Selection of the Dominant Endophytes Based on Illumina Sequencing Analysis for Controlling Bacterial Wilt of Patchouli Caused by Ralstonia solanacearum.

Bacterial wilt caused by Ralstonia solanacearum (RS) is one of the most devastating diseases in patchouli (Pogostemon cablin [Blanco] Benth.), which results in low yield and quality of patchouli. However, no stable and effective control methods have been developed yet. To evaluate the potential of dominant bacterial endophytes in biocontrol, the endophytic bacterial diversity of patchouli was investigated based on Illumina sequencing analysis, and the ability of isolates belonging to the dominant bacterial genera to control RS wilt of patchouli was explored in pot experiments. A total of 245 bacterial genera were detected in patchouli plants, with the highest relative abundance of operational taxonomic units belonging to the genus Pseudomonas detected in roots, leaves, and stems. The Pseudomonas isolates S02, S09, and S26 showed antagonistic activity against RS in vitro and displayed many plant growth-promoting characteristics, including production of indole-3-acetic acid, siderophores, and 1-aminocyclopropane-1-carboxylic acid deaminase and phosphate- and potassium-solubilizing capability. Inoculation of patchouli plants with the isolates S02, S09, and S26 significantly improved shoot growth and decreased the incidence of bacterial wilt caused by RS. The results suggest that screening of dominant bacterial endophytes for effective biocontrol agents based on Illumina sequencing analysis is more efficient than random isolation and screening procedures.

Evaluation of the potential of endophytic Trichoderma sp. isolated from medicinal plant Ampelopsis japonica against MRSA and bioassay-guided separation of the anti-MRSA compound.

Endophytic fungi have been recognized as a valuable source for the production of biologically active compounds with potential applications in various domains. This study aimed to isolate endophytic fungi from Ampelopsis japonica (Thunb.) Makino and assess their anti-MRSA activity. Meanwhile, chromatographic separation techniques were applied to analyze the constituents of endophytic fungal secondary metabolites. The isolate BLR24, which exhibited strong inhibition activity against MRSA, was identified as Trichoderma virens based on morphological characteristics and ITS sequence analyses. The ethyl acetate extract of BLR24 (EA-BLR24) showed good anti-MRSA activity with the MIC and MBC values of 25 µg/mL and 50 µg/mL, separately. The inhibition of biofilm formation was up to 34.67% under MIC concentration treatment. Meanwhile, EA-BLR24 could significantly reduce the expression of biofilm-related genes (icaA, sarA, and agrA) of MRSA. Based on LC-MS/MS analysis, twenty compounds in EA-BLR24 could be annotated using the GNPS platform, mainly diketopiperazines. The anti-MRSA compound (Fr.1.1) was obtained from EA-BLR24 by bioassay-guided fractionation and determined as gliotoxin. The results indicated that endophytic Trichoderma virens BLR24 isolated from the medical plant A. japonica roots could be a promising source of natural anti-MRSA agents. Endophytic fungal secondary metabolites are abundant in biologically active compounds. Endophytic fungi from medicinal plants could be a source yielding bioactive metabolites of pharmaceutical importance.

Study on skin infection model of Staphylococcus aureus based on analytic hierarchy process and Delphi method.

Purpose: Infectious skin diseases are a type of inflammatory skin lesions caused by pathogenic microorganisms. Because of the uncertainty of methodology, the skin infection model usually have low replication rate and lack of good evaluation system. We aimed to establish multi-index and comprehensive evaluation method for Staphylococcus aureus (S.aureus) skin-infection models through Analytic hierarchy process (AHP) and Delphi method, and screen high quality animal models through it. Materials and methods: Firstly, the evaluation indicators of skin infection were collected basing on literature research. The weight of the evaluation indicators were decided according to AHP and Delphi method. Then different ulcer models (mouse or rat) infected by S. aureus were selected as the research objects. Results: The evaluation indicators were classified into four groups of criteria (including ten sub-indicators) and given different weights, physical sign changes (0.0518), skin lesion appearance (0.2934), morphological observation (0.3184), etiological examination (0.3364). Through the evaluation system, we screened and found that the mouse ulcer model which caused by a round wound and 1.0 × 1010 CFU/mL (0.1 mL) bacterial concentration got the highest comprehensive score, and also found that the model which caused by a 1.5 cm-round wound and 1.0 × 1010 CFU/mL (0.2 mL) maybe the best rat ulcer model. Conclusions: This study has established an evaluation system based on AHP and Delphi method, also provided the best skin ulcer models selected by this system, the models are