微生物专题英文文献共24页文档
食品微生物外文文献2017.1.19
食品微生物外文文献2017.1.19脉冲强光对诺如病毒和沙门及o157的灭活作用Pulsed light (PL) inactivation of two human norovirus (HuNoV) surrogates, murine norovirus (MNV-1) and Tulane virus (TV), and two bacterial pathogens, Escherichia coli O157:H7 and Salmonella, were evaluated. The viruses and bacteria were suspended in phosphate buffered saline (PBS) to final populations of ～6 log PFU/mL and ～6 log CFU/mL, respectively. Both viral and bacterial suspensions were then irradiated by PL for different durations and the reductions of each microorganisms were determined. MNV-1 and TV were significantly (P < 0.05) more resistant to PL treatment than Salmonella and E. coli O157:H7 in PBS suspension. MNV-1, Salmonella and E. coli O157:H7 were also inoculated on strawberries and blueberries and the PL inactivation of each microorganism was determined. Lower inactivation of each microorganism was achieved on berry surfaces than in PBS suspension. This study shows that PL can induce rapid inactivation of MNV-1, TV, Salmonella and E. coli O157:H7 in clear suspension with viruses more resistant to PL treatment than bacteria. The efficacy of PL treatment is substantially influenced by food surface structure.乳酸菌基因组学和代谢组学的研究进展The Lactobacillus genus represents the largest and most diverse genera of all the lactic acid bacteria (LAB), encompassing species with applications in industrial, biotechnological and medical fields. The increasing number of available Lactobacillus genome sequences has allowed understanding of genetic and metabolic potential of this LAB group. Pangenome and coregenome studies are available for numerous species, demonstrating the plasticity of the Lactobacillus genomes and providing the evidence of niche adaptability. Advancements in the application of lactobacilli in the dairy industry lie in exploring the genetic background of their commercially important characteristics, such as flavour development potential or resistance to the phage attack. The integration of available genomic and metabolomic data through the generation of genome scale metabolic models has enabled the development of computational models that predict the behaviour of organisms under specific conditions and present a route to metabolic engineering. Lactobacilli are recognised as potential cell factories, confirmed by the successful production of many compounds. In this review, we discuss the current knowledge of genomics, metabolomics and metabolic engineering of the prevalent Lactobacillus species associated with the production of fermented dairy foods. In-depth understanding of their characteristics opens the possibilities for their future knowledge-based applications.牛肉冷冻状态下的微生物The primary objective of this study was to characterise (microbiology and physical parameters) beef carcasses and primals during chilled storage. A minor aim was to compare observed growth of key spoilage bacteria on carcasses with that predicted by ComBase and the Food Safety Spoilage Predictor (FSSP). Total viable count (TVC), total Enterobacteriacae count (TEC), Pseudomonas spp., lactic acid bacteria (LAB), Brochothrixthermosphacta and Clostridium spp. were monitored on beef carcasses (n = 30) and primals (n = 105) during chilled storage using EC Decision 2001/471/EC and ISOsampling/laboratory procedures. The surface and/or core temperature, pH and water activity (aw) were also recorded. Clostridium (1.89 log10 cfu/cm2) and Pseudomonas spp. (2.12 log10 cfu/cm2) were initially the most prevalent bacteria on carcasses and primals, respectively. The shortest mean generation time (G) was observed on carcasses with Br. thermosphacta (20.3 h) and on primals with LAB (G = 68.8 h) and Clostridium spp. (G = 67 h). Over the course of the experiment the surface temperature dec reased from 37 °Cto 0 °C, pH from 7.07 to 5.65 and aw from 0.97 to 0.93 The observed Pseudomonas spp. and Br. thermosphacta growth was more or less within the range of predictions of Combase. In contrast, the FSSP completely overestimated the growth of LAB. This study contributes to the very limited microbiological data on beef carcasses and primals during chilling.冰箱中牡蛎气调包装的微生物组分变化As filter-feeding bivalves, oysters can accumulate microorganisms into their gills, causing spoilage and potential safety issues. This study aims to investigate the changes in the gill microbiota of oysters packed under air and modified atmospheres (MAs, 50% CO2: 50% N2, 70% CO2: 30% O2, and 50% CO2: 50% O2) during storage at 4 °C. The diversity of bacterial microbiota in oyster gills was profiled through polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) analysis on the 16S rRNA gene V3 region to describe the variation during the entire storage period. The DGGE profile revealed high bacterial diversity in the air- and MA-packaged oyster gills, and the spoilage bacterial microbiota varied in the MA-packaged oyster gills. Results indicated that CO2:O2 (70%:30%) was suitable for oyster MA packaging andthat high bacterial loads in oyster gills need to be considered during storage. In addition, Lactobacillus and Lactococcus species were found to grow dominantly in fresh oyster gills under MA packaging, which supports the potential application of MA packaging for oyster storage.干腌制猪腰猪腿的猪链球菌的生存Dry-cured hams, shoulders and loins of Iberian pigs are highly appreciated in national and international markets. Salting, additive addition and dehydration are the main strategies to produce these ready-to-eat products. Although the dry curing process is known to reduce the load of well-known food borne pathogens, studies evaluating the viability of other microorganisms in contaminated pork have not been performed. In this work, the efficacy of the dry curing process to eliminate three swine pathogens associated with pork carcass condemnation, Streptococcus suis, Streptococcus dysgalactiae and Trueperellapyogenes, was evaluated. Results of this study highlight that the dry curing process is a suitable method to obtain safe ready-to-eat products free of these microorganisms. Although salting of dry-cured shoulders had a moderate bactericidal effect, results of this study suggest that drying and ripening were the most important stages to obtain dry-cured products free of these microorganisms.使用噬菌体提高贝壳类中大肠杆菌的去除率The present study investigated the potential application of the bacteriophage (or phage) phT4A, ECA2 and the phage cocktail phT4A/ECA2 to decrease the concentration of Escherichia coli during the depuration of natural and artificially contaminated cockles. Depuration in static seawater at multiplicity of infection (MOI) of 1 with single phage suspensionsof phT4A and ECA2 was the best condition, as it decreased by ～2.0 log CFU/g the concentration of E. coli in artificially contaminated cockles after a 4 h of treatment. When naturally contaminated cockles were treated in static seawater with single phage suspensions and the phage cocktail, similar decreases in the concentration of E. coli (～0.7 log CFU/g) were achieved. However, when employing the phage cocktail, a longer treatment time was required to obtain comparable results to those achieved when using single phage suspensions. When naturally contaminated cockles were depurated with phage phT4A in a recirculated seawater system (mimicking industrial depuration conditions), a 0.6 log CFU/g reduction of E. coli was achieved after a 2 h of treatment. When the depuration process was performed without phage addition, a 4 h treatment was necessary toobtain a similar decrease. By combining phage therapy and depuration procedures, a reduction in bivalves depuration period can be achieved for, thus decreasing the cost associated with this procedure and even enhance the quality and safety of depurated bivalves destined for human consumption.乳制品中诺如病毒的三种提取方法比较Noroviruses (NoV) are currently the most common cause of viral foodborne diseases and RT-qPCR is widely used for their detection in food because of its sensitivity, specificity and rapidity. The ISO/TS (15216-1, 15216-2) procedures for detecting NoV and HAV in high-risk food categories such as shellfish, bottled water and vegetables were published in 2013. Milk products are less implicated in foodborne viral outbreaks but they can be contaminated with fruit added to these products or by the food handler. Thus, the development of sensitive and reliabletechniques for the detection of NoV in dairy products is needed to ensure the safety of these products. The aim of this study was to develop a RT-qPCR based method for the detection of NoV in milk products. Three methods were tested to recover NoV from artificially contaminated milk and cottage cheese. The selected method was based on the use of proteinase K and the recovery efficiencies ranged from 54.87% to 98.87% for NoV GI, 61.16%–96.50% for NoV GII. Murine norovirus and mengovirus were used as process controls and their recovery efficiencies were respectively 60.59% and 79.23%. The described method could be applied for detecting NoV in milk products for routine diagnosis needs.溶解氧和培养基对空肠弯曲菌形成生物膜的影响Campylobacter jejuni survival in aerobic environments has been suggested to be mediated by biofilm formation. Biofilm formation by eight C. jejuni strains under both aerobic and microaerobic conditions in different broths (Mueller-Hinton (MH), Bolton and Brucella) was quantified. The dissolved oxygen (DO) content of the broths under both incubation atmospheres was determined. Biofilm formation for all strains was highest in MH broth under both incubation atmospheres. Four strains had lower biofilm formation in MH under aerobic as compared to microaerobic incubation, while biofilm formation by the other four strains did not differ under the 2 atm. Two strains had higher biofilm formation under aerobic as compared to microaerobic atmospheres in Bolton broth. Biofilm formation by all other strains in Bolton, and all strains in Brucella broth, did not differ under the 2 atm. Under aerobic incubation DO levels in MH >Brucella> Bolton broth. Under microaerobic conditions levels in MH = Brucella> Bolton broth. Levels of DO in MH andBrucella broth were lower under microaerobic conditions but those of Bolton did not differ under the 2 atm. Experimental conditions and especially the DO of broth media confound previous conclusions drawn about aerobic biofilm formation by C. jejuni. 热-紫外处理对液体食品的灭菌效果优化The combination of ultraviolet radiation and heat (UV-H treatment) has been demonstrated as a promising strategy to overcome the limited UV germicidal effect in fruit juices. Nonetheless, there are so far no data regarding the efficacy of the combined process for the inactivation of bacterial foodborne pathogens in other liquid foods with different pH and composition. In this investigation, the optimum UV-H processing conditions for the inactivation of Escherichia coli, Salmonella Typhimurium, Listeria monocytogenes, and S. aureus in chicken and vegetable broth, in addition to juices, were determined. From these data models that accurately predict the most advantageous UV-H treatment temperature and the expected synergistic lethal effect from UV and heat resistance data separately were constructed. Equations demonstrated that the optimumUV-H treatment temperature mostly depended on heat resistance, whereas the maximum synergistic lethal effect also was affected by the UV resistance of the microorganism of concern in a particular food.肠炎沙门鞭毛对侵袭的作用Nontyphoidal Salmonella strains are the main source of pathogenic bacterial contamination in the poultry industry. Recently, Salmonella entericaserovar Kentucky has been recognized as the most prominent serovar on carcasses in poultry-processing plants. Previous studies showed that flagellaare one of the main factors that contribute to bacterial attachment to broiler skin. However, the precise role of flagella and the mechanism of attachment are unknown. There are two different flagellar subunits (fliC and fljB) expressed alternatively in Salmonella entericaserovars using phase variation. Here, by making deletions in genes encoding flagellar structural subunits (flgK, fliC, and fljB), and flagellar motor (motA), we were able to differentiate the role of flagella and their rotary motion in the colonization of broiler skin and cellular attachment. Utilizing a broiler skin assay, we demonstrated that the presence of FliC is necessary for attachment to broiler skin. Expression of the alternative flagellar subunit FljB enables Salmonella motility, but this subunit is unable to mediate tight attachment. Deletion of the flgK gene prevents proper flagellar assembly, making Salmonella significantly less adherent to broiler skin than the wild type. S. Kentucky with deletions in all three structural genes, fliC, fljB, and flgK, as well as a flagellar motor mutant (motA), exhibited less adhesion and invasion of Caco-2 cells, while an fljB mutant was as adherent and invasive as the wild-type strain.IMPORTANCE In this work, we answered clearly the role of flagella in S. Kentucky attachment to the chicken skin and Caco-2 cells. We demonstrated that the presence of FliC is necessary for attachment to broiler skin. Expression of the alternative flagellar subunit FljB enables Salmonella motility, but this subunit is unable to mediate strong attachment. Deletion of the flgK gene prevents proper flagellar assembly, making Salmonella significantly less adherent to broiler skin than the wild type. S. Kentucky with deletions in all three structural genes, fliC, fljB, and flgK, as well as a flagellar motor mutant (motA), exhibited less adhesion and invasion of Caco-2 cells, while an fljB mutant wasas adherent and invasive as the wild-type strain. We expect these results will contribute to the understanding of the mechanisms of Salmonella attachment to food products.单增李斯特菌可能是通过胃液和体外细胞侵染的主要细菌Various Listeria monocytogenes strains may contaminate a single food product, potentially resulting in simultaneous exposure of consumers to multiple strains. However, due to bias in strain recovery, L. monocytogenes strains isolated from foods by selective enrichment (SE) might not always represent those that can better survive the immune system of a patient. We investigated the effect of cocultivation in tryptic soy broth with 0.6% yeast extract (TSB-Y) at 10°C for 8 days on (i) the detection of L. monocytogenes strains during SE with the ISO 11290-1:1996/Amd 1:2004 protocol and (ii) the in vitro virulence of strains toward the Caco-2 human colon epithelial cancer cell line following exposure to simulated gastric fluid (SGF; pH 2.0)-HCl (37°C). We determined whether the strains which were favored by SE would be effective competitors under the conditions of challenges related to gastrointestinal passage of the pathogen. Interstrain competition of L. monocytogenes in TSB-Y determined the relative population of each strain at the beginning of SE. This in turn impacted the outcome of SE (i.e., favoring survival of competitors with better fitness) and the levels exposed subsequently to SGF.However, strong growth competitors could be outcompeted after SGF exposure and infection of Caco-2 cells by strains outgrown in TSB-Y and underdetected (or even missed) during enrichment. Our data demonstrate a preferential selection of certain L. monocytogenes strains during enrichments, often not reflecting a selective advantage of strains during infection. Thesefindings highlight a noteworthy scenario associated with the difficulty of matching the source of infection (food) with the L. monocytogenes isolate appearing to be the causative agent during listeriosis outbreak investigations.