基于高通量测序技术的柄扁桃根围AMF群落季节动态研究

【目的】研究柄扁桃(Prunus pedunculata)根围丛枝菌根真菌(arbuscular mycorrhizal fungi,AMF)群落组成的季节动态变化,为柄扁桃天然林的保护和恢复提供理论依据。【方法】以春季(5月)、夏季(7月)和秋季(10月)采集的27个柄扁桃根围土壤样本为研究对象,利用AMV4.5NF/AMDGR引物扩增核糖体小亚基(SSU)rDNA特定片段,Illumina MiSeq测序研究AMF群落,分析其组成和多样性的季节差异,并测定土壤理化性质,调查植物群落和植被盖度,利用冗余分析探究季节、植被和土壤因子对AMF群落的影响。【结果】从柄扁桃根围土壤中共检测到153个AMF操作分类单元(operational taxonomic unit,OTU),其中春季和秋季AMF丰富度显著高于夏季,春季与秋季间不存在显著差异,Shannon多样性、Simpson多样性和Pielou均匀度指数在季节间不存在显著差异。春季与夏季、夏季与秋季间AMF群落组成存在显著差异,但在春季和秋季间差异不显著。从特有和指示AMF OTU数目来看,夏季最多,春季次之,秋季最少。季节差异会影响特定AMF类群,但不会显著影响AMF丰富度和多样性。AMF丰富度与土壤速效磷和土壤含水量呈极显著负相关;AMF多样性与土壤速效氮、速效磷和土壤含水量呈显著负相关,与植物多样性呈极显著正相关。【结论】柄扁桃根围土壤中AMF丰富度及群落组成呈季节性动态变化,且这种变化可能主要受土壤和植被因子季节变化的影响。     

A comprehensive analysis of the response of the fungal community structure to long-term continuous cropping in three typical upland crops

Certain agricultural management practices are known to affect the soil microbial community structure; however, knowledge of the response of the fungal community structure to the long-term continuous cropping and rotation of soybean, maize and wheat in the same agroecosystem is limited. We assessed the fungal abundance, composition and diversity among soybean rotation, maize rotation and wheat rotation systems and among long-term continuous cropping systems of soybean, maize and wheat as the effect of crop types on fungal community structure. We compared these fungal parameters of same crop between long-term crop rotation and continuous cropping systems as the effect of cropping systems on fungal community structure. The fungal abundance and composition were measured by quantitative real-time PCR and Illumina MiSeq sequencing. The results revealed that long-term continuous soybean cropping increased the soil fungal abundance compared with soybean rotation, and the fungal abundance was decreased in long-term continuous maize cropping compared with maize rotation. The long-term continuous soybean cropping also exhibited increased soil fungal diversity. The variation in the fungal community structure among the three crops was greater than that between long-term continuous cropping and rotation cropping. Mortierella, Guehomyces and Alternaria were the most important contributors to the dissimilarity of the fungal communities between the continuous cropping and rotation cropping of soybean, maize and wheat. There were 11 potential pathogen and 11 potential biocontrol fungi identified, and the relative abundance of most of the potential pathogenic fungi increased during the long-term continuous cropping of all three crops. The relative abundance of most biocontrol fungi increased in long-term continuous soybean cropping but decreased in long-term continuous maize and wheat cropping. Our results indicate that the response of the soil fungal community structure to long-term continuous cropping varies based upon crop types.     

4个不同苯甲醇含量茶树品种的转录组分析

香气是茶叶品质决定因子之一,苯甲醇作为一种芳香族醇,参与茶叶香气物质形成。本研究选用4个苯甲醇含量不同的茶树品种,分别是‘薮北’(苯甲醇含量低)、‘湘妃翠’、‘牛皮茶’(苯甲醇含量中等)和‘铁香茗’(苯甲醇含量高),通过Illumina Hiseq^TM2000高通量测序技术对上述4种茶树一芽二叶的转录组进行测序,通过Blast搜索比对,共有151 198条Unigene获得了基因注释。4种茶树在新陈代谢路径中注释的基因数最多,其中次生代谢物生物合成和萜类和聚酮化合物代谢途径可能与茶树苯甲醇合成有关,通过对4种茶树样品的Unigene与NR数据库比对,发现5条可能编码茶树苯甲醇合成途径的关键酶β-葡萄糖苷酶基因,这些相关研究为分析茶树香气功能基因提供理论指导,为香气相关候选基因的发掘提供重要依据。     

