Cloning, sequencing, and disruption of a levanase gene of Bacillus polymyxa CF43.

The Bacillus polymyxa CF43 lelA gene, expressing both sucrose and fructan hydrolase activities, was isolated from a genomic library of B. polymyxa screened in Bacillus subtilis. The gene was detected as expressing sucrose hydrolase activity; B. subtilis transformants did not secrete the lelA gene product (LelA) into the extracellular medium. A 1.7-kb DNA fragment sufficient for lelA expression in Escherichia coli was sequenced. It contains a 548-codon open reading frame. The deduced amino acid sequence shows 54% identity with mature B. subtilis levanase and is similar to other fructanases and sucrases (beta-D-fructosyltransferases). Multiple-sequence alignment of 14 of these proteins revealed several previously unreported features. LelA appears to be a 512-amino-acid polypeptide containing no canonical signal peptide. The hydrolytic activities of LelA on sucrose, levan, and inulin were compared with those of B. subtilis levanase and sucrase, confirming that LelA is indeed a fructanase. The lelA gene in the chromosome of B. polymyxa was disrupted with a chloramphenicol resistance gene (cat) by "inter-gramic" conjugation: the lelA::cat insertion on a mobilizable plasmid was transferred from an E. coli transformant to B. polymyxa CF43, and B. polymyxa transconjugants containing the lelA::cat construct replacing the wild-type lelA gene in their chromosomes were selected directly. The growth of the mutant strain on levan, inulin, and sucrose was not affected.     

Alterations in plant growth and in root hormone levels of lodgepole pines inoculated with rhizobacteria.

The presence of other soil microorganisms might influence the ability of rhizobacterial inoculants to promote plant growth either by reducing contact between the inoculant and the plant root or by interfering with the mechanism(s) involved in rhizobacterially mediated growth promotion. We conducted the following experiments to determine whether reductions in the extent of growth promotion of lodgepole pine mediated by Paenibacillus polymyxa occur in the presence of a forest soil isolate (Pseudomonas fluorescens M20) and whether changes in plant growth promotion mediated by P. polymyxa (i) are related to changes in P. polymyxa density in the rhizosphere or (ii) result from alterations in root hormone levels. The extent of plant growth, P. polymyxa rhizosphere density, and root hormone concentrations were determined for lodgepole pine treated with (i) a single growth-promoting rhizobacterial strain (P. polymyxa L6 or Pw-2) or (ii) a combination of bacteria: strain L6 + strain M20 or strain Pw-2 + strain M20. There was no difference in the growth of pines inoculated with strain L6 and those inoculated with strain L6 + strain M20. However, seedlings inoculated with strain Pw-2 had more lateral roots and greater root mass at 12 weeks after inoculation than plants inoculated with strain Pw-2 + strain M20. The extent of growth promotion mediated by P. polymyxa L6 and Pw-2 in each treatment was not correlated to the average population density of each strain in the rhizosphere. Bacterial species-specific effects were observed in root hormone levels: indole-3-acetic acid concentration was elevated in roots inoculated with P. polymyxa L6 or Pw-2, while dihydrozeatin riboside concentration was elevated in roots inoculated with P. fluorescens M20.     

Colonization of Wheat Roots by an Exopolysaccharide-Producing Pantoea agglomerans Strain and Its Effect on Rhizosphere Soil Aggregation

