Enhanced Epiphytic Coexistence of Near-Isogenic Salicylate-Catabolizing and Non-Salicylate-Catabolizing Pseudomonas putida Strains after Exogenous Salicylate Application

The hypothesis that epiphytic bacterial populations can coexist through nutritional resource partitioning was tested with the near-isogenic bacterial strain pair Pseudomonas putida R20 and R20(pNAH7). The plasmid pNAH7 conferred upon R20 the ability to catabolize salicylate as a sole carbon source in vitro. P. putida R20(pNAH7) also catabolized exogenously applied salicylate in planta and reached a significantly larger epiphytic population size than the near-isogenic parental strain R20 under the same conditions. This supports previous observations that epiphytic populations on plants grown under nitrogen-sufficient conditions are limited by carbon availability. In the absence of exogenous salicylate, R20 and R20(pNAH7) competed for and partitioned endogenous carbon according to their inoculum proportion in replacement series experiments, exhibiting a low level of coexistence. In the presence of exogenous salicylate, however, R20(pNAH7) was solely able to catabolize the additional carbon and achieved a higher level of coexistence with R20 than was possible in the absence of exogenous carbon.     

矿区分离丛枝菌根真菌对万寿菊吸Cd潜力影响

盆栽试验研究土壤不同施Cd水平(0、5、20、50μg/g)下,接种矿区污染土壤中丛枝菌根真菌对万寿菊根系侵染率、植株生物量及Cd吸收与分配的影响。结果表明:接种丛枝菌根真菌显著提高Cd胁迫下万寿菊的根系侵染率和植株生物量;随着施Cd水平提高,各处理植株Cd浓度显著增加。各施Cd水平下万寿菊地上部Cd吸收量远远高于根系Cd吸收量,在土壤施Cd量达到50μg/g时,接种处理地上部Cd吸收量是根系的3.48倍,对照处理地上部Cd吸收量是根系的1.67倍;同一施Cd水平下接种处理植株Cd吸收量要显著高于对照。总体上,试验条件下污染土壤中分离的丛枝菌根真菌促进了万寿菊对土壤中Cd的吸收,并在一定程度上增加Cd向地上部分的运转,表现出植物提取的应用潜力。     

Effect of applying an arsenic-resistant and plant growth-promoting rhizobacterium to enhance soil arsenic phytoremediation by Populus deltoides LH05-17

Aims: Bioremediation of highly arsenic (As)‐contaminated soil is difficult because As is very toxic for plants and micro‐organisms. The aim of this study was to investigate soil arsenic removal effects using poplar in combination with the inoculation of a plant growth–promoting rhizobacterium (PGPR). Methods and Results: A rhizobacterium D14 was isolated and identified within Agrobacterium radiobacter. This strain was highly resistant to arsenic and produced indole acetic acid and siderophore. Greenhouse pot bioremediation experiments were performed for 5 months using poplar (Populus deltoides LH05‐17) grown on As‐amended soils, inoculated with strain D14. The results showed that P. deltoides was an efficient arsenic accumulator; however, high As concentrations (150 and 300 mg kg^?1) inhibited its growth. With the bacterial inoculation, in the 300 mg kg^?1 As‐amended soils, 54% As in the soil was removed, which was higher than the uninoculated treatments (43%), and As concentrations in roots, stems and leaves were significantly increased by 229, 113 and 291%, respectively. In addition, the As translocation ratio [(stems + leaves)/roots = 0·8] was significantly higher than the uninoculated treatments (0·5). About 45% As was translocated from roots to the above‐ground tissues. The plant height and dry weight of roots, stems and leaves were all enhanced; the contents of chlorophyll and soluble sugar, and the activities of superoxide dismutase and catalase were all increased; and the content of a toxic compound malondialdehyde was decreased. Conclusions: The results indicated that the inoculation of strain D14 could contribute to the increase in the As tolerance of P. deltoides, promotion of the growth, increase in the uptake efficiency and enhancement of As translocation. Significance and Impact of the Study: The use of P. deltoides in combination with the inoculation of strain D14 provides a potential application for efficient soil arsenic bioremediation.     

