长江干流浮游细菌群落结构及影响因素

为研究长江干流浮游细菌群落结构的空间分布规律及其环境影响因素，针对细菌16S rRNA基因，采用定量PCR和高通量测序技术，对2015年和2016年采集的长江干流自四川泸州到上海吴淞口共计40个样点进行分析。结果表明：水体中浮游细菌16S rRNA基因丰度为2.64×106~1.44×109 copies/L。上游（泸州至秭归）、中游（宜昌至湖口）和下游水体（湖口至上海入海口）的α多样性在3组间都没有呈现出显著差异性。长江干流浮游细菌在门水平上主要由变形菌门（Proteobacteria，7.49%~86.53%）、放线菌门（Actinobacteria，0.27%~54.72%）、厚壁菌门（Firmicutes，0.03%~90.95%）、拟杆菌门（Bacteroidetes，0.50%~45.36%）和异常球菌-栖热菌门（Deinococcus-Thermus，0.00%~23.96%）构成。在属或科水平上主要由丛毛单胞菌科（Comamonadaceae，0.03%~65.67%）、CL500-29_marine_group（0.00%~24.23%）、hgcI_clade（0.00%~28.82%）、芽孢杆菌属（Bacillus，0.00%~90.88%）和氢噬胞菌属（Hydrogenophaga，0.03%~38.55%）组成。从上中下游水体的浮游细菌群落组成来看，上游水体相对丰度最高的类群为Bacillus（0.00%~90.88%），中游和下游为Comamonadaceae（分别为0.60%~65.67%和2.87%~50.64%）。造成上中下游水体中浮游细菌群落组成差异的环境因子各不相同：影响上游的主要为pH和悬浮颗粒物（SS），影响中游的主要为溶解氧（DO），影响下游的主要为DO、水温（WT）和总磷（TP）。     

2,4,6-三氯苯酚降解菌的筛选及降解特性

为了研究氯酚类环境激素2,4,6-三氯苯酚的生物降解性,从活性污泥中分离筛选出一株能以2,4,6-三氯苯酚为唯一碳源生长的降解性菌株T10,通过16S rRNA测序鉴定其为一种芽孢杆菌(Bacillus sp.)。在实验室条件下探讨pH值、温度等环境因素对菌株T10降解性的影响,实验结果表明该菌株在pH为7.0,30⁴0℃范围内具有高降解活性,底物浓度为25 mg/L时,120 h的降解率均能达到90%以上,在30℃条件下的降解半衰期为1.26 d。初始底物浓度达到30 mg/L时,2,4,6-TCP的抑制作用开始起主导作用,影响菌株降解活性。     

温度分化对APBR反应器性能及产甲烷菌群落的影响

为了研究温度分化对固定床厌氧反应器(anaerobic packed bed reactor,APBR)牛粪发酵处理效果及产甲烷菌群落的影响,反应器发酵温度从室温(22℃±1℃)阶梯式分化到低温(15℃±1℃)、中温(37℃±1℃)和高温(55℃±1℃).温度变化的过程中,温度越高COD(chemical oxygen demand)去除率和日总产气量越高,分化后COD去除率分别为25%、45%、60%,相应的日产气量为2.3、4.0、8.5 L·d-1,但是甲烷含量基本保持不变(~60%);温度突然变化造成挥发性脂肪酸含量骤然增加,并处于波动状态.16S r RNA基因克隆文库法分析表明,室温时包含广古菌门中的常见重要产甲烷菌MBT(甲烷杆菌目)、Mst(甲烷鬃菌科)、Msc(甲烷八叠球菌科)和MMB(甲烷微菌目),以及嗜热菌,也有少部分泉古生菌门,发酵温度分化后,产甲烷菌多样性减少,中温条件下产甲烷菌种类相对较少.定量PCR表明Mst、MMB和Msc总基因浓度都有所减少,并且温度越高减少越多,各菌数量相对比例变化较大,但Mst仍为优势产甲烷菌.     

