Detection of Phaeocystis globosa using sandwich hybridization integrated with nuclease protection assay (NPA-SH)

Phaeocystis globosa Scherffel is one of the common harmful algae species in coastal waters of the southeastern China. In this study, sandwich hybridization integrated with nuclease protection assay (NPA-SH) was used to qualitatively and quantitatively detect P. globosa. Results showed that this method had good applicability and validity in analyzing the samples from laboratory cultures and from fields. The linear regression equation for P. globosa was obtained, and the lowest detection number of cells was 1.8×104 cells. Statistics showed that there was no distinct difference between the results of detecting the microalgae by NPA-SH and traditional microscopy. This technique has good reliability, accuracy, and can give a remarkably high sample processing rate. Sandwich hybridization integrated with nuclease protection assay will provide an efficient alternative to microscopic method for monitoring and investigating the bloom of P. globosa.     

应用电致化学发光分子探针技术对微小原甲藻的检测

以双特异分子探针技术(NPA-SH)为基础,运用电致化学发光技术(ECL)和磁性微球分选技术对其进行改进,成功建立了用于赤潮藻类定性定量分析的电致化学发光分子探针(ECL-MP)检测新技术.设计微小原甲藻特异性NPA探针并对其进行联吡啶钌和生物素标记,优化磁性微球使用量,在ECL检测装置中启动电化学反应,建立光信号与微小原甲藻细胞数目间的ECL-MP分析曲线并对其进行验证.结果表明,标记后的NPA探针具有一定的特异性和实用性;4μg是检测20μL目标藻杂交混合液的最适磁性微球使用量;在最适条件下,微小原甲藻细胞数的线性分析范围6.25×102～4×104个;比较ECL-MP和显微计数方法的样品检测结果,在95%置信区间内二者无显著差异(t-检验).ECL-MP方法为实现微小原甲藻现场样品的快速准确检测提供了一种新方法.     

二氯异氰脲酸钠和三氯异氰脲酸对棕囊藻细胞去除的研究

研究了二氯异氰脲酸钠和三氯异氰脲酸对球形棕囊藻的灭杀和控制作用,及其投药时间对除藻效果的影响,并比较2种药剂的除藻效果,初步探讨了除藻机理.结果表明,这2种除藻剂可以缓释次氯酸,提高其稳定性,有较长的药效期.当有效氯浓度达到4.5mg/L时,2种药剂均能有效地控制和灭杀球形棕囊藻,具有高效、低毒的优点.有效氯浓度为5.0mg/L,藻细胞去除率在24h可达到90%以上.在等于和大于5.0mg/L有效氯浓度时,三氯异氰脲酸的除藻效果优于二氯异氰脲酸钠,通过对2种药剂除藻率的t检验进一步证实了这一结论.     

Growth responses of the newly-discovered Cd-hyperaccumulator Rorippa globosa and its accumulation characteristics of Cd and As under joint stress of Cd and As

Rorippa globosa has been identified as a newly-found Cd-hyperaccumulating species. In the present study, growth responses of Rorippa globosa and its accumulation characteristics of Cd and As were examined under joint stress of Cd and As. The results showed that Cd and As had an antagonistic effect on enhancing the growth of Rorippa globosa plants and Cd uptake and accumulation under the low concentration Cd and As treatments. When the concentration of Cd in the soil was 10 mg/kg and the concentration of As was 50 mg/kg, the highest growing height of the plant was up to 35.9 cm and the dry weight of the shoots was up to 2.2 g/pot, respectively. Meanwhile, the accumulation of Cd in the leaves under the joint stress was higher than that at the same level under single Cd pollution. However, there were synergic adverse effects on plant growth and Cd uptake under the combined pollution from a high concentration of Cd and As. Meanwhile, the accumulation of As in the roots was greater than that in the shoots, the translocation factor (TF) was ⩽0.3 and the bioaccumulation factor (BF) was ⩽0.6, thus showing that Rorippa globosa had an excluding effect on As uptake. These results confirmed that Rorippa globosa had a strong tolerant ability to the joint stress of Cd and As, and the potential for phytoremediation of soils contaminated by Cd and As. Translated from Environmental Science, 2007, 28(6) [译自: 环境科学]     