Multispecies and monoculture rhizoremediation of polycyclic aromatic hydrocarbons (PAHs) from the soil

In this study, we investigated the potential of multispecies rhizoremediation and monoculture rhizoremediation in decontaminating polycyclic aromatic hydrocarbon (PAH) contaminated soil Plant-mediated PAH dissipation was evaluated using monoplanted soil microcosms and soil microcosms vegetated with several different grass species (Brachiaria serrata and Eleusine corocana). The dissipation of naphthalene and fluorene was higher in the "multispecies" vegetated soil compared to the monoplanted and nonplanted control soil. The concentration of naphthalene was undetectable in the multispecies vegetated treatment compared to 96% removal efficiencies in the monoplanted treatments and 63% in the nonplanted control after 10 wk of incubation. Similar removal efficiencies were obtained for fluorene. However, there was no significant difference in the dissipation of pyrene in both the mono- and multispecies vegetated treatments. There also was no significant difference between the dissipation of PAHs in the monoplanted treatments with different grass species. Principle component analysis (PCA) and cluster analysis were used to evaluate functional diversity of the different treatments during phytoremediation of PAHs. Both PCA and cluster analysis revealed differences in the metabolic fingerprints of the PAH contaminated and noncontaminated soils. However, the differences in metabolic diversity between the multispecies vegetated and monoplanted treatments were not clearly revealed. The results suggest that multispecies rhizoremediation using tolerant plant species rather than monoculture rhizoremediation have the potential to enhance pollutant removal in moderately contaminated soils.     

Potential of wheat (Triticum aestivum L.) and pea (Pisum sativum) for remediation of soils contaminated with bromides and PAHs

The aim of the research was to study a removal of polycyclic aromatic hydrocarbons (PAHs) and phytoextraction of bromine (Br) from contaminated soils. The experiments using pea and wheat seedlings as potential candidates for soil remediation were performed. The soil for the experiments was collected from a site slightly contaminated by some PAHs. Before planting, the soil was exposed to 20 mg of Br/kg of soil. In the soil taken from rhizosphere of pea and wheat, the concentrations of many PAHs decreased up to 7 times compared to the concentrations of the compounds in the initial soil. Pea was capable of more effectively influencing the soil PAHs than wheat. The growth of pea and wheat in the soil spiked with Br resulted in a significant increase of Br concentration in a plant. Concentration of Br in roots of pea and wheat increased 21 and 3 times, respectively. Bromine content in leaves of wheat and pea increased 10 and 4.5 times. This accumulation of Br in the plants led to a decrease of its concentration in the rhizosphere soil. The experimental results demonstrated a good ability of the plants to cleanup the soils contaminated with organic and inorganic compounds.     

Biodegradation of polycyclic aromatic hydrocarbons by an acidophilic Stenotrophomonas maltophilia strain AJH1 isolated from a mineral mining site in Saudi Arabia

The present study aims at analyzing the degradation of polycyclic aromatic hydrocarbons (PAHs) at acidic conditions (pH = 2) by acidophilic Stenotrophomonas maltophilia strain AJH1 (KU664513). The strain AJH1 was obtained from an enrichment culture obtained from soil samples of mining area in the presence of PAH as sole sources of carbon and energy. Strain AJH1was able to degrade low (anthracene, phenanthrene, naphthalene, fluorene) and high (pyrene, benzo(e)pyrene and benzo(k)fluoranthene) molecular weight PAHs in acidophilic mineral salt medium at pH 2, with removal rates of up to 95% (LMW PAH) and 80% (HMW PAH), respectively. In addition, strain AJH1 treated petroleum wastewater with 89 ± 1.1% COD removal under acidic condition (pH 2) in a continuously stirred reactor. Acidophilic S. maltophilia strain AJH1, hence holds the promise as an effective degrader for biological treatment of PAHs contaminated wastewater at acidic pH.

     

Use of bacterial acc deaminase to increase oil (especially poly aromatic hydrocarbons) phytoremediation efficiency for maize (zea mays) seedlings

Oil presence in soil, as a stressor, reduces phytoremediation efficiency through an increase in the plant stress ethylene. Bacterial 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase, as a plant stress ethylene reducer, was employed to increase oil phytoremediation efficiency. For this purpose, the ability of ACC deaminase-producing Pseudomonas strains to grow in oil-polluted culture media and withstand various concentrations of oil and also their ability to reduce plant stress ethylene and enhance some growth characteristics of maize and finally their effects on increasing phytoremediation efficiency of poly aromatic hydrocarbons (PAHs) in soil were investigated. Based on the results, of tested strains just P9 and P12 were able to perform oil degradation. Increasing oil concentration from 0 to 10% augmented these two strains population, 15.7% and 12.9%, respectively. The maximum increase in maize growth was observed in presence of P12 strain. Results of high-performance liquid chromatography (HPLC) revealed that PAHs phytoremediation efficiency was higher for inoculated seeds than uninoculated. The highest plant growth and PAHs removal percentage (74.9%) from oil-polluted soil was observed in maize inoculated with P12. These results indicate the significance of ACC deaminase producing bacteria in alleviation of plant stress ethylene in oil-polluted soils and increasing phytoremediation efficiency of such soils.     

