Efficient Reduction of Iron Oxides by Paenibacillus spp. Strains Isolated from Tropical Soils

The genus Paenibacillus was hardly described as a Fe(III)-reducing agent, only limited to reduce soluble forms or Fe inserted in poorly crystallized structures. In this study, three Paenibacillus strains capable of reducing manganese oxides in addition to iron oxides were isolated from Cameroonian and Brazilian soils. These strains reduced iron minerals from poorly crystallized 2-line ferrihydrite to well-crystallized Al-substituted and pure goethite with a significant production of soluble ferrous iron. These Paenibacillus strains, inhabitants from ferralitic temporarily waterlogged soils, could play an important role in the bioweathering of minerals and metal mobility in soils.     

Phylogeny and Characterization of Three nifH-Homologous Genes from Paenibacillus azotofixans

In this paper, we report the cloning and characterization of three Paenibacillus azotofixans DNA regions containing genes involved in nitrogen fixation. Sequencing analysis revealed the presence of nifB1H1D1K1 gene organization in the 4,607-bp SacI DNA fragment. This is the first report of linkage of a nifB open reading frame upstream of the structural nif genes. The second (nifB2H2) and third (nifH3) nif homologues are confined within the 6,350-bp HindIII and 2,840-bp EcoRI DNA fragments, respectively. Phylogenetic analysis demonstrated that NifH1 and NifH2 form a monophyletic group among cyanobacterial NifH proteins. NifH3, on the other hand, clusters among NifH proteins of the highly divergent methanogenic archaea.     

Phylogeny and characterization of three nifH-homologous genes from Paenibacillus azotofixans

In this paper, we report the cloning and characterization of three Paenibacillus azotofixans DNA regions containing genes involved in nitrogen fixation. Sequencing analysis revealed the presence of nifB1H1D1K1 gene organization in the 4,607-bp SacI DNA fragment. This is the first report of linkage of a nifB open reading frame upstream of the structural nif genes. The second (nifB2H2) and third (nifH3) nif homologues are confined within the 6,350-bp HindIII and 2,840-bp EcoRI DNA fragments, respectively. Phylogenetic analysis demonstrated that NifH1 and NifH2 form a monophyletic group among cyanobacterial NifH proteins. NifH3, on the other hand, clusters among NifH proteins of the highly divergent methanogenic archaea.     

Genetic Diversity of nifH Gene Sequences in Paenibacillus azotofixans Strains and Soil Samples Analyzed by Denaturing Gradient Gel Electrophoresis of PCR-Amplified Gene Fragments

The diversity of dinitrogenase reductase gene (nifH) fragments in Paenibacillus azotofixans strains was investigated by using molecular methods. The partial nifH gene sequences of eight P. azotofixans strains, as well as one strain each of the close relatives Paenibacillus durum, Paenibacillus polymyxa, and Paenibacillus macerans, were amplified by PCR by using degenerate primers and were characterized by DNA sequencing. We found that there are two nifH sequence clusters, designated clusters I and II, in P. azotofixans. The data further indicated that there was sequence divergence among the nifH genes of P. azotofixans strains at the DNA level. However, the gene products were more conserved at the protein level. Phylogenetic analysis showed that all nifH cluster II sequences were similar to the alternative (anf) nitrogenase sequence. A nested PCR assay for the detection of nifH (cluster I) of P. azotofixans was developed by using the degenerate primers as outer primers and two specific primers, designed on the basis of the sequence information obtained, as inner primers. The specificity of the inner primers was tested with several diazotrophic bacteria, and PCR revealed that these primers are specific for the P. azotofixans nifH gene. A GC clamp was attached to one inner primer, and a denaturing gradient gel electrophoresis (DGGE) protocol was developed to study the genetic diversity of this region of nifH in P. azotofixans strains, as well as in soil and rhizosphere samples. The results revealed sequence heterogeneity among different nifH genes. Moreover, nifH is probably a multicopy gene in P. azotofixans. Both similarities and differences were detected in the P. azotofixans nifH DGGE profiles generated with soil and rhizosphere DNAs. The DGGE assay developed here is reproducible and provides a rapid way to assess the intraspecific genetic diversity of an important functional gene in pure cultures, as well as in environmental samples.     