系统模型的建立来描述动物粪便是果蔬感染致病菌的主要途径The majority of foodborne outbreaks in the United States associated with the consumption of leafy greens contaminated with Escherichia coli O157:H7 have been reported during the period of July to November. A dynamic system model consisting of subsystems and inputs to the system (soil, irrigation, cattle, wild pig, and rainfall) simulating a hypothetical farm was developed. The model assumed two crops of lettuce in a year and simulated planting, irrigation, harvesting, ground preparation for the new crop, contamination of soil and plants, and survival of E. coli O157:H7. As predicted by the baseline model for crops harvested in different months from conventional fields, an estimated 13 out of 257 (5.05%) first crops harvested in July would have at least one plant with at least 1 CFU of E. coli O157:H7. Predictions indicate that no first crops would be contaminated with at least 1 CFU of E. coli O157:H7 for other months (April to June). The maximum E. coli O157:H7 concentration in a plant was higher in the second crop (27.10 CFU) than in the first crop (9.82 CFU). For the second crop, the probabilities of having at least one plant with at least 1 CFU of E. coli O157:H7 in a crop were predicted as 15/228 (6.6%), 5/333 (1.5%), 14/324 (4.3%), and 6/115 (5.2%) in August, September, October, and November, respectively. For organic fields, the probabilities of having at least one plant with ≥1 CFU of E. coli O157:H7 in a crop (3.45%) were predicted to be higher than those for the conventional fields (2.15%).O157迫于环境压力的嗜热和苏醒加速进化The development of resistance in foodborne pathogens to food preservation techniques is an issue of increasing concern, especially in minimally processed foods where safety relies on hurdle technology. In this context, mild heat can be used in combination with so-called nonthermal processes, such as high hydrostatic pressure (HHP), at lower individual intensities to better retain the quality of the food. However, mild stresses may increase the risk of (cross-)resistance development in the surviving population, which in turn might compromise food safety. In this investigation, we examined the evolution of Escherichia coli O157:H7 strain ATCC 43888 after recurrent exposure to progressively intensifying mild heat shocks (from 54.0°C to 60.0°C in 0.5°C increments) with intermittent resuscitation and growth of survivors. As such, mutant strains were obtained after 10 cycles of selection with ca. 106-fold higher heat resistance than that for the parental strain at 58.0°C, although this resistance did not extend to temperatures exceeding 60.0°C. Moreover, th ese mutant strains typically displayed cross-resistance against HHP shock and displayed signs of enhanced RpoS and RpoH activity. Interestingly, additional cycles of selection maintaining the intensity of the heat shock constant (58.5°C) selected for mutan t strains in which resuscitation speed, rather than resistance, appeared to be increased. Therefore, it seems that resistance and resuscitation speed are rapidly evolvable traits in E. coli ATCC 43888 that can compromise food safety.肉桂油抑制大肠0157噬菌体的入侵和增殖This study evaluated the inhibitory effect of cinnamon oil against Escherichia coli O157:H7 Shiga toxin (Stx) production andfurther explored the underlying mechanisms. The MIC and minimum bactericidal concentration (MBC) of cinnamon oil against E. coli O157:H7 were 0.025% and 0.05% (vol/vol), respectively. Cinnamon oil significantly reduced Stx2 production and the stx2 mRNA expression that is associated with diminished Vero cell cytotoxicity. Consistently, induction of the Stx-converting phage where the stx2 gene is located, along with the total number of phages, decreased proportionally to cinnamon oil concentration. In line with decreased Stx2 phage induction, cinnamon oil at 0.75× and 1.0× MIC eliminated RecA, a key mediator of SOS response, polynucleotide phosphorylase (PNPase), and poly(A) polymerase (PAP I), which positively regulate Stx-converting phages, contributing to reduced Stx-converting phage induction and Stx production. Furthermore, cinnamon oil at 0.75× and 1.0× MIC strongly inhibited the qse BC and luxS expression associated with decreased AI-2 production, a universal quorum sensing signaling molecule. However, the expression of oxidative stress response genes oxyR, soxR, and rpoS was increased in response to cinnamon oil at 0.25× or 0.5× MIC, which may contribute to stunted bacterial growth and reduced Stx2 phage induction and Stx2 production due to the inhibitory effect of OxyR on prophage activation. Collectively, cinnamon oil inhibits Stx2 production and Stx2 phage induction in E. coli O157:H7 in multiple ways.长时间接触导致增加了产气肠杆菌的食品感染Bacterial cross-contamination from surfaces to food can contribute to foodborne disease. The cross-contamination rate of Enterobacteraerogenes on household surfaces was evaluated by using scenarios that differed by surface type, food type, contact time (<1, 5, 30, and 300 s), and inoculum matrix (trypticsoy broth or peptone buffer). The surfaces used were stainless steel, tile, wood, and carpet. The food types were watermelon, bread, bread with butter, and gummy candy. Surfaces (25 cm2) were spot inoculated with 1 ml of inoculum and allowed to dry for 5 h, yielding an approximate concentration of 107 CFU/surface. Foods (with a 16-cm2 contact area) were dropped onto the surfaces from a height of 12.5 cm and left to rest as appropriate. Posttransfer, surfaces and foods were placed in sterile filter bags and homogenized or massaged, diluted, and plated on tryptic soy agar. The transfer rate was quantified as the log percent transfer from the surface to the food. Contact time, food, and surface type all had highly significant effects (P < 0.000001) on the log percent transfer of bacteria. The inoculum matrix (tryptic soy broth or peptone buffer) also had a significant effect on transfer (P = 0.013), and most interaction terms were significant. More bacteria transferred to watermelon (～0.2 to 97%) than to any other food, while the least bacteria transferred to gummy candy (～0.1 to 62%). Transfer of bacteria to bread (～0.02 to 94%) was similar to transfer of bacteria to bread with butter (～0.02 to 82%), and these transfer rates under a given set of conditions were more variable than with watermelon and gummy candy.。
微生物专题英文文献
班级：生物工程 学生：马春玲 2013年12月13日
LOGO
试验内容
1. Purpose and meaning 2. Introduction 3. Materials and methods
3、Materials and methods
供试菌种 黑曲霉A3（褐黑色）
培养基 孢子培养基：马铃薯、葡萄糖、琼脂、去离子水 菌种培养基：PDA培养基 摇瓶发酵培养基：玉米穗、麸皮、（NH4）2SO4、NaNO3 分离培养基：蛋白胨、琼脂、麸皮
矿质元素、生长因子：KH2PO4、MgSO4、FeSO4、CoCl2
通过对出发菌株注入不同剂量的氮离子，低能氮离子 束对菌体细胞均有一定程度的致死和损伤作用，细胞及其 损伤DNA又在其修复系统的作用下得到不同程度的修复， 从而导致黑曲霉孢子的存活率先下降，后上升，然后又下 降，并且菌种的修复出错会使其突变率大大提高，从而提 高了菌株的正突变率，从而确定了氮离子最佳注入参数。 以上试验可以得出最优培养基的组成（即各组分的最 适浓度），而且在以上培养得到了黑曲霉N212（表2），当 它发酵60个小时后酶活达到600IU/ml，比之前未优化的菌 株减少了12个小时，而且相对于原出发菌株酶活增加了100 ％。 试验证明离子注入对微生物进行诱变改良是一种行之 有效的诱变技术。
4.6 Analysis 根据表5中的极差值，四个因素对木聚糖酶的影 响强度顺序是A>B>C>D, 根据表2,黑曲霉发酵产木聚 糖酶其碳源玉米芯的浓度对产酶量有着直接的影响. 在合适的玉米芯浓度范围内，随着浓度的升高酶活 也相应升高，与此同时发酵液的黏度升高，溶氧性 能降低，因此产酶能力也降低。
微生物英文文献及翻译—翻译
A/O法活性污泥中氨氧化菌群落的动态与分布摘要：我们研究了在厌氧—好氧序批式反应器（SBR）中氨氧化菌群落（AOB）和亚硝酸盐氧化菌群落（NOB）的结构活性和分布。
在研究过程中，分子生物技术和微型技术被用于识别和鉴定这些微生物。
污泥微粒中的氨氧化菌群落结构大体上与初始的接种污泥中的结构不同。
与颗粒形成一起，由于过程条件中生物选择的压力，AOB的多样性下降了。
DGGE测序表明，亚硝化菌依然存在，这是因为它们能迅速的适应固定以对抗洗涤行为。
DGGE更进一步的分析揭露了较大的微粒对更多的AOB种类在反应器中的生存有好处。
在SBR反应器中有很多大小不一的微粒共存，颗粒的直径影响这AOB和NOB的分布。
中小微粒（直径<0.6mm）不能限制氧在所有污泥空间的传输。
大颗粒（直径>0.9mm）可以使含氧量降低从而限制NOB的生长。
所有这些研究提供了未来对AOB微粒系统机制可能性研究的支持。
关键词：氨氧化菌（AOB），污泥微粒，菌落发展，微粒大小，硝化菌分布，发育多样性1.简介在浓度足够高的条件下，氨在水环境中对水生生物有毒，并且对富营养化有贡献。
因此，废水中氨的生物降解和去除是废水处理工程的基本功能。
硝化反应，将氨通过硝化转化为硝酸盐，是去除氨的一个重要途径。
这是分两步组成的，由氨氧化和亚硝酸盐氧化细菌完成。
好氧氨氧化一般是第一步，硝化反应的限制步骤：然而，这是废水中氨去除的本质。
对16S rRNA的对比分析显示，大多数活性污泥里的氨氧化菌系统的跟ß-变形菌有关联。
然而，一系列的研究表明，在氨氧化菌的不同代和不同系有生理和生态区别，而且环境因素例如处理常量，溶解氧，盐度，pH，自由氨例子浓度会影响氨氧化菌的种类。
因此，废水处理中氨氧化菌的生理活动和平衡对废水处理系统的设计和运行是至关重要的。
由于这个原因，对氨氧化菌生态和微生物学更深一层的了解对加强处理效果是必须的。
当今，有几个进阶技术在废水生物处理系统中被用作鉴别、刻画微生物种类的有价值的工具。
微生物英文文献及翻译—原文
微生物英文文献及翻译—原文本期为微生物学的第二讲，主要讨论炭疽和蛔虫病这两种既往常见而当今社会较为罕见的疾病。
炭疽是由炭疽杆菌所致的一种人畜共患的急性传染病。
人因接触病畜及其产品及食用病畜的肉类而发生感染。
临床上主要表现为皮肤坏死、溃疡、焦痂和周围组织广泛水肿及毒血症症状；似蚓蛔线虫简称蛔虫，是人体内最常见的寄生虫之一。
成虫寄生于小肠，可引起蛔虫病。
其幼虫能在人体内移行，引起内脏幼虫移行症。
案例分析Case 1：A local craftsman who makes garments from the hides of goats visits his physician because over the past few days he has developed several black lesions on his hands and arms. The lesions are not painful, but he is alarmed by their appearance. He is afebrile and his physical examination is unremarkable.案例1：一名使用鹿皮做皮衣的当地木匠来就医，主诉过去几天中手掌和手臂上出现几个黑色皮肤损害。
皮损无痛，但是外观较为骇人。
患者无发热，体检无异常发现。
1. What is the most likely diagnosis?Cutaneous anthrax, caused by Bacillus anthracis. The skin lesions are painless and dark or charred ulcerations known as black eschar. It is classically transmitted by contact with thehide of a goat at the site of a minor open wound.皮肤炭疽：由炭疽杆菌引起，皮损通常无痛、黑色或称为焦痂样溃疡。
Microbiology微生物学分类相关中英文对照
Microbiology微生物学分类相关中英文对照Microbiology 微生物学分类相关中英文对照微生物学microbiology病毒学virology噬菌体学bacteriophagology细菌学bacteriology鉴定细菌学determinative bacteriology系统细菌学systematic bacteriology真菌学mycology原生生物学protistology原生动物学protozoology普通微生物学general microbilogy微生物分类学microbial taxonomy微生物生理学microbial physiology微生物生物化学microbial biochemistry 微生物遗传学microbial genetics微生物生态学microbial ecology古微生物学paleomicrobiology土壤微生物学soil microbiology水生微生物学aquatic microbiology海洋微生物学marine microbiology悉生生物学gnotobiology医学微生物学medical microbiology兽医微生物学veterinary microbiology农业微生物学agricultural microbiology工业微生物学industrial microbiology石油微生物学petroleum microbiology食品微生物学food microbiology乳品微生物学diary microbiology瘤胃微生物学rumen microbiology诊断微生物学diagnostic microbiology病原学etiology国际微生物学会联合会International Union of Microbiological Societies, IUMS中国微生物学会Chinese Society for Microbiology, CSM世界培养物保藏协会World Federation for Culture Collection, WFCC中国微生物菌种保藏管理委员会China Committee for Culture Collection of Microorganisms,CCCCM美国模式培养物保藏所American Type Culture Collection, ATCC 自然发生说，无生源说spontaneous generation, abiogenesis 原界urkingdom始祖生物progenote古始生物界archetista古细菌archaebacteria原生生物protista原生动物protozoan原生植物protophyte真核生物eukaryote原核生物prokaryote裂殖植物schizophyte微生物microorganism数值分类法numerical taxonomy模式目type order模式科type family模式属type genus模式种type species模式株type strain真菌fungi捕食真菌predacious fungi虫道真菌ambrosia fungi地下真菌hypogeal fungi虫生真菌entomogenous fungi 菌根真菌mycorrhizal fungi 木腐菌wood-decay fungi霉菌mold, mould半知菌imperfect fungi子囊菌ascomycetes粘菌slime mold, slime mould 壶菌chytrid卵菌oomycetes接合菌zygomycetes担子菌basidiomycetes核菌pyrenomycetes盘菌cup fungi块菌truffles锈菌rust fungi蘑菇mushrooms毒蘑菇poisonous mushroom酵母菌yeast无孢子酵母菌asporogenous yeasts 有孢子酵母菌sporogenous yeasts 黑粉菌smut fungi双态性真菌dimorphic fungi毛外癣菌ectothrix毛内癣菌endothrix完全真菌perfect fungi黑粉病smut disease锈病rust disease菌丝hypha菌髓trama假菌丝体pseudomycelium气生菌丝体aerial mycelium基内菌丝体substrate mycelium球拍状菌丝体racquet mycelium结节状菌丝nodular mycelium梳状菌丝pectinafe mycelium螺旋菌丝spiral mycelium匍匐菌丝stolon次生菌丝体secondary mycelium有隔菌丝septate hypha无隔菌丝nonseptate hypha生殖菌丝体reproductive mycelium 营养菌丝体vegetative mycelium不育菌丝体sterile mycelium菌丝体mycelium黄癣菌丝favic chandelier mycelium 产囊丝ascogenous hypha 产囊体ascogonium原植体thallus粘菌体aethalium合胞体syncytium虫菌体hyphal body盾状体clypeus子实体fruiting body产孢体gleba子实层体hymenophore 子实层hymenium子实下层subhymenium 菌丝层subiculum菌丝段hyphal fragment 菌丝束coremium菌丝索funiculus菌核sclerotium器菌核pycnosclerotium 菌环annulus菌裙indusium菌盖pileus顶体apicle藏卵器oogonium雄器antheridium[锈菌]性孢子器pycnium锈子器aecium精子器spermogonium囊状体cystidium粉孢子梗oidiophore小梗sterigma接合孢子柄zygosporophore 孢囊柄sporangiophore 配囊柄suspensor孢子梗sporophore分生孢子梗conidiophore雄器柄androphore帚状枝penicillus瓶梗phialide梗基metulae芽孔germ pore芽管germ tube芽缝germ slit孢丝capillitium周丝periphysis类周丝periphysoid侧丝paraphysis拟侧丝pseudoparaphysis类侧丝paraphysoid[孢子]外壁exosporium外生菌根ectomycorrhiza内生菌根endomycorrhiza内外生菌根ectendomycorrhiza泡囊丛枝菌根vesicular-arbuscular mycorrhiza刺突spike弹丝elater刚毛seta微体microbody泡囊vesicle隔膜septum假隔膜pseudoseptum分生孢子盘acervulus分生孢子座sporodochium 精子团spermatium囊基膜hypothallus囊层基hypothecium囊层被epithecium囊间丝hamathecium囊托apophysis囊领collarette囊轴columella孔口ostiole菌托volva孢子角cirrus孢子球spore ball孢子印spore print聚簇cluster[菌丝]融合anastomosis [孢子]切落abjunction [孢子]缢断abstriction多态[现象] polymorphism 缢缩[作用] constriction 粉孢子oidium孢子spore厚壁孢子chlamydospore 环痕孢子annellospore节孢子arthrospore卷旋孢子helicospore腊肠形孢子allantospore孔出孢子porospore星形孢子staurospore线形孢子scolecospore砖格孢子dictyospore侧生孢子aleuriospore芽生孢子blastospore瓶梗孢子phialospore无梗孢子thallospore分生孢子conidium大分生孢子macroconidium 小分生孢子microconidium 节分生孢子arthroconidium 芽分生孢子blastoconidium 器孢子pycnidiospore无隔孢子amerospore双胞孢子didymospore多隔孢子phragmospore休眠孢子hypnospore顶生孢子acrospore顶生厚壁孢子fuseau内分生孢子endoconidium担孢子basidiospore双孢担孢子dispore同形孢子isospore柄生孢子stylospore[锈菌]性孢子pycniospore产雄器孢子androspore夏孢子urediniospore, aeciospore 冬孢子teliospore四分孢子tetraspore粘孢子myxospore多核孢子coenospore孢囊孢子sporangiospore子囊孢子ascospore多核细胞coenocyte分生孢子果conidiocarp分生孢子器pycnidium孢[子]囊sporangium柱孢子囊merosporangium四分孢子囊tetrasporangium原孢子囊prosporangium多核孢子囊coenosporangium 休眠孢子囊hypnosporangium 子囊ascus接合孢子zygospore拟接合孢子azygospore原囊壁子囊prototunicate ascus 单囊壁子囊unitunicate ascus 双囊壁子囊bitunicate ascus子囊果ascocarp子囊壳perithecium闭囊壳cleistothecium闭囊果cleistocarp盘状子囊果discocarp孢囊果sporangiocarp [接]合子zygote单性合子azygote多核合子coenozygote异形合子heterozygote合子核zygotonucleus游动合子planozygote担子basidium半担子hemibasidium隔担子heterobasidium无隔担子holobasidium有隔担子phragmobasidium 内生担子endobasidium原担子protobasidium上担子epibasidium下担子hypobasidium同担子homobasidium担子果basidiocarp担子体basidiophore配子gamete原配子progamete雄配子androgamete雄核发育androgenesis同形配子isogamete异形配子heterogamete游动配子zoogamete多核配子coenogamete配子囊gametangium配子母细胞gametocyte同形配子囊isogametangium 原配子囊progametangium 小孢子囊sporangiole微包囊microcyst足细胞foot cell脚胞foot cell固着器holdfast附着枝hyphopodium吸盘sucker锁状细胞clamp cell锁状联合clamp connection 偶核细胞zeugite卵球oosphere卵质ooplasm孢原质sporoplasm卵配子oogamete卵孢子oospore球状胞sphaerocyst子囊腔locule子囊盘apothecium子囊座ascostroma缝裂壳hysterothecium下子座hypostroma包被peridium子座stroma壳心centrum拟包被pseudoperidium无融合生殖apomixis同宗配合homothallism准性生殖parasexuality异宗配合heterothallism同配生殖isogamy异配生殖heterogamy无配生殖apogamy配囊交配gametangial copulation 交配型mating type 全型holomorph夏孢子期uredostage冬孢子堆teleutosorus, telium 夏孢子堆uredinium子囊孢子形成ascosporulation 孢子形成sporulation 细菌bacteria薄壁[细]菌类gracilicutes硬壁[细]菌类fermicutes疵壁[细]菌类mendosicutes无壁[细]菌类tenericutes柔膜细菌mollicutes真细菌eubacteria暗细菌scotobacteria无氧光细菌anoxyphotobacteria 生氧光细菌oxyphotobacteria 放线菌actinomycetes螺[旋]菌spirilla粘细菌slime bacteria。
微生物发酵中英文对照外文翻译文献
中英文对照外文翻译文献葡萄栽培过程中产生废弃物的侧耳属菇类生物降解：一种微生物和人类食物的来源及其在动物养殖中的潜在用途在通过侧耳属菌（平菇）程序进行葡萄园剪枝和葡萄皮渣的生物转化过程中，使用固态发酵技术受到了高度评价。
我们对水果实体的生产和收获之后被酶作用物的化学变化进行了测量计算，发现生物学效率和生物转化率各自都发生了变化，分别从37.2% 上升至78.7%和16.7%上升至 38.8%。
对于菌丝生长和蘑菇产量提高最有益的基质是与葡萄园剪枝项目相混合操作。
葡萄园修剪产生的枝条与葡萄皮渣相比具有较高的酚类成分、总糖、更好的c/n比值、天然脂肪和总氨。
与之相反，在纯葡萄皮渣的实验中，菌丝生长得非常缓慢甚至是不会生长。
葡萄皮渣比例较高的混合物中水分、蛋白质、脂肪和木质素含量一般较高，然而修剪产生的葡萄枝中，中性洗涤剂纤维、半纤维素、纤维素含量较高。
侧耳菌株的生长可能依赖于基质中纤维成分的可获取情况，而且其消化过程中发生的动态变化可能随着这些纤维在真菌生长过程中的改变而发生。
通过以侧耳属菌为媒介的SSF技术对葡萄栽培残基进行回收利用的潜力巨大，可以生产出人类所需的食物以及在反刍动物饲养中还有限使用的高纤维饲料。
