Improved efficiency of Sedum lineare (Crassulaceae) in remediation of arsenic-contaminated soil by phosphate-dissolving strain P-1 in association with phosphate rock.

The selection of appropriate plants and growth strategies is a key factor in improving the efficiency and universal applicability of phytoremediation. Sedum lineare grows rapidly and tolerates multiple adversities. The effects of inoculation of Acinetobacter sp. phosphate solubilizing bacteria P-1 and application of phosphate rock (PR) as additives on the remediation efficiency of As-contaminated soil by S. lineare were investigated. Compared with the control, both the single treatment and the combination of inoculation with strain P-1 and application of PR improved the biomass by 30.7-395.5%, chlorophyll content by 48.1-134.8%, total protein content by 12.5-92.4% and total As accumulation by 45.1-177.5%, and reduced the As-induced oxidative damage. Inoculation with strain P-1 increased the activities of superoxide dismutases and catalases of S. lineare under As stress, decreased the accumulation of reactive oxygen species in plant tissues and promoted the accumulation of As in roots. In contrast, simultaneous application of PR decreased As concentration in S. lineare tissues, attenuated As-induced lipid peroxidation and improved As transport to shoots. In addition, the combined application showed the best performance in improving resistance and biomass, which significantly increased root length by 149.1%, shoot length by 33%, fresh weight by 395.5% and total arsenic accumulation by 159.2%, but decreased the malondialdehyde content by 89.1%. Our results indicate that the combined application of strain P-1 and PR with S. lineare is a promising bioremediation strategy to accelerate phytoremediation of As-contaminated soils.

Biochar loaded with bacteria enhanced Cd/Zn phytoextraction by facilitating plant growth and shaping rhizospheric microbial community.

Biochar and metal-tolerant bacteria have been widely used in the remediation of heavy metal contaminated soil. However, the synergistic effect of biochar-functional microbes on phytoextraction by hyperaccumulators remains unclear. In this study, the heavy metal-tolerant strain Burkholderia contaminans ZCC was selected and loaded on biochar to produce biochar-resistant bacterial material (BM), and the effects of BM on Cd/Zn phytoextraction by Sedum alfredii Hance and rhizospheric microbial community were explored. The results showed that, BM application significantly enhanced the Cd and Zn accumulation of S. alfredii by 230.13% and 381.27%, respectively. Meanwhile, BM alleviated metal toxicity of S. alfredii by reducing oxidative damage and increasing chlorophyll and antioxidant enzyme activity. High-throughput sequencing revealed that BM significantly improved soil bacterial and fungal diversity, and increased the abundance of genera with plant growth promoting and metal solubilizing functions such as Gemmatimonas, Dyella and Pseudarthrobacter. Co-occurrence network analysis showed that BM significantly increased the complexity of the rhizospheric bacterial and fungal network. Structural equation model analysis revealed that soil chemistry property, enzyme activity and microbial diversity contributed directly or indirectly to Cd and Zn extraction by S. alfredii. Overall, our results suggested that biochar- B. contaminans ZCC was able to enhance the growth and Cd/Zn accumulation by S. alfredii. This study enhanced our understanding on the hyperaccumulator-biochar-functional microbe interactions, and provided a feasible strategy for promoting the phytoextraction efficiency of heavy metal contaminated soils.

Wood vinegar facilitated growth and Cd/Zn phytoextraction of Sedum alfredii Hance by improving rhizosphere chemical properties and regulating bacterial community.

Soil Cd and Zn contamination has become a serious environmental problem. This work explored the performance of wood vinegar (WV) in enhancing the phytoextraction of Cd/Zn by hyperaccumulator Sedum alfredii Hance. Rhizosphere chemical properties, enzyme activities and bacterial community were analyzed to determine the mechanisms of metal accumulation in this process. Results demonstrated that, after 120 days growth, different times dilution of WV increased the shoot biomass of S. alfredii by 85.2%-148%. In addition, WV application significantly increased soil available Cd and Zn by lowing soil pH, which facilitated plant uptake. The optimal Cd and Zn phytoextraction occurred from the 100 times diluted WV (D100), which increased the Cd and Zn extraction by 188% and 164%, compared to CK. The 100 and 50 times diluted WV significantly increased soil total and available carbon, nitrogen and phosphorus, and enhancing enzyme activities of urease, acid phosphatase, invertase and protease by 10.1-21.4%, 29.1-42.7%,12.2-38.3% and 26.8-85.7%, respectively, compared to CK. High-throughput sequencing revealed that the D 100 significantly increased the bacterial diversity compared to CK. Soil bacterial compositions at phylum, family and genera level were changed by WV addition. Compared to CK, WV application increased the relative abundances of genus with plant growth promotion and metal mobilization function such as, Bacillus, Gemmatimonas, Streptomyces, Sphingomonas and Polycyclovorans, which was positively correlated to biomass, Cd/Zn concentrations and extractions by S. alfredii. Structural equation modeling analysis showed that, soil chemical properties, enzyme activities and bacterial abundance directly or indirectly contributed to the biomass promotion, Cd, and Zn extraction by S. alfredii. To sum up, WV improved phytoextraction efficiency by enhancing plant growth, Cd and Zn extraction and increasing soil nutrients, enzyme activities, and modifying bacterial community.