海洋微藻对游离氨基酸的利用特性研究

分别以4种游离氨基酸——丙氨酸、甘氨酸、谷氨酸和天冬氨酸为唯一氮源,采用实验室一次性培养的方法,研究典型赤潮藻——东海原甲藻(Prorocentrum donghaiense)、球形棕囊藻(Phaeoecystis globosa)和米氏凯伦藻(Karenia mikimotoi)及常见种类中肋骨条藻(Skeletonema costatum)的生长特性、光合特征及对氨基酸的吸收动力学特征,以无机氮源硝氮、氨氮作为对照.研究结果表明,在无菌条件下,东海原甲藻和球形棕囊藻可以利用多种游离氨基酸快速生长,而米氏凯伦藻和中肋骨条藻在以游离氨基酸为唯一氮源条件下不能维持生长.东海原甲藻和球形棕囊藻在丙氨酸中的生长速率最高,天冬氨酸和谷氨酸次之,甘氨酸最低.氨基酸处理组的Fv/Fm值在培养后期降幅小、降速慢,且高于无机氮处理组.东海原甲藻和球形棕囊藻对丙氨酸的最大吸收速率和半饱和常数分别为3.32、0.41pmol·cell-1·h-1和6.99和20.54pmol·L-1.因此,海洋微藻对游离氨基酸的吸收利用具有显著的种间差异,东海原甲藻和球形棕囊藻具有更广的氮营养生态位,在近海有机污染不断加剧的背景下,更容易形成优势甚至暴发赤潮.     

溶藻活性物质对棕囊藻溶藻及其脂肪酸影响的模拟

为了探讨芽孢杆菌B1分泌的胞外溶藻活性物质对球形棕囊藻的溶藻特性和藻毒素物质脂肪酸的影响,比较了模拟自然水体中叶绿素a、p H、溶解氧DO、高锰酸盐指数和营养元素N、P浓度在溶藻前后的变化,并利用GC-MS检测了球形棕囊藻脂肪酸的成分和含量.用体积比1∶100的芽孢杆菌B1胞外活性物质处理模拟水体14 d,发现水体中叶绿素a、p H值和高锰酸盐指数随处理时间的增加而降低,DO和N、P浓度随处理时间的增加而增加.在第14 d时,处理组水体中p H值由8.50降低到7.51,叶绿素a降低82.3%(P0.05),DO增加29.5%(P0.05),高锰酸盐指数降低55.2%(P0.01).NH+4-N、NO-2-N、NO-3-N和PO3-4-P浓度分别增加了0.46、1.50、6.24和1.30倍.投加活性物质处理14 d后,球形棕囊藻藻毒素中的主要3种脂肪酸C18:2、C16:0和C18:1分别降低了100%、97.7%和85.4%(P0.01),总脂肪酸含量降低83.4%(P0.01).结果表明,芽孢杆菌B1胞外溶藻活性物质在模拟自然水体中能有效抑制球形棕囊藻的生长,并降低藻毒素脂肪酸的含量.研究结果为芽孢杆菌B1胞外活性物质的生态安全性应用提供理论基础.     

水环境中大肠杆菌多特征抗原及抗体的制备研究

为了建立用于水环境中大肠杆菌的酶联免疫快速检测方法,针对水中大肠杆菌属多种血清型的特征,制备了水环境样品中大肠杆菌的多特征抗原,包括全菌体抗原、破碎全菌体抗原、菌体抗原、鞭毛抗原和菌毛抗原;采用这5种大肠杆菌抗原分别免疫新西兰大白兔,获得了5种大肠杆菌多克隆抗体,抗体效价高、纯度好,具有较强且稳定的特异性结合抗原的功能;间接ELISA方法表明,全菌体抗体和破碎全菌体抗体的效价均大于1×10⁵,性能优良.基于上述抗体,建立了水中大肠杆菌间接ELISA检测方法,实际水样的检测结果表明,检测限可达10⁴个/L.     

一株产电菌嗜根考克氏菌(Kocuria rhizophila)的分离及其产电性能优化

本研究以天津泰达污水处理厂污泥浓缩间的污泥为接种物,启动并运行了微生物燃料电池(MFCs).从富集的阳极生物膜上分离得到了一株纯培养的微生物菌种,命名为P2-A-5.研究发现,菌株P2-A-5的16S rDNA序列与菌株Kocuria rhizophila DC2201具有100%的同源性,结合该菌的形态特征和生理生化实验,将其归属为嗜根考克氏菌(Kocuria rhizophila).通过化学剂处理、底物种类和浓度的优化,进一步提高其在微生物燃料电池中的产电性能.结果表明,菌株K.rhizophila P2-A-5经0.5 mg·L-1溶菌酶处理45 min后,接种到以2.0 g·L-1海藻糖为底物的阳极液中运行MFCs,其功率密度达到314.8 m W·m-2,比优化前(74.9 m W·m-2)提高了320.3%.这是首次对K.rhizophila种内微生物产电性能及其在微生物燃料电池中应用的报道,其成果对于丰富产电微生物的多样性,挖掘更多具有高电化学活性的微生物菌种,提高其产电性能具有重要的理论意义.     