关键词：生物转化酶作用；侧耳属菌；回收利用；固态发酵；葡萄栽培过程的副产品引言：葡萄种植是墨西哥西北部一项重要的生产活动，在墨西哥西北部有33500公顷的土地栽培了数类不同品种的葡萄。
这么大规模的生产活动每年大约产生了大约27万吨的工农业废料，而这其中有大约93%是葡萄园修剪掉的枝条。
这些废料一般直接在田间进行焚烧处理，以防止种植物病原菌的扩散，从而引起环境和生态问题以及危害人类健康的风险。
木质素是工农业废料中所有碳含量的主要组成部分，当它在遇热降解过程中会产生多环芳香烃成分，如苯并芘、邻苯二酚、对苯二酚菲和萘。
所有这些化合物可以抑制DNA 合成，并可能诱发动物和人类的肝脏、肺、喉和子宫颈产生癌变肿瘤。
微生物英文文献阅读
Increase minerals (N, P, K and Mg), and both Chl inhibiting spore biosynthesis and germination the photosynthetic activity ， accumulation of metabolites. 8h S+T > Ex1 +Ex2
Influence of Plant Extracts and Microbioagents on Physiological Traits of Faba Bean Infected with Botrytis fabae(蚕豆葡萄孢盘菌)
CONTENTS
1 2 ABSTRACT BACKGROUND
16h Ex1 +Ex2 > S+T The efficiency test+Ex2 biocontrol agents seemed to be in the 4 days S+T of > Ex1 > T > Ex2 > Ex1 = S
order: T+S > Ex1 +Ex2 > T > S > Ex2 > Ex1.
3.2 Preparation of B. fabae Spore Suspension
Isolated from the infected faba bean leaves—Identified—PDA medium—the spores（4 x 104 CFU m-1）
吴冰月
3 MATERIALS AND METHODS
Greenhouse experiments Laboratory experiments Reduce the activities of peroxidase(过氧化物酶), catalase
微生物英文文献及翻译—原文
Dynamic and distribution of ammonia-oxidizing bacteria communities during sludge granulation in an anaerobic e aerobic sequencing batch reactorZhang Bin a ,b ,Chen Zhe a ,b ,Qiu Zhigang a ,b ,Jin Min a ,b ,Chen Zhiqiang a ,b ,Chen Zhaoli a ,b ,Li Junwen a ,b ,Wang Xuan c ,*,Wang Jingfeng a ,b ,**aInstitute of Hygiene and Environmental Medicine,Academy of Military Medical Sciences,Tianjin 300050,PR China bTianjin Key Laboratory of Risk Assessment and Control for Environment and Food Safety,Tianjin 300050,PR China cTianjin Key Laboratory of Hollow Fiber Membrane Material and Membrane Process,Institute of Biological and Chemical Engineering,Tianjin Polytechnical University,Tianjin 300160,PR Chinaa r t i c l e i n f oArticle history:Received 30June 2011Received in revised form 10September 2011Accepted 10September 2011Available online xxx Keywords:Ammonia-oxidizing bacteria Granular sludgeCommunity development Granule sizeNitrifying bacteria distribution Phylogenetic diversitya b s t r a c tThe structure dynamic of ammonia-oxidizing bacteria (AOB)community and the distribution of AOB and nitrite-oxidizing bacteria (NOB)in granular sludge from an anaerobic e aerobic sequencing batch reactor (SBR)were investigated.A combination of process studies,molecular biotechniques and microscale techniques were employed to identify and characterize these organisms.The AOB community structure in granules was substantially different from that of the initial pattern of the inoculants sludge.Along with granules formation,the AOB diversity declined due to the selection pressure imposed by process conditions.Denaturing gradient gel electrophoresis (DGGE)and sequencing results demonstrated that most of Nitrosomonas in the inoculating sludge were remained because of their ability to rapidly adapt to the settling e washing out action.Furthermore,DGGE analysis revealed that larger granules benefit more AOB species surviving in the reactor.In the SBR were various size granules coexisted,granule diameter affected the distribution range of AOB and NOB.Small and medium granules (d <0.6mm)cannot restrict oxygen mass transfer in all spaces of the rger granules (d >0.9mm)can result in smaller aerobic volume fraction and inhibition of NOB growth.All these observations provide support to future studies on the mechanisms responsible for the AOB in granules systems.ª2011Elsevier Ltd.All rights reserved.1.IntroductionAt sufficiently high levels,ammonia in aquatic environments can be toxic to aquatic life and can contribute to eutrophica-tion.Accordingly,biodegradation and elimination of ammonia in wastewater are the primary functions of thewastewater treatment process.Nitrification,the conversion of ammonia to nitrate via nitrite,is an important way to remove ammonia nitrogen.It is a two-step process catalyzed by ammonia-oxidizing and nitrite-oxidizing bacteria (AOB and NOB).Aerobic ammonia-oxidation is often the first,rate-limiting step of nitrification;however,it is essential for the*Corresponding author .**Corresponding author.Institute of Hygiene and Environmental Medicine,Academy of Military Medical Sciences,Tianjin 300050,PR China.Tel.:+862284655498;fax:+862223328809.E-mail addresses:wangxuan0116@ (W.Xuan),jingfengwang@ (W.Jingfeng).Available online atjournal homepage:/locate/watresw a t e r r e s e a r c h x x x (2011)1e 100043-1354/$e see front matter ª2011Elsevier Ltd.All rights reserved.doi:10.1016/j.watres.2011.09.026removal of ammonia from the wastewater(Prosser and Nicol, 2008).Comparative analyses of16S rRNA sequences have revealed that most AOB in activated sludge are phylogeneti-cally closely related to the clade of b-Proteobacteria (Kowalchuk and Stephen,2001).However,a number of studies have suggested that there are physiological and ecological differences between different AOB genera and lineages,and that environmental factors such as process parameter,dis-solved oxygen,salinity,pH,and concentrations of free ammonia can impact certain species of AOB(Erguder et al., 2008;Kim et al.,2006;Koops and Pommerening-Ro¨ser,2001; Kowalchuk and Stephen,2001;Shi et al.,2010).Therefore, the physiological activity and abundance of AOB in waste-water processing is critical in the design and operation of waste treatment systems.For this reason,a better under-standing of the ecology and microbiology of AOB in waste-water treatment systems is necessary to enhance treatment performance.Recently,several developed techniques have served as valuable tools for the characterization of microbial diversity in biological wastewater treatment systems(Li et al., 2008;Yin and Xu,2009).Currently,the application of molec-ular biotechniques can provide clarification of the ammonia-oxidizing community in detail(Haseborg et al.,2010;Tawan et al.,2005;Vlaeminck et al.,2010).In recent years,the aerobic granular sludge process has become an attractive alternative to conventional processes for wastewater treatment mainly due to its cell immobilization strategy(de Bruin et al.,2004;Liu et al.,2009;Schwarzenbeck et al.,2005;Schwarzenbeck et al.,2004a,b;Xavier et al.,2007). Granules have a more tightly compact structure(Li et al.,2008; Liu and Tay,2008;Wang et al.,2004)and rapid settling velocity (Kong et al.,2009;Lemaire et al.,2008).Therefore,granular sludge systems have a higher mixed liquid suspended sludge (MLSS)concentration and longer solid retention times(SRT) than conventional activated sludge systems.Longer SRT can provide enough time for the growth of organisms that require a long generation time(e.g.,AOB).Some studies have indicated that nitrifying granules can be cultivated with ammonia-rich inorganic wastewater and the diameter of granules was small (Shi et al.,2010;Tsuneda et al.,2003).Other researchers reported that larger granules have been developed with the synthetic organic wastewater in sequencing batch reactors(SBRs)(Li et al., 2008;Liu and Tay,2008).The diverse populations of microor-ganisms that coexist in granules remove the chemical oxygen demand(COD),nitrogen and phosphate(de Kreuk et al.,2005). However,for larger granules with a particle diameter greater than0.6mm,an outer aerobic shell and an inner anaerobic zone coexist because of restricted oxygen diffusion to the granule core.These properties of granular sludge suggest that the inner environment of granules is unfavorable to AOB growth.Some research has shown that particle size and density induced the different distribution and dominance of AOB,NOB and anam-mox(Winkler et al.,2011b).Although a number of studies have been conducted to assess the ecology and microbiology of AOB in wastewater treatment systems,the information on the dynamics,distribution,and quantification of AOB communities during sludge granulation is still limited up to now.To address these concerns,the main objective of the present work was to investigate the population dynamics of AOB communities during the development of seedingflocs into granules,and the distribution of AOB and NOB in different size granules from an anaerobic e aerobic SBR.A combination of process studies,molecular biotechniques and microscale techniques were employed to identify and char-acterize these organisms.Based on these approaches,we demonstrate the differences in both AOB community evolu-tion and composition of theflocs and granules co-existing in the SBR and further elucidate the relationship between distribution of nitrifying bacteria and granule size.It is ex-pected that the work would be useful to better understand the mechanisms responsible for the AOB in granules and apply them for optimal control and management strategies of granulation systems.2.Material and methods2.1.Reactor set-up and operationThe granules were cultivated in a lab-scale SBR with an effective volume of4L.The effective diameter and height of the reactor was10cm and51cm,respectively.The hydraulic retention time was set at8h.Activated sludge from a full-scale sewage treat-ment plant(Jizhuangzi Sewage Treatment Works,Tianjin, China)was used as the seed sludge for the reactor at an initial sludge concentration of3876mg LÀ1in MLSS.The reactor was operated on6-h cycles,consisting of2-min influent feeding,90-min anaerobic phase(mixing),240-min aeration phase and5-min effluent discharge periods.The sludge settling time was reduced gradually from10to5min after80SBR cycles in20days, and only particles with a settling velocity higher than4.5m hÀ1 were retained in the reactor.The composition of the influent media were NaAc(450mg LÀ1),NH4Cl(100mg LÀ1),(NH4)2SO4 (10mg LÀ1),KH2PO4(20mg LÀ1),MgSO4$7H2O(50mg LÀ1),KCl (20mg LÀ1),CaCl2(20mg LÀ1),FeSO4$7H2O(1mg LÀ1),pH7.0e7.5, and0.1mL LÀ1trace element solution(Li et al.,2007).Analytical methods-The total organic carbon(TOC),NHþ4e N, NOÀ2e N,NOÀ3e N,total nitrogen(TN),total phosphate(TP) concentration,mixed liquid suspended solids(MLSS) concentration,and sludge volume index at10min(SVI10)were measured regularly according to the standard methods (APHA-AWWA-WEF,2005).Sludge size distribution was determined by the sieving method(Laguna et al.,1999).Screening was performed with four stainless steel sieves of5cm diameter having respective mesh openings of0.9,0.6,0.45,and0.2mm.A100mL volume of sludge from the reactor was sampled with a calibrated cylinder and then deposited on the0.9mm mesh sieve.The sample was subsequently washed with distilled water and particles less than0.9mm in diameter passed through this sieve to the sieves with smaller openings.The washing procedure was repeated several times to separate the gran-ules.The granules collected on the different screens were recovered by backwashing with distilled water.Each fraction was collected in a different beaker andfiltered on quantitative filter paper to determine the total suspended solid(TSS).Once the amount of total suspended solid(TSS)retained on each sieve was acquired,it was reasonable to determine for each class of size(<0.2,[0.2e0.45],[0.45e0.6],[0.6e0.9],>0.9mm) the percentage of the total weight that they represent.w a t e r r e s e a r c h x x x(2011)1e10 22.2.DNA extraction and nested PCR e DGGEThe sludge from approximately8mg of MLSS was transferred into a1.5-mL Eppendorf tube and then centrifuged at14,000g for10min.The supernatant was removed,and the pellet was added to1mL of sodium phosphate buffer solution and aseptically mixed with a sterilized pestle in order to detach granules.Genomic DNA was extracted from the pellets using E.Z.N.A.äSoil DNA kit(D5625-01,Omega Bio-tek Inc.,USA).To amplify ammonia-oxidizer specific16S rRNA for dena-turing gradient gel electrophoresis(DGGE),a nested PCR approach was performed as described previously(Zhang et al., 2010).30m l of nested PCR amplicons(with5m l6Âloading buffer)were loaded and separated by DGGE on polyacrylamide gels(8%,37.5:1acrylamide e bisacrylamide)with a linear gradient of35%e55%denaturant(100%denaturant¼7M urea plus40%formamide).The gel was run for6.5h at140V in 1ÂTAE buffer(40mM Tris-acetate,20mM sodium acetate, 1mM Na2EDTA,pH7.4)maintained at60 C(DCodeäUniversal Mutation Detection System,Bio-Rad,Hercules,CA, USA).After electrophoresis,silver-staining and development of the gels were performed as described by Sanguinetti et al. (1994).These were followed by air-drying and scanning with a gel imaging analysis system(Image Quant350,GE Inc.,USA). The gel images were analyzed with the software Quantity One,version4.31(Bio-rad).Dice index(Cs)of pair wise community similarity was calculated to evaluate the similarity of the AOB community among DGGE lanes(LaPara et al.,2002).This index ranges from0%(no common band)to100%(identical band patterns) with the assistance of Quantity One.The Shannon diversity index(H)was used to measure the microbial diversity that takes into account the richness and proportion of each species in a population.H was calculatedusing the following equation:H¼ÀPn iNlogn iN,where n i/Nis the proportion of community made up by species i(bright-ness of the band i/total brightness of all bands in the lane).Dendrograms relating band pattern similarities were automatically calculated without band weighting(consider-ation of band density)by the unweighted pair group method with arithmetic mean(UPGMA)algorithms in the Quantity One software.Prominent DGGE bands were excised and dissolved in30m L Milli-Q water overnight,at4 C.DNA was recovered from the gel by freeze e thawing thrice.Cloning and sequencing of the target DNA fragments were conducted following the estab-lished method(Zhang et al.,2010).2.3.Distribution of nitrifying bacteriaThree classes of size([0.2e0.45],[0.45e0.6],>0.9mm)were chosen on day180for FISH analysis in order to investigate the spatial distribution characteristics of AOB and NOB in granules.2mg sludge samples werefixed in4%para-formaldehyde solution for16e24h at4 C and then washed twice with sodium phosphate buffer;the samples were dehydrated in50%,80%and100%ethanol for10min each. Ethanol in the granules was then completely replaced by xylene by serial immersion in ethanol-xylene solutions of3:1, 1:1,and1:3by volume andfinally in100%xylene,for10min periods at room temperature.Subsequently,the granules were embedded in paraffin(m.p.56e58 C)by serial immer-sion in1:1xylene-paraffin for30min at60 C,followed by 100%paraffin.After solidification in paraffin,8-m m-thick sections were prepared and placed on gelatin-coated micro-scopic slides.Paraffin was removed by immersing the slide in xylene and ethanol for30min each,followed by air-drying of the slides.The three oligonucleotide probes were used for hybridiza-tion(Downing and Nerenberg,2008):FITC-labeled Nso190, which targets the majority of AOB;TRITC-labeled NIT3,which targets Nitrobacter sp.;TRITC-labeled NSR1156,which targets Nitrospira sp.All probe sequences,their hybridization condi-tions,and washing conditions are given in Table1.Oligonu-cleotides were synthesized andfluorescently labeled with fluorochomes by Takara,Inc.