Nutritional Requirements of Acinetobacter Strains Isolated from Soil, Water, and Sewage

One hundred five strains of Acinetobacter were isolated from water, soil, and sewage on nonselective complex media, and their nutritional properties were studied. Only one of these strains requires growth factors in order to grow in a mineral medium containing a single carbon source.     

Frequency and Biodiversity of 2,4-Diacetylphloroglucinol-Producing Bacteria Isolated from the Maize Rhizosphere at Different Stages of Plant Growth

A Pseudomonas 2,4-diacetylphloroglucinol (DAPG)-producing population that occurred naturally on the roots, in rhizosphere soil of Zea mays and in the nonrhizosphere soil was investigated in order to assess the microbial diversity at five stages of plant growth. A total of 1,716 isolates were obtained, and 188 of these isolates were able to produce DAPG. DAPG producers were isolated at each stage of plant growth, indicating that the maize rhizosphere is colonized by natural DAPG producers throughout development. The frequency of DAPG producers was very low in the first stage of plant growth and increased over time. An analysis of the level of biodiversity of the DAPG producers at the species level was performed by comparing the AluI restriction patterns of the 16S ribosomal DNAs (rDNAs) amplified by PCR from 167 isolates. This comparison allowed us to cluster the isolates into four amplified rDNA restriction analysis (ARDRA) groups, and the main group (ARDRA group 1) contained 89.8% of the isolates. The diversity of the 150 isolates belonging to ARDRA group 1 was analyzed by the random amplified polymorphic DNA (RAPD) technique. An analysis of RAPD patterns by a molecular variance method revealed that there was a high level of genetic diversity in this population and that the genetic diversity was related to plant age. Finally, we found that some of the DAPG producers, which originated from all stages of plant growth, had the same genotype. These DAPG producers could be exploited in future screening programs for biocontrol agents.     

两个玉米品系叶片光合速率差异的生理机制初探

田间生长的两个玉米品系"双105"和"40×44"叶片净光合速率(Pn)存在显著差异。在不同的生长阶段,"40×44"Pn值均比"双105"高10%~32%,而且PSII光化学效率(Fv/Fm和△F/Fm')也较高,表现为植株较高,叶片较大,但叶绿素含量较低。尽管"双105"品系具有较低的光合速率和较低的气孔导度(Gs),但其胞间CO₂浓度(Ci)却略高于"40×44"。因此,这两个玉米品系叶片光合速率的差异并非气孔因素,而可能源于光合机构的光反应系统。     

Biocontrol and PGPR Features in Native Strains Isolated from Saline Soils of Argentina

A bacterial collection of approximately one thousand native strains, isolated from saline soils of Cordoba province (Argentina), was established. From this collection, a screening to identify those strains showing plant growth promotion and biocontrol activities, as well as salt tolerance, was performed. Eight native strains tolerant to 1 M NaCl and displaying plant growth promotion and/or biocontrol features were selected for further characterization. Strains MEP₂ 18, MRP₂ 26, MEP₂ 11a, MEP₃ 1, and MEP₃ 3b significantly increased the growth of maize seedlings under normal and saline conditions, whereas isolates ARP₂ 3, AEP₁ 5, and ARP₂ 6 were able to increase the root dry weight of agropyre under saline conditions. On the other hand, strains MEP₂ 18 and ARP₂ 3 showed antagonistic activity against phytopathogenic fungi belonging to Sclerotinia and Fusarium genus. Antifungal activity was found in cell-free supernatants, and it was heat and protease resistant. Strains MEP₂18 and ARP₂3 were identified as Bacillus sp. and strains MEP₂11a and MEP₃3b as Ochrobactrum sp. according to the sequence analysis of 16S rRNA gene.     

