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。     

马铃薯健株与黄萎病株根际土壤真菌群落结构及其对碳源利用特征

[目的]通过研究马铃薯健康植株与黄萎病株的根际土壤真菌群落结构与功能多样性的差异,明确土壤真菌群落结构与黄萎病发生之间的关系,为最终从微生物生态学的角度解释马铃薯黄萎病的发生原因及其生态防控提供理论依据.[方法]以河北省坝上地区马铃薯健株与黄萎病株的根际土壤为研究对象,分别利用实时荧光定量PCR(real-time P...     

Biocontrol of soil diseases and soil profile associated with rhizosphere of rice (Oryza sativa Subsp. Japonica) growing paddy fields in Kansai region,Japan

ABSTRACT

This research focused on in vitro and in vivo biocontrol of two major rice diseases, Bacterial Leaf Blight caused by Xanthomonus oryzae pv oryzae (Xoo) and Sheath Blight caused by Rhizoctonia solani. Furthermore, the soil of paddy fields in Kansai region, Japan was analysed for the presence of these pathogens, and their microbial communities were evaluated according to the location and management systems. For biocontrol, the rhizospheric bacteria were isolated and molecularly identified. The results indicated that DAR17225040 and DAR17225017 were 99% similar to Bacillus aryabhattai and Bacillus megaterium. The isolated bacteria were able to suppress Bacterial Leaf Blight and Sheath Blight disease 78% and 86% respectively, under greenhouse conditions. The 16S amplicon sequencing (Illumina MiSeq 250bp) was used to analyse the bacterial community in paddy fields of soil taken from five locations under conventional and organic systems. Xoo and R. solani were detected in sampled soil, however, no clear difference was noted, regarding systems. From amplicon sequencing data, Chloroflexi, Acidobacteria, Proteobacteria, Actinobacteria, Planctomycetes were the dominant phyla of rice soil in Kansai region. The phylum Kazan-3B-28 was found more in organic system than conventional. The bacterial community was more affected by location and soil type than by management systems.     

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.     

不同地点和复垦方式煤矸石山土壤中丛枝菌根真菌群落变化

以山西省大同市晋华宫人工复垦和自然恢复煤矸石山以及忻州窑自然恢复煤矸石山为研究对象,采用IlluminaMiSeq高通量测序技术对三座煤矸石山土壤AM真菌群落组成进行分析,并测定其土壤理化性质,以探究不同地点煤矸石山不同复垦方式下土壤中丛枝菌根真菌（Arbuscular mycorrhizal fungi, AM真菌）群落组成的差异及其土壤影响因子。结果表明：晋华宫人工复垦煤矸石山和忻州窑自然恢复煤矸石山AM真菌多样性指数（Chao指数和香农-威纳指数）存在显著差异,但晋华宫人工复垦和自然恢复煤矸石山AM真菌群落的多样性指数差异不显著。试验共检测到67个AM真菌OTUs,可鉴定到3个AM真菌属级分类单元,其中球囊霉属（Glomus）在晋华宫人工复垦煤矸石山、晋华宫自然恢复煤矸石山和忻州窑自然恢复煤矸石山中均为最优势菌,多样孢囊霉属（Diversispora）、类球囊霉属（Paraglomus）仅出现在晋华宫自然恢复煤矸石山。多元置换方差分析和多维尺度分析结果表明,晋华宫两种恢复方式的煤矸石山之间AM真菌群落组成差异不显著,而晋华宫与忻州窑两个地点的煤矸石山AM真菌群落组成差异显著。AM...     

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.     

应用焦磷酸测序鉴定马铃薯感染的马铃薯Y病毒株系

为了检验是否可以通过焦磷酸测序法进行短序列测序,来鉴别马铃薯Y病毒(PVY)株系,从NCBI库中选取了代表9个PVY株系的全基因组序列30个,通过比对P1和CP蛋白的氨基酸序列,分别找出一个差异区段,针对这2个区域设计了反转录引物、PCR引物和测序引物。分别从黑龙江省克山县和讷河市各采集了一份马铃薯病叶样品,通过提取总RNA,反转录,再进行PCR扩增获得扩增产物,并对扩增产物进行了焦磷酸测序。测序结果显示,尽管两份马铃薯病叶症状不同,但均感染了PVY(N-Wi株系)(PVYN-Wi)。上述研究表明,利用本试验设计的引物,通过焦磷酸测序法进行短序列测序,可以鉴定马铃薯感染的PVY株系。     

不同种植方式大豆根际土壤细菌多样性分析

利用Illumina MiSeq第二代高通量测序平台, 对黑龙江省不同地区轮作及连作大豆根际土壤细菌16S rDNA基因组测序, 初步分析在不同种植方式下受大豆胞囊线虫侵染的大豆根际土壤细菌群落结构的变化。从6个土壤样本中共获得25 419个OTUs, 鉴定到细菌的47个门, 147个纲, 709个属。变形菌门、放线菌门、酸杆菌门、芽单胞菌门、浮霉菌门是供试土壤细菌的优势菌门, 占所有细菌群落总数90%以上。连作4年总OTUs及丰富度最高, 连作20年最低。不同轮作方式土壤细菌丰富度差异不显著(P > 0.05), 短期连作与长期连作细菌丰富度及多样性差异显著(P < 0.05)。不同轮作方式下放线菌门相对丰度低于连作方式, 芽单胞菌门和拟杆菌门相对丰度高于同地区连作方式。土壤功能细菌根瘤菌(Bradyrhizobium)、链霉菌属(Streptomyces)、芽孢杆菌属(Bacillus)、溶杆菌属(Lysobacter)、土微菌属(Pedomicrobium)的相对丰度在不同年限连作下高于轮作。长期连作土壤优势细菌丰度与轮作土壤相似性更高。     

The diazotrophic community in oat rhizosphere: effects of legume intercropping and crop growth stage

In this study, the abundance, diversity and structure of the diazotrophic community in oat rhizosphere soil in three cropping systems and at two oat growth stages were investigated using real-time PCR and Illumina MiSeq sequencing. The nifH gene abundance in oat-soybean intercropping (OSO) and oat-mungbean intercropping (OMO) was significantly greater than that in sole oat (O), but the nifH gene abundance significantly decreased at the later stage in all the treatments. Alpha diversity indices in OSO and OMO were higher at the heading stage, but lower at the maturity stage than that in O. Bradyrhizobium and Skermanella were the dominant genera identified in all samples, with an average proportion of 35.8% and 12.4%, respectively. The proportion of dominant genera showed significant differences and varied with cropping system and growth stage. Principal component analysis showed that growth stage had a stronger effect than intercropping on the diazotrophic community structure. However, Mantel test and redundancy analysis showed there was no environmental factor significantly correlated to the diazotrophic community structure. Our results demonstrate that intercropping had a weaker effect than growth stage on the abundance, diversity and structure of the diazotrophic community in oat rhizosphere soil.