(Dalian,China).Hybridizations were performed at46 C for2h with a hybridization buffer(0.9M NaCl,formamide at the percentage shown in Table1,20mM Tris/HCl,pH8.0,0.01% SDS)containing each labeled probe(5ng m LÀ1).After hybrid-ization,unbound oligonucleotides were removed by a strin-gent washing step at48 C for15min in washing buffer containing the same components as the hybridization buffer except for the probes.For detection of all DNA,4,6-diamidino-2-phenylindole (DAPI)was diluted with methanol to afinal concentration of1ng m LÀ1.Cover the slides with DAPI e methanol and incubate for15min at37 C.The slides were subsequently washed once with methanol,rinsed briefly with ddH2O and immediately air-dried.Vectashield(Vector Laboratories)was used to prevent photo bleaching.The hybridization images were captured using a confocal laser scanning microscope (CLSM,Zeiss710).A total of10images were captured for each probe at each class of size.The representative images were selected andfinal image evaluation was done in Adobe PhotoShop.w a t e r r e s e a r c h x x x(2011)1e1033.Results3.1.SBR performance and granule characteristicsDuring the startup period,the reactor removed TOC and NH 4þ-N efficiently.98%of NH 4þ-N and 100%of TOC were removed from the influent by day 3and day 5respectively (Figs.S2,S3,Supporting information ).Removal of TN and TP were lower during this period (Figs.S3,S4,Supporting information ),though the removal of TP gradually improved to 100%removal by day 33(Fig.S4,Supporting information ).To determine the sludge volume index of granular sludge,a settling time of 10min was chosen instead of 30min,because granular sludge has a similar SVI after 60min and after 5min of settling (Schwarzenbeck et al.,2004b ).The SVI 10of the inoculating sludge was 108.2mL g À1.The changing patterns of MLSS and SVI 10in the continuous operation of the SBR are illustrated in Fig.1.The sludge settleability increased markedly during the set-up period.Fig.2reflects the slow andgradual process of sludge granulation,i.e.,from flocculentsludge to granules.3.2.DGGE analysis:AOB communities structure changes during sludge granulationThe results of nested PCR were shown in Fig.S1.The well-resolved DGGE bands were obtained at the representative points throughout the GSBR operation and the patterns revealed that the structure of the AOB communities was dynamic during sludge granulation and stabilization (Fig.3).The community structure at the end of experiment was different from that of the initial pattern of the seed sludge.The AOB communities on day 1showed 40%similarity only to that at the end of the GSBR operation (Table S1,Supporting information ),indicating the considerable difference of AOB communities structures between inoculated sludge and granular sludge.Biodiversity based on the DGGE patterns was analyzed by calculating the Shannon diversity index H as204060801001201401254159738494104115125135147160172188Time (d)S V I 10 (m L .g -1)10002000300040005000600070008000900010000M L S S (m g .L -1)Fig.1e Change in biomass content and SVI 10during whole operation.SVI,sludge volume index;MLSS,mixed liquid suspendedsolids.Fig.2e Variation in granule size distribution in the sludge during operation.d,particle diameter;TSS,total suspended solids.w a t e r r e s e a r c h x x x (2011)1e 104shown in Fig.S5.In the phase of sludge inoculation (before day 38),H decreased remarkably (from 0.94to 0.75)due to the absence of some species in the reactor.Though several dominant species (bands2,7,10,11)in the inoculating sludge were preserved,many bands disappeared or weakened (bands 3,4,6,8,13,14,15).After day 45,the diversity index tended to be stable and showed small fluctuation (from 0.72to 0.82).Banding pattern similarity was analyzed by applying UPGMA (Fig.4)algorithms.The UPGMA analysis showed three groups with intragroup similarity at approximately 67%e 78%and intergroup similarity at 44e 62%.Generally,the clustering followed the time course;and the algorithms showed a closer clustering of groups II and III.In the analysis,group I was associated with sludge inoculation and washout,group IIwithFig.3e DGGE profile of the AOB communities in the SBR during the sludge granulation process (lane labels along the top show the sampling time (days)from startup of the bioreactor).The major bands were labeled with the numbers (bands 1e15).Fig.4e UPGMA analysis dendrograms of AOB community DGGE banding patterns,showing schematics of banding patterns.Roman numerals indicate major clusters.w a t e r r e s e a r c h x x x (2011)1e 105startup sludge granulation and decreasing SVI 10,and group III with a stable system and excellent biomass settleability.In Fig.3,the locations of the predominant bands were excised from the gel.DNA in these bands were reamplified,cloned and sequenced.The comparative analysis of these partial 16S rRNA sequences (Table 2and Fig.S6)revealed the phylogenetic affiliation of 13sequences retrieved.The majority of the bacteria in seed sludge grouped with members of Nitrosomonas and Nitrosospira .Along with sludge granula-tion,most of Nitrosomonas (Bands 2,5,7,9,10,11)were remained or eventually became dominant in GSBR;however,all of Nitrosospira (Bands 6,13,15)were gradually eliminated from the reactor.3.3.Distribution of AOB and NOB in different sized granulesFISH was performed on the granule sections mainly to deter-mine the location of AOB and NOB within the different size classes of granules,and the images were not further analyzed for quantification of cell counts.As shown in Fig.6,in small granules (0.2mm <d <0.45mm),AOB located mainly in the outer part of granular space,whereas NOB were detected only in the core of granules.In medium granules (0.45mm <d <0.6mm),AOB distributed evenly throughout the whole granular space,whereas NOB still existed in the inner part.In the larger granules (d >0.9mm),AOB and NOB were mostly located in the surface area of the granules,and moreover,NOB became rare.4.Discussion4.1.Relationship between granule formation and reactor performanceAfter day 32,the SVI 10stabilized at 20e 35mL g À1,which is very low compared to the values measured for activated sludge (100e 150mL g À1).However,the size distribution of the granules measured on day 32(Fig.2)indicated that only 22%of the biomass was made of granular sludge with diameter largerthan 0.2mm.These results suggest that sludge settleability increased prior to granule formation and was not affected by different particle sizes in the sludge during the GSBR operation.It was observed,however,that the diameter of the granules fluctuated over longer durations.The large granules tended to destabilize due to endogenous respiration,and broke into smaller granules that could seed the formation of large granules again.Pochana and Keller reported that physically broken sludge flocs contribute to lower denitrification rates,due to their reduced anoxic zone (Pochana and Keller,1999).Therefore,TN removal efficiency raises fluctuantly throughout the experiment.Some previous research had demonstrated that bigger,more dense granules favored the enrichment of PAO (Winkler et al.,2011a ).Hence,after day 77,removal efficiency of TP was higher and relatively stable because the granules mass fraction was over 90%and more larger granules formed.4.2.Relationship between AOB communities dynamic and sludge granulationFor granule formation,a short settling time was set,and only particles with a settling velocity higher than 4.5m h À1were retained in the reactor.Moreover,as shown in Fig.1,the variation in SVI 10was greater before day 41(from 108.2mL g À1e 34.1mL g À1).During this phase,large amounts of biomass could not survive in the reactor.A clear shift in pop-ulations was evident,with 58%similarity between days 8and 18(Table S1).In the SBR system fed with acetate-based synthetic wastewater,heterotrophic bacteria can produce much larger amounts of extracellular polysaccharides than autotrophic bacteria (Tsuneda et al.,2003).Some researchers found that microorganisms in high shear environments adhered by extracellular polymeric substances (EPS)to resist the damage of suspended cells by environmental forces (Trinet et al.,1991).Additionally,it had been proved that the dominant heterotrophic species in the inoculating sludge were preserved throughout the process in our previous research (Zhang et al.,2011).It is well known that AOB are chemoau-totrophic and slow-growing;accordingly,numerous AOBw a t e r r e s e a r c h x x x (2011)1e 106populations that cannot become big and dense enough to settle fast were washed out from the system.As a result,the variation in AOB was remarkable in the period of sludge inoculation,and the diversity index of population decreased rapidly.After day 45,AOB communities’structure became stable due to the improvement of sludge settleability and the retention of more biomass.These results suggest that the short settling time (selection pressure)apparently stressed the biomass,leading to a violent dynamic of AOB communities.Further,these results suggest that certain populations may have been responsible for the operational success of the GSBR and were able to persist despite the large fluctuations in pop-ulation similarity.This bacterial population instability,coupled with a generally acceptable bioreactor performance,is congruent with the results obtained from a membrane biore-actor (MBR)for graywater treatment (Stamper et al.,2003).Nitrosomonas e like and Nitrosospira e like populations are the dominant AOB populations in wastewater treatment systems (Kowalchuk and Stephen,2001).A few previous studies revealed that the predominant populations in AOB communities are different in various wastewater treatment processes (Tawan et al.,2005;Thomas et al.,2010).Some researchers found that the community was dominated by AOB from the genus Nitrosospira in MBRs (Zhang et al.,2010),whereas Nitrosomonas sp.is the predominant population in biofilter sludge (Yin and Xu,2009).In the currentstudy,Fig.5e DGGE profile of the AOB communities in different size of granules (lane labels along the top show the range of particle diameter (d,mm)).Values along the bottom indicate the Shannon diversity index (H ).Bands labeled with the numbers were consistent with the bands in Fig.3.w a t e r r e s e a r c h x x x (2011)1e 107sequence analysis revealed that selection pressure evidently effect on the survival of Nitrosospira in granular sludge.Almost all of Nitrosospira were washed out initially and had no chance to evolve with the environmental changes.However,some members of Nitrosomonas sp.have been shown to produce more amounts of EPS than Nitrosospira ,especially under limited ammonia conditions (Stehr et al.,1995);and this feature has also been observed for other members of the same lineage.Accordingly,these EPS are helpful to communicate cells with each other and granulate sludge (Adav et al.,2008).Therefore,most of Nitrosomonas could adapt to this challenge (to become big and dense enough to settle fast)and were retained in the reactor.At the end of reactor operation (day 180),granules with different particle size were sieved.The effects of variation in granules size on the composition of the AOBcommunitiesFig.6e Micrographs of FISH performed on three size classes of granule sections.DAPI stain micrographs (A,D,G);AOB appear as green fluorescence (B,E,H),and NOB appear as red fluorescence (C,F,I).Bar [100m m in (A)e (C)and (G)e (I).d,particle diameter.(For interpretation of the references to colour in this figure legend,the reader is referred to the web version of this article.)w a t e r r e s e a r c h x x x (2011)1e 108were investigated.As shown in Fig.5,AOB communities structures in different size of granules were varied.Although several predominant bands(bands2,5,11)were present in all samples,only bands3and6appeared in the granules with diameters larger than0.6mm.Additionally,bands7and10 were intense in the granules larger than0.45mm.According to Table2,it can be clearly indicated that Nitrosospira could be retained merely in the granules larger than0.6mm.Therefore, Nitrosospira was not present at a high level in Fig.3due to the lower proportion of larger granules(d>0.6mm)in TSS along with reactor operation.DGGE analysis also revealed that larger granules had a greater microbial diversity than smaller ones. This result also demonstrates that more organisms can survive in larger granules as a result of more space,which can provide the suitable environment for the growth of microbes(Fig.6).4.3.Effect of variance in particle size on the distribution of AOB and NOB in granulesAlthough an influence of granule size has been observed in experiments and simulations for simultaneous N-and P-removal(de Kreuk et al.,2007),the effect of granule size on the distribution of different biomass species need be revealed further with the assistance of visible experimental results, especially in the same granular sludge reactors.Related studies on the diversity of bacterial communities in granular sludge often focus on the distribution of important functional bacteria populations in single-size granules(Matsumoto et al., 2010).In the present study,different size granules were sieved,and the distribution patterns of AOB and NOB were explored.In the nitrification processes considered,AOB and NOB compete for space and oxygen in the granules(Volcke et al.,2010).Since ammonium oxidizers have a higheroxygen affinity(K AOBO2<K NOBO2)and accumulate more rapidly inthe reactor than nitrite oxidizers(Volcke et al.,2010),NOB are located just below the layer of AOB,where still some oxygen is present and allows ready access to the nitrite produced.In smaller granules,the location boundaries of the both biomass species were distinct due to the limited existence space provided by granules for both microorganism’s growth.AOB exist outside of the granules where oxygen and ammonia are present.Medium granules can provide broader space for microbe multiplying;accordingly,AOB spread out in the whole granules.This result also confirms that oxygen could penetrate deep into the granule’s core without restriction when particle diameter is less than0.6mm.Some mathematic model also supposed that NOBs are favored to grow in smaller granules because of the higher fractional aerobic volume (Volcke et al.,2010).As shown in the results of the batch experiments(Zhang et al.,2011),nitrite accumulation temporarily occurred,accompanied by the more large gran-ules(d>0.9mm)forming.This phenomenon can be attrib-uted to the increased ammonium surface load associated with larger granules and smaller aerobic volume fraction,resulting in outcompetes of NOB.It also suggests that the core areas of large granules(d>0.9mm)could provide anoxic environment for the growth of anaerobic denitrificans(such as Tb.deni-trificans or Tb.thioparus in Fig.S7,Supporting information).As shown in Fig.2and Fig.S3,the removal efficiency of total nitrogen increased with formation of larger granules.5.ConclusionsThe variation in AOB communities’structure was remarkable during sludge inoculation,and the diversity index of pop-ulation decreased rapidly.Most of Nitrosomonas in the inocu-lating sludge were retained because of their capability to rapidly adapt to the settling e washing out action.DGGE anal-ysis also revealed that larger granules had greater AOB diversity than that of smaller ones.Oxygen penetration was not restricted in the granules of less than0.6mm particle diameter.However,the larger granules(d>0.9mm)can result in the smaller aerobic volume fraction and inhibition of NOB growth.Henceforth,further studies on controlling and opti-mizing distribution of granule size could be beneficial to the nitrogen removal and expansive application of granular sludge technology.AcknowledgmentsThis work was supported by grants from the National Natural Science Foundation of China(No.51108456,50908227)and the National High Technology Research and Development Program of China(No.2009AA06Z312).Appendix.Supplementary dataSupplementary data associated with this article can be found in online version at doi:10.1016/j.watres.2011.09.026.r e f e r e n c e sAdav,S.S.,Lee, D.J.,Show,K.Y.,2008.Aerobic granular sludge:recent advances.Biotechnology Advances26,411e423.APHA-AWWA-WEF,2005.Standard Methods for the Examination of Water and Wastewater,first ed.American Public Health Association/American Water Works Association/WaterEnvironment Federation,Washington,DC.de Bruin,L.M.,de Kreuk,M.,van der Roest,H.F.,Uijterlinde,C., van Loosdrecht,M.C.M.,2004.Aerobic granular sludgetechnology:an alternative to activated sludge?Water Science and Technology49,1e7.de Kreuk,M.,Heijnen,J.J.,van Loosdrecht,M.C.M.,2005.Simultaneous COD,nitrogen,and phosphate removal byaerobic granular sludge.Biotechnology and Bioengineering90, 761e769.de Kreuk,M.,Picioreanu,C.,Hosseini,M.,Xavier,J.B.,van Loosdrecht,M.C.M.,2007.Kinetic model of a granular sludge SBR:influences on nutrient removal.Biotechnology andBioengineering97,801e815.Downing,L.S.,Nerenberg,R.,2008.Total nitrogen removal ina hybrid,membrane-aerated activated sludge process.WaterResearch42,3697e3708.Erguder,T.H.,Boon,N.,Vlaeminck,S.E.,Verstraete,W.,2008.Partial nitrification achieved by pulse sulfide doses ina sequential batch reactor.Environmental Science andTechnology42,8715e8720.w a t e r r e s e a r c h x x x(2011)1e109。