Conjugal transfer and mobilization capacity of the completely sequenced naphthalene plasmid pNAH20 from multiplasmid strain Pseudomonas fluorescens PC20

The complete 83 042-bp nucleotide sequence of the IncP-9 naphthalene degradation plasmid pNAH20 from Pseudomonas fluorescens PC20 exhibits striking similarity in size and sequence to another naphthalene (NAH) plasmid pDTG1. However, the positions of insertion sequence (IS) elements significantly alter both catabolic and backbone functions provided by the two plasmids. In pDTG1, insertion of a pCAR1 IS Pre1 -like element disrupts expression of the lower naphthalene operon and this strain utilizes the chromosomal pathway for complete naphthalene degradation. In pNAH20, this operon is intact and functional. The transfer frequency of pNAH20 is 100 times higher than that of pDTG1 probably due to insertion of the pCAR1 IS Pre2 -like element into the mpfR gene coding for a putative repressor of the mpf operon responsible for mating pilus formation. We also demonstrate in situ plasmid transfer – we isolated a rhizosphere transconjugant strain of pNAH20, P. fluorescens NS8. The plasmid pNS8, a derivative of pNAH20, lacks the ability to self-transfer as a result of an additional insertion event of IS Pre2 -like element that disrupts the gene coding for VirB2-like major pilus protein MpfA. The characteristics of the strain PC20 and the conjugal transfer/mobilization capacity of pNAH20 (or its backbone) make this strain/plasmid a potentially successful tool for bioremediation applications.     

Naphthalene degradation via salicylate and gentisate by Rhodococcus sp. strain B4.

Rhodococcus sp. strain B4, isolated from a soil sample contaminated with polycyclic aromatic hydrocarbons, grows with naphthalene as the sole source of carbon and energy. Salicylate and gentisate were identified as intermediates in the catabolism of naphthalene. In contrast to the well-studied catabolic pathway encoded by the NAH7 plasmid of Pseudomonas putida, salicylate does not induce the genes of the naphthalene-degradative pathway in Rhodococcus sp. strain B4. The key enzymes of naphthalene degradation in Rhodococcus sp. strain B4 have unusual cofactor requirements. The 1,2-dihydroxynaphthalene oxygenase activity depends on NADH and the salicylate 5-hydroxylase requires NADPH, ATP, and coenzyme A.     

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.     

Revegetation of a lakeside barren area by the application of plant growth-promoting rhizobacteria

The growth stimulation of wild plants by several bacterial species showing plant growth-promoting capabilities was examined in a barren lakeside area at Lake Paro, Korea. Microbial numbers and activities in the field soil were monitored for 73 days after inoculation of the bacteria. The acridine orange direct counts for the total soil bacterial populations ranged between 2.0-2.3x10(9) cells/g soil and 1.4-1.8x10(9) cells/g soil in the inoculated and uninoculated soils, respectively. The numbers of Pseudomonas spp., which is known as a typical plant growth-promoting rhizobacteria, and the total microbial activity were higher in the inoculated soil compared to those in the uninoculated soil. The average shoot and root lengths of the wild plants grown in the inoculated soil were 17.3 cm and 12.4 cm, respectively, and longer than those of 11.4 cm and 8.5 cm in the uninoculated soil. The total dry weight of the harvested wild plants was also higher in the inoculated soil (42.0 g) compared to the uninoculated soil (35.1 g). The plant growth-promoting capabilities of the inoculated bacteria may be used for the rapid revegetation of barren or disturbed land, and as biofertilizer in agriculture.     

Physiologo-biochemical properties of a strain of Beijerinckia mobilis 1phi Phn+--a degrader of polycyclic aromatic hydrocarbons

Beijerinckia mobilis 1f capable of degrading polycyclic aromatic hydrocarbons (PAHs) was isolated from a soil contaminated with creosote. Strain 1f could utilize phenanthrene and naphthalene as the sole sources of carbon. The mean rate of phenanthrene degradation during culture growth was 7-8 micrograms/(ml h). After cultivation under nonselective conditions, strain 1f retained its ability to degrade phenanthrene. Cometabolism considerably widened the range of PAHs that could be transformed by strain 1f. The strain was able to grow in a mineral medium with creosote as the sole source of carbon. After 30 days of cultivation in this medium, the total concentration of PAHs decreased from 665.5 mg/l to 170 mg/l.     

