Isolation and characterization of bacteria associated with two sand dune plant species, Calystegia soldanella and Elymus mollis

Little is known about the bacterial communities associated with the plants inhabiting sand dune ecosystems. In this study, the bacterial populations associated with two major sand dune plant species, Calystegia soldanella (beach morning glory) and Elymus mollis (wild rye), growing along the costal areas in Tae-An, Chungnam Province, were analyzed using a culture-dependent approach. A total of 212 bacteria were isolated from the root and rhizosphere samples of the two plants, and subjected to further analysis. Based on the analysis of the 16S rDNA sequences, all the bacterial isolates were classified into six major phyla of the domain Bacteria. Significant differences were observed between the two plant species, and also between the rhizospheric and root endophytic communities. The isolates from the rhizosphere of the two plant species were assigned to 27 different established genera, and the root endophytic bacteria were assigned to 21. Members of the phylum Gammaproteobacteria, notably the Pseudomonas species, comprised the majority of both the rhizospheric and endophytic bacteria, followed by members of Bacteroidetes and Firmicutes in the rhizosphere and Alphaproteobacteria and Bacteroidetes in the root. A number of isolates were recognized as potentially novel bacterial taxa. Fifteen out of 27 bacterial genera were commonly found in the rhizosphere of both plants, which was comparable to 3 out of 21 common genera in the root, implying the host specificity for endophytic populations. This study of the diversity of culturable rhizospheric and endophytic bacteria has provided the basis for further investigation aimed at the selection of microbes for the facilitation of plant growth.     

Cultivation-dependent characterization of rhizobacterial communities from field grown Chinese cabbage Brassica campestris ssp pekinensis and screening of traits for potential plant growth promotion

The composition of the bacterial community associated with plant roots is influenced by a variety of plant, environmental factors and also management practices. Our study aimed at detecting the root associated bacterial communities of Chinese cabbage under different fertilization regimes using cultivation dependent methods. The cultivable population was studied using plate count assay, fatty acid methyl ester (FAME) analysis and carbon substrate utilization␣(SU)using BIOLOG™ plates. Taxonomical identification of the isolates by FAME resulted in about 83% identification and they represented 9 and 14 different known bacterial genera from the rhizosphere and root interior respectively from Proteobacteria (α, β, and γ), firmicutes (actinobacteria and the Bacillus groups) and Bacteroidetes. Pseudomonas and Bacillus were associated with the plants grown under all the fertilized conditions and actinobacteria could be observed only in rhizosphere of plants grown on unfertilized plots. FAME and BIOLOG profiles of the rhizosphere and endophytic isolates could separate them with reference to fertilization. Principal component analysis (PCA) on the BIOLOG SU revealed that the isolates were metabolically dissimilar. The diversity, as revealed by the diversity indices was greater among the isolates obtained from unfertilized samples than that of fertilized ones. The isolates analyzed for different traits related to plant growth promotion revealed differences between rhizosphere and endophytic isolates and also with reference to the treatments. The highest percentage of phosphate solubilizing bacteria (PSB) and 1-aminocyclopropane-1-carboxylic acid (ACC) utilizers was recorded in chemical fertilizer treated samples, followed by the organic fertilizer treated. The results from this study indicate that fertilizers have an effect on the root associated bacterial communities of Chinese cabbage and also on their physiological characteristics related to plant growth promotion.     

Isolation and characterization of pyrene metabolizing microbial consortia from the plant rhizoplane

Most research on the ecology of PAH degrading bacteria in the rhizosphere has focused on individual strains that grow on specific PAHs. Thus, there are fundamental questions as to importance of microbial consortia for PAH degradation in the plant rhizosphere. The study reported here characterized cultivable pyrene degrading rhizoplane microbial communities from two different plant species using a root printing technique on agar plates. Colonies were revealed by formation of clearing zones on medium containing a thin film of pyrene on the surface of a mineral nutrient agar. Prints of the rhizoplane colonies were obtained from roots of Melilotus officinalis (sweet yellow clover) and Andropogon gerardii (big bluestem) plants. Phylogenetic characterizations of selected pyrene degrading colonies were assessed by PCR-DGGE and DNA sequencing. Results showed that different populations of cultivable pyrene degraders were obtained from representative consortia that were examined. Many of the PAH degrading consortia consisted of mixtures of bacterial species that were unable to degrade pyrene by themselves. While this study focused on culturable PAH degraders, the results suggest that pyrene degradation in the rhizosphere commonly involves the activity of bacterial consortia in which various species of bacteria interact to achieve PAH degradation.     