Cadmium phytoextraction through Brassica juncea L. under different consortia of plant growth-promoting bacteria from different ecological niches.

Combined bioaugmentation inoculants composed of two or more plant growth-promoting bacteria (PGPB) were more effective than single inoculants for plant growth and cadmium (Cd) removal in contaminated soils. However, the principles of consortia construction still need to be discovered. Here, a pot experiment with Cd natural polluted soil was conducted and PGPB consortia with different ecological niches from hyperaccumulator Sedum alfredii Hance were used to compare their effects and mechanisms on plant growth condition, Cd phytoextraction efficiency, soil enzymatic activities, and rhizospheric bacterial community of Brassica juncea L. The results showed that both rhizospheric and endophytic PGPB consortia inoculants promoted plant growth (6.9%-22.1%), facilitated Cd uptake (230.0%-350.0%) of oilseed rape, increased Cd phytoextraction efficiency (343.0%-441.0%), and enhanced soil Cd removal rates (92.0%-144.0%). PGPB consortia inoculants also enhanced soil microbial carbon by 22.2%-50.5%, activated the activities of soil urease and sucrase by 74.7%-158.4% and 8.4%-61.3%, respectively. Simultaneously, PGPB consortia inoculants increased the relative abundance of Flavobacterium, Rhodanobacter, Kosakonia, Pseudomonas and Paraburkholderia at the genus level, which may be beneficial to plant growth promotion and bacterial phytopathogen biocontrol. Although the four PGPB consortia inoculants promoted oilseed growth, amplified Cd phytoextraction, and changed bacterial community structure in rhizosphere soil, their original ecological niches were not a decisive factor for the efficiency of PGPB consortia. therefore, the results enriched the present knowledge regarding the significant roles of PGPB consortia as bioaugmentation agents and preliminarily explored construction principles of effective bioaugmentation inoculants, which will provide insights into the microbial responses to combined inoculation in the Cd-contaminated soils.

Transcriptome analysis of Penicillium italicum in response to the flavonoids from Sedum aizoon L.

Natural products extracted from plants are an alternative method for controlling postharvest citrus blue mold, caused by Penicillium italicum (P. italicum). In this study, RNA sequence analysis was used to investigate the underlying anti-fungal mechanism of flavonoids from Sedum aizoon L. (FSAL) on P. italicum. Significant differences in 3592 genes were observed, including 2507 up-regulated and 1085 down-regulated genes between the FSAL-treated and the control groups. Furthermore, the GO and KEGG analysis results indicated that FSAL inhibited genes related to the integral components of membrane, oxidation-reduction process, mitochondrion, ribosome, and amino acid metabolism. In the presence of FSAL, the cellular constituents, including DNA and RNA were leaked from hyphae of P. italicum. Reactive oxygen species (ROS) production in P. italicum was also determined with a significant concentration-effect under the treatment of FSAL. Thus, we speculate that the inhibitory activity of FSAL on P. italicum is mainly achieved by damaging the structure of the cell membrane and cell wall, disrupting the process of mitochondrial respiratory metabolism, protein biosynthesis, and amino acid metabolism, leading to cell death. The present study provided a global perspective on the molecular mechanism of FSAL on P. italicum through transcriptome analysis, which may help develop a novel plant-derived anti-fungal agent for the blue mold of citrus.

Effects of EDTA and plant growth-promoting rhizobacteria on plant growth and heavy metal uptake of hyperaccumulator Sedum alfredii Hance.