Genotypic and Phenotypic Diversity of phlD-Containing Pseudomonas Strains Isolated from the Rhizosphere of Wheat

Production of 2,4-diacetylphloroglucinol (2,4-DAPG) in the rhizosphere by strains of fluorescent Pseudomonas spp. results in the suppression of root diseases caused by certain fungal plant pathogens. In this study, fluorescent Pseudomonas strains containing phlD, which is directly involved in the biosynthesis of 2,4-DAPG, were isolated from the rhizosphere of wheat grown in soils from wheat-growing regions of the United States and The Netherlands. To assess the genotypic and phenotypic diversity present in this collection, 138 isolates were compared to 4 previously described 2,4-DAPG producers. Thirteen distinct genotypes, one of which represented over 30% of the isolates, were differentiated by whole-cell BOX-PCR. Representatives of this group were isolated from eight different soils taken from four different geographic locations. ERIC-PCR gave similar results overall, differentiating 15 distinct genotypes among all of the isolates. In most cases, a single genotype predominated among isolates obtained from each soil. Thirty isolates, representing all of the distinct genotypes and geographic locations, were further characterized. Restriction analysis of amplified 16S rRNA gene sequences revealed only three distinct phylogenetic groups, one of which accounted for 87% of the isolates. Phenotypic analyses based on carbon source utilization profiles revealed that all of the strains utilized 49 substrates and were unable to grow on 12 others. Individually, strains could utilize about two-thirds of the 95 substrates present in Biolog SF-N plates. Multivariate analyses of utilization profiles revealed phenotypic groupings consistent with those defined by the genotypic analyses.     

两株乙型肝炎病毒基因组结构分析及其复制和抗原表达特性的比较

为从全基因组水平研究乙型肝炎病毒（ＨＢＶ）的核苷酸结构及复制和抗原表达特性,分别从２例ＨＢｓＡｇ和ＨＢｅＡｇ阳性、ＨＢＶ ＤＮＡ滴度为１０＾１４和１０＾１３拷贝／ｍｌ的慢性乙型肝炎患者（编号为５６和２－１８）血清中扩增、克隆了ＨＢＶ全基因组,并测定了核苷酸序列。两株病毒基因组转染ＨｅｐＧ２细胞的培养上清中ＨＢｓＡｇ水平基本相同,但＃５６毒株表达的ＨＢｅＡｇ约为＃２－１８株的３倍,Ｓｏｕｔｈｅｒｍ印     

Identification of Yeast Strains Isolated from Agricultural Soils for Releasing Potassium-bearing Minerals

Thirty-five yeast strains were isolated from soil samples that were collected from different locations in Upper Egypt. The purified isolates were screened for the release potassium from mica on Aleksandrov agar medium. Two yeast isolates (KSY-29 and KSY-33) showed an ability to solubilize potassium by inducing clear zones around their colonies. They were identified as Pichia anomala and Rhodotorula glutinis, respectively, based on PCR analysis of the ITSI-26S region that was amplified by NL1/NL4 species-specific primers. The amount of K released from muscovite mica in the broth culture of the yeast isolates was measured after 5, 10, 15 and 20 days of the incubation at 25°C. Both yeast isolates were very effective in releasing K of muscovite in broth culture, recording 8.11–13.21 μg/ml that were released from muscovite mica after 20 days of incubation. The inoculation of maize (Zea maize) plants with these yeast isolates under different K levels (25, 50 and 100% of recommended dose of potassium, RDK) as potassium sulfate was tested on growth and K uptake by these plants in the greenhouse. Significant increases (p < 0.05) in plant height, root and shoot dry weights as well as K uptakes by shoots and roots maize plants occurred through the inoculation with KSY-29 or KSY-33 isolates.     