Inoculation with a Native Soil Community Advances Succession in a Grassland Restoration

Restoration on post‐agricultural land may be hindered by the degradation of the soil community, which has been shown to contribute to structuring plant communities and driving succession. Our experiment tested the effect of inoculation with remnant grassland whole soil with or without nurse plants on the survival and growth of uninoculated early and late successional plant species. In 2007 and 2008, we planted uninoculated early, mid, and late successional plant species 0.25–2 m away from a central point of inoculated nurse plants. We found a negative response to inoculation on early successional plants and a positive response to inoculation on mid to late successional plants. This work suggests that the restoration of the soil community is critical to establishing a late successional plant community and that the benefit of inoculated plants can spread to neighbors.     

Effect of native and introduced arbuscular mycorrhizal fungi on growth and nutrient uptake ofLygeum spartum andAnthyllis cytisoides

The interaction between native and introduced fungi and their effect on plant growth and mineral uptake were studied. The host plants wereLygeum spartum andAnthyllis cytisoides, the introduced fungus wasGlomus fasciculatum. The four soils used were selected from disturbed and contaminated by mining activities areas. Inoculated and uninoculated plants were grown in the unsterilized and sterilized soils (with and withouth native microflora, respectively). Plants inoculated withG. fasciculatum were higher and had higher tissue P concentration than uninoculated plants, especially inA. cytisoides. However, this inoculation was not effective in unsterilized substrates, suggesting a competition between introduced and native fungi. Concentration of mineral elements other than P varied depending on the host plant and soil. Decrease in Fe, Cu, Mn, Zn and Pb was observed in mycorrhizalA. cytiosides plants and a slight increase in Zn concentration was noted in mycorrhizalL. spartum plants. The study showed that the type of soil and their populations of native endophytes have a considerable effect on plant response to mycorrhizal symbiosis, especially in disturbed soils.     

Response of Eucalyptus tereticornis to inoculation with indigenous AM fungi in a semiarid alfisol achieved with different concentrations of available soil P

Eucalyptus tereticornis was grown in a green house in a low phosphorus (0.67 ppm Olsen's P) soil (Typic Haplustalf) inoculated with mixed indigenous arbuscular mycorrhizal (AM) fungi. Soil was amended to achieve P levels of 10, 20, 25, 30 and 40 ppm to evaluate the growth response and dependence of E. tereticornis to inoculation with AM fungi. A positive response to mycorrhizal inoculation was evident at the first two levels of soil P, i.e., at 0.67 and 10 ppm but not at the higher levels of soil P. Dry matter yield of inoculated plants beyond 20 ppm soil P was similar or even less compared to their uninoculated counterparts. Inoculated plants produced maximum dry matter (root and shoot) at 10 ppm soil P, whereas uninoculated plants did not produce until the level reached 20 ppm. The percentage root length colonized by AM fungi decreased from 31% to 3% as the concentration of P increased beyond 10 ppm soil P. Higher levels of soil P depressed the AM colonization significantly. Inoculated plants had higher shoot P and N contents compared to their uninoculated counterparts at all levels of soil P. However, at the first two lower levels of soil P, inoculated plants showed significantly higher shoot P and N contents over their respective uninoculated counterparts. The increasing shoot P accumulation beyond 10 ppm did not enhance dry matter yields. Inoculated plants had lower values of phosphorus utilization efficiency (PUE) and nitrogen utilization efficiency (NUE) at all levels of soil P except at the unamended level (0.67 ppm) where the inoculated plants showed higher values of NUE compared to uninoculated control plants. Taking dry matter yield into consideration, Eucalyptus plants were found to be highly dependent on 10 ppm of soil P for maximum dry matter production. Any further amendment of P to soil was not beneficial neither for AM symbiosis nor plant growth.     