Phytoremediation is a cost-effective and environment-friendly strategy for decontaminating heavy-metal-contaminated soil. However, the practical use of phytoremediation is constrained by the low biomass of plants and low bioavailability of heavy metals in soil. A pot experiment was conducted to investigate the effects of the metal chelator ethylenediaminetetraacetic acid (EDTA) and EDTA in combination with plant growth-promoting rhizobacteria (Burkholderia sp. D54 or Burkholderia sp. D416) on the growth and metal uptake of the hyperaccumulator Sedum alfredii Hance. According to the results, EDTA application decreased shoot and root biomass by 50% and 43%, respectively. The soil respiration and Cd, Pb, Zn uptake were depressed, while the photosynthetic rate, glutathione and phytochelatin (PC) contents were increased by EDTA application. Interestingly, Burkholderia sp. D54 and Burkholderia sp. D416 inoculation significantly relieved the inhibitory effects of EDTA on plant growth and soil respiration. Compared with the control, EDTA + D416 treatment increased the Cd concentration in shoots and decreased the Pb concentration in shoots and roots, but did not change the Zn concentration in S. alfredii plants. Furthermore, EDTA, EDTA + D54 and EDTA + D416 application increased the cysteine and PC contents in S. alfredii (p < 0.05); among all tested PCs, the most abundant species was PC2, and compared with the control, the PC2 content was increased by 371.0%, 1158.6% and 815.6%, respectively. These results will provide some insights into the practical use of EDTA and PGPR in the phytoremediation of heavy-metal-contaminated soil by S. alfredii.

Ochrobactrum intermedium and saponin assisted phytoremediation of Cd and B[a]P co-contaminated soil by Cd-hyperaccumulator Sedum alfredii.

Pot-culture experiments were conducted to investigate the potential of microorganism-saponin assisted phytoremediation of cadmium (Cd) and benzo(a)pyrene (B[a]P) co-contaminated soil using Cd-hyperaccumulator Sedum alfredii. Results showed that B[a]P-degrading bacterium (Ochrobactrum intermedium B[a]P-16) inoculation significantly increased root (by 22.1-24.1%) and shoot (by 20.5-23.4%) biomass of S. alfredii, whereas the application of saponin had no effect on the growth of S. alfredii. The saponin solution at 2 g L-1 extracted more Cd and B[a]P than water, saponin enhanced Cd and B[a]P bioavailability in soil and thus promoted their uptake and accumulation in S. alfredii. The activity of B[a]P-16, dehydrogenase and polyphenol oxidase in co-contaminated soil was promoted by growing S. alfredii, and the application of B[a]P-16 and saponins caused a significant (P < 0.05) increase in both enzyme activities. The maximum B[a]P removal rate (82.0%) and Cd phytoextraction rate (19.5%) were obtained by co-application of S. alfredii with B[a]P-16 and saponin. The B[a]P-16 and plant promoted biodegradation were the predominant contributors towards removal of B[a]P from soil. A significant (P < 0.05) synergistic effect of B[a]P-16 and saponin on B[a]P and Cd removal efficiency was observed in this study. It is suggested that planting S. alfredii with application of B[a]P-16 and saponin would be an effective method for phytoremediation of soil co-contaminated with Cd and PAHs.

Inoculation of plant growth promoting bacteria from hyperaccumulator facilitated non-host root development and provided promising agents for elevated phytoremediation efficiency.

Plant growth promoting bacteria (PGPB) have been reported to have the ability to promote plant growth, development and increase heavy metals (HMs) uptake. Therefore, PGPB inoculation as soil remediation agents into plants with larger biomass and potential of phytoextraction is of great importance to increase bioremediation efficiency. In this study, 12 PGPB strains isolated from a cadmium (Cd)/zinc hyperaccumulator Sedum alfredii Hance were inoculated into non-host plant Brassica juncea and their effects on plant growth and Cd uptake were determined. The results showed that inoculation of most PGPB strains promoted plant growth, boosted root development and improved chlorophyll content in the absence of Cd. Inoculation of PGPB strains promoted plant growth up to 111% in shoot and 358% in root when treated with 2 µM Cd. In addition, PGPB inoculation not only ameliorated plant root morphology including the total root length (RL), total surface area (SA), total root volume (RV) and number of root tips (RT), but also facilitated Cd uptake up to 126%. Furthermore, inoculation of PGPB strains promoted plant Cd accumulation up to 261% in shoot and up to 8.93-fold increase in root. Among all the 12 PGPB strains, Burkholdria SaMR10 and Sphingomonas SaMR12 were identified as the promising microbes for improving phytoremediation efficiency of Cd contaminated soils. These results not only provided useful findings for further investigation of interacting mechanisms between different bacterial strains and plants, but also facilitated the development of microbe-assisted phytoremediation application for HM contaminated soil.