The effect of bacterial secretion of an exopolysaccharide (EPS) on rhizosphere soil physical properties was investigated by inoculating strain NAS206, which was isolated from the rhizosphere of wheat (Triticum durum L.) growing in a Moroccan vertisol and was identified as Pantoea aglomerans. Phenotypic identification of this strain with the Biotype-100 system was confirmed by amplified ribosomal DNA restriction analysis. After inoculation of wheat seedlings with strain NAS206, colonization increased at the rhizoplane and in root-adhering soil (RAS) but not in bulk soil. Colonization further increased under relatively dry conditions (20% soil water content; matric potential, −0.55 MPa). By means of genetic fingerprinting using enterobacterial repetitive intergenic consensus PCR, we were able to verify that colonies counted as strain NAS206 on agar plates descended from inoculated strain NAS206. The intense colonization of the wheat rhizosphere by these EPS-producing bacteria was associated with significant soil aggregation, as shown by increased ratios of RAS dry mass to root tissue (RT) dry mass (RAS/RT) and the improved water stability of adhering soil aggregates. The maximum effect of strain NAS206 on both the RAS/RT ratio and aggregate stability was measured at 24% average soil water content (matric potential, −0.20 MPa). Inoculated strain NAS206 improved RAS macroporosity (pore diameter, 10 to 30 μm) compared to the noninoculated control, particularly when the soil was nearly water saturated (matric potential, −0.05 MPa). Our results suggest that P. agglomerans NAS206 can play an important role in the regulation of the water content (excess or deficit) of the rhizosphere of wheat by improving soil aggregation.     

Improvement in bioavailability of tricalcium phosphate to Cymbopogon martinii var. motia by rhizobacteria, AMF and Azospirillum inoculation

The interactive effects of phosphate solubilizing bacteria, N2 fixing bacteria and arbuscular mycorrhizal fungi (AMF) were studied in a low phosphate alkaline soil amended with tricalcium insoluble source of inorganic phosphate on the growth of an aromatic grass palmarosa (Cymbopogon martinii). The microbial inocula consisted of the AM fungus Glomus aggregatum, phosphate solubilizing rhizobacteria Bacillus polymyxa and N2 fixing bacteria Azospirillum brasilense. These rhizobacteria behaved as "mycorrhiza helper" and enhanced root colonization by G. aggregatum in presence of tricalcium phosphate at the rate of 200 mg kg(-1) soil (P1 level). Dual inoculation of G. aggregatum and B. polymyxa yielded 21.5 g plant dry weight (biomass), while it was 21.7 g in B. polymyxa and A. brasilense inoculated plants as compared to 14.9 g of control at the same level. Phosphate content was maximum (0.167%) in the combined treatment of G. aggregatum, B. polymyxa and A. brasilense at P1 level, however acid phosphatase activity was recorded to be 4.75 pmol mg(-1) min(-1) in G. aggregatum, B. polymyxa and A. brasilense treatment at P0 level. This study indicates that all microbes inoculated together help in the uptake of tricalcium phosphate which is otherwise not used by the plants and their addition at 200 mg kg(-1) of soil gave higher productivity to palmarosa plants.     

Direct selection of cloned DNA in Bacillus subtilis based on sucrose-induced lethality.

Expression of the Bacillus subtilis or Bacillus amyloliquefaciens sacB gene in the presence of sucrose is lethal for a variety of bacteria. Sucrose-induced lethality can be used to select for inactivation of sacB by insertion of heterologous DNA in sensitive bacteria. This procedure has not been applicable to B. subtilis heretofore because expression of wild-type sacB is not detrimental to B. subtilis. The W29 mutation in the B. amyloliquefaciens sacB gene interferes with processing of the levansucrase signal peptide. The W29 mutation does not affect growth of B. subtilis in media lacking sucrose. However, this mutation inhibited growth of B. subtilis in media containing sucrose. Inactivation of the fructose polymerase activity encoded by sacB indicated that levan production was essential for sucrose-induced lethality. As a result, it was possible to select for cloned DNA in B. subtilis by insertional inactivation of the mutant sacB gene located on a multicopy plasmid vector in medium containing sucrose.     