关于微生物的文献
MINI-REVIEWInteractions of microorganisms with rare earth ions and their utilization for separation and environmental technology Hiroshi Moriwaki&Hiroki YamamotoReceived:5September2012/Revised:10October2012/Accepted:14October2012#Springer-Verlag Berlin Heidelberg2012Abstract In recent years,rare earth elements(REEs)have been widely used in various modern technological devices and the global demand for REE has been increasing.The increased demand for REEs has led to environmental expo-sure or water pollution from rare earth metal mines and various commercial products.Therefore,the development of a safe technology for the separation and adsorption of REEs is very important from the perspective of green chem-istry and environmental pollution.In this review,the appli-cation and mechanisms of microorganisms for the removal and extraction of REEs from aqueous solutions are de-scribed.In addition,the advantages in using microorgan-isms for REE adsorption and future studies on this topic are discussed.Keywords Rare earth elements.Adsorption. Microorganisms.Mechanism.Teichoic acid IntroductionThe rare earth elements(REEs)are a group of metals com-prising a set of15or16elements in the periodic table, specifically the14lanthanides(Pm,whose isotopes are radioactive with short half-lives,is generally excluded.), yttrium,and sometimes scandium.REEs are widely used in various technological devices,including superconductors (Y,La,and Lu),magnets(Ce,Nd,Sm,Gd,Tb,and Dy), fluorescent materials(Sc,Y,Eu,Tb,and Tm),and catalysts (Y,La,and Ce;Andrianov et al.2011;Molander and Romero2002).The demand for REEs is rapidly increasing worldwide.China has played a dominant role in REE mining pro-duction since1990(Du and Graedel2011).Recently,high concentrations of REEs have been found in the deep-sea mud at numerous sites throughout the eastern South and central North Pacific(Kato et al.2011).However,the REE resources are not exploited commercially at the present time because of the technical difficulty and high cost of mining. The importance of REEs as a resource has been tremen-dously growing,and as a result their monetary value has been increasing.Techniques for the separation and purification of REEs have gained in importance with the increasing demand for REEs.Various kinds of phosphates or functionalized poly-mers have been widely used for these purposes(Chen et al. 2012;Sun et al.2012;Buchmeiser et al.1998;Zahir and Masuda1997).However,most of these reagents are expen-sive and may be toxic themselves.Under these circumstan-ces,it is very important to develop low-cost and environmentally friendly procedures for the extraction of rare earth ions from water.It is well-known that REEs exhibit hepatotoxic and neurotoxic effects(Pałasz and Czekaj2000;Basu et al.1982).However,up to now,carci-nogenicity of REEs in animals has not been reported (Hirano and Suzuki1996).More and more REEs diffuse into the environment because they are being rapidly exploited and widely used in modern industry and daily life. In fact,in one report,high levels of rare earth metals were detected in hair from the scalp of children living in a rare earth metals mining area in China(Tong et al.2005).It is of great concern that the spreading of REEs has adverse effects on living organisms.Therefore,it is very important to de-velop a safe adsorbent and a procedure for its use in remov-ing REEs from the aqueousenvironment.H.Moriwaki(*):H.YamamotoFaculty of Textile Science and Technology,Division of Applied Biology,Shinshu University,3-15-1,Tokida,Ueda386-8567,Japane-mail:moriwaki@shinshu-u.ac.jpAppl Microbiol BiotechnolDOI10.1007/s00253-012-4519-9It is beneficial to use bioresource materials as adsorbents for environmental pollutants because they are economic and eco-friendly(Babel and Kurniawan2003;Sud et al.2008).Recent-ly,various bioresource materials have been used as adsorbents for pollutants such as oil,pesticides,and metal ions(Moriwaki et al.2009;Senthilkumaar et al.2010;Ghimire et al.2003; Demirbas2008).In particular,applications of microorganisms for the removal of heavy metal ions from aqueous environ-ments have received much attention,and intensive studies have been performed on a variety of bacteria to improve these procedures(Kuroda and Ueda2010;Wu et al.2012).The processes for the removal of heavy metal ions by using microorganisms may be divided into four categories, as shown in Fig.1.An efficient and widely applied tech-nique involves the adsorption of metal ions on the surface of microbial cells.Another well-known technique involves the adsorption of metal ions by extracellular biopolymers,such as polyglutamic acid and polysaccharide.A third technique of metal ion removal from solution involves their absorption into microbial cells(Klaus-Joerger et al.2001)and a forth technique involves adsorption by bio-minerals such as man-ganese oxide(Tani et al.2004).Adsorption of metal ions by the microbial cell surface is well understood.There are various advantages of using microbial cell surfaces as adsorbents,which include high efficiency of adsorption of dissolved metals because micro-bial cells have a high surface area per unit weight and metal adsorption by them has a relatively small impact on the environment(Wu et al.2012).Several studies dealing with REE adsorption by bacterial cells were published recently(Merroun et al.2003;Takahashi et al.2005;Ozaki et al.2006).In particular,interactions between the bacterial cell wall and REEs were studied to gain insights into the mechanism of REE adsorption and to shed light on their potential utility of bacterial cell walls as adsorb-ents for REEs.Understanding interactions between the bacte-rial cell wall and REEs provides useful information about the environmental behavior and geological distribution of REEs.Andres et al.(2003)published a review on the removal of REEs by microbial bio-sorption.The interaction of the surface of a bacterium with REEs at the molecular level has not been studied until recently;however,the selective accumulation of REEs by bacteria has been the focus of several recent studies.In this review,we summarize the results of REE adsorption onto bacterial cells from studies conducted in the last decade.In particular,we describe in detail the mechanism of the interaction between the micro-bial cell wall and REEs at the molecular level.REE adsorption by microorganismsTable1summarizes some of the major published REE sorption experiments on bacteria.REE adsorption by vari-ous bacterial strains was examined.In particular,many researchers studied the adsorption onto the cell wall of gram-positive bacteria,such as Bacillus subtilis,and gram-negative bacteria,such as Escherichia coli.For example,the adsorption behaviors of Eu(III)on three kinds of gram-negative bacteria were compared,and the coordination en-vironment of Eu(III)on the bacteria was found to be differ-ent(Ozaki et al.2005).Moreover,the adsorption behavior of Eu(III)on Halobacterium salinarum was also investigat-ed(Ozaki et al.2002a).The authors concluded that the coordination states of Eu(III)adsorbed on the B.subtilis and H.salinarum had different characteristics.Various microbial cells can produce extracellular poly-meric substances(EPS),which lead to floc formation due to the agglomeration of bacteria(Sponza2002).EPS providea) Surface adsorptionM n+b) Adsorption on extracellular biopolymer M n+: metal iond) Adsorption on extracellular biomineralFig.1Pattern diagrams for theinteraction of heavy metal ionswith bacteriaAppl Microbiol BiotechnolT a b l e 1P u b l i c a t i o n s d e a l i n g w i t h R E E a d s o r p t i o n b y m i c r o o r g a n i s mE n t r y M i c r o o r g a n i s m sM e c h a n i s mM e t a l i o n R e s u l t s C o m m e n t s R e f e r e n c e1B a c i l l u s s u b t i l i s A d s o r p t i o n o n t o c e l l w a l l E uT h e c o o r d i n a t i o n s t a t e s o f E u a d s o r b e d o n H .s a l i n a r u m w a s m o r e o u t e r -s p h e r i c a l t h a n E u a d s o r b e d o n B .s u b t i l i s .T h e c o o r d i n a t i o n s t a t e o f E u a d s o r b e d o n t h e m i c r o o r g a n i s m s w a s s t u d i e d b y T i m e -r e s o l v e d l a s e r -i n d u c e d f l u o r e s c e n c e s p e c t r o s c o p y (T R L F S ).O z a k i e t a l .(2002a )H a l o b a c t e r i u m s a l i n a r u m2B .s u b t i l i s A d s o r p t i o n o n t o c e l l w a l l E uT h e E u (I I I )/B .s u b t i l i s e q u i l i b r i u m w a s r e v e r s i b l e a t p H 5.T R L F S w a s u s e d t o s t u d y l a n t a n i d e a d s o r p t i o n .M a r k a i e t a l .(2003)3M y x o c o c c u s x a n t h u sA d s o r p t i o n t o e x t r a c e l l u l a r b i o p o l y m e r a n d c e l l w a l lL a L a w e r e a d s o r b e d o n t o t h e p o l y s a c c h a r i d e p r o d u c e d b y M .x a n t h u s .L a f i x a t i o n o n t h e e x t r a c e l l u l a r p o l y m e r i c s u b s t a n c e s w a s c o n f i r m e d b y T E M .M e r r o u n e t a l .(2003)4G a l l i o n e l l a f e r r u g i n e aA d s o r p t i o n t o e x t r a c e l l u l a r m i n e r a l14k i n d s o f R E E s B a c t e r i o g e n i c i r o n o x i d e s c a n a c c u m u l a t e R E E s .B a c t e r i o g e n i c F e o x i d e s c a n b e e n r i c h e d i n h e a v y R E E s c o m p a r e d t o i n o r g a n i c m a t e r i a l s .A n d e r s o n a n d P e d e r s o n (2003)5B .s u b t i l i s A d s o r p t i o n o n t o c e l l w a l l 15k i n d s o f R E E s T m ,Y b ,a n d L u a r e e n r i c h e d o n t h e c e l l w a l l o f B .s u b t i l i s a n d E .c o l i c o m p a r e d t o o t h e r R E E s .T h e R E E p a t t e r n s o f n a t u r a l m i c r o b i a l m a t w e r e s i m i l a r t o t h a t o b t a i n e d i n t h e l a b o r a t o r y e x p e r i m e n t s u s i n g p u r e b a c t e r i a l s t r a i n s .T a k a h a s h i e t a l .(2005)E s c h e r i c h i a c o l i6A l c a l i g e n e s f a e c a l i s A d s o r p t i o n o n t o c e l l w a l l E u T R L F S s h o w e d t h a t t h e c o o r d i n a t i o n o f E u o n t h e b a c t e r i a d i f f e r e d ,t h o u g h t h e y a r e c a t e g o r i z e d a s G r a m -n e g a t i v e b a c t e r i a .T h e a s s o c i a t i o n s o f E u w i t h G r a m -n e g a t i v e b a c t e r i a b y b a t c h m e t h o d a n d T R L F S w e r e s t u d i e d O z a k i e t a l .(2005)S c h w a n e l l a p u t r e f a c i e n sP a r a c o c c u s d e n i t r i f i c a n s7P s e u d o m o n a s f l u o r e s c e n sA d s o r p t i o n o n t o c e l l w a l l E uC i r i c a c i d r e d u c e d t h e s o r p t i o n o f E u o n t h e b a c t e r i a .T h e R E E c o m p l e x a t i o n w i t h o r g a n i c a c i d w o u l d b e i m p o r t a n t f o r e s t i m a t i n g t h e e n v i r o n m e n t a l b e h a v i o r o f R E E s .S u z u k i e t a l .(2005)8A r t h o b a c t e r n i c o t i a n a e A d s o r p t i o n o n t o c e l l w a l l 15k i n d s o f R E E sB o t h s t r a i n c o u l d e f f i c i e n t l y a c c u m u l a t e S m a n d E u .S m w a s s e l e c t i v e l y a c c u m u l a t e d b y A .n i c o t i n a e i n t h e p r e s e n c e o f C u ,M n ,C o ,N i ,Z n ,a n d C d .T s u r u t a (2006)S t r e p t o m y c e s a l b u s9S .a c i d i s c a b i e s W -12A d s o r p t i o n o n t o c e l l w a l l 14k i n d s o f R E E sA l l i n v e s t i g a t e d s t r a i n s d i s p l a y e d t h e s o r p t i o n c h a r a c t e r i s t i c s f o r R E E s .T h r e e b a c t e r i a l s t r a i n c o l l e c t e d f r o m a c i d m i n e d r a i n a g e w a s u s e d f o r t h e a d s o r b e n t o f R E E s .H a f e r b u r g e t a l .(2007)M .l u t e u s W -20B a c i l l u s s p .W -2810B .s u b t i l i s A d s o r p t i o n o n t o c e l l w a l l 14k i n d s o f R E E sR E E d i s t r i b u t i o n p a t t e r n s b e t w e e n b a c t e r i a a n d w a t e r f o r f i v e d i f f e r e n t b a c t e r i a l s t r a i n s e x h i b i t a s t e e p i n c r e a s e i n t h e h e a v y R E E .T h e R E E d i s t r i b u t i o n p a t t e r n s a r e s i m i l a r t o t h o s e o b s e r v e d f o r b i o f i l m s i n t h e B u d o P o n d .T a k a h a s h i e t a l .(2007)E .c o l iA l c a l i g e n e f a e c a l i sS h e w a n e l l a p u t r e f a c i e n sP s e u d o m o n a s f l u o r e s c e n s11P a n t o e a a g g l o m e r a n sA d s o r p t i o n o n t o c e l l w a l lL a ,N d ,S m ,G d ,E r ,Y b E x t e n d e d X -r a y a b s o r p t i o n f i n e s t r u c t u r e s p e c t r o m e t r y (E X A F S )w a s a p p l i e d t o c o m p a r e t h e s o r p t i o n o f R E E s o n b a c t e r i a .S u r f a c e c o m p l e x a t i o n m o d e l i n g c o n s i s t e d o f l i g h t R E E s a d s o r b i n g t o p h o s p h a t e s i t e s a n d t h e m i d d l e a n d h e a v y R E E s a d s o r b i n g t o c a r b o x y l a n d p h o s p h a t e s i t e s .N g w e n y a e t a l .(2009)Appl Microbiol Biotechnolan extensive surface area per unit volume for adsorption of heavy metal ions,such as Pb (Rudd et al.1984).This suggests that the EPS could protect the bacterium in its habitat because these substances facilitate heavy metal fixa-tion surrounding the cell rather than inside the cells (Merroun et al.1998).Merroun et al.