Rhizocompartments and environmental factors affect microbial composition and variation in native plants.

Molecular analysis based on large-scale sequencing of the plant microbiota has revealed complex relationships between plants and microbial communities, and environmental factors such as soil type can influence these relationships. However, most studies on root-associated microbial communities have focused on model plants such as Arabidopsis, rice or crops. Herein, we examined the microbiota of rhizocompartments of two native plants, Sedum takesimense Nakai and Campanula takesimana Nakai, using archaeal and bacterial 16S rRNA gene amplicon profiling, and assessed relationships between environmental factors and microbial community composition. We identified 390 bacterial genera, including known plant-associated genera such as Pseudomonas, Flavobacterium, Bradyrhizobium and Rhizobium, and uncharacterized clades such as DA101 that might be important in root-associated microbial communities in bulk soil. Unexpectedly, Nitrososphaera clade members were abundant, indicating functional association with roots. Soil texture/type has a greater impact on microbial community composition in rhizocompartments than chemical factors. Our results provide fundamental knowledge on microbial diversity, community and correlations with environmental factors, and expand our understanding of the microbiota in rhizocompartments of native plants.

Promotion of the root development and Zn uptake of Sedum alfredii was achieved by an endophytic bacterium Sasm05.

Endophyte-assisted phytoremediation has gained increasing attention. However, the interacting mechanisms of endophytes and metal hyperaccumulators are still not clear. An endophytic bacterium Pseudomonas fluorescens Sasm05 inoculation promoted Sedum alfredii Hance rooting and root development, in which the specific root length (SRL) and average number of root tips (ART) increased to 2.09- and 3.35-fold, respectively. Sasm05 inoculation promoted plant growth, increased the chlorophyll content, and elevated Zn uptake of plant at excess Zn supply. At 200 µM Zn treatment level, Sasm05 inoculation increased plant biomass and the chlorophyll content by more than 40%, and root Zn content by 40%. Furthermore, Sasm05 inoculation upregulated the expression of the Zn transporter SaIRT1 to 3.43-fold in the roots, while another transporter SaNramp1 expression was increased to 38.66-fold in the roots and 7.53-fold in the shoots. Time course study showed the best effects of Sasm05 on plant biomass and the chlorophyll content were detected at 30 d, while for Zn content at 3 d. These results firstly provided molecular evidences of endophytic bacteria in facilitating host plant Zn uptake, which will absolutely benefit the understanding of interacting mechanisms between hyperaccumulators and their endophytes.

Effects of CO2 application and endophytic bacterial inoculation on morphological properties, photosynthetic characteristics and cadmium uptake of two ecotypes of Sedum alfredii Hance.

Plant uptake of cadmium (Cd) is affected by soil and environmental conditions. In this study, hydroponic experiments were conducted to investigate the effects of elevated CO2 coupled with inoculated endophytic bacteria M002 on morphological properties, gas exchange, photosynthetic pigments, chlorophyll fluorescence, and Cd uptake of S. alfredii. The results showed that bio-fortification processes (elevated CO2 and/or inoculated with endophytic bacteria) significantly (p < 0.05) promoted growth patterns, improved photosynthetic characteristics and increased Cd tolerance of both ecotypes of S. alfredii, as compared to normal conditions. Net photosynthetic rate (Pn) in intact leaves of hyperaccumulating ecotype (HE) and non-hyperaccumulating ecotype (NHE) were increased by 73.93 and 32.90%, respectively at the low Cd (2 µM), 84.41 and 57.65%, respectively at the high Cd level (10 µM). Superposition treatment increased Cd concentration in shoots and roots of HE, by 50.87 and 82.12%, respectively at the low Cd and 46.75 and 88.92%, respectively at the high Cd level. Besides, superposition treatment declined Cd transfer factor of NHE, by 0.85% at non-Cd rate, 17.22% at the low Cd and 22.26% at the high Cd level. These results indicate that elevated CO2 coupled with endophytic bacterial inoculation may effectively improve phytoremediation efficiency of Cd-contaminated soils by hyperaccumulator, and alleviate Cd toxicity to non-hyperaccumulator ecotype of Sedum alfredii.