The root growth of wheat plants,the water conservation and fertility status of sandy soils influenced by plant growth promoting rhizobacteria

Plant growth promoting rhizobacteria were isolated and characterized from sandy soils in Pakistan. The role of the rhizobacteria, in association with plant growth regulators, was studied on the roots of wheat grown under water stressed conditions. The plant growth promoting rhizobacteria were characterized on the basis of colony morphology, biochemical traits and identified on the basis of 16S-rRNA gene sequencing which identified the selected isolates Planomicrobium chinense, Bacillus cereus and Pseudomonas fluorescens. Antibacterial and antifungal activities were determined. The fresh cultures (24 h old) of isolates were used to soak the seeds for 2–3 h prior to sowing. The growth regulators salicylic acid and putrescine were applied to the plant as foliar spray at three leaf stage. The plant growth promoting rhizobacteria produced exopolysaccharides that formed soil aggregation around roots of the plants and significantly enhanced water holding capacity of sandy soil. The relative water content (80%) of leaves and root fresh (80%) and dry weight (68%) were higher in plant growth promoting rhizobacteria inoculated plants. The nutrient content of rhizosphere soil of treated plants was also enhanced (Ca 35%, K 34%, Mg 52% and Na 42%) over stressed controls. Integrative use of effective plant growth promoting rhizobacteria in combination with salicylic acid appears to be an effective eco-friendly approach to increase drought tolerance in wheat plants to combat desertification.     

Fluorescent pseudomonads in the rhizosphere of plants and their relation to root exudates

Fluorescent pseudomonads were present in chernozem soil not influenced by plant roots (10(3)-10(4) per g dry soil) in the rhizosphere soil of various plants (10(4)-10(5) per g soil) and on roots (10(3) to 10(7) per g fresh roots), depending on the species and age of the plant. Relative species representation of fluorescent pseudomonads changed on the roots and in the plant rhizosphere as compared with free soil. Pseudomonas fluorescens, representing 60-93% of the population of fluorescent pseudomonads predominated on the roots of all plants investigated. Somewhat different results were obtained in rhizosphere soil. Relatively higher numbers of P. fluorescens were detected in the rhizosphere soil of cucumber and maize, numbers in the rhizosphere soil of wheat were practically the same as in free soil and higher numbers of P. putida were found in the rhizosphere soil of barley. Almost all components contained in the root exudates of the plants studied, including beta-pyrazolylalanine from the root exudates of cucumbers were utilized as carbon and energy sources. Root exudates of wheat and maize were utilized by the strain P. putida K2 with an efficiency of 73-91%, depending on species and age of the plant.     

猪传染性胸膜肺炎放线杆菌毒素apxH基因无药物抗性标记突变株HBC-/GFP+的构建及其生物学特性研究

通过使用枯草芽胞杆菌一个蔗糖敏感sacB基因发展了一种依靠蔗糖的负向筛选系统,这种方法允许没有标记突变的基因进入胸膜肺炎放线杆菌染色体。首先,构建了猪传染性胸膜肺炎放线杆菌毒素apxⅡ基因GFP插入失活型的重组质粒pOSAKCG,其中一个表达盒含有氨苄青霉素基因和以外膜蛋白omlA作为启动子表达sacB基因。重组质粒pOSAKCG通过电穿孔转化,它的突变apxⅡCA基因与野生型亲本菌株胸膜肺炎放线杆菌HB03染色体上野生型apxⅡCA基因发生同源交换,两步法筛选获得了apxⅡ基因突变株HBC^-／GFP^ ,PCR和Southern blot对突变株进行初步鉴定,进一步对突变株的一些生物学特性,包括它的溶血活性、免疫原性、生长特性及其对小鼠的安全性进行了研究。结果表明,无药物抗性标记突变株的构建是成功的。该突变株的构建为进一步研究突变株作为载体和疫苗奠定了坚实的基础。     

Arbuscular mycorrhizae formed by Penicillium pinophilum improve the growth, nutrient uptake and photosynthesis of strawberry with two inoculum-types

Penicillium pinophilum was isolated from the soil in a commercial strawberry field. The strain readily formed arbuscular mycorrhizae (AM) with the roots of strawberry 'Zoji' (Fragaria x ananassa Duch. CV.) when plants were inoculated with either fresh cultured hyphae or root/soil mixtures. Fresh hyphae, however, resulted in higher amounts of colonization than root/soil inoculum. Compared with uninoculated strawberries, inoculation increased plant dry weight by 31%, as well as nitrogen content (47%), phosphorus content (57%), and photosynthetic rate (71%). AM inoculation also shortened the blossom and ripening date by 3 and 4 days, respectively. This is the first report of a P. pinophilum strain resulting in mycorrhiza with strawberry roots. The significant advantages of this strain are that it is easy to culture and inoculation of plants results in significant growth benefits that may be useful in strawberry production.     