含vgb基因的重组毕赤酵母菌对重金属离子的吸附解吸特征

酵母因为其比表面积大、密度高被广泛用于水体中重金属的吸附.本文研究比较野生毕赤酵母KM71及含有vgb基因的重组毕赤酵母KM71对重金属Ni²⁺、Cd²⁺和Pb²⁺的吸附解吸特性,并研究混合金属离子对酵母菌吸附性能的影响.不同金属离子均在2 h达到吸附解吸平衡,酵母菌对不同种类金属离子的吸附具有选择性,温度和pH变化对毕赤酵母吸附重金属影响较小.重组毕赤酵母菌对重金属的抗性更强.通过对环境因素及复合剂的优化,得到对重金属镉、镍、铅具有高效吸附性能的重组酵母菌,重组酵母菌对单元素溶液中Ni²⁺、Cd²⁺和Pb²⁺的吸附量分别为10.56、17.68和20.14 mg·g^-1,对三元金属混合溶液中的Ni²⁺、Cd²⁺和Pb²⁺的吸附量分别为8.28、9.89和19.99 mg·g^-1.通过条件优化重组毕赤酵母对Ni²⁺、Cd²⁺和 Pb²⁺的吸附率分别提高了15.14%、21.04%、18.47%,吸附量分别提高了11.26、13.43、22.90 mg·g^-1.本试验为促进酵母菌在处理重金属污染废水中的广泛应用提供了科学依据.     

水华束丝藻对汞的吸附-解吸特征

通过室内模拟实验,探究不同丰度的活、死水华束丝藻对Hg²⁺的吸附动力学特征和等温吸附模型以及解吸特征。结果表明,水华束丝藻对Hg²⁺有较好的吸附效果,能在短时间内吸附大量Hg²⁺,120 min左右达到吸附平衡,且活藻对Hg²⁺的吸附效果比死藻好;活藻和死藻吸附Hg²⁺的动力学过程符合准一级、准二级动力学模型,且准二级动力学模型拟合效果更好;活、死水华束丝藻对Hg²⁺的吸附分别符合Langmuir模型、D-R模型,最大吸附量分别为2.07×10^-2 ng/（10⁶ cells）、3.56×10^-2 ng/（10⁶ cells）;水华束丝藻对Hg²⁺的单位吸附量随着藻丰度的增加而减少,吸附总量随着藻丰度的增加而增加。反应初期（0～5 min）,活、死水华束丝藻对Hg²⁺进行生物吸附,吸附速度快且效率高;随后活藻依靠新陈代谢将Hg²⁺转移至细胞内进行生物富集,因而活藻的单位吸附量高于死藻。活藻和死藻对Hg²⁺的解吸量随藻丰度的增加而增加,且死藻变化更明显。     

A bioaugmentation failure caused by phage infection and weak biofilm formation ability

Two biological aerated filters (BAF) were setup for ammonia removal treatment of the circulation water in a marine aquaculture. One of the BAFs was bioaugmented with a heterotrophic nitrifying bacterium, Lutimonas sp. H10, where the ammonia removal was not improved and the massive inoculation was even followed by a nitrification breakdown from day 9 to 18. The nitrification was remained stable in control BAF operated under the same conditions. Fluorescent in situ hybridization (FISH) with rRNA-targeted probes and cultivable method revealed that Lutimonas sp. H10 almost disappeared from the bioaugomented BAF within 3 d, and this was mainly due to the infection of a specific phage as revealed by flask experiment, plaque assay and transmission electron observation. Analyses of 16S rRNA gene libraries showed that bacterial groups from two reactors evolved di erently and an overgrowth of protozoa was observed in the bioaugmented BAF. Therefore, phage infection and poor biofilm forming ability of the inoculated strain are the main reasons for bioaugmentation failure. In addition, grazing by protozoa of the bacteria might be the reason for the nitrification breakdown in bioaugmented BAF during day 9–18.     