Molecular characterization and toxin quantification of Microcystis panniformis: A microcystin producer in Lake Taihu, China

Microcystis panniformis is a bloom forming species with flat panniform-like colonies. This species was recently found in Lake Taihu, China. To specifically characterize M. panniformis based on isolated strains, morphological examination on colonial transition and genetic examination are needed. Three M. panniformis strains isolated from a water bloom sample in Lake Taihu were characterized by molecular analysis and toxin quantification. Phylogenetic analysis based on both 16S rRNA gene and internal transcribed spacer(ITS) between 16S and 23S rRNA genes were performed and compared to facilitate easy identification of the species.Relatively high similarities(98%–99%) were shown in 16S rDNA sequences between the strains of M. panniformis and those of other Microcystis species, whereas the similarities for ITS sequences were 88%–95%. In the phylogenetic tree based on the 16S rDNA sequences, the M. panniformis and M. aeruginosa strains were intermixed together with no clear division,whereas all of the M. panniformis strains were clustered together in a single clade based on the ITS sequences based phylogenyetic tree. The mcyE gene was detected in all three strains, and microcystin was determined by high-performance liquid chromatography. The molecular detection and toxin production of M. panniformis strains are of great significance for the environmental risk assessment of Microcystis blooms.     

利用16S-23S rDNA RFLP及16S rRNA基因序列分析方法对湖北省饭豆(Vigna umbellata L.)根瘤菌系统发育的研究

采用16S-23S rDNA间隔区段(IGS)PCR-RFLP与16S rRNA基因部分序列分析的方法对饭豆根瘤菌进行了遗传多样性及系统发育分析.由16S-23S rDNA IGS PCR-RFLP分析可知,所有菌株在52%的相似性水平上聚在一起,形成了慢生型菌群与快生型菌群这两大菌群.群Ⅰ中,在79%相似性的水平上分为ⅠA与ⅠB两个分支.群Ⅱ中,在62%相似性的水平上分为ⅡA与ⅡB两个分支,分支ⅡA在72%的相似性水平上进一步分为ⅡA1、ⅡA2和ⅡA3三簇;分支ⅡB中的饭豆根瘤菌与标准菌株USDA205T聚在一起,表现的差异并不大.由16SrRNA基因部分序列分析结果可知,供试的4个代表菌株分别位于不同的系统发育分支中.CYR4243与Sinorhizobium fredii的模式菌株USDA205T的序列相似性达到了99.87%.HCY1101与Rhizobium leguminosarum中的三叶草生物型(bv.trifolii)和豌豆生物型(bv.viceae)这两个生物型的参比菌株亲缘关系最近,序列相似性为100%.HCY5202与R.galegae亲缘关系最近,序列相似性为99.86%.CYY3302与Bradyrhizobium elkanii的参比菌株USDA86有最近的亲缘关系,序列相似性近似于100%.图4表1参13     

Start-up of the anammox process from the conventional activated sludge in a hybrid bioreactor

The anaerobic ammonium oxidation (anammox) process was successfully started up from conventional activated sludge using a hybrid bioreactor within 2 months. The average removal efficiencies of ammonia and nitrite were both over 80%, and the maximum total nitrogen removal rate of 1.85 kg N/(m³.day) was obtained on day 362 with the initial sludge concentration of 0.7 g mixed liquor suspended solids (MLSS)/L. Scanning electron microscope (SEM) observation of the granular sludge in the hybrid reactor clearly showed a high degree of compactness and cell sphericity, and the cell size was quite uniform. Transmission electron microscope photos showed that cells were round or oval, the cellular diameter was 0.6--1.0 μupm, and the percentage of the anammoxosome compartment was 51%--85% of the whole cell volume. Fluorescence in situ hybridization analysis (FISH) indicated that anammox bacteria became the dominant population in the community (accounting for more than 51% of total bacteria on day 250). Seven planctomycete 16S rRNA gene sequences were present in the 16S rRNA gene clone library generated from the biomass and affiliated to Candidatus Kuenenia stuttgartiensis and Candidatus Brocadia sp., a new anammox species. In addition, the average effluent suspended solid (MLSS) concentrations of outlets I (above the non-woven carrier) and II (below the non-woven carrier) were 0.0009 and 0.0035 g/L, respectively. This showed that the non-woven carrier could catch the biomass effectively, which increased biomass and improved the nitrogen removal rate in the reactor.     

