高。在CLSI （Clinical and Laboratory Standards Institute）和EUCAST （The European Committee on Antimicrobial Susceptibility Testing）缺乏敏感性评判标准的情况下,本研究可以直观地识别非野生型菌株的出现,有利于耐药性监测工作的开展,对副猪嗜血杆菌的治疗和防控具有一定的参考价值。     

Molecular characterization of benzimidazole-resistant B. cinerea field isolates with reduced or enhanced sensitivity to zoxamide and diethofencarb

Sensitivity profiles of Botrytis cinerea field isolates to zoxamide and the molecular basis of the resistance mechanism involved in cross-resistance relationships between benzamides, benzimidazoles and N-phenylcarbamates were investigated. B. cinerea isolates collected from southern, central and northern Greece were characterized based on their sensitivity to zoxamide, the benzimidazole carbendazim and the N-phenylcarbamate diethofencarb. Isolates exhibiting baseline sensitivity to carbendazim and zoxamide but no sensitivity to diethofencarb were considered wild type (S phenotype) and accounted for 44% of the total strains sampled. Thirty-three percent of the isolates had increased sensitivity (HS phenotype) to zoxamide and diethofencarb and were highly resistant to carbendazim compared to S isolates. Eight percent of the sample was highly resistant (HR phenotype) to all anti-tubulin agents studied. The rest of the isolates were moderately resistant to zoxamide (MR phenotype) and equally sensitive to benzimidazoles and N-phenylcarbamates compared to isolates of the S phenotype. Fungitoxicity tests with botrycides belonging to other chemical classes revealed no cross-resistance relationships between zoxamide and the phenylpyrrole fludioxonil, the dicarboximide iprodione, the hydroxyanilide fenhexamid, the anilinopyrimidine cyprodinil, the carboxamide boscalid and the strobilurin-type fungicide pyraclostrobin. Study of fitness characteristics did not show any significant difference between zoxamide resistant and sensitive isolates with respect to the parameters tested. PCR-RFLP analysis of a part of the β-tubulin gene sequence detected mutations in position 198 for both HS and HR zoxamide-sensitivity phenotypes. DNA sequence analysis of the B. cinerea β-tubulin gene revealed two previously described benzimidazole-resistance-conferring mutations. The first one was the glutamic acid (GAG) to alanine (GCG) change at position 198 (E198A), which was identified in all HS isolates. The second mutation (E198K) was a GAG-to-AAG substitution resulting in the replacement of glutamic acid with lysine present in all B. cinerea isolates highly resistant to all three anti-tubulin classes of fungicides. A number of mutations in other positions of the β-tubulin gene were detected in the moderately zoxamide-resistance phenotype.     

产β-葡萄糖苷酶的菌种的筛选,鉴定及其酶学特性

β-葡萄糖苷酶来源广泛,几乎存在于所有的生物体中,而不同来源的β-葡萄糖苷酶其性质也各不同。本文利用七叶苷分离培养基从土样中分离筛选出产6种β-葡萄糖苷酶时间较快的菌种,其中发现菌种WGEA1酶活性较高,随后对菌种WGEA1进行初步的鉴定并且采用DNS法测该菌株所产粗酶液的酶学特性。酶学特性表明,WGEA1产的β-葡萄糖苷酶最适温度是在50～55℃之间,最适pH在6～7之间;在低于50℃条件下,pH为5～8时,酶活较稳定,同时在最适反应时间30min下,金属离子和有机溶剂都对酶活性影响很大,这些发现都为在非水相体系中酶法合成烷基糖苷奠定了一定的基础。     

β-胡萝卜素对绵羊瘤胃微生物发酵参数的影响

试验采用自身对照法试验设计,选用6只安装有永久性瘤胃瘘管的去势蒙古公羊,分3组,即0.000%β-胡萝卜素(对照组)、0.005%β-胡萝卜素和0.008%β-胡萝卜素组,研究在日粮中添加β-胡萝卜素对绵羊瘤胃内环境及微生物发酵参数的影响。结果表明:日粮中添加β-胡萝卜素对绵羊瘤胃中pH值没有影响;日粮中添加β-胡萝卜素有降低绵羊瘤胃中氨态氮含量的趋势;日粮中添加β-胡萝卜素使绵羊瘤胃中菌体蛋白含量降低,且存在随β-胡萝卜素添加比例的增加菌体蛋白含量有降低的趋势;日粮中添加β-胡萝卜素有促进瘤胃中TVFA含量升高和降低乙丙比的趋势。     