(2003)reported that Myxococcus xanthus ,a soil bacterium of the myxobacteria group,could accumulate 0.6mmol of La/g of wet biomass and/or 0.99mmol/g of dry biomass,and a substantial amount of La was fixed in the EPS and in the cell wall.Bacterial oxidation results in the precipitation of biogenic minerals such as silica,iron oxides (Fortin and Langley 2005),and manganese oxide (Tani et al.2003).The adsorp-tion of REEs on biogenic Mn oxide produced by Acremo-nium sp.strain KR21-2has been reported (Tanaka et al.2010).In addition,it was reported that organic molecules,such as siderophore,iminoacetic acid,nitrotriacetic acid,and ethylenediamine tetraacetic acid affect the sorption behavior of REEs on cell surfaces by forming complexes with REEs (Yoshida et al.2004;Takahashi et al.2005).These results indicate that the complexation of REEs with organic mole-cules released from bacterial cells affect the behavior of REEs in the environment.Selective accumulation of REEs using microorganisms In order to separate REEs by use bacteria,the capacity of the bacteria to achieve separation of REEs and of other metals has to be assessed.Several studies have reported the prefer-ential adsorption of REEs over that of other metals.For example,the adsorptive removal of Tm(III)from a Fe(II)and Tm(III)binary solution (20mL,20μg/mL,pH 3)by the cell walls of B .subtilis was studied (Moriwaki et al.2012).The percentage of Tm(III)removed (89%)from the solu-tion was higher than the percentage of Fe(II)removed (5.3%).This finding indicates that the cell wall of the B .subtilis strain can selectively accumulate REEs from a so-lution containing Fe(II).The unique properties of REEs include their strong Lewis acidity and their affinity for heteroatoms such as oxygen.Rare earth ions are known to interact with phosphate groups.Their adsorption onto the oxygen atom of the phosphate group of bacterial cell walls is more powerful than that of Fe(II).This explains the selective adsorption of rare earth ions onto the cell walls.It has also been reported that the relative degree of the metal accumulation by Arthrobacter nicotianae cells was Sm 3+>>Cu 2+,Mn 2+,Co 2+,Ni 2+,Zn 2+,and Cd 2+,indicating that A .nicotianae can accumulate Sm 3+more than the other type of metal ions (Tsuruta 2006).It is very difficult to separate REEs from each other because their physicochemical properties,such as ionic radii,valence,and magnetic properties,are very similarT a b l e 1(c o n t i n u e d )E n t r y M i c r o o r g a n i s m sM e c h a n i s mM e t a l i o n R e s u l t s C o m m e n t s R e f e r e n c e12B .s u b t i l i s A d s o r p t i o n o n t o c e l l w a l l15k i n d s o f R E E sE X AF S d a t a i n d i c a t e d t h a t h e a v y R E E s f o r m e d c o m p l e x e s w i t h m u l t i p l e p h o s p h a t e s i t e a t l o w e r R E E -b a c t e r i a r a t i o s .I t w a s c l e a r t h a t t h e R E E p r i m a r y b o u n d t o t h e p h o s p h a t e s i t e a n d s u b s e q u e n t l y t o t h e c a r b o x y l a t e s i t e o n t h e b a c t e r i a l c e l l s u r f a c e .T a k a h a s h i e t a l .(2010)13A c r e m o n i u m s p .s t r a i n K R 21-2A d s o r p t i o n t o e x t r a c e l l a r b i o m i n e r a l 15k i n d s o f R E E s T h e b i o g e n i c M n o x i d e p r o d u c e d b y t h e b a c t e r i a a d s o r b e d R E E s .O x i d a t i o n o f C e (I I I )t o C e (I V )b y t h e b i o g e n i c M n o x i d e w a s c o n f i r m e d .T a n a k a e t a l .(2010)14B .s u b t i l i s A d s o r p t i o n o n t o c e l l w a l lL a ,E u ,T mR R E s w e r e e f f i c i e n t l y a d s o r b e d o n m i c r o b i a l c e l l w a l l .R E E r e m o v a l (%)o f l i p o t e i c h o i c a c i d -d e f e c t i v e s t r a i n w a s l o w e r t h a n t h a t o f w i l d t y p e .M o r i w a k i e t a l .(2012)Appl Microbiol Biotechnol(Miyawaki and Nakai 1993).At present,the separation of REEs from each other is predominantly achieved by solvent extraction procedures (Fontana and Pietrelli 2009),but the methods generically require large amounts of organic sol-vent.Therefore,adsorbents that exhibit selectivity for a specific REE need to be developed.Bacteria that adsorb REEs selectively may satisfy such a need.The distribution coefficient K d has been frequently used to assess the selectivity of adsorbents for different REEs.The distribution coefficient K d between a liquid and solid phase for [REE],expressed in L/g,is defined by the follow-ing equation:K d ¼REE ½ init ÀREE ½ dis ÀÁc REE ½ disð1Þwhere [REE]dis is the concentration of REEs in the aqueous phase and [REE]init is the initial concentration of each dis-solved REE.The variable c (g/L)is the ratio of the solid to the solution.The REE distribution patterns for the adsorption of the REE onto bacterial cells such as B .subtilis and E .coli have been studied (Takahashi et al.2005).The K d of the REE between the bacterial cell surface and the solution showed a pattern with a prominent enrichment of heavy REEs such as Tm,Yb,and Lu,and included a maximum enrichment around Sm and Eu.Enrichment was also observed around Pr and was accompanied by a decline for Nd.A similar K d pattern for REE adsorption was observed for other bacteria,Alcaligenes faecalis ,Shewanella putrefaciens ,and Pseudomonas fluores-cens (Takahashi et al.2007).Tanaka et al.(2010)reported the sorption of REEs on biogenic Mn oxide produced by Acre-monium sp.strain KR21-2,and a large positive Ce anomaly in the distribution coefficient of the REE between the adsorbent and solution.They concluded that the Ce adsorption was due to Ce oxidation by biogenic Mn oxide.Selectivity in REE adsorption by bacteria affects the geological and environmental distribution of REEs.In fact,Takahashi et al.(2007)reported that the REE patterns from biofilms formed in a groundwater discharge area in the Budo pond in Hiroshima were similar to the pattern obtained from the culture-based experiments.Mechanism of REE adsorption on cell wall of microorganismsIn order to understand how bacteria are able to adsorb REEs selectively,the mechanism by which they adsorb REEs needs to be understood.Many researchers have investigated REE-binding sites on microbial cell walls.Several experi-mental approaches have been used to understand microbial REE-binding sites.Time-resolved laser-induced fluorescence spectroscopy (TRLFS)is a very sensitive,selective,and fast method for the analysis of fluorescent lanthanides that has been exten-sively used to determine the coordination structure of metals adsorbed by microorganisms (Texier et al.2000).A simul-taneous determination of emission wavelength and fluores-cence lifetime provides structural information about fluorescing ions.The Eu(III)ion,which exhibits a strong fluorescence,is particularly suitable for such investigations.The fluorescence lifetime of Eu(III)is related to the number of water molecules in the primary coordination sphere (N H2O ).The determination of N H2O for Eu(III)in a complex is useful to predict the coordination structure in the inner-sphere.The relative intensity of fluorescence spectra (R E/M )serves as a good indicator for examining the symmetry around Eu(III),which is well correlated with the degree of interaction including both the inner and outer spheres (Ozaki et al.2002b ).The coordination states of Eu(III)adsorbed on the B .subtilis and H .salinarum cells were investigated by TRLFS (Ozaki et al.2002a ).The result indicated that H .salinarum exhibited more outer-sphere interaction with Eu(III)than B .subtilis .Furthermore,TRLFS confirmed that Eu(III)complexed with both carbox-ylic and phosphate groups on the cell wall of B .subtilis (Markai et al.2003).Extended X-ray absorption fine structure (EXAFS),resulting from the interference of the photoelectrons scat-tered by the surrounding atoms,provides information about the local structure.Several researchers applied the technique to identify the REE-binding sites on the bacterial cell surfa-ces.The coordination number between the REE and carbox-ylic or phosphate group,REE-O distance,and the influence of water molecules on the interaction between the REE and phosphate can be estimated using EXAFS.Using EXAFS measurement,Ngwenya et al.(2009)reported coordination of lanthanide with gram-negative cells.Their findings indicate a predominance of phosphate binding of REEs at low pH and an increase of REE-carboxylate binding as the pH increased.On a stoichiomet-ric basis,inner-sphere complexation involving REEs was the most likely reaction that led to proton exchange.The REE binding sites at the cell surface of B .subtilis ,which were implicated in the enrichment of heavy REEs on the basis of distribution coefficient studies,were studied using EXAFS (Takahashi et al.2010).The EXAFs data showed that the heavy REEs form complexes with multiple phosphate sites with a large coordination number,whereas at pH 3.5,REEs of light and middle REEs form complexes at the phosphate site that exhibit a lower coordination num-ber.On the basis of these results,the structure and number of phosphate sites of bacteria determine the partitioning of REE between bacteria and the aqueous phase.The results of TRLFS and EXAFS experiments suggest that the REE-binding sites of bacteria are phosphate and carboxyl groups present in the cell wall.Gram-negativeAppl Microbiol Biotechnolbacteria contain phosphate groups at N -acetylglucosamine phosphates,which are structural components of Lipid A in the outer membrane.Macroscopic analysis with X-ray ab-sorption spectroscopic measurements and EXAFS sug-gested that the phosphate sites located on N -acetylglucosamine phosphate are the binding site of REEs (Ngwenya et al.2009).On the other hand,the major structural compounds in the cell wall of gram-positive bacteria are teichoic acids and peptidoglycans (Neuhaus and Baddiley 2003;Schirner et al.2009).Teichoic acids are bacterial polysaccharides that con-sist of glycerol phosphate or ribitol phosphate (Fig.2a ),and they are present in two distinct forms depending on whether they are linked to the head groups of membrane lipids(lipoteichoic acids)or to the peptidoglycan wall (wall tei-choic acids;Fig.2b ).In order to evaluate the distribution of lipoteichoic acid relative to the REE adsorption of B .subtilis cell wall,the REE adsorption of the freeze-dried cell powder of a wild type (WT powder)and lipoteichoic acid-defective strain (ΔLTA powder)were compared (Moriwaki et al.2012).It was found that the percentage removal of La,Eu,and Tm ions by using WT powder from water was greater than that by ΔLTA powder (Fig.3).This result indicates that lipoteichoic acid contributes to the adsorption of REEs by the cell wall of B .subtilis .Conclusions and perspectivesVarious bacteria have been studied for their use as REE adsorbents.Such microbes have excellent capacity and se-lectivity for REE adsorption.The REE partitioning between the bacterial cell wall and the aqueous solution frequently exhibits anomalous enrichment in the heavy REE part.Furthermore,several studies have shown that microorgan-isms that were used as adsorbent materials selectively adsorbed REEs from mixtures of REE and other metal ing TRLFS,EXAFS,and genetically modified strains,the REE adsorption sites were studied in considerable detail,and the adsorption mechanisms were elucidated.The bind-ing sites of the bacterial cell wall are phosphate and carbox-yl groups,and it is strongly suggested that the interaction between these sites and REEs determine the selectivity of the microbial materials.However,many problems remain when bacteria are used to remove or collect REEs from the environment.At this stage,it is difficult to use bacteria as REE adsorbents be-cause of the high costs.Desorption of REEs from microbial adsorbents needs to be developed for recovery of REEs as a resource.Desorption of REEs from soil has been actively studied (Shan et al.2002).Establishing an elution method by exploiting these techniques would make it possible to reuse the microbial adsorbent.Wall teichoic acid (WTA)b)OP OP O-O-Teichoic acidFig.2a Molecular structure of teichoic acid.b Schematic structure of the cell wall of gram-positive bacteriaWild type LTA∇Fig.3Removal (percentage)of La,Eu,and Tm ions from water using Bacillus subtilis powders (initial REE concen-tration,20μg/mL;pH 03;ad-sorbent loading:20mg/20mL)Appl Microbiol BiotechnolB.subtilis168is the gram-positive model organism and its genome databases are quite substantial.In addition,the bio-synthetic pathways of both wall-and lipo-teichoic acids are well understood(Lazarevic et al.2002;Schirner et al.2009; Wörmann et al.2011;Reichmann and Gründling2011).Gene inactivation procedure is fully established in this organism. Therefore,in order to improve REE adsorption,approaches using genetic modification would be possible.For example, the freeze-dried cell powder of the lipoteichoic acid-defective strain(LTA powder)has an advantage for REE removal compared with the wild-type strain.TheΔLTA powder readily coagulated and sedimented in the presence of REEs.On the other hand,such coagulation and sedimentation did not occur with the WT powder and rare earth ion aqueous solution under the same conditions(Moriwaki et al.2012).The length of time required for the filtration of the test solution(20mL)by using 5A filtration paper were105and43s for the WT powder and ΔLTA powder,respectively.This result suggests thatΔLTA powder is useful for the extraction of rare earth ions from water.It will be possible to construct the mutant strains,which have a high selectivity or unique adsorption behavior and to use them in environmental applications in the future,when the mechanistic theory of the REE adsorption on the micro-bial cells is advanced further.Additional work is required to develop a better application for bacteria as REE adsorbents for separation and in the environment.Acknowledgments A part of this work was supported by Grants-in-Aid for Scientific Research(C;19580085and23580107)and grants from the Kurata Memorial Hitachi Science and Technology Founda-tion,the Nagase Science and Technology Foundation,the Research Foundation for the Electrotechnology of Chubu,and Institute for Fermentation,Osaka to H.Y.ReferencesAnderson CR,Pederson K(2003)In situ growth of Gallionella bio-films and partitioning of lanthanides and actinides between bio-logical material and ferric oxyhydroxides.Geobiol1:169–178 Andres Y,Texier AC,Cloirec PL(2003)Rare earth elements removal by microbial biosorption:a review.Environ Technol24:1367–1375Andrianov A V,Savel’eva OA,Bauer E,Staunton JB(2011)Squeezing the crystalline lattice of the heavy rare earth metals to change their magnetic order:experiment and ab initio 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微生物学【英文版】(全套课件499P)
Concepts
Microorganisms are responsible for many of the changes observed in organic and inorganic matter (e.g., fermentation and the carbon, nitrogen and sulfur cycles that occurred in nature.
Microbiology is a large discipline, which has a
1.1 What is a microbe?
The word microbe (microorganism) is used to describe an organism that is so small that can not be seen without the use of a microscope. Viruses, bacteria, fungi, protozoa and some algae are all included in this category.