Molecular genetic identification of yeast strains isolated from egyptian soils for solubilization of inorganic phosphates and growth promotion of corn plants

Forty yeast strains isolated from soils taken from different locations in Egypt were tested for their P-solubilizing activities on the basis of analyzing the clear zone around colonies growing on a tricalcium phosphate medium after incubation for 5 days at 25degreesC, denoted as the solubilization index (SI). Nine isolates that exhibited P-solubilization potential with an SI ranging from 1.19 to 2.76 were genetically characterized as five yeasts belonging to the genus Saccharomyces cerevisiae and four non-Saccharomyces, based on a PCR analysis of the ITS1-26S region amplied by SC1/SC2 species-specific primers. The highest Psolubilization efficiency was demonstrated by isolate PSY- 4, which was identified as Saccharomyces cerevisiae by a sequence analysis of the variable D1/D2 domain of the 26S rDNA. The effects of single and mixed inoculations with yeast PSY-4 and Bacillus polymyxa on the P-uptake and growth of corn were tested in a greenhouse experiment using different levels of a phosphorus chemical fertilizer (50, 100, and 200 kg/ha super phosphate 15.5% P2O5). The results showed that inoculating the corn with yeast PSY-4 or B. polymyxa caused significant increases in the shoot and root dry weights and P-uptake in the shoots and roots. The P-fertilization level also had a significant influence on the shoot and root dry weights and P-uptake in the shoots and roots when increasing the P-level from 50 up to 200 kg/ha. Dual inoculation with yeast strain PSY-4 and B. polymyxa at a P-fertilization level of 200 kg/ha gave higher values for the shoot and root dry weights and P-uptake in the shoots and roots, yet these increases were nonsignificant when compared with dual inoculation with yeast strain PSY-4 and B. polymyxa at a P-fertilization level of 100 kg/ha. The best increases were obtained from dual inoculation with yeast strain PSY-4 and B. polymyxa at a P-fertilization level of 100 kg/ha, which induced the following percentage increases in the shoot and root dry weights, and P-uptake in the shoots and roots; 16.22%, 46.92%, 10.09%, and 31.07%, respectively, when compared with the uninoculated control (fertilized with 100 kg/ha).     

The exopolysaccharide of Rhizobium sp. YAS34 is not necessary for biofilm formation on Arabidopsis thaliana and Brassica napus roots but contributes to root colonization

Microbial exopolysaccharides (EPSs) play key roles in plant–microbe interactions, such as biofilm formation on plant roots and legume nodulation by rhizobia. Here, we focused on the function of an EPS produced by Rhizobium sp. YAS34 in the colonization and biofilm formation on non-legume plant roots ( Arabidopsis thaliana and Brassica napus ). Using random transposon mutagenesis, we isolated an EPS-deficient mutant of strain YAS34 impaired in a glycosyltransferase gene ( gta ). Wild type and mutant strains were tagged with a plasmid-born GFP and, for the first time, the EPS produced by the wild-type strain was seen in the rhizosphere using selective carbohydrate probing with a fluorescent lectin and confocal laser-scanning microscopy. We show for the fist time that Rhizobium forms biofilms on roots of non-legumes, independently of the EPS synthesis. When produced by strain YAS34 wild type, EPS is targeted at specific parts of the plant root system. Nutrient fluctuations, root exudates and bacterial growth phase can account for such a production pattern. The EPS synthesis in Rhizobium sp. YAS34 is not essential for biofilm formation on roots, but is critical to colonization of the basal part of the root system and increasing the stability of root-adhering soil. Thus, in Rhizobium sp. YAS34 and non-legume interactions, microbial EPS is implicated in root–soil interface, root colonization, but not in biofilm formation.     