基于高通量测序技术的酿酒葡萄产区土壤微生物多样性

【目的】研究新疆4大酿酒葡萄产区土壤中的微生物群落结构及多样性,为构建新疆酿酒葡萄主产区的微生物菌库奠定基础。【方法】分别从新疆酿酒葡萄主产区采集土壤样品,提取总DNA,通过Illumina MiSeq高通量测序结合生物信息学分析,得到新疆酿酒葡萄主产区的土壤微生物群落结构及多样性。【结果】在12份土壤样品中,伊犁河谷土壤中细菌的群落多样性最高,天山北麓土壤中细菌群落多样性最低;吐哈盆地中真菌群落多样性最低,焉耆盆地中真菌的群落多样性最高。节杆菌属细菌和耐冷酵母属真菌是吐哈盆地和天山北麓葡萄园土壤中的优势菌属;芽单胞菌属细菌和刚毛四枝孢属真菌是伊犁河谷葡萄园土壤中的优势菌属;节杆菌属细菌和赤霉菌属真菌是焉耆盆地葡萄园土壤中的优势菌属。土壤pH值对细菌群落组成的影响最显著,Mn元素含量对真菌群落组成的影响最显著。【结论】揭示了新疆酿酒葡萄主产区土壤中细菌和真菌群落组成,新疆酿酒葡萄产区的微生物多样性存在差异。     

Microbial communities in rice rhizosphere altered by intermittent and continuous flooding in fields with long-term arsenic application

Rice cultivated on arsenic (As)-contaminated soils can, under some conditions, accumulate high concentrations of As in grain, mostly as a result of the continuous flooding practices commonly used for rice cultivation. Intermittent flooding, as opposed to continuous flooding, might reduce soluble As concentrations in the rice rhizosphere, but it might also alter soil microbial populations that may impact As chemistry. A field-scale study was conducted to analyze As concentrations and microbial populations in the rice rhizosphere, in response to intermittent and continuous flooding in plots that were historically amended with “As-containing” pesticide and unamended soil. Rhizosphere, pore-water and grain As concentrations were quantified, and microbial populations in the rhizosphere were characterized using community quantitative-PCR and 16S rRNA gene sequencing. Pore-water As concentrations decreased by 41-81% and grain As by 31-48% in the intermittently flooded plots relative to the continuously flooded plots. The relative abundance of bacteria increased over the course of the growing season, while archaeal and fungal gene abundances decreased. Bacterial community structure and composition were significantly different between As amended and unamended plots, as well as between the flooding treatments. Proteobacteria was the predominant phylum detected in most treatments with relative abundance of 24-29%. The relative abundance of iron-reducing bacteria was higher with the continuous flood compared to the intermittent-flood treatment, implying greater relative iron reduction and possibly As release from the iron oxides under the continuously flooded conditions. These differences in rhizosphere-microbial communities may have contributed to the lower pore-water arsenic concentrations in the intermittently flooded conditions.     

复合菌剂防控香蕉枯萎病的效果及其微生物学机制

由病原菌尖孢镰刀菌古巴专化型（Fusarium oxysporum f. sp. cubense (Foc)）侵染引起的香蕉枯萎病对全世界香蕉产业带来了毁灭性的影响,且目前尚无广泛采用的防治方法。研究复合生防真菌制剂对香蕉枯萎病的防治效果,以期为大田香蕉枯萎病的防治提供依据。设置3组不同的菌剂处理,分别为对照组CK、复合菌剂NFP、复合菌剂NFPT,通过两季的盆栽试验,研究复合菌剂对香蕉枯萎病的防治效果及其对土壤微生物多样性的影响;利用 Illumina Miseq 高通量测序平台对细菌16S rRNA基因和真菌ITS区域进行测序分析,采用实时荧光qPCR定量分析各处理病原菌的数量。结果表明：（1）复合菌剂处理（NFP和NFPT）对香蕉枯萎病有较好的防治效果,其防效分别为43%和48%。（2）施用复合菌剂增加了细菌和真菌群落丰富度和多样性。基于Bray-curtis距离矩阵的主坐标分析（PCoA）结果表明NFP和NFPT改变了细菌和真菌群落结构。NFP和NFPT处理增加了潜在有益微生物中与香蕉枯萎病病情指数呈显著负相关的大理石雕菌属Marmoricola、类诺卡氏菌属Nocardioides、野野村式菌属Nonomuraea、 norank_c__Acidobacteria、DS-100属和norank_f__Blastocatellaceae__Subgroup_4属的相对丰度,显著减少了病原菌尖孢镰刀菌的数量,重塑了土壤微生物结构和功能,增强其抗病性。     