Microbial world
Organisms
(living)
Infectious agents (non-living) viruses viroids prions
Prokaryotes (unicellular)
eukaryotes
Eubacteria
Archaea
Algae (unicellular or multicellular)
Chapter 1
Microorganisms and Microbiology
pter outline
1.1 What is a microbe? 1.2 The importance of Microbiology 1.3 Microbes in our lives 1.4 The history of microbiology 1.5 Important events in the development of microbiology
肠道微生物英文文献pig
Animal(2012),6:10,pp1620–1626&The Animal Consortium2012doi:10.1017/S1751731112000481The effect of chitooligosaccharide supplementation on intestinal morphology,selected microbial populations,volatile fatty acid concentrations and immune gene expression in the weaned pig A.M.Walsh,T.Sweeney,B.Bahar,B.Flynn and J.V.O’Doherty-School of Agriculture,Food Science and Veterinary Medicine,University College Dublin,Lyons Research Farm,Newcastle,Co.Dublin,Ireland(Received24March2011;Accepted29January2012;First published online2March2012)An experiment(complete randomised design)was conducted to investigate the effects of supplementing different molecular weights (MW)of chitooligosaccharide(COS)on intestinal morphology,selected microbial populations,volatile fatty acid(VFA)concentrations and the immune status of the weaned pig.A total of28piglets(24days of age,9.1kg(6s.d.0.80)live weight)were assignedto one of four dietary treatments for8days and then sacrificed.The treatments were(1)control diet(0ppm COS),(2)control diet plus5to10kDa COS,(3)control diet plus10to50kDa COS and(4)control diet plus50to100kDa COS.The COS was included in dietary treatments at a rate of250mg/kg.Tissue samples were taken from the duodenum,jejunum and ileum for morphological measurements.Digesta samples were taken from the proximal colon to measure lactobacilli and Escherichia coli populations and digesta samples were taken from the caecum and proximal colon for VFA analysis.Gene expression levels for specific cytokines were investigated in colonic tissue of the pig.Supplementation of different MW of COS had no significant effect on pig performance during the post-weaning period(days0to8;P.0.05).The inclusion of COS at all MW in the diet significantly reduced faecal scores compared with the control treatment(P,0.01).Pigs fed the10to50kDa COS had a higher villous height(P,0.05)and villous height:crypt depth ratio(P,0.05)in the duodenum and the jejunum compared with the control treatment.Pigs fed the5to10kDa COS had a lower lactobacilli population(P,0.05)and E.coli population(P,0.05)in the colon compared with the control group.Pigs offered the5to10kDa COS had significantly lower levels of acetic acid and valeric acid compared with the control group(P,0.05). The inclusion of different MW of COS had no significant effect on the expression of the cytokines tumour necrosis factor-a,Interleukin (IL)-6,IL-8and IL-10in the gastro-intestinal tract of the weaned pig.The current results indicate that a lower MW of5to10kDa COS possessed an antibacterial activity,while the higher MW of10to50kDa was optimum for enhancing the intestinal structure. Keywords:chitooligosaccharide,pig,microbiology,intestinal morphologyImplicationOur results indicate that the inclusion of chitooligosaccharides (COSs)in piglet diets may moderate several gut health para-meters that contribute to some of the common problems that occur after weaning in the absence of in-feed antibiotics.It was observed that COSs with a molecular weight(MW)of5to 10kDa were more effective in reducing Escherichia coli populations while a MW of10to50kDa enhanced the intestinal structure.IntroductionThe weaning period imposes profound social and environ-mental stresses on the piglet such as removal from the sow,change in diet and mixing of piglets from different litters. Numerous studies have reported that there is a reduction in villous height(villous atrophy)and an increase in crypt depth (crypt hyperplasia)after weaning,which leads to increased susceptibility to intestinal gut dysfunction(Spreeuwenberg et al.,2001;Pierce et al.,2006).The post-weaning period is characterised by a reduction in feed intake,poor growth rates,diarrhoea and an increased risk of disease(Lalles et al., 2007).These negative effects on piglet growth during the weaning period were managed by growth-promoting anti-biotics.However,the European Union placed a total ban on the use of in-feed antibiotic growth promoters on the1st January2006due to public concerns regarding bacterial resistant and human health issues. Chitooligosaccharides(COS)may be a potential viable alternative to traditional antimicrobials in animal production.-E-mail:john.vodoherty@ucd.ie 1620Chitosan is a natural biopolymer derived by alkaline deacety-lation of chitin,which is the principal component of protective cuticles of crustaceans such as crabs,shrimps,prawns,lobsters and cell walls of some fungi such as aspergillus(Qin et al., 2006).Both chitin and chitosan are biopolymers composed of glucosamine and N-acetylated glucosamine(2-acetylamino-2-deoxy-D-glucopyranose)units linked by b(1to4)glycosidic bonds(Koide,1998).Low molecular weight(MW)COS is a water-soluble derivative of chitosan due to shorter chain lengths(Kim and Rajapakse,2005).Recently,both chitosan and its derivatives have generated considerable interest due to their biological activities,including antimicrobial,antitumour, immunoenhancing effects and the acceleration of wound healing(No et al.,2002;Liu et al.,2006)There is considerable variation in the literature on the biological properties of COS (Jeon et al.,2001;Liu et al.,2006).Most of this variation is partly due to the widely different MW used across studies.It is hypothesised that the biological properties of COS may be influenced by its MW and COS will enhance selected indices of health in weaned piglets.Material and methodsAll procedures described in this experiment were conducted under an experimental licence from the Irish Department of Health in accordance with the cruelty to Animals Act1876 and the European Communities(Amendments of the Cruelty to Animals Act1976)Regulations.Experimental dietsThe experiment was designed as a complete randomised block design and comprised four dietary treatments.Thedietary treatments were as follows:(1)control diet(0ppm COS),(2)control diet plus5to10kDa COS,(3)control diet plus10to50kDa COS and(4)control diet plus50to100kDa COS.The COS was sourced from Kitto Life Co.Ltd(Kyungki-do,Seoul,Korea)and was supplemented in the experimental diets at a concentration of250ppm.The diets were fed for 8days ad libitium,after which time the pigs were humanely sacrificed.The diets were formulated to have similar diges-tible energy(16MJ/kg)and standardised ileal digestible (SID)lysine(14g/kg)contents.All amino acids requirements were met relative to SID lysine(National Research Council, 1998).The ingredient composition and chemical analysis of the dietary treatments are presented in Table1.Animals and managementA total of28piglets(progeny of large white3(large white3landrace sows))were selected from a commercial pig unit at24days of age.The piglets had a weaning weight of9.1kg(s.d.50.80)and were blocked on the basis of litter of origin and live weight(n57).The piglets were individu-ally housed in fully slated pens(1.7m31.2m).They were individually fed and had ad libitum access to feed and water. The house temperature was thermostatically controlled at 308C throughout the experiment.This study was not a growth performance study but some performance data were recorded.The piglets were weighed at the beginning of the experiment(day0)and at the end of the experiment(day8). Food was available up to thefinal weighing and all remaining food was weighed back for the purpose of cal-culating feed efficiency.Pigs were observed for clinical signs of diarrhoea and a scoring system was applied to indicate the presence and severity of this as described by Pierce et al. (2006).Faeces scores were assigned daily for individual pigs from day0and continued until day8.The following faeces scoring system was used:15hard faeces,25slightly soft faeces in the pen,35soft,partially formed faeces,45loose, semi-liquid faeces and55watery,mucous-like faeces.Gut morphological analysisThe piglets were humanely sacrificed on day8by a lethal injection of Euthatal(pentobarbitone sodium BP–Merial Animal Ltd,Sandringham House,Essex,UK)at a rate of1ml/ 1.4kg BW.On removal of the digestive tract,sections of the duodenum(10cm from the stomach),the jejunum(60cm from stomach)and the ileum(15cm from caecum)were excised andfixed in10%phosphate-buffered formalin.The preserved segments were prepared using standard paraffin-embedding techniques.The samples were sectioned at5m m Table1Composition and chemical analysis of experimental diets (as-fed basis)Items Starter diet* Ingredient(g/kg)Whey permeate125.0 Wheat444.2 Soya bean meal142.5 Whey protein isolate130.0 Full-fat soybean80.0 Soya oil65.0 Vitamins and minerals 5.0 Lysine HCL 4.5 DL-methionine 1.6L-threonine 2.2 Analysis(g/kg,unless otherwise stated)DM892.5 CP(N36.25)224.2 GE(MJ/kg)18.2 Ash43.7 NDF110.3 Lysine-16.5 Methionine and cysteine-9.9 Threonine-10.7 Tryptophan- 2.5 Calcium-8.0 Phosphorous- 6.0 DM5dry matter;GE5gross energy.Starter diet provided(mg/kg completed diet):Cu,175;Fe,140;Mn,47;Zn, 120;I,0.6;Se,0.3;retinol,1.8;cholecalciferol,0.025;alpha-tocopherol,67; phytylmenaquinone,4;cyanocobalamin,0.01;riboflavin,2;nicotinic acid,12; pantothenic acid,10;choline chloride,250;thiamine,2;pyridoxine,0.015.*COS was included in dietary treatments T2–T4at a rate of250mg/kg.-Calculated for tabulated nutritional composition(Sauvant et al.,2004).Chitooligosaccharide in piglet diets1621thickness and stained with haemotoxylin and eosin(Pierce et al.,2006).Villous height and crypt depth were measured on the stained sections(43objective)using a light micro-scopefitted with an image analyser(Image Pro Plus,Media Cybernetics,Buckinghamshire,UK).Measurements of15well oriented and intact villi and crypts were taken for each seg-ment.Villous height was measured from the crypt–villous junction to the tip.Crypt depth was measured from the crypt–villous junction to the base.Results were expressed as the mean villous height or crypt depth in micrometres. Intestinal microfloraFor microbial analysis,digesta samples(,1061g)were aseptically recovered from the proximal colon of each pig immediately post slaughter.Digesta samples were stored in sterile containers(Sarstedt,Wexford,Ireland),placed on ice and transported to the laboratory within2h.A1.0g sample was removed from the digesta sample,serially diluted (1:10)in9.0ml aliquots of maximum recovery diluents (Oxoid,Basingstoke,UK)and spread plated(0.1ml aliquots) onto selective agars,as follows:Lactobacillus spp.were isolated on de Man,Rogosa and Sharp(MRS)agar(Oxoid) with an overnight(18to24h)incubation at378C in an atmosphere enriched with5%CO2,as recommended by the manufacturers(Oxoid).The Escherichia coli species were isolated on MacConkey agar(Oxoid)following aerobic incubation at378C for18to24h(O’Doherty et al.,2010). Target colonies of Lactobacilli and E.coli were identified by Gram stains and colony morphology(Salanitro et al.,1977). The API50CHL(BioMerieux,Biomerieux,Craponne,France) kit was used to confirm suspect Lactobacilli spp.Suspect E. coli colonies were confirmed with API20E(BioMerieux, France).This API system identifies the suspect colonies by measuring their ability to produce cytochrome oxidase. Typical colonies of each bacteria on each agar were counted, log transformed and the numbers of bacteria were expressed per gram of digesta after being serially diluted.Volatile fatty acid(VFA)analysisSamples of digesta from individual pigs were taken from the caecum and the proximal colon to measure the VFA concentration and molar proportions of VFAs.The VFA con-centrations in the digesta were determined using gas liquid chromatography according to the method described by Pierce et al.(2007).A1-g sample was diluted with distilled water (2.53weight of sample)and centrifuged at14003g for4min(Sorvall GLC–2B laboratory centrifuge,Dupont, Wilmington,DE,USA).Then,1ml of the subsequent super-natant and1m l of internal standard(0.5g3-methyl-n-valeric acid in1l of0.15mol/l oxalic acid)were mixed with3ml of distilled water.Following centrifugation to remove the precipitate,the sample wasfiltered through Whatman 0.45m m polyethersulphone membranefilters into a chromato-graphic sample vial.A1-m l sample was injected into a model 3800Varian gas chromatograph with a25m30.53mm i.d. megabore column(coating CP-Wax58(FFAP)–CB(no. CP7614))(Varian,Middelburg,the Netherlands).RNA extraction and complementary DNA(cDNA)synthesis Tissue samples were collected from the mesenteric side of the colon,rinsed with ice-cold sterile phosphate-buffered saline(Oxoid)and stripped of overlying smooth muscle cells. Approximately1to2g of the porcine colon tissue was cut into small pieces and placed in tubes containing15ml of RNAlater(Applied Biosystems,Foster City,CA,USA)and immediately stored at2208C pending RNA extraction.Total RNA was extracted from colon tissue samples(25mg)using a GenElute Mammalian Total RNA Miniprep Kit(RTN70, Sigma-Aldrich,St Louis,MO,USA)according to the manu-facturer’s instructions.To eliminate possible genomic DNA contamination,total RNA samples were subjected to DNAse I(AMPD1,Sigma-Aldrich)treatment according to the man-ufacturer’s protocol.Then RNA purification was performed using a phenol–chloroform extraction method(Chomczynski and Sacchi,2006).The total RNA was quantified using a NanoDrop-ND1000Spectrophotometer(Thermo Fisher Scien-tific,Wilmington,DE,USA)and the purity was assessed by determining the ratio of the absorbance at260and280nm. All total RNA samples had260/280nm ratios above1.8.In addition,RNA integrity was verified by visualisation of the18 and28S ribosomal RNA bands stained with ethidium bromide after gel electrophoresis on1.2%agarose gels(Egel,Invitro-gen Inc.,Carlsbad,CA,USA).Total RNA(1m g)was reverse transcribed(RT)using the RevertAid H minusfirst strand cDNA synthesis kit(Fermentas GmbH,St Leon-Rot,Germany)with oligo dT primers.Thefinal RT product was adjusted to a volume of120m l using nuclease-free water.Real-time quantitative PCRAll primers for the selected cytokines,genes such as Inter-leukin-1a(IL-1a),IL-6,IL-10,tumour necrosis factor(TNF-a) and the reference genes b-actin(ACTB),b2-microglobin (B2M),glyceraldehyde-3-phosphate dehydrogenase(GAPDH) and peptidylprolyl isomerise A(PPIA)are presented in Table2. Amplification was carried out in a reaction volume of20m l containing10m l SYBR Green Fast PCR Mastermix(Applied Biosystem),forward and reverse primer mix(1m l),8m l DEPC treated water and1m l of template cDNA.Quantitative real-time PCR was carried out using an ABI PRISM7500Fast sequence detection system for96-well plates(Applied Biosys-tem).The thermal cycling conditions were as follows:an initial denaturation step at958C for10min,40cycles of958C for15s, followed by608C for1min.Dissociation analyses of the PCR product were performed to confirm the specificity of the resulting PCR products.All samples were run in triplicate.The cycle threshold value(C t)is defined as the fractional cycle number at whichfluorescence passes thefixed threshold.The mean C t values of triplicates of each sample were used for calculations.Normalisation of quantitative PCR dataNormalisation of the C t values obtained from real-time PCR was performed by(i)transforming the raw C t values into relative quantities using the formula,relative quantities5 (PCR efficiency)D C t,where D C t is the change in the C t valuesWalsh,Sweeney,Bahar,Flynn and O’Doherty 1622of the sample relative to the highest expression (minimum C t