Effect of microbial inoculants on the indigenous actinobacterial endophyte population in the roots of wheat as determined by terminal restriction fragment length polymorphism

The effect of single actinobacterial endophyte seed inoculants and a mixed microbial soil inoculant on the indigenous endophytic actinobacterial population in wheat roots was investigated by using the molecular technique terminal restriction fragment length polymorphism (T-RFLP). Wheat was cultivated either from seeds coated with the spores of single pure actinobacterial endophytes of Microbispora sp. strain EN2, Streptomyces sp. strain EN27, and Nocardioides albus EN46 or from untreated seeds sown in soil with and without a commercial mixed microbial soil inoculant. The endophytic actinobacterial population within the roots of 6-week-old wheat plants was assessed by T-RFLP. Colonization of the wheat roots by the inoculated actinobacterial endophytes was detected by T-RFLP, as were 28 to 42 indigenous actinobacterial genera present in the inoculated and uninoculated plants. The presence of the commercial mixed inoculant in the soil reduced the endophytic actinobacterial diversity from 40 genera to 21 genera and reduced the detectable root colonization by approximately half. The results indicate that the addition of a nonadapted microbial inoculum to the soil disrupted the natural actinobacterial endophyte population, reducing diversity and colonization levels. This was in contrast to the addition of a single actinobacterial endophyte to the wheat plant, where the increase in colonization level could be confirmed even though the indigenous endophyte population was not adversely affected.     

Effect of Microbial Inoculants on the Indigenous Actinobacterial Endophyte Population in the Roots of Wheat as Determined by Terminal Restriction Fragment Length Polymorphism

The effect of single actinobacterial endophyte seed inoculants and a mixed microbial soil inoculant on the indigenous endophytic actinobacterial population in wheat roots was investigated by using the molecular technique terminal restriction fragment length polymorphism (T-RFLP). Wheat was cultivated either from seeds coated with the spores of single pure actinobacterial endophytes of Microbispora sp. strain EN2, Streptomyces sp. strain EN27, and Nocardioides albus EN46 or from untreated seeds sown in soil with and without a commercial mixed microbial soil inoculant. The endophytic actinobacterial population within the roots of 6-week-old wheat plants was assessed by T-RFLP. Colonization of the wheat roots by the inoculated actinobacterial endophytes was detected by T-RFLP, as were 28 to 42 indigenous actinobacterial genera present in the inoculated and uninoculated plants. The presence of the commercial mixed inoculant in the soil reduced the endophytic actinobacterial diversity from 40 genera to 21 genera and reduced the detectable root colonization by approximately half. The results indicate that the addition of a nonadapted microbial inoculum to the soil disrupted the natural actinobacterial endophyte population, reducing diversity and colonization levels. This was in contrast to the addition of a single actinobacterial endophyte to the wheat plant, where the increase in colonization level could be confirmed even though the indigenous endophyte population was not adversely affected.     

禾谷多粘菌对小麦梭条斑花叶病毒的传播及其土壤侵染潜力的测定

从大田侵染小麦梭条斑花叶病毒的小麦病根中挑取禾谷多粘菌休眠孢子堆,接种受侵染小麦品种扬麦４号,经砂培养纯化,获得５个禾谷多粘菌分离物,但都为无毒。无毒多粘菌休眠孢子堆接种表现ＷＳＳＭＶ症状的小麦,经培养可饲获病毒,并可经接咱后将病毒传播给无病小麦,供试的４个大小麦禾谷多粘菌分离物都可对大小进行交叉侵染,产生同样数量的游动孢子产量。供试５个病土和２个无病土样品,都具有强大持多粘菌侵染潜力,即使稀释放     

Differences between maize and wheat in growth-related nutrient demand and uptake of potassium and phosphorus at suboptimal root zone temperatures