铜绿假单胞菌对铜和铅的吸附

研究了铜绿假单胞菌(Pseudomonas aeruginosa)对Cu和Pb的吸附特性.结果表明,相同条件下,该菌株对Cu~(2+)的吸附率低于Pb~(2+).对于单一重金属体系,吸附率均随时间的延长先上升后平稳变化,2 h达到稳定.吸附率随投菌量的增加先迅速增加,之后趋于平稳.对于Cu~(2+),投菌量为1 g·L~(-1)时吸附率达到稳定,而Pb~(2+)的吸附效果达到平稳时的投菌量为0.5 g·L~(-1).单位质量菌体对Cu~(2+)、Pb~(2+)的吸附量随投菌量的增加而下降.pH为3时,菌体的吸附效果较差,当pH为5~8时,2种重金属的吸附效果较高.对于活菌,Pb~(2+)对菌体吸附Cu~(2+)有抑制作用,而Cu~(2+)对菌体吸附Pb~(2+)的影响无明显规律.对于失活菌,P.aeruginosa吸附Pb~(2+)和Cu~(2+)的效果均随共存重金属浓度的增大而降低,但Cu~(2+)对Pb~(2+)的影响比Pb~(2+)对Cu~(2+)的影响更显著.扫描电镜观察发现,吸附后的菌体较吸附前聚集性更好.总体而言,P.aeruginosa能对水体中共存的Cu~(2+)和Pb~(2+)有较好的吸附效果.     

Effects of Cd2+ on the nucleolus in root tip cells of Allium cepa

The effects of different concentrations of cadmium chloride ranging from 0.5 to 20 ppm on the nucleolus in root tip cells of Allium cepa were studied using the silver staining technique. The results indicated that after the treatment with Cd²⁺,different changes in nucleolar morphology appeared. The nucleolar material was extruded from the nucleus into the cytoplasm,and the nucleoli at mitotic metaphase did not disappear. Apparently,cadmium showed a specific effects on the nucleoli in root tip cells of Allium cepa. The possible mechanism behind this phenomenon is also briefly discussed.     

FeCl3-生化耦合技术调控未知诱因的污泥膨胀

丝状菌污泥膨胀是影响污水处理厂出水水质的常见问题.现有的调控技术存在见效慢、耗时长和通用性差等弊端.提高COD和DO浓度,投加FeCl_3和交替曝气是目前调控丝状菌膨胀的主要方法,这些方法的耦合技术调控未知诱因的污泥膨胀的效果鲜有报道.实验设计三段A/O反应器应急性运行SBR工艺,交替曝气时提高DO至(7. 45±0. 49) mg·L~(-1),COD初始浓度提高至(332. 73±106. 06) mg·L~(-1),沉淀工序中投加FeCl_3,使之在混合液中质量浓度为120 mg·L~(-1),形成的FeCl_3-生化耦合技术快速调控未知诱因的Candidatus Saccharibacteria为主要菌属的污泥膨胀,污泥体积指数14 d从274 m L·g~(-1)稳定至56 m L·g~(-1). FeCl_3-生化耦合技术有效抑制Candidatus Saccharibacteria的繁殖,其属水平相对丰度从97. 64%降低至32. 67%.调控过程出水COD和PO_4~(3-)-P浓度均稳定达到一级A标准,NH_4~+-N去除率从65. 33%增长至74. 65%.表明FeCl_3-生化耦合技术调控未知诱因的污泥膨胀具有良好效果.     

Detection of Escherichia coli in wastewater based on enzyme immunoassay

This research describes a fast detection method on the basis of enzyme-linked immunosorbent assay (ELISA) for Escherichia coli in drainage of wastewater treatment plants. Optimized conditions such as the reaction format (sandwich or direct), the concentrations of diluted horseradish peroxidase (HRP)-E. coli conjugate, and anti-HPR antibody and pretreatment of E. coli were studied. Those results showed that the linear range of detection for E. coli was 10 cfu/mL-6 × 10⁴ cfu/mL. Compared with conventional methods, it is a convenient and sensitive detection method with low cost. Translated from Environmental Science, 2005, 26(5): 128–131 [译自: 环境科学]     

贵州万山汞矿区苔藓植物对汞的吸附和富集特征

在所设定的贵州万山汞矿区调查范围内,有6种苔藓植物,其中常见种为东亚褶叶藓和大羽藓.结果表明,苔藓能高度富集汞,东亚褶叶藓和大羽藓的汞含量随污染源距离和相对高度的增加而降低;不同苔藓对汞的吸附等温线具共同性,吸附量随平衡液浓度的增加而增大,东亚褶叶藓对Hg的吸附量比大羽藓大,吸附特征都能用Liner、Freundlic...     