Potential of Siderophore Production by Bacteria Isolated from Heavy Metal: Polluted and Rhizosphere Soils

Recently, heavy metals have been shown to have a stimulating effect on siderophore biosynthesis in various bacteria. In addition, several studies have found that siderophore production is greater in bacteria isolated from soil near plant roots. The aim of this study was to compare the production of siderophores by bacterial strains isolated from heavy metal-contaminated and uncontaminated soils. Chrome azurol sulphonate was used to detect siderophore secretion by several bacterial strains isolated from heavy metal-contaminated and rhizosphere-uncontaminated soils with both a qualitative disc diffusion method and a quantitative ultraviolet spectrophotometric method. Siderophore production by rhizosphere bacteria was significantly greater than by bacteria isolated from contaminated soil. The Pearson’s correlation test indicated a positive correlation between the amount of siderophore produced by bacteria isolated from the rhizosphere using the quantitative and qualitative detection methods and the amount of heavy metal in the soil. However, a significant negative correlation was observed between the amount of siderophore produced by bacteria isolated from heavy metal-contaminated soil and the amount of heavy metal (r value of ?0.775, P < 0.001).     

两种菌株来源的glyA基因的克隆、表达及酶活性检测

采用PCR方法,分别从大肠杆菌和嗜热链球菌基因组DNA中扩增获得glyA基因,分别克隆入载体pET-28 a(+)中并进行表达,分离和纯化得到两种不同来源的SHMT,分别检测两种SHMT的逆向酶活。比较来源于大肠杆菌K12与嗜热链球菌AS1.2471中的glyA基因表达的丝氨酸羟甲基转移酶(SHMT)的活性,以获得高活性的SHMT。结果成功获得两种菌中的glyA基因,并表达出具有较高活性的SHMT,其中嗜热链球菌中glyA基因表达出的SHMT的酶活性大约为大肠杆菌的两倍。从嗜热链球菌中克隆表达的SHMT具有更高的催化活性及良好的工业应用前景。     

Complete Genome Sequence of the Bacteriophages ECBP1 and ECBP2 Isolated from Two Different Escherichia coli Strains

Escherichia coli is recognized as one of the most abundant avian bacterial pathogens. In this study, we report the sequencing by the traditional Sanger method of ECBP1 and ECBP2: bacteriophages that infected two different E. coli strains which might be used as therapeutic agents in combination with alternative antibiotics.     

Soil Type Dependent Rhizosphere Competence and Biocontrol of Two Bacterial Inoculant Strains and Their Effects on the Rhizosphere Microbial Community of Field-Grown Lettuce

Rhizosphere competence of bacterial inoculants is assumed to be important for successful biocontrol. Knowledge of factors influencing rhizosphere competence under field conditions is largely lacking. The present study is aimed to unravel the effects of soil types on the rhizosphere competence and biocontrol activity of the two inoculant strains Pseudomonas jessenii RU47 and Serratia plymuthica 3Re4-18 in field-grown lettuce in soils inoculated with Rhizoctonia solani AG1-IB or not. Two independent experiments were carried out in 2011 on an experimental plot system with three soil types sharing the same cropping history and weather conditions for more than 10 years. Rifampicin resistant mutants of the inoculants were used to evaluate their colonization in the rhizosphere of lettuce. The rhizosphere bacterial community structure was analyzed by denaturing gradient gel electrophoresis of 16S rRNA gene fragments amplified from total community DNA to get insights into the effects of the inoculants and R. solani on the indigenous rhizosphere bacterial communities. Both inoculants showed a good colonization ability of the rhizosphere of lettuce with more than 10⁶ colony forming units per g root dry mass two weeks after planting. An effect of the soil type on rhizosphere competence was observed for 3Re4-18 but not for RU47. In both experiments a comparable rhizosphere competence was observed and in the presence of the inoculants disease symptoms were either significantly reduced, or at least a non-significant trend was shown. Disease severity was highest in diluvial sand followed by alluvial loam and loess loam suggesting that the soil types differed in their conduciveness for bottom rot disease. Compared to effect of the soil type of the rhizosphere bacterial communities, the effects of the pathogen and the inoculants were less pronounced. The soil types had a surprisingly low influence on rhizosphere competence and biocontrol activity while they significantly affected the bottom rot disease severity.     