The effects of low root zone temperatures (RZT) on nutrient demand for growth and the capacity for nutrient acquisition were compared in maize and wheat growing in nutrient solution. To differentiate between direct temperature effects on nutrient uptake and indirect effects via an altered ratio of shoot to root growth, the plants were grown with their shoot base including apical shoot meristem either within the root zone (low SB), i.e. at RZT (12°, 16°, or 20°C) or, above the root zone (high SB), i.e. at uniformly high air temperature (20°/16° day/night).At low SB, suboptimal RZT reduced shoot growth more than root growth in wheat, whereas the opposite was true in maize. However, in both species the shoot growth rate per unit weight of roots, which was taken as parameter for the shoot demand for mineral nutrients per unit of roots, decreased at low RZT. Accordingly, the concentrations of potassium (K) and phosphorus (P) remained constant or even increased at low RZT despite reduced uptake rates.At high SB, shoot growth at low RZT in both species was higher than at low SB, whereas root growth was not increased. At high SB, the shoot demand per unit of roots was similar for all RZT in wheat, but increased with decreasing RZT in maize. Uptake rates of K at high SB and low RZT adapted to shoot demand within four days, and were even higher in maize than in wheat. Uptake rates of P adapted more slowly to shoot demand in both species, resulting in reduced concentrations of P in the shoot, particularly in maize.In conclusion, the two species did not markedly differ in their physiological capacity for uptake of K and P at low RZT. However, maize had a lower ability than wheat to adapt morphologically to suboptimal RZT by increasing biomass allocation towards the roots. This may cause a greater susceptibility of maize to nutrient deficiency, particularly if the temperatures around the shoot base are high and uptake is limited by nutrient transport processes in the soil towards the roots.     

Importance of internal hydraulic redistribution for prolonging the lifespan of roots in dry soil

Redistribution of water within plants could mitigate drought stress of roots in zones of low soil moisture. Plant internal redistribution of water from regions of high soil moisture to roots in dry soil occurs during periods of low evaporative demand. Using minirhizotrons, we observed similar lifespans of roots in wet and dry soil for the grapevine 'Merlot' (Vitis vinifera) on the rootstock 101-14 Millardet de Gramanet (Vitis riparia x Vitis rupestris) in a Napa County, California vineyard. We hypothesized that hydraulic redistribution would prevent an appreciable reduction in root water potential and would contribute to prolonged root survivorship in dry soil zones. In a greenhouse study that tested this hypothesis, grapevine root systems were divided using split pots and were grown for 6 months. With thermocouple psychrometers, we measured water potentials of roots of the same plant in both wet and dry soil under three treatments: control (C), 24 h light + supplemental water (LW) and 24 h light only (L). Similar to the field results, roots in the dry side of split pots had similar survivorship as roots in the wet side of the split pots (P = 0.136) in the C treatment. In contrast, reduced root survivorship was directly associated with plants in which hydraulic redistribution was experimentally reduced by 24 h light. Dry-side roots of plants in the LW treatment lived half as long as the roots in the wet soil despite being provided with supplemental water (P < 0.0004). Additionally, pre-dawn water potentials of roots in dry soil under 24 h of illumination (L and LW) exhibited values nearly twice as negative as those of C plants (P = 0.034). Estimates of root membrane integrity using electrolyte leakage were consistent with patterns of root survivorship. Plants in which nocturnal hydraulic redistribution was reduced exhibited more than twice the amount of electrolyte leakage in dry roots compared to those in wet soil of the same plant. Our study demonstrates that besides a number of ecological advantages to protecting tissues against desiccation, internal hydraulic redistribution of water is a mechanism consistent with extended root survivorship in dry soils.     