高铵条件下绿狐尾藻的生理与氮磷吸收特征

为了探讨水生植物绿狐尾藻(Myriophyllum aquaticum)在生态湿地系统对高铵态氮(NH_4~+-N)的耐受及氮(N)、磷(P)吸收能力,试验设置3个高NH_4~+水平(70、210、420 mg·L~(-1))的营养液,对绿狐尾藻培养21 d,研究高浓度NH_4~+对绿狐尾藻生理及N、P吸收特征的影响.结果表明,70 mg·L~(-1)高NH_4~+处理,绿狐尾藻生长健壮,培养21 d后,茎高和生物量分别达到40.56 cm和17.82 g·穴-1;与70 mg·L~(-1)高NH_4~+对照相比,210 mg·L~(-1)高NH_4~+处理下,绿狐尾藻丙二醛含量显著增加,但叶绿素和可溶性糖含量仍较高,说明其受到一定NH_4~+的胁迫,但不影响其正常生长,茎和生物量的生长速率与对照无显著差异.当NH_4~+水平达到420 mg·L~(-1)时,丙二醛含量增加了2倍,茎高和生物量生长速率仅为对照的27.4%和17.9%,植物受到严重胁迫,生长受阻甚至死亡.3种高NH_4~+处理下,绿狐尾藻总N和总P含量的变化范围为30.7~53.4 mg·g~(-1)和3.8~7.7mg·g~(-1),表现出对N、P超高的吸收能力.鉴于绿狐尾藻在高浓度NH_4~+-N中的高耐受性,及很强的N、P吸收能力,绿狐尾藻可作为高铵/氨废水生物处理中理想的湿地生物,将具有良好的应用前景.     

新型氧化石墨烯生物纳米材料对印染废水中镉和孔雀石绿的去除性能及机制

印染废水因其成分复杂,并且难以被自然去除,己成为环境的重点污染源之一.本研究采用甘蔗渣还原液还原氧化石墨烯,并负载伯克霍尔德菌成功制备了新型生物纳米材料MNMs.采用SEM-EDS-Mapping和Raman分析证实该材料具有较好的结构稳定性和良好的吸附效果,并对印染废水的典型污染物Cd²⁺和孔雀石绿的联合去除率均达到80%以上.GC-MS检测结果证实该复合纳米材料可将孔雀石绿降解为N,N-二甲基苯胺和4-（二甲基氨基）二苯甲酮.动力学拟合分析表明Cd²⁺和孔雀石绿被MNMs吸附过程遵循伪二级速率模型（R²>0.99）,而MG的降解过程更遵循伪一级速率模型（R²>0.99）.MNMs对Cd²⁺和MG的理论最大吸附量分别为84.03 mg·g^-1和45.25 mg·g^-1.研究结果表明,该生物纳米材料对水溶液中Cd²⁺和孔雀石绿有较好的去除效果,未来可应用于工业印染废水的污染控制.     

基于脱氧核酶的水中铀酰离子光纤倏逝波生物传感检测技术研究

为了满足铀的高通量快速筛查及环境突发事件快速检测的需要,本研究利用脱氧核酶的特异性催化反应,结合本课题组自主研制开发的倏逝波光纤生物传感器,通过设计两步法的反应策略,即"均相反应,固相杂交"的检测方案,建立了光纤倏逝波生物传感检测铀酰离子的新型技术方法.结果发现,该方法对于5~100 nmol·L~(-1)内的铀酰离子具有良好的线性检测区间,检测限低至0.5 nmol·L~(-1),整个检测过程仅需16 min即可完成,铅和汞等10种金属离子对于本检测方法无明显干扰,表现出了对铀酰离子的良好选择性.传感光纤可重复使用100次以上,不会有信号的明显衰减,可有效降低检测成本.对丹江口水库水进行的实际水样加标回收实验结果显示,丹江口水库实际水样的加标回收率为91.6%~94.6%.本研究可为利用生物传感器法检测环境水体中的铀酰离子提供技术支撑.     

发光菌生物毒性测试方法的改进

针对发光菌发光强度本底差异较大,检测期间发光变化幅度宽的问题,在传统发光菌生物毒性测试实验中,引入校正因子(P^I_丙酮),研究了磁力搅拌时间、培养时间、培养代数和培养温度等条件对EC₅₀测定值的影响.结果表明改进后的方法实验重现性好,相对标准偏差(RSD)在2.1%～13.1%之间.发光菌生物毒性测试方法的改进,为获得可靠的毒性实验数据,进行多组分有机化合物定量结构-活性相关(QSARs)的研究奠定了一定的实验基础.