小蓬竹根际土壤微生物及内生真菌多样性分析

为了解喀斯特典型物种-小蓬竹根际土壤微生物及不同部位内生真菌多样性,采用沿等高线等距离取样法采集小蓬竹根际土壤及健康植株,通过可培养对根际土微生物及内生菌进行分离,利用分子技术对其进行鉴定,根据鉴定结果构建系统发育树,并计算小蓬竹根际土壤微生物和根茎叶内生真菌多样性。结果如下：（1）共从根际土壤、根、茎、叶分离得到139个真菌菌株,隶属于27属,其中根际土壤分离得到34个真菌菌株隶属于12属,根部分离得到的63个内生真菌菌株隶属于17个属,茎部分离得到的14个内生真菌菌株隶属于8个属,叶部分离得到28个内生真菌菌株隶属于9个属;（2）根际土壤共分离得到41株细菌菌株,隶属于7个属26个种,20株放线菌菌株,隶属于1属15种;从Shannon-Wiener多样性指数、均匀度指数、Simpson指数排序来看,真菌主要表现为根 > 根际土壤 > 茎 > 叶,细菌和放线菌多样性均较低。（3）按层次聚类分析可分别将真菌、细菌、放线菌聚为3支。小蓬竹根际土壤、根、茎和叶具有丰富的微生物多样性,不同部位菌群组成存在差异性（P<0.05）,且存在以假单胞菌属、芽孢杆菌属等为优势属的抗盐耐旱菌群,这有助于揭示小蓬竹对喀斯特生境的适应性,以及为微生物-植物群落之间相互关系提供一定基础数据,为后期寻找小蓬竹相关耐性功能菌奠定基础。     

Bulk and rhizosphere soil bacterial communities studied by denaturing gradient gel electrophoresis: plant-dependent enrichment and seasonal shifts revealed

The bacterial rhizosphere communities of three host plants of the pathogenic fungus Verticillium dahliae, field-grown strawberry (Fragaria ananassa Duch.), oilseed rape (Brassica napus L.), and potato (Solanum tuberosum L.), were analyzed. We aimed to determine the degree to which the rhizosphere effect is plant dependent and whether this effect would be increased by growing the same crops in two consecutive years. Rhizosphere or soil samples were taken five times over the vegetation periods. To allow a cultivation-independent analysis, total community DNA was extracted from the microbial pellet recovered from root or soil samples. 16S rDNA fragments amplified by PCR from soil or rhizosphere bacterium DNA were analyzed by denaturing gradient gel electrophoresis (DGGE). The DGGE fingerprints showed plant-dependent shifts in the relative abundance of bacterial populations in the rhizosphere which became more pronounced in the second year. DGGE patterns of oilseed rape and potato rhizosphere communities were more similar to each other than to the strawberry patterns. In both years seasonal shifts in the abundance and composition of the bacterial rhizosphere populations were observed. Independent of the plant species, the patterns of the first sampling times for both years were characterized by the absence of some of the bands which became dominant at the following sampling times. Bacillus megaterium and Arthrobacter sp. were found as predominant populations in bulk soils. Sequencing of dominant bands excised from the rhizosphere patterns revealed that 6 out of 10 bands resembled gram-positive bacteria. Nocardia populations were identified as strawberry-specific bands.     

Chemical characterization of root exudates from rice (Oryza sativa) and their effects on the chemotactic response of endophytic bacteria

Root exudates represent an important source of nutrients for microorganisms in the rhizosphere and seem to participate in early colonization inducing chemotactic responses of rhizospheric bacteria. We characterized the root exudates collected from rice plantlets cultured under hydroponic conditions and assessed their effects on the chemotaxis of two strains of endophytic bacteria, Corynebacterium flavescens and Bacillus pumilus, collected from the rice rhizosphere. We compared these chemotactic effects on endophytic bacteria with those on two strains of plant-growth-promoting bacteria, Azospirillum brasilense (isolated from the corn rhizosphere) and Bacillus sp. (from the rice rhizosphere). The root exudates were collected at different time intervals. The highest concentration and diversity of amino acids and carbohydrates were found during the first 2 weeks after seeding. Histidine, proline, valine, alanine, and glycine were the main amino acid residues identified during the 4 weeks of culture. The main carbohydrates identified were glucose, arabinose, mannose, galactose, and glucuronic acid. The chemotactic responses of the analyzed endophytic bacteria to root exudates were 3.9 to 5.1 times higher than those of A. brasilense and 2.2 to 2.8 times higher than Bacillus sp. Our results indicate that rice exudates may induce a higher chemotactic response for endophytic bacteria than for other bacterial strains present in the rice rhizosphere.     