一株乐果降解菌的筛选、鉴定及其降解特性研究

采用自制驯化装置,从土壤中分离纯化出一株能以乐果为单一碳源生长的菌株,命名为菌株LPx。根据生理生化特征和16S rRNA(GenBank Accession No.HM488993)基因序列分析,初步将该菌株鉴定为假单胞菌属(Pseudomonas sp.)。通过对其降解乐果特性研究,结果显示,菌株LPx降解乐果的最适pH为7.5、最适温度为30℃、最适接种量为10%(体积分数)。最适条件下,100 mg/L乐果可在120 h内基本被降解。菌株对乐果的降解属于高浓度底物抑制的酶促反应,vmax(不存在抑制剂时最大酶促反应速率)=0.734 d-1,km(米氏常数)=21.700 mg/L,k1(底物抑制系数)=259.215 mg/L。     

自然生境中氨氧化细菌的分子生物学研究进展

摘要 硝化过程是自然界氨氮污染降解的主要生化机制,氨氧化细菌对于富营养的河流、海洋及土壤环境等自然生境具有重要作用.氨氧化细菌的群落结构很复杂,且在不同研究区的时空上存在很大差异性.随着分子生物技术的发展,各种自然环境中存在的氨氧化细菌得到了研究.目前,中国对氨氧化细菌的研究多集中于污水处理、土壤等方面,采用非培养方法对自然环境中氨氧化细菌的研究较少.首先介绍了氨氧化细菌的发现过程和生理特性,其次论述了不同的分子生物学方法对土壤和水生生态环境等自然生境中氨氧化细菌研究进展和研究方法,最后提出了自然生境中氨氧化细菌研究的问题和展望.     

基于454和Illumina高通量测序平台对长江口邻近海域沉积物微生物群落的比较分析

作为第二代测序技术的主要平台, 454和Illumina测序方法在微生物核酸分析方面有着广泛应用。本研究以16S rRNA基因为标靶, 分别采用454和Illumina测序技术对长江口邻近海域表层沉积物中的微生物DNA进行分析, 通过对序列长度、微生物群落多样性及组成进行比较, 分析两种方法在沉积物微生物群落结构研究方面的适用性。结果表明, Illumina测序获得的序列数和OTU数要多于454, 其中获得的OTUs中以非优势菌群为主。Illumina与454相比能够检出更多微生物菌群, 且检测到的稀有菌群占有较大比例。综上可知, Illumina测序技术在分析微生物群落多样性及物种组成方面要优于454测序技术, 454的长序列优势并未能体现出来, 且454测序技术可能低估了微生物组成及其多样性。     

高温冲击对亚硝酸盐氧化过程中微生物菌群结构影响

为了进一步了解在亚硝酸盐氧化过程中,高温冲击对活性污泥微生物菌群结构的影响,本研究中,以在不同NO~-_2-N浓度条件下富集的硝化活性污泥为研究对象,利用16S rRNA高通量测序技术分析方法,通过改变环境温度考察活性污泥微生物菌群丰度变化及结构特征.高通量测序分析结果表明:25℃时易于微生物生长,系统活性污泥微生物菌群多样性最丰富.随着高温冲击试验进行,系统内菌群丰富度、均匀度和多样性呈下降趋势.此外,分析发现本系统主要硝化功能菌为Nitrospirae的Nitrospira,更适宜在35℃生长.且高温冲击试验同样引起了活性污泥中非硝化功能微生物(Bacteroidetes、Chloroflexi、Halomonas和Pseudomonas等)的菌群结构差异.试验结果可为高温冲击条件下亚硝酸盐氧化过程中微生物菌群分布特点的研究提供部分理论参考,并为相关高温冲击试验给予部分借鉴.