Production of the Antibiotic Phenazine-1-Carboxylic Acid by Fluorescent Pseudomonas Species in the Rhizosphere of Wheat

Pseudomonas fluorescens 2-79 and P. aureofaciens 30-84 produce the antibiotic phenazine-1-carboxylic acid and suppress take-all, an important root disease of wheat caused by Gaeumannomyces graminis var. tritici. To determine whether the antibiotic is produced in situ, wheat seeds were treated with strain 2-79 or 30-84 or with phenazine-nonproducing mutants or were left untreated and then were sown in natural or steamed soil in the field or growth chamber. The antibiotic was isolated only from roots of wheat colonized by strain 2-79 or 30-84 in both growth chamber and field studies. No antibiotic was recovered from the roots of seedlings grown from seeds treated with phenazine-nonproducing mutants or left untreated. In natural soils, comparable amounts of antibiotic (27 to 43 ng/g of root with adhering soil) were recovered from roots colonized by strain 2-79 whether or not the pathogen was present. Roots of plants grown in steamed soil yielded larger bacterial populations and more antibiotic than roots from natural soils. In steamed and natural soils, roots from which the antibiotic was recovered had significantly less disease than roots with no antibiotic, indicating that suppression of take-all is related directly to the presence of the antibiotic in the rhizosphere.     

Piriformospora indica,a plant-root-colonising fungus enhances growth and tolerance of wheat to seedlings damping-off caused by Fusarium oxysporum

In greenhouse experiments, we investigated the potentials of Piriformospora indica (Pi) to penetrate and colonise roots of wheat and to induce beneficial effects on growth as well as to reduce seedlings damping-off disease of wheat caused by the soil-borne fungus Fusarium oxysporum (Fo). By microscopy we observed the intracellular hyphae within root cortex cells of 7–14 day-old plants and chlamydospores within root cortex cells of 14–28 day-old plants and within root hair cells of 21–28 day-old plants. Moreover, diagnostic PCR based on the β-tubulin gene marker confirmed the presence of Pi in roots of inoculated plants. Also, we found that plants inoculated with Pi exhibited a better growth of roots and shoots as well as early flowering as compared with non-treated plants. Moreover, Pi conferred tolerance of wheat to seedlings damping-off i.e. by reducing the harmful effects of Fo on infected plants.     

Draft genome sequence of the Paenibacillus polymyxa type strain (ATCC 842T), a plant growth-promoting bacterium

Paenibacillus polymyxa is an endospore-forming Gram-positive soil bacterium that is well-known for its ability to promote plant growth. Here we report the draft genome sequence of P. polymyxa ATCC 842(T), the type strain of the species P. polymyxa, and the family Paenibacillaceae. The P. polymyxa genome contains a repertoire of biosynthetic genes for antibiotics and hydrolytic enzymes that account for its beneficial effects in the rhizosphere to the host plants it associates with.     

The knockout of the lprG-Rv1410 operon produces strong attenuation of Mycobacterium tuberculosis

P27 lipoprotein was previously described as an antigen in the Mycobacterium tuberculosis complex, encoded by the lprG gene, also named Rv1411 in the TubercuList (http://genolist.pasteur.fr/TubercuList) gene bank. It forms an operon with Rv1410 that encodes for an efflux pump, P55. A mutant of the H37Rv strain of M. tuberculosis not producing P27 (strain DeltaP27) was obtained by two-step mutagenesis using the counterselectable marker sacB and a thermosensitive origin of replication in the shuttle plasmid pPR27. By RT-PCR, we observed no lprG or Rv1410 mRNA in the DeltaP27 mutant strain compared with the wild type and complemented strains. Western blot experiments using anti-P27 polyclonal sera showed that the P27 protein was present both in the parental and in a complemented strain, in which the entire lprG-Rv1410 operon was reintroduced, but absent in the mutant strain. The three strains showed similar growth kinetics and characteristics in culture broth. To study the effect of the lprG mutation on M. tuberculosis virulence, BALB/c mice were inoculated to determine bacterial loads in spleens. At days 15 and 35 after infection, decreases of 1.5 and 2.5 logs in the bacterial load were found, respectively, in animals inoculated with the DeltaP27 mutant strain or with the wild type. This attenuation was reverted in the complemented strain. These results demonstrated that lprG gene is required for growth of M. tuberculosis in immunocompetent mice. The reversion of attenuation in the complemented strain indicates that the attenuated phenotype resulted from disruption of the lprG-Rv1410 operon.