Endophytic bacterium Buttiauxella sp. SaSR13 improves plant growth and cadmium accumulation of hyperaccumulator Sedum alfredii.

Inoculation with endophytic bacterium has been considered as a prospective application to improve the efficiency of phytoextraction. In this study, the effect of Buttiauxella sp. SaSR13 (SaSR13), a novel endophytic bacterium isolated from the root of hyperaccumulator Sedum alfredii, on plant growth and cadmium (Cd) accumulation in S. alfredii was investigated. Laser scanning confocal microscopic (LSCM) images showed that SaSR13 was mainly colonized in the root elongation and mature zones. The inoculation with SaSR13 to Cd-treated plants significantly enhanced plant growth (by 39 and 42% for shoot and root biomass, respectively), chlorophyll contents (by 38%), and Cd concentration in the shoot and root (by 32 and 22%, respectively). SaSR13 stimulated the development of roots (increased root length, surface area, and root tips number) due to an increase in the indole-3-acid (IAA) concentrations and a decrease in the concentrations of superoxide anion (O2.-) in plants grown under Cd stress. Furthermore, inoculation with SaSR13 enhanced the release of root exudates, especially malic acid and oxalic acid, which might have facilitated the uptake of Cd by S. alfredii. It is suggested that inoculation with endophytic bacterium SaSR13 is a promising bioaugmentation method to enhance the Cd phytoextraction efficiency by S. alfredii.

Structural and functional variability in root-associated bacterial microbiomes of Cd/Zn hyperaccumulator Sedum alfredii.

Interactions between roots and microbes affect plant's resistance to abiotic stress. However, the structural and functional variation of root-associated microbiomes and their effects on metal accumulation in hyperaccumulators remain poorly understood. Here, we characterize the root-associated microbiota of a hyperaccumulating (HP) and a non-hyperaccumulating (NHP) genotype of Sedum alfredii by 16S ribosomal RNA gene profiling. We show that distinct microbiomes are observed in four spatially separable compartments: the bulk soil, rhizosphere, rhizoplane, and endosphere. Both the rhizosphere and rhizoplane were preferentially colonized by Proteobacteria, and the endosphere by Actinobacteria. The rhizosphere and endophytic microbiomes were dominated by the family of Sphingomonadaceae and Streptomycetaceae, respectively, which benefited for their survival and adaptation. The bacterial α-diversity decreases along the spatial gradient from the rhizosphere to the endosphere. Soil type and compartment were strongest determinants of root-associated community variation, and host genotype explained a small, but significant amount of variation. The enrichment of Bacteroidetes and depletion of Firmicutes and Planctomycetes in the HP endosphere compared with that of the NHP genotype may affect metal hyperaccumulation. Program PICRUSt predicted moderate functional differences in bacterial consortia across rhizocompartments and soil types. The functional categories involved in membrane transporters (specifically ATP-binding cassette transporters) and energy metabolism were overrepresented in endosphere of HP in comparison with NHP genotypes. Taken together, our study reveals substantial variation in structure and function of microbiomes colonizing different compartments, with the endophytic microbiota potentially playing an important role in heavy metal hyperaccumulation.

Enhanced Cd extraction of oilseed rape (Brassica napus) by plant growth-promoting bacteria isolated from Cd hyperaccumulator Sedum alfredii Hance.

Four plant growth-promoting bacteria (PGPB) were used as study materials, among them two heavy metal-tolerant rhizosphere strains SrN1 (Arthrobacter sp.) and SrN9 (Bacillus altitudinis) were isolated from rhizosphere soil, while two endophytic strains SaN1 (Bacillus megaterium) and SaMR12 (Sphingomonas) were identified from roots of the cadmium (Cd)/zinc (Zn) hyperaccumulator Sedum alfredii Hance. A pot experiment was carried out to investigate the effects of these PGPB on plant growth and Cd accumulation of oilseed rape (Brassica napus) plants grown on aged Cd-spiked soil. The results showed that the four PGPB significantly boosted oilseed rape shoot biomass production, improved soil and plant analyzer development (SPAD) value, enhanced Cd uptake of plant and Cd translocation to the leaves. By fluorescent in situ hybridization (FISH) and green fluorescent protein (GFP), we demonstrated the studied S. alfredii endophytic bacterium SaMR12 were able to colonize successfully in the B. napus roots. However, all four PGPB could increase seed Cd accumulation. Due to its potential to enhance Cd uptake by the plant and to restrict Cd accumulation in the seeds, SaMR12 was selected as the most promising microbial partner of B. napus when setting up a plant-microbe fortified remediation system.