花鲈irak4基因cDNA的克隆与表达分析

该研究基于Illumina MiSeq高通量测序技术,对周期性缺氧应激下花鲈(Lateolabrax maculatus)肠道菌群结构的变化进行分析,为研究其幼鱼肠道菌群对环境缺氧的适应机制提供参考依据。结果显示,周期性缺氧导致花鲈肠道菌群多样性和丰富度显著增加(P<0.05),群落结构复杂化,缺氧组和常氧组群落组成存在较大差异。缺氧组肠道菌群分类操作单元相比常氧组显著增加(P<0.05),其特有操作分类单元 (operational taxonomic unit, OTU)数达1 003个。门分类水平上,变形菌门、厚壁菌门和拟杆菌门为2组肠道菌群的主要组成菌门。与常氧组相比,缺氧组变形菌门相对丰度显著降低(P<0.05),而拟杆菌门相对丰度显著升高(P<0.05);纲水平上,缺氧组α变形菌纲和芽孢杆菌纲相对丰度显著降低(P<0.05),梭菌纲、γ变形菌纲和拟杆菌纲相对丰度显著升高(P<0.05)。此外,周期性缺氧应激还引起花鲈肠道内厌氧绳菌科、毛螺菌科、瘤胃菌科等厌氧或兼性厌氧菌和绿硫菌科等光合产氧菌类相对丰度升高。

     

不同时期的柑橘果实中真菌种类分析

柑橘采后真菌病害发生普遍,对柑橘产业造成了极大的威胁。柑橘体内存在丰富的真菌类群,为了了解柑橘成熟和衰老过程中果实内生真菌的多样性和变化动态,本论文通过MiSeq技术解读真菌ITS区域序列,分别对生长期、成熟期和贮藏期的晚蜜一号果实的果皮和果肉的内生真菌种类进行了分析。发现相比于果肉,果皮中真菌在目和属水平的多样性更为丰富,而成熟期果肉中真菌种类多于果皮。不同生长阶段的两种组织中优势的真菌属也不同,果皮在生长期以Medicopsis和刺盘孢属(Colletotrichum)占绝对优势,成熟期为刺盘孢属,贮藏期以葡萄孢属(Botrytis)、Erythrobasidium和Strelitziana较为常见;而果肉中生长期主要为青霉属(Penicillium)和枝孢菌属(Cladosporium),约占1/4,成熟期半数以上为葡萄孢属,及至贮藏期约90%为链格孢属(Alternaria)和青霉属,其中链格孢属约占2/3。常见植物病原真菌如枝孢菌属(Cladosporium)、巨座壳属(Magnaporthe)、核盘菌属(Sclerotinia)、葡萄孢属(Botrytis)、白粉菌属(Erysiphe)、青霉属(Penicillium)、链格孢属(Alternaria)和镰孢霉属(Fusarium)等在果肉中的数量高于果皮。果肉中存在较大数量的常见植物病原真菌,不同属真菌在不同样品中呈不同的变化动态,暗示采后病变是多种真菌交互作用的结果,采后病害的防治可以提前至开花期。     

Bioprospecting from Marine Sediments of New Brunswick,Canada: Exploring the Relationship between Total Bacterial Diversity and Actinobacteria Diversity

Actinomycetes are an important resource for the discovery of natural products with therapeutic properties. Bioprospecting for actinomycetes typically proceeds without a priori knowledge of the bacterial diversity present in sampled habitats. In this study, we endeavored to determine if overall bacterial diversity in marine sediments, as determined by 16S rDNA amplicon pyrosequencing, could be correlated with culturable actinomycete diversity, and thus serve as a powerful tool in guiding future bioprospecting efforts. Overall bacterial diversity was investigated in eight marine sediments from four sites in New Brunswick, Canada, resulting in over 44,000 high quality sequences (x = 5610 per sample). Analysis revealed all sites exhibited significant diversity (H’ = 5.4 to 6.7). Furthermore, statistical analysis of species level bacterial communities (D = 0.03) indicated community composition varied according to site and was strongly influenced by sediment physiochemical composition. In contrast, cultured actinomycetes (n = 466, 98.3% Streptomyces) were ubiquitously distributed among all sites and distribution was not influenced by sediment composition, suggesting that the biogeography of culturable actinomycetes does not correlate with overall bacterial diversity in the samples examined. These actinomycetes provide a resource for future secondary metabolite discovery, as exemplified by the antimicrobial activity observed from preliminary investigation.