Enhanced tolerance to naphthalene and enhanced rhizoremediation performance for Pseudomonas putida KT2440 via the NAH7 catabolic plasmid

In this work, we explore the potential use of the Pseudomonas putida KT2440 strain for bioremediation of naphthalene-polluted soils. Pseudomonas putida strain KT2440 thrives in naphthalene-saturated medium, establishing a complex response that activates genes coding for extrusion pumps and cellular damage repair enzymes, as well as genes involved in the oxidative stress response. The transfer of the NAH7 plasmid enables naphthalene degradation by P. putida KT2440 while alleviating the cellular stress brought about by this toxic compound, without affecting key functions necessary for survival and colonization of the rhizosphere. Pseudomonas putida KT2440(NAH7) efficiently expresses the Nah catabolic pathway in vitro and in situ, leading to the complete mineralization of [(14)C]naphthalene, measured as the evolution of (14)CO(2), while the rate of mineralization was at least 2-fold higher in the rhizosphere than in bulk soil.     

Effect of arbuscular mycorrhizal fungi, organic fertilizer and soil sterilization on maize growth

The effects of inoculation with arbuscular mycorrhizal (AM) fungi, organic fertilizer (F) applications, and soil sterilization on maize growth were evaluated in a pot experiment. The experiment was in a completely randomized factorial design (2 × 4 × 2) with six replicates for each treatment. There were two soil treatments (sterilized soil, SS and unsterilized soil, US), four organic fertilizer treatments (0.0, 0.5, 1.0 and 2.0 g kg^?1 soil), and two AM fungi treatments (inoculation with Glomus mosseae, +AM and uninoculated control, ?AM). Inoculated plants generally had greater AM colonization, plant height, dry weight and phosphorus (P) uptake than uninoculated controls, and these parameters were significantly increased as the organic fertilizer application increased up to 0.5 g kg^?1 but decreased or had no significant effect compared to the uninoculated plants at the highest fertilizer rate (2.0 g kg^?1). Plant growth, P uptake and AM colonization of root system were significantly higher in sterilized soil compared to the unsterilized control. Our results indicated that the inoculation of AM fungi in field soil with optimal organic fertilizer application greatly improved maize growth and nutrient uptake, and the effect was greater under sterilized soil condition.     

The effect on yield in courgette and marrow of the mild strain of zucchini yellow mosaic virus used for cross-protection

The effects on yield in courgette and marrow (Cucurbita pepo) crops resulting from inoculation with the mild strain of zucchini yellow mosaic virus (ZYMV:WK), have been determined in polythene-house trials and in three years of outdoor, commercial field trials. In polythene-house trials ZYMV:WK inoculated plants were up to 10 days later in flowering than uninoculated plants and their cumulative yields were between 5% and 26% less than uninoculated plants depending on the cultivar. In most field trials cumulative yields from inoculated plants were between 4% and 38% less than uninoculated plants depending on the site and cultivar, but in one trial the yield was 7% higher from inoculated plants. In all experiments, courgette and marrow fruits harvested from ZYMV:WK inoculated plants were symptomless and indistinguishable from fruit harvested from uninoculated plants. The mild leaf symptoms induced by ZYMV:WK infection did not intensify to severe leaf symptoms and where there were natural outbreaks of severe ZYMV infection, fruits from inoculated plants remained symptomless whilst those from uninoculated plants were severely affected and unmarketable.     

Quantitative analysis of experiments on bacterial chemotaxis to naphthalene

A mathematical model was developed to quantify chemotaxis to naphthalene by Pseudomonas putida G7 (PpG7) and its influence on naphthalene degradation. The model was first used to estimate the three transport parameters (coefficients for naphthalene diffusion, random motility, and chemotactic sensitivity) by fitting it to experimental data on naphthalene removal from a discrete source in an aqueous system. The best-fit value of naphthalene diffusivity was close to the value estimated from molecular properties with the Wilke-Chang equation. Simulations applied to a non-chemotactic mutant strain only fit the experimental data well if random motility was negligible, suggesting that motility may be lost rapidly in the absence of substrate or that gravity may influence net random motion in a vertically oriented experimental system. For the chemotactic wild-type strain, random motility and gravity were predicted to have a negligible impact on naphthalene removal relative to the impact of chemotaxis. Based on simulations using the best-fit value of the chemotactic sensitivity coefficient, initial cell concentrations for a non-chemotactic strain would have to be several orders of magnitude higher than for a chemotactic strain to achieve similar rates of naphthalene removal under the experimental conditions we evaluated. The model was also applied to an experimental system representing an adaptation of the conventional capillary assay to evaluate chemotaxis in porous media. Our analysis suggests that it may be possible to quantify chemotaxis in porous media systems by simply adjusting the model's transport parameters to account for tortuosity, as has been suggested by others.