value),(ii)using geNorm,a normalisation factor was obtained from the relative quantities of four most stable housekeeping genes (GAPDH,B2M,ACTB and PPIA)and (iii)the normalised fold change or the relative abundance of each of the target genes was calculated by dividing their relative quantities by the normalisation factor.Statistical analysisThe experimental data were analysed as a randomised block design using the GLM procedure of SAS (2004).The individualpig served as the experimental unit.Food intake was inclu-ded as a covariate in the model for villous height,crypt depth and villous height to crypt depth ratio in the digestive tract.The microbial counts were log transformed.The data were checked for normality using the Proc Univariate function of SAS.The means were separated using the Tukey–Kramer Test.Probability values of ,0.05were used as the criterion of statistical significance.All results are presented in the tables as least square means 6standard error of the means (s.e.).ResultsPerformance and faecal scoringThe average faecal scores of the pigs are presented in Table 3.The supplementation of different MW of COS had no significant effect on the growth performance of the pig during the 8-day experimental period (P .0.05).However,the inclusion of COS at all MW in the diet significantly reduced faecal scores com-pared with the control treatment (P ,0.01).MicrobiologyThe effect of COS supplementation at different MW on selected microbial populations in the colon of the pig is shown in Table 3.Pigs offered diets containing 5to 10kDa COS had a lower E.coli number compared with the control (P ,0.05)and the 50to 100kDa COS (P ,0.05)treatments.The 10to 50kDa treatment had a neumerically lower E.coli number compared with the control group (P 50.09).Pigs offered diets containing 5to 10kDa COS had a significantly lower population of lacto-bacilli in the colon compared with the control group (P ,0.05)and the 50to 100kDa COS diet (P ,0.01).Pigs offered 50to 100kDa COS had a higher lactobacilli number than pigs offered 10to 50kDa COS (P ,0.05).The supplementation of different MW of COS had no significant dietary effect on the lacto-bacilli :E.coli ratio in the colon of the pig.Table 2Porcine-specific primers used for real-time PCR 1.Forward primer sequence (50-30)Gene 2.Reverse primer sequence (50-30)T m (8C)IL-6 1.AGACAAAGCCACCACCCCTAA59.82.CTCGTTCTGTGACTGCAGCAGCTTATC 62.7IL-8 1.TGCACTTACTCTTGCCAGAGAACTG 61.92.CAAACTGGCTGTTGCCTTCTT 61.7IL-10 1.GCCTTCGGCCCAGTGAA 57.62.AGAGACCCGGTCAGCAACAA 59.4TNF-a 1.TGGCCCCTTGAGCATCA55.22.CGGGCTTATCTGAGGTTTGAGA 60.3GAPDH 1.CAGCAATGCCTCCTGTACCA 62.22.ACGATGCCGAAGTTGTCATG 62.1B2M 1.CGGAAAGCCAAATTACCTGAAC 59.02.TCTCCCCGTTTTTCAGCAAAT 60.0ACTB 1.CAAATGCTTCTAGGCGGACTGT 59.02.TCTCATTTTCTGCGCAAGTTAGG 60.0PPIA1.CGGGTCCTGGCATCTTGT58.02.TGGCAGTGCAAATGAAAAACTG56.5IL 5interleukin;TNF 5tumour necrosis factor;GAPDH 5glyceraldehyde-3-phosphate dehydrogenase;B2M 5b 2-microglobin;ACTB 5genes b -actin;PPIA 5peptidylprolyl isomerise A.Primers were designed using Primer Express TM software and were synthesisedby MWG Biotech (Milton Keynes,UK).Table 3Effect of COS supplementation at different MW on faecal scoring,selected microbial populations in the proximal colon and the total VFA concentration and the proportions of VFAs in the caecum of the weaned pig (least square means and s.e.;n 57)Dietary treatmentsControl 5to 10kDa 10to 50kDa50to 100kDas.e.SignificanceFaeces scoring Days 0to 84.06b 3.31a 3.44a 3.38a 0.124**Proximal colonic bacterial population (log cfu/g of digesta)Escherichia coli5.94b 4.34a 4.71a 5.81b 0.477*Lactobacilli spp.7.39bc 6.24a 6.56ab 7.56c 0.347*VFA concentrations in the caecum Total VFA (mmol/g of digesta)95.8770.25103.00116.7913.006ns Acetic acid 67.36b 45.77a 70.30b 79.20b 8.543*Propionic acid 19.8416.5123.3927.05 3.616ns Isobutyric acid 0.770.490.810.790.157ns Butyric acid 6.45 6.14 6.817.51 1.575ns Isovaleric acid 0.67b 0.45a 0.68b 0.83b 0.092*Valeric acid0.790.881.011.410.282nsCOS 5chitooligosaccharide;MW 5molecular weight;VFA 5volatile fatty acid.Probability of significance:*P ,0.05;**P ,0.01;ns,P ,0.05.Means with the same superscript alphabets within rows are not significantly different (P .0.05).Chitooligosaccharide in piglet diets1623Volatile fatty acidsThe effects of COS supplementation at different MW on the VFA concentrations in the caecum are shown in Table3.The supplementation of different MW of COS had a significant effect on the concentrations of acetic acid(P,0.05)and isovaleric acid(P,0.05)in the caecum.Pigs fed5to10kDa COS had lower levels of acetic acid and isovaleric acid compared with the control(P,0.05),10to50kDa COS (P,0.05)and50to100kDa COS(P,005).There was no significant effect of MW on VFA concentrations(P.0.05)in the proximal colon(data not shown).Gut morphologyThe effects of varying COS MW on villous height,crypt depth and the villous height:crypt depth ratios in the gastro-intestinal tract are shown in Table4.Pigs fed the10to 50kDa COS had a higher villous height in the duodenum and the jejunum compared with the control group(P,0.05), 5to10kDa COS(P,0.01)and50to100kDa COS diets (P,0.05).There was no effect of dietary treatment on crypt depth in the duodenum(P.0.05).Pigs offered the10to 50kDa COS had a higher villous height:crypt ratio in the duodenum and the jejunum compared with the control group(P,0.05)and the5to10kDa COS diet(P,0.01).Cytokine gene expression analysisThe effects of COS supplementation on the immune response in colon tissues of the pig are shown in Table5.The supplementation of different MW of COS had no significant effect on the expression of the cytokines TNF-a,IL-6,IL-8 and IL-10(P.0.05)in the gastro-intestinal tract of the pig. DiscussionThe hypothesis of the current experiment is that the biolo-gical properties of COS may be influenced by its MW and COS will enhance selected indices of health in weaned pig-lets.It was demonstrated in the current study that the lower MW of5to10kDa possessed antibacterial activity while the higher MW of10to50kDa was optimum for enhancing intestinal structure.Dietary supplementation of COS at the low MW of5to 10kDa decreased both lactobacilli and E.coli counts,while the10to50kDa COS numerically decreased E.coli popula-tions in the colon of the pig.In a study by Liu et al.(2008), COS supplementation at different concentrations reduced E. coli concentrations in the caecum of the weanling pig.E.coli is considered to be one of the most important causes of post-weaning diarrhoea in weaned pigs;therefore,a reduction inTable4Effect of COS supplementation at different MW on villous height,crypt depth and the villous height:crypt depth ratio in the gastro-intestinal tract of the weaned pig(least square means and s.e.)Dietary treatments Control5to10kDa10to50kDa50to100kDa s.e.Significance Covariate(intake) Villous height(m m)Duodenum284.0a256.0a326.3b266.2a17.38*ns Jejunum271.6a270.7a316.5b260.8a16.15*ns Ileum239.8268.3251.5242.915.07ns ns Crypt depth(m m)Duodenum305.7330.2280.1311.718.98ns ns Jejunum294.1298.4281.6268.420.93ns ns Ileum207.5242.8228.2239.811.29ns ns Villous:crypt depth ratioDuodenum 1.0a0.8a 1.2b0.9a0.08*ns Jejunum0.9a0.9a 1.2b 1.0ab0.06*ns Ileum 1.2 1.1 1.1 1.00.06ns nsCOS5chitooligosaccharide;MW5molecular weight.Probability of significance:*P,0.05;**P,0.01;ns,P,0.05.Means with the same superscript alphabets within rows are not significantly different(P.0.05).Table5Effect of COS supplementation at different MW on the immune response in unchallenged proximal colon tissues(leastsquare means of fold change in normalised relative gene expression with their s.e.;n57animals)Dietary treatments Control5to10kDa10to50kDa50to100kDa s.e.SignificanceColonTNF-a0.3660.3530.3760.3620.0568nsIL-60.2480.3590.3180.3220.0645nsIL-80.3850.5440.3700.3580.0797nsIL-100.3640.3420.3110.3070.0614ns COS5chitooligosaccharide;MW5molecular weight;TNF5tumour necrosis factor;IL5interleukin.Probability of significance:*P,0.05;**P,0.01;ns,P,0.05.Walsh,Sweeney,Bahar,Flynn and O’Doherty1624E.coli populations may reduce the incidence of diarrhoea in post-weaned pigs(Fairbrother et al.,2005).Although many species of E.coli are commensal,high levels of specific E.coli (like ETEC)will increase the risk of disease.Unfortunately, ETEC numbers were not measured in the current study.In the current study,the faecal score was decreased in pigs fed the COS diets compared with the control.These results suggest that the supplementation of the5to10kDa and10 to50kDa COS reduces E.coli populations in the colon, resulting in a lower faecal score in the post-weaning period. The50to100kDa COS led to a reduced diarrhoea score but no reduction in E.coli populations;therefore,this MW of COS may be working as a bulking agent to affect the faecal score.The50to100kDa COS may retard the rate of passage through the intestine and may have the ability to absorb water.In the current study,it was demonstrated that supple-mentation of5to10kDa COS had the strongest antimicrobial effect against both lactobacilli and E.coli.This is in agreement with other studies in which low MW COS(5to10kDa)were shown to possess strong antibacterial properties compared with higher MW COS and the antibacterial properties of COS increased at a low MW of,5kDa against Gram-negative such as E.coli(Zheng and Zhu,2003;Kittur et al.,2005).In a study by Liu et al.(2010),COS supplementation decreased E.coli populations compared with the control in the caecum of weaned pigs,while Jeon et al.(2001)observed a anti-microbial effect of COS against Gram-positive bacteria such as Lactobacilli under in-vitro conditions.To explain COS antibacterial activity,two mechanisms have been proposed.Thefirst mechanism is that the posi-tively charged COS reacts with negatively charged molecules at the microbial cell surface,thereby altering cell perme-ability(Chung and Chen,2008).Therefore,COS may interact with the membrane of the cell to alter cell permeability. However,as evident from the current study,this activity may differ with varying MW as the50to100kDa group had no inhibitive effect on the selected microbial populations,while the MWs of5to10kDa and10to50kDa COS had the strongest inhibitive effect.The other antibacterial mechan-ism is the binding of COS with DNA to inhibit RNA synthesis (Liu et al.,2004).It has been proposed that COS penetrates the nuclei of the bacteria and interferes with RNA and pro-tein synthesis.It is noteworthy that all the COS samples used in the current study were soluble in aqueous solutions.Kim and Rajapakse(2005)found that COS with a MW of .30kDa were not effective as antibacterial agents due to their poor solubility in aqueous solutions at a neutral pH. Volatile fatty acids are the major end products of bacterial metabolism in the large intestine(Macfarlane and Macfarlane, 2003).Both protein and carbohydrate fermentation contribute to the production of acetic acid;however,branched-chain fatty acids such as isovaleric acid are produced from protein fermentation(Mackie et al.,1998).In the current study,the5 to10kDa group had the lowest selected microbial populations while also reducing isovaleric acid and acetic acid concen-trations in the caecum.The shift in the production of the fermentation end products is reflected in the reduction of the selected microbial populations.The quantity of VFA produced depends on the amount and composition of the substrate and on the type of microbes present in the large intestine (van Beers-Schreurs et al.,1998).Reduced VFA concentrations indicate that lower amounts of substrate were fermented as a result of a lower microbial activity in the caecum(Htoo et al.,2007).Villous height is generally reduced and crypt depth is increased,which may explain the increased occurrence of diarrhoea and reduced growth after weaning(Pluske et al., 1996).The inclusion of10to50kDa COS in the present study was found to increase the villous height and villous:crypt depth ratio in the duodenum and also in the jejunum com-pared with the control group.Very little data have been published on the effects of COS MW on gut morphology in weaned piglets;thus,the exact mechanism for the increase in villous height and villous:crypt depth ratio is unclear.It may be hypothesised that low MW COS has the potential to promote intestinal morphology through cell proliferation. The COS has been shown to influence colonic cell prolifera-tion,crypt depth and crypt circumference in mice(Torzsas et al.,1996).A study carried out by Liu et al.(2008),on different con-centrations of COS,demonstrated that200mg/kg of COS increased villous height and villous:crypt ratio in the jeju-num and ileum(Liu et al.,2008).The possible explanation for this improved intestinal structure was that COS is com-posed of N-acetyl glucosamine(Kim and Rajapakse,2005), which may bind to certain types of bacteria and possibly interfere with their adhesion to the gut tissue of host animals (Ofek et al.,2003;Liu et al.,2008).This result is in agree-ment with Moura˜o et al.(2006),who reported that an increase in villi length in the ileum of weaned rabbits was correlated to a lower intestinal microflora.A decrease in bacteria load has been shown to increase the proliferation of epithelial cells,which leads to an improved intestinal mor-phology and increased villous height(Moura˜o et al.,2006). In the present study,in pigs fed the lower MW of5to10kDa COS,a strong antimicrobial effect on both Lactobacilli and E.coli populations was observed,with no effect on villous structure,while the higher MW of10to50kDa resulted in a reduction in E.coli numbers in comparison with the control and was optimum for improving villous integrity.There were no effects of COS supplementation in colon tissue on any of the cytokines analysed.This overall lack of an effect on these inflammatory cytokines implies that COS inclusion in the diet had no effects on immune gene expression of the pigs.Mori et al.(1997)also demonstrated that chitin and its derivatives do not stimulate the production of IL-6,IL-1and TNF-a byfibroblasts.In our study,no dif-ferences were observed on growth performance between days0and8post-weaning.In conclusion,MW is an important factor to consider when investigating the biological properties of COS.On the basis of the current study,the lower MW of5to10kDa possessed antibacterial activity while the higher MW of10to50kDaChitooligosaccharide in piglet diets1625。
(英文)生物外文文献
FIG. 1
15.5%
FIG. 2
GRN knockdown reduced the mRNA expression of these genes, similar to the effects of STAT3 knockdown
58%
染色质免疫共沉淀技术（ chromatin immunoprecipitation assay, CHIP ）
• 皮尔森相关系数（Pearson correlation coefficient）也称皮尔森积矩 相关系数(Pearson product-moment correlation coefficient) ，是一种 线性相关系数。皮尔森相关系数是用来反映两个变量线性相关程 度的统计量。相关系数用r表示，其中n为样本量，分别为两个变 量的观测值和均值。r描述的是两个变量间线性相关强弱的程度。 r的绝对值越大表明相关性越强。
2.2倍
Indicate that in primary breast cancers, GRN expression specifically correlates with enhanced STAT3 transcriptional activity in the presence of tyrosine-phosphorylated STAT3
5
Suggesting that inhibiting GRN preferentially affects cells with activated STAT3 These findings suggest that the effect of granulin depletion is mediated by decreased STAT3 transcriptional activity.
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11、战争满足了，或曾经满足过人的 好斗的 本能， 但它同 时还满 足了人 对掠夺 ，破坏 以及残 酷的纪 律和专 制力的 欲望。 ——查·埃利奥 特 12、不应把纪律仅仅看成教育的手段 。纪律 是教育 过程的 结果， 首先是 学生集 体表现 在一切 生活领 域—— 生产、 日常生 活、学 校、文 化等领 域中努 力的结 果。— —马卡 连柯(名 言网)
13、遵守纪律的风气的培养，只有领 导者本 身在这 方面以 身作则 才能收 到成效 。—— 马卡连 柯 14、劳动者的组织性、纪律性、坚毅 精神以 及同全 世界劳 动者的 团结一 致，是 取得最 后胜利 的保证 。—— 列宁 摘自名言网
15、机会是不守纪律的。——雨果
61、奢侈是舒适的，否则就不是奢侈 。——CocoCha nel 62、少而好学，如日出之阳；壮而好学 ，如日 中之光 ；志而 好学， 如炳烛 之光。 ——刘 向 63、三军可夺帅也，匹夫不可夺志也。 ——孔 丘 64、人生就是学校。在那里，与其说好 的教师 是幸福 ，不如 说好的 教师是 不幸。 ——海 贝尔 65、接受挑战，就可以享受胜利的喜悦 。——杰纳勒 尔·乔治·S·巴顿