饲用葡甘露低聚糖的酶法制取

葡甘露低聚糖是一种重要的双歧因子,近年来已被作为一种功能性饲料添加剂,成为动物营养研究的新方向。试验通过对黑曲霉酸性β-甘露聚糖酶酶解魔芋粉制备甘露寡糖进行条件研究,获得了以葡甘露低聚糖为主的酶解产物。酶解条件为:用煮沸的pH值3.5的磷酸氢二钠-柠檬酸缓冲液配制180 g/l的魔芋粉溶液,加酶量按30 IU/g魔芋粉计算,酶解水浴温度为65℃,酶解时间为4 h。酶解结束后,将酶解液进行高温灭菌并接种酿酒酵母进行发酵,发酵温度为30℃,摇床转速200 r/min,发酵20 h后将发酵液离心,所得上清液即为不含单糖的葡甘露低聚糖,得率可以达到35.73%(以魔芋粉计)。     

奶山羊IL-1β在Sf9细胞中的表达

将CalL-1β基因整个编码区cDNA亚克隆到杆状病毒载体pFastBac-HTb中,构建转移载体pFastBac-HTb-CaIL-1β,转座到宿主菌DH10 Bac中,在Kan+/Gm+/Tet+/X-gal/IPTG LB平板上筛选白色菌落,碱裂解法提取质粒即为穿梭载体Baemid-CaIL-1β,转染Sf9细胞...     

The intestinal microbiome: A separate organ inside the body with the metabolic potential to influence the bioactivity of botanicals

For many years, it was believed that the main function of the large intestine was the resorption of water and salt and the facilitated disposal of waste materials. However, this task definition was far from complete, as it did not consider the activity of the microbial content of the large intestine. Nowadays it is clear that the complex microbial ecosystem in our intestines should be considered as a separate organ within the body, with a metabolic capacity which exceeds the liver with a factor 100. The intestinal microbiome is therefore closely involved in the first-pass metabolism of dietary compounds. This is especially true for botanical supplements, which are now marketed for various health applications. Being of natural origin, their structural building blocks, such as polyphenols, are often highly recognized by the human and especially the intestinal microbial metabolism machinery. Intensive metabolism results in often low circulating levels of the original products, with the consequence that final health effects of botanicals are often related to specific active metabolites which are produced in the body rather than being related to the product's original composition. Understanding how such metabolic processes contribute to the in situ exposure is therefore crucial for the proper interpretation of biological responses. A multidisciplinary approach, characterizing the food and phytochemical intake as well as the metabolic potency of the gut microbiota, while measuring biomarkers of both exposure and response in target tissues, is therefore of critical importance. With polyphenol metabolism as example, this review describes how the incorporation of microbial metabolism as an important variable in the evaluation of the final bioactivity of botanicals strongly increases the relevance and predictive value of the outcome. Moreover, knowledge about intestinal processes may offer innovative strategies for targeted product development.     

Flavonoid glycosides from Microtea debilis and their cytotoxic and anti-inflammatory effects

Two new 5-O-glucosylflavones, 5-O-β-d-glucopyranosyl cirsimaritin (1) and 5, 4′-O-β-d-diglucopyranosyl cirsimaritin (2), four known flavonoids, cirsimarin (3), cirsimaritin (4), salvigenin (5), 4′, 5-dihydroxy-7-methoxyflavone (6), and a norisoprenoid, vomifoliol (7), have been isolated from the aerial parts of Microtea debilis. All isolates were tested for cytotoxicity in human cancer cell lines (Hep G2, COLO 205, and HL-60) and anti-inflammatory activities in LPS-treated RAW264.7 macrophages. Compound 6 was found to be a potent inhibitor to nitrite production in macrophages. Compounds 2, 4, 6, and 7 showed moderate anti-proliferative activity against COLO-205 cells with IC₅₀ values of 7.1, 13.1, 6.1, and 6.8 μM, respectively.