Botrytis pyriformis sp. nov., a novel and likely saprophytic species of Botrytis.

A novel species of Botrytis from Sedum sarmentosum was described based on morphology and analyses of DNA sequences of nuc rDNA ITS regions and three nuclear genes (G3PDH, HSP60, RPB2). Meanwhile pathogenicity in 32 plant species, response to temperature for growth and conidial germination for the species were determined. The Botrytis species was named Botrytis pyriformis sp. nov. It was characterized by formation of grayish mycelia, brownish conidia and melanized sclerotia on PDA. The conidia are pear-shaped, melanized and covered with abundant villiform appendages on the conidial surface. Comparison of the ITS sequences confirmed its placement in the genus Botrytis Phylogenetic analysis based on DNA sequences of G3PDH, HSP60 and RPB2 genes indicated that B. pyriformis and other 30 Botrytis species form a monophyletic clade, which was further divided into three subclades. Subclade I comprised B. pyriformis alone, whereas subclades II and III comprised six and 24 Botrytis species, respectively. Botrytis pyriformis could not infect 32 plant species including S. sarmentosum, possibly due to deficiency in formation of infection cushions. This study presents a formal description and illustrations for B. pyriformis and provides experimental evidence, indicating that B. pyriformis might be a saprophytic species.

Endophytic bacterium Sphingomonas SaMR12 promotes cadmium accumulation by increasing glutathione biosynthesis in Sedum alfredii Hance.

A hydroponic experiment was conducted to verify the effects of inoculation with endophytic bacteria Sphingomonas SaMR12 on root growth, cadmium (Cd) uptake, reactive oxygen species (ROS), antioxidases, glutathione (GSH) and the related gene expression of Sedum alfredii Hance under different levels of Cd such as 0, 10, 25, 100 and 400 µM. The results showed that inoculation of SaMR12 improved Cd accumulation and upregulated glutathione synthase (GS) expression, but slightly reduced malondialdehyde (MDA) concentration and alleviated Cd-induced damage in roots. However it didn't alter the activities of antioxidant enzymes. When Cd concentration exceeded 25 µM, SaMR12 increased the concentration of GSH and the expression level of GSH1. At high Cd treatment levels (100 and 400 µM), SaMR12 significantly reduced H2O2 concentration and enhanced expression level of 1-Cys peroxiredoxin PER1 and ATPS genes. These results indicate that although SaMR12 has no significant effects on antioxidases activities, it reduces H2O2 concentration by enhancing GSH concentration and relevant genes expression, and subsequently improves Cd tolerance and accumulation.

Application of succulent plant leaves for Agrobacterium infiltration-mediated protein production.

When expressing plant cell wall degrading enzymes in the widely used tobacco (Nicotiana benthamiana) after Agrobacterium infiltration, difficulties arise due to the thin leaf structure. Thick leaved succulents, Kalanchoe blossfeldiana and Hylotelephium telephium, were tested as alternatives. A xyloglucanase, as well as a xyloglucanase inhibitor protein was successfully produced.

An endophytic bacterium Acinetobacter calcoaceticus Sasm3-enhanced phytoremediation of nitrate-cadmium compound polluted soil by intercropping Sedum alfredii with oilseed rape.

Intensive agricultural system with high input of fertilizer results in high agricultural output. However, excessive fertilization in intensive agricultural system has great potential to cause nitrate and heavy metal accumulation in soil, which is adverse to human health. The main objective of the present study was to observe the effects of intercropping and inoculation of endophytic bacterium Acinetobacter calcoaceticus Sasm3 on phytoremediation of combined contaminated soil in oilseed rape (Brassica napus L.). The results showed that with Sasm3 inoculation, the biomass of rape was increased by 10-20% for shoot, 64% for root, and 23-29% for seeds while the nitrate accumulation in rape was decreased by 14% in root and by 12% in shoot. The cadmium concentration in rape increased significantly with mono-inoculating treatment, whereas it decreased significantly after intercropping treatment. By denaturing gradient gel electrophoresis (DGGE) and real-time quantitative PCR analysis, the diversity of bacterial community and the number of nirS and nirK gene copies increased significantly with inoculation or/and intercropping treatment. In conclusion, the endophytic bacterium Sasm3-inoculated intercropping system not only improved the efficiency of clearing cadmium from soil without obstructing crop production, but also improved the quality of crop.