两株筛自大规模生产跑道池的节旋藻性能比较研究

温度是影响节旋藻大规模培养中生物质产率的重要因素。筛选高产和耐受温度胁迫的优良藻株对提高节旋藻产量具有重要意义。从生产规模跑道池中分离到形态特征差异显著而亲缘关系很近的两株节旋藻Arthrospira sp.DICP-D(D)和Arthrospira sp.DICP-F(F),对其生物质积累性能和对低温、高温胁迫的耐受能力进行了评估。结果显示,尽管藻株D和藻株F在正常条件下拥有相似的生物质组成,在N胁迫和低温胁迫下拥有相同的碳水化合物积累能力,然而藻株D的生物质产率比相同条件下的藻株F高33%~230%。藻株D对高温胁迫的耐受能力是藻株F的2.1倍。藻株D在41℃和15℃下的生物质产率分别维持在正常温度下的73%和61%,显示其对温度胁迫的良好耐受能力。藻株D比藻株F拥有更有效的光保护机制,使其能够更好地适应胁迫环境。藻株D在生物质产率和胁迫耐受能力上的优良性状使其在户外大规模培养中具有良好的应用前景。     

Functional Potential of Soil Microbial Communities in the Maize Rhizosphere

Microbial communities in the rhizosphere make significant contributions to crop health and nutrient cycling. However, their ability to perform important biogeochemical processes remains uncharacterized. Here, we identified important functional genes that characterize the rhizosphere microbial community to understand metabolic capabilities in the maize rhizosphere using the GeoChip-based functional gene array method. Significant differences in functional gene structure were apparent between rhizosphere and bulk soil microbial communities. Approximately half of the detected gene families were significantly (p<0.05) increased in the rhizosphere. Based on the detected gyrB genes, Gammaproteobacteria, Betaproteobacteria, Firmicutes, Bacteroidetes and Cyanobacteria were most enriched in the rhizosphere compared to those in the bulk soil. The rhizosphere niche also supported greater functional diversity in catabolic pathways. The maize rhizosphere had significantly enriched genes involved in carbon fixation and degradation (especially for hemicelluloses, aromatics and lignin), nitrogen fixation, ammonification, denitrification, polyphosphate biosynthesis and degradation, sulfur reduction and oxidation. This research demonstrates that the maize rhizosphere is a hotspot of genes, mostly originating from dominant soil microbial groups such as Proteobacteria, providing functional capacity for the transformation of labile and recalcitrant organic C, N, P and S compounds.     

High Salt and High pH Tolerance of New Isolated Rhizobium etli Strains from Egyptian Soils

Saline and alkaline soils are major problems contributing to the low productivity of common bean (Phaseolus vulgaris) in arid and semi-arid regions such as Egypt. Therefore our study was directed toward selecting strains more tolerant to these environmental stresses. Among seven Rhizobium etli strains isolated from Egyptian soils, we found a high degree of diversity. Strains EBRI 21 and EBRI 26 are highly tolerant to a salt concentration up to 4% NaCl. A positive correlation was found between the salt tolerance and the adaptation to alkaline pH (9). Strains EBRI 2 and EBRI 26 were adapted to elevated temperatures (42°C). The minimum level of low pH for the majority of Rhizobium etli strains from Egypt was pH 4.7 while the Colombian strain Rhizobium tropici CIAT 899 survived well at pH 4. At 0.4% NaCl, the symbiotic efficiency of the salt-tolerant strain EBRI 26 was superior in cultivar Giza 6 compared with the salt-sensitive strain EBRI 2 (18.2 compared with 13.9 nM C₂H₄ h^–1 mg^–1 nodule fresh weight). In the bean cultivar Saxa, nitrogen fixation was much more affected by high salt concentration (0.4% NaCl) than in the cultivar Giza 6 with both strains (3.9 and 3.8 nM C₂H₄ h^–1 mg^–1 nodule fresh weight, respectively). In general, stress of alkalinity had a less detrimental effect on nodulation and N₂ fixation than stress of salinity.