Fatty acid methyl ester (FAME) profiles as a tool to investigate community structure of two agricultural soils

Soil microbiological parameters may be the earliest predictors of soil quality changes. Recently, molecular techniques such as fatty acid methyl ester (FAME) profiles have been used to characterize soil microbial communities. Fatty acid methyl ester (FAME) from whole soil may be derived from live cells, dead cells, humic materials, as well as plant and root exudates. Our objective was to verify differences in FAME profiles from two agricultural soils with different plants. Soil samples were collected from Ritzville and Palouse silt loams for fatty acid analysis. Soil samples from wheat (Triticum aestivum L.), barley (Hordeum vulgare L.), pea (Pisum sativum L.), jointed goatgrass ( Aegilops cylindrica L.) and downy brome (Bromus tectorum L.) rhizospheres were also collected for fatty acid analysis. Principal component analysis (PCA) of the two soils explained 42% of the variance on PC1, which accounted for Palouse soil. Ritzville soil accounted for 19% of the variance on PC2. Factor analysis showed that rhizosphere microbial communities from various plant species may differ depending on the plant species. Presence of Gram-positive bacteria as identified by a15:0, i15:0, a17:0 and i17:0 peaks were similar between rhizosphere and nonrhizosphere soils. Gram-negative bacteria characterized by short chain hydroxy acids (10:03OH and 12:03OH) as well as cyclopropane acids (cy17:0) were higher in rhizosphere soil than nonrhizosphere. This indicates a possible shift in the bacterial community to more Gram-negative bacteria and fewer Gram-positive bacteria in the rhizospheres of the plants species studied.     

Characterization of rhizosphere and endophytic bacterial communities from leaves,stems and roots of medicinal Stellera chamaejasme L.

A diverse array of bacteria that inhabit the rhizosphere and different plant organs play a crucial role in plant health and growth. Therefore, a general understanding of these bacterial communities and their diversity is necessary. Using the 16S rRNA gene clone library technique, the bacterial community structure and diversity of the rhizosphere and endophytic bacteria in Stellera chamaejasme compartments were compared and clarified for the first time. Grouping of the sequences obtained showed that members of the Proteobacteria (43.2%), Firmicutes (36.5%) and Actinobacteria (14.1%) were dominant in both samples. Other groups that were consistently found, albeit at lower abundance, were Bacteroidetes (2.1%), Chloroflexi (1.9%), and Cyanobacteria (1.7%). The habitats (rhizosphere vs endophytes) and organs (leaf, stem and root) structured the community, since the Wilcoxon signed rank test indicated that more varied bacteria inhabited the rhizosphere compared to the organs of the plant. In addition, correspondence analysis also showed that differences were apparent in the bacterial communities associated with these distinct habitats. Moreover, principal component analysis revealed that the profiles obtained from the rhizosphere and roots were similar, whereas leaf and stem samples clustered together on the opposite side of the plot from the rhizosphere and roots. Taken together, these results suggested that, although the communities associated with the rhizosphere and organs shared some bacterial species, the associated communities differed in structure and diversity.     

Distinct Microbial Communities within the Endosphere and Rhizosphere of Populus deltoides Roots across Contrasting Soil Types

The root-rhizosphere interface of Populus is the nexus of a variety of associations between bacteria, fungi, and the host plant and an ideal model for studying interactions between plants and microorganisms. However, such studies have generally been confined to greenhouse and plantation systems. Here we analyze microbial communities from the root endophytic and rhizospheric habitats of Populus deltoides in mature natural trees from both upland and bottomland sites in central Tennessee. Community profiling utilized 454 pyrosequencing with separate primers targeting the V4 region for bacterial 16S rRNA and the D1/D2 region for fungal 28S rRNA genes. Rhizosphere bacteria were dominated by Acidobacteria (31%) and Alphaproteobacteria (30%), whereas most endophytes were from the Gammaproteobacteria (54%) as well as Alphaproteobacteria (23%). A single Pseudomonas-like operational taxonomic unit (OTU) accounted for 34% of endophytic bacterial sequences. Endophytic bacterial richness was also highly variable and 10-fold lower than in rhizosphere samples originating from the same roots. Fungal rhizosphere and endophyte samples had approximately equal amounts of the Pezizomycotina (40%), while the Agaricomycotina were more abundant in the rhizosphere (34%) than endosphere (17%). Both fungal and bacterial rhizosphere samples were highly clustered compared to the more variable endophyte samples in a UniFrac principal coordinates analysis, regardless of upland or bottomland site origin. Hierarchical clustering of OTU relative abundance patterns also showed that the most abundant bacterial and fungal OTUs tended to be dominant in either the endophyte or rhizosphere samples but not both. Together, these findings demonstrate that root endophytic communities are distinct assemblages rather than opportunistic subsets of the rhizosphere.     

应用LH-PCR研究党参、麻黄和独一味内生细菌多样性

通过直接提取药用植物样品的总DNA,采用长度多态片段PCR (length heterogeneity PCR, LH-PCR)技术研究四川省甘孜藏族自治州的党参、麻黄和独一味3种药用植物内生细菌多样性.结果表明： 同种植物根、茎、叶的LH-PCR图谱相似度很高,条带丰富度差别不大;但不同植物样品之间的差异较大.党参的植物样品条带丰富度最大,麻黄次之,独一味最低.3种药用植物中474 bp长度的细菌是绝对的优势菌群.植物内生细菌多样性与土壤速效磷呈负相关,而与土壤pH值呈正相关.海拔和土壤总氮是影响植物样品内生细菌多样性分布的两个重要环境因子.LH-PCR所得的种群信息能较直观地反映不同植物样品间细菌多样性的差异.因此LH-PCR适用于分析药用植物内生菌多样性.     

南京小龙山钾矿区植物根际可培养细菌的遗传多样性分析

摘要:【目的】矿区优势植物可培养细菌生物多样性研究将有助于了解植物根际细菌与矿物,植物根系相互作用及对矿物风化和土壤形成的重要影响。【方法】采用纯培养法分离南京小龙山废钾矿区野生植物野塘蒿,千金子和栽培植物甘薯根内与根周围土壤的可培养细菌, 通过16S rDNA限制性酶切多态性分析(amplified rDNA restriction analysis, ARDRA)和16S rDNA序列分析研究了可培养细菌的多样性。【结果】分离纯化到60株具不同菌落形态的可培养细菌, 在60%相似水平上可分为18个OTU. 19株代表菌株分别属于3个门, 10个科, 11个属。多数菌株属于变形菌门(α-proteobacteria, 4株, 21.1%; β-proteobacteria, 2株, 10.5%; γ-proteobacteria, 6株, 31.6%)。假单胞菌属(Pseudomonas), 泛菌属(Pantoea)和根瘤菌属(Rhizobium)为优势种群。【结论】小龙山废矿区优势植物根围具有丰富的微生物种群多样性。     

Distribution of Endophytic Bacteria in Alopecurus aequalis Sobol and Oxalis corniculata L. from Soils Contaminated by Polycyclic Aromatic Hydrocarbons

The distributions of endophytic bacteria in Alopecurus aequalis Sobol and Oxalis corniculata L. grown in soils contaminated with different levels of polycyclic aromatic hydrocarbons (PAHs) were investigated with polymerase chain reaction followed by denaturing gradient gel electrophoresis technology (PCR-DGGE) and cultivation methods. Twelve types of PAHs, at concentrations varying from 0.16 to 180 mg·kg⁻¹, were observed in the roots and shoots of the two plants. The total PAH concentrations in Alopecurus aequalis Sobol obtained from three different PAH-contaminated stations were 184, 197, and 304 mg·kg⁻¹, and the total PAH concentrations in Oxalis corniculata L. were 251, 346, and 600 mg·kg⁻¹, respectively. The PCR-DGGE results showed that the endophytic bacterial communities in the roots and shoots of the two plants were quite different, although most bacteria belonged to Firmicutes, Proteobacteria, Actinobacteria and Bacteroidetes. A total of 68 endophytic bacterial strains were isolated from different tissues of the two plants and classified into three phyla: Firmicutes, Proteobacteria and Bacteroidetes. In both plants, Bacillus spp. and Pseudomonas spp. were the dominant cultivable populations. With an increase in the PAH pollution level, the diversity and distribution of endophytic bacteria in the two plants changed correspondingly, and the number of cultivable endophytic bacterial strains decreased rapidly. Testing of the isolated endophytic bacteria for tolerance to each type of PAH showed that most isolates could grow well on Luria-Bertani media in the presence of different PAHs, and some isolates were able to grow rapidly on a mineral salt medium with a single PAH as the sole carbon and energy source, indicating that these strains may have the potential to degrade PAHs in plants. This research provides the first insight into the characteristics of endophytic bacterial populations under different PAH pollution levels and provides a species resource for the isolation of PAH-degrading endophytic bacteria.     

Comparison of aerobic heterotrophic taxa isolated from four root domains of mature sugar beet (Beta vulgaris)

Abstract: The distribution of bacteria in the rhizosphere, rhizoplane, interior root tissues (core) and lower root (all tissues) of mature sugar beet roots ( Beta vulgaris ) was compared. Of 556 isolates, 102 species from 40 genera were identified by fatty acid methyl ester gas-chromatographic (FAME-GC) analysis. The ten most common genera ( Bacillus , 14%; Arthrobacter , 12%; Pseudomonas , 11%; Aureobacterium , 9%; Micrococcus , 6%; Xanthomonas , 5%; Alcaligenes , 4%; Flavobacterium , 3%; Agrobacterium , 3%; Microbacterium , 3%) accounted for 70% of isolates, and were found in each of three root domains (rhizosphere, rhizoplane and interior root tissues) on the two principal sampling occasions. Gram-positive strains were more abundant in the rhizosphere than the rhizoplane. Compared to the rhizoplane, rhizosphere bacterial communities were represented by a less diverse, more hierarchical distribution of species where twice as many isolates formed late developing colonies on isolation plates. Between October and January, the bacteria isolated from root interior tissues acquired a distinct change in taxonomic pattern, with decreased diversity and increased hierarchy. A bacterial continuum of similar taxa was observed which extended from the rhizosphere to interior root tissues.     

Bacterial Biosynthesis of 1-Aminocyclopropane-1-Carboxylate (ACC) Deaminase and Indole-3-Acetic Acid (IAA) as Endophytic Preferential Selection Traits by Rice Plant Seedlings

In this study, bacteria were isolated from the rhizosphere and inside the roots and nodules of berseem clover plants grown in the field in Iran. Two hundred isolates were obtained from the rhizosphere (120 isolates), interior roots (57 isolates), and nodules (23 isolates) of clover plants grown in rotation with rice plants. Production of chitinase, pectinase, cellulase, siderophore, salicylic acid, hydrogen cyanide, indole acetic acid (IAA), 1-aminocyclopropane-1-carboxylate (ACC) deaminase, solubilization of phosphate, antifungal activity against various rice plant pathogen fungi, N₂ fixation, and colonization assay on rice seedlings by these strains was evaluated and compared (endophytic isolates vs. rhizosphere bacteria). The results showed both the number and the ability of plant growth-promoting (PGP) traits were different between endophytic and rhizosphere isolates. A higher percentage of endophytic isolates were positive for production of IAA, ACC deaminase, and siderophore than rhizosphere isolates. Therefore, it is suggested that clover plant may shape its own associated microbial community and act as filters for endophyte communities, and rhizosphere isolates with different (PGP) traits. We also studied the PGP effect of the most promising endophytic and rhizosphere isolates on rice seedlings. A significant relationship among IAA and ACC deaminase production, the size of root colonization, and plant growth (root elongation) in comparison with siderophore production and phosphate solubilization for the isolates was observed. The best bacterial isolates (one endophytic isolate and one rhizosphere isolate), based on their ability to promote rice growth and colonize rice roots, were identified. Based on 16S rDNA sequence analysis, the endophytic isolate CEN7 and the rhizosphere isolate CEN8 were closely related to Pseudomonas putida and Pseudomonas fluorescens, respectively. It seems that PGP trait production (such as IAA, ACC deaminase) may be required for endophytic and rhizosphere competence as compared to other PGP traits in rice seedlings under constant flooded conditions. The study also shows that the presence of diverse rhizobacteria with effective growth-promoting traits associated with clover plants may be used for sustainable crop management under field conditions.     

Isolation of plant-growth-promoting Bacillus strains from soybean root nodules

Endophytic bacteria reside within plant tissues and have often been found to promote plant growth. Fourteen strains of putative endophytic bacteria, not including endosymbiotic Bradyrhizobium strains, were isolated from surface-sterilized soybean (Glycine max. (L.) Merr.) root nodules. These isolates were designated as non-Bradyrhizobium endophytic bacteria (NEB). Three isolates (NEB4, NEB5, and NEB17) were found to increase soybean weight when plants were co-inoculated with one of the isolates and Bradyrhizobium japonicum under nitrogen-free conditions, compared with plants inoculated with B. japonicum alone. In the absence of B. japonicum, these isolates neither nodulated soybean, nor did they affect soybean growth. All three isolates were Gram-positive spore-forming rods. While Biolog tests indicated that the three isolates belonged to the genus Bacillus, it was not possible to determine the species. Phylogenetic analysis of 16S rRNA gene hypervariant region sequences demonstrated that both NEB4 and NEB5 are Bacillus subtilis strains, and that NEB17 is a Bacillus thuringiensis strain.     

新疆北部主要盐生植物根部内生细菌群落结构的高通量分析

【目的】揭示同一盐渍环境中不同种盐生植物根部内生细菌群落多样性特征和分布规律,结合根际土壤理化因子探讨其对内生细菌群落结构的影响。【方法】通过罗氏454高通量测序获得内生细菌16S r RNA片段,然后进行生物信息分析。【结果】研究的16种盐生植物其内生细菌群落主要由Proteobacteria、Tenericutes、Actinobacteria和Firmicutes 4个门的细菌组成。从植物"种"的水平来看,不同种盐生植物内生细菌群落存在差异;从植物"属"的水平来看,同一属的盐生植物内生细菌相似;从植物"科"的水平来看,藜科盐生植物内生细菌以Actinobacteria和Proteobacteria门为主;蒺藜科盐生植物内生细菌以Proteobacteria门为主;柽柳科盐生植物内生细菌以Tenericutes门为主;白花丹科盐生植物内生细菌以Proteobacteria、Fimicutes和Actinobacteria门为主。根际土壤中Cl~–含量对盐生植物内生细菌群落变化具有显著影响;而Cl~–、Mg~(2+)和总氮组成的集合与内生细菌群落结构相关性最高。【结论】盐生植物内生细菌多样性丰富。在同一盐渍生境中,盐生植物内生细菌群落分布呈现宿主的种属特异性,根际土壤中Cl~–是影响其内生细菌群落变化的主要驱动因素之一。     

Diversity of Culturable Bacteria Isolated from Root Domains of Moso Bamboo (Phyllostachys edulis)

The distribution of culturable bacteria in the rhizosphere, rhizoplane, and interior root tissues of moso bamboo plants was investigated in this study. Of the 182 isolates showing different colony characteristics on Luria–Bertani and King B plates, 56 operational taxonomic units of 22 genera were identified by 16S ribosomal RNA gene sequence analysis. The majority of root endophytic bacteria were Proteobacteria (67.5%), while the majority of rhizospheric and rhizoplane bacteria were Firmicutes (66.3% and 70.4%, respectively). The most common genus in both the rhizosphere and on the rhizoplane was Bacillus (42.4% and 44.4%, respectively), while Burkholderia was the most common genus inside the roots, comprising 35.0% of the isolates from this root domain. The endophytic bacterial community was less diverse than the rhizoplane and rhizospheric bacterial communities. Members of Lysinibacillus, Bacillus, and Burkholderia were found in all three root domains, whereas many isolates were found in only a single domain. Our results show that the population diversity of culturable bacteria is abundant in the root domains of moso bamboo plants and that obvious differences exist among the rhizospheric, rhizoplane, and endophytic bacterial communities.     

Bulk and Rhizosphere Soil Bacterial Communities Studied by Denaturing Gradient Gel Electrophoresis: Plant-Dependent Enrichment and Seasonal Shifts Revealed

The bacterial rhizosphere communities of three host plants of the pathogenic fungus Verticillium dahliae, field-grown strawberry (Fragaria ananassa Duch.), oilseed rape (Brassica napus L.), and potato (Solanum tuberosum L.), were analyzed. We aimed to determine the degree to which the rhizosphere effect is plant dependent and whether this effect would be increased by growing the same crops in two consecutive years. Rhizosphere or soil samples were taken five times over the vegetation periods. To allow a cultivation-independent analysis, total community DNA was extracted from the microbial pellet recovered from root or soil samples. 16S rDNA fragments amplified by PCR from soil or rhizosphere bacterium DNA were analyzed by denaturing gradient gel electrophoresis (DGGE). The DGGE fingerprints showed plant-dependent shifts in the relative abundance of bacterial populations in the rhizosphere which became more pronounced in the second year. DGGE patterns of oilseed rape and potato rhizosphere communities were more similar to each other than to the strawberry patterns. In both years seasonal shifts in the abundance and composition of the bacterial rhizosphere populations were observed. Independent of the plant species, the patterns of the first sampling times for both years were characterized by the absence of some of the bands which became dominant at the following sampling times. Bacillus megaterium and Arthrobacter sp. were found as predominant populations in bulk soils. Sequencing of dominant bands excised from the rhizosphere patterns revealed that 6 out of 10 bands resembled gram-positive bacteria. Nocardia populations were identified as strawberry-specific bands.     

再生水灌溉对草坪根际可培养细菌组成影响

【目的】为了了解再生水灌溉对草坪根际可培养细菌群落组成的影响,【方法】采用稀释平板法,对北京市陶然亭公园内再生水灌区及其对照自来水灌区草坪根际细菌进行了分离,并对其16S rDNA序列进行了分析。【结果】16S rDNA序列分析表明自来水样品分离得到的菌落分属于15个属的20个种,而再生水样品分离得到的菌落分属于18个属的24个种。自来水和再生水灌区草坪根际细菌主要包括变形菌门α亚群(Alphaproteobacteria,分别为9.7%和13.4%)、变形菌门β亚群(Betaproteobacteria,分别为8.1%和12.3%)、变形菌门γ亚群(Gammaproteobacteria,分别为17.9%和42.0%)、拟杆菌门(Bacteroidetes,分别为13.0%和2.9%)、厚壁菌门(Firmicutes,分别为23.6%和10.1%)和放线菌门(Actinobacteria,分别为27.6%和19.6%)其中,芽孢杆菌属(Bacillus sp.)是自来水灌区草坪根际优势菌属(23.6%),而不动杆菌属(Acinetobacter sp.)是再生水灌区根际优势菌属(17.4%)。从不同类群优势菌属看,除变形菌门γ亚群受再生水影响优势菌属在两灌区表现出一定的差异外,其余各亚群优势菌属均未受再生水影响,其中,不动杆菌属是再生水灌区变形菌门γ亚群的优势菌属(41.3%),肠杆菌属(Enterobacter sp.)是自来水灌区变形菌门γ亚群的优势菌属(45.4%)。【结论】这表明,再生水灌溉未改变细菌群落组成类型,但改变了不同类型多度分布状况。具体表现为优势种多度值增加及部分非优势种有无。再生水灌区特有机会性致病菌、植物致病菌和重金属耐性细菌的出现表明再生水灌溉中病原微生物及重金属的控制工作还有待于进一步严格。     

Application of terminal restriction fragment length polymorphism (T-RFLP) analysis to monitor effect of biocontrol agents on rhizosphere microbial community of hot pepper (<Emphasis Type="Italic">Capsicum annuum</Emphasis> L.)

Microbial communities in hot pepper (Capsicum annuum L.) cultivation fields under different cultivation methods were investigated by terminal restriction fragment length polymorphism (T-RFLP) analysis. Rhizosphere soil and leaf samples were collected from control, conventional and nature-friendly cultivation fields between May and July, 2009. Two Bacillus subtilis strains were applied to nature-friendly cultivation fields as biocontrol agents during the sampling period. Relative abundances of bacteria and plant pathogenic fungi related T-RFs were also measured to monitor the effect of biocontrol agents on potential plant pathogenic fungi. In the principal component analysis (PCA) based on T-RFLP profiles, the microbial communities from rhizosphere soil samples in July, including bacteria and fungi, showed distinct difference between nature-friendly cultivation fields and other cultivation fields. However, there was no correlation between cultivation methods and leaf microbial communities at any sampling period. Changes in the abundance of bacteria related T-RF in the rhizosphere of nature-friendly cultivation fields were observed clearly two months after application of biocontrol agent, while the abundance of plant pathogenic fungi related T-RFs significantly decreased.