Inoculation with Metal-Mobilizing Plant-Growth-Promoting Rhizobacterium Bacillus sp. SC2b and Its Role in Rhizoremediation.

A plant growth-promoting bacterial (PGPB) strain SC2b was isolated from the rhizosphere of Sedum plumbizincicola grown in lead (Pb)/zinc (Zn) mine soils and characterized as Bacillus sp. based on (1) morphological and biochemical characteristics and (2) partial 16S ribosomal DNA sequencing analysis. Strain SC2b exhibited high levels of resistance to cadmium (Cd) (300 mg/L), Zn (730 mg/L), and Pb (1400 mg/L). This strain also showed various plant growth-promoting (PGP) features such as utilization of 1-aminocyclopropane-1-carboxylate, solubilization of phosphate, and production of indole-3-acetic acid and siderophore. The strain mobilized high concentration of heavy metals from soils and exhibited different biosorption capacity toward the tested metal ions. Strain SC2b was further assessed for PGP activity by phytagar assay with a model plant Brassica napus. Inoculation of SC2b increased the biomass and vigor index of B. napus. Considering such potential, a pot experiment was conducted to assess the effects of inoculating the metal-resistant PGPB SC2b on growth and uptake of Cd, Zn and Pb by S. plumbizincicola in metal-contaminated agricultural soils. Inoculation with SC2b elevated the shoot and root biomass and leaf chlorophyll content of S. plumbizincicola. Similarly, plants inoculated with SC2b demonstrated markedly higher Cd and Zn accumulation in the root and shoot system, indicating that SC2b enhanced Cd and Zn uptake by S. plumbizincicola through metal mobilization or plant-microbial mediated changes in chemical or biological soil properties. Data demonstrated that the PGPB Bacillus sp. SC2b might serve as a future biofertilizer and an effective metal mobilizing bioinoculant for rhizoremediation of metal polluted soils.

The hyperaccumulator Sedum plumbizincicola harbors metal-resistant endophytic bacteria that improve its phytoextraction capacity in multi-metal contaminated soil.

Endophyte-assisted phytoremediation has recently been suggested as a successful approach for ecological restoration of metal contaminated soils, however little information is available on the influence of endophytic bacteria on the phytoextraction capacity of metal hyperaccumulating plants in multi-metal polluted soils. The aims of our study were to isolate and characterize metal-resistant and 1-aminocyclopropane-1-carboxylate (ACC) utilizing endophytic bacteria from tissues of the newly discovered Zn/Cd hyperaccumulator Sedum plumbizincicola and to examine if these endophytic bacterial strains could improve the efficiency of phytoextraction of multi-metal contaminated soils. Among a collection of 42 metal resistant bacterial strains isolated from the tissues of S. plumbizincicola grown on Pb/Zn mine tailings, five plant growth promoting endophytic bacterial strains (PGPE) were selected due to their ability to promote plant growth and to utilize ACC as the sole nitrogen source. The five isolates were identified as Bacillus pumilus E2S2, Bacillus sp. E1S2, Bacillus sp. E4S1, Achromobacter sp. E4L5 and Stenotrophomonas sp. E1L and subsequent testing revealed that they all exhibited traits associated with plant growth promotion, such as production of indole-3-acetic acid and siderophores and solubilization of phosphorus. These five strains showed high resistance to heavy metals (Cd, Zn and Pb) and various antibiotics. Further, inoculation of these ACC utilizing strains significantly increased the concentrations of water extractable Cd and Zn in soil. Moreover, a pot experiment was conducted to elucidate the effects of inoculating metal-resistant ACC utilizing strains on the growth of S. plumbizincicola and its uptake of Cd, Zn and Pb in multi-metal contaminated soils. Out of the five strains, B. pumilus E2S2 significantly increased root (146%) and shoot (17%) length, fresh (37%) and dry biomass (32%) of S. plumbizincicola as well as plant Cd uptake (43%), whereas Bacillus sp. E1S2 significantly enhanced the accumulation of Zn (18%) in plants compared with non-inoculated controls. The inoculated strains also showed high levels of colonization in rhizosphere and plant tissues. Results demonstrate the potential to improve phytoextraction of soils contaminated with multiple heavy metals by inoculating metal hyperaccumulating plants with their own selected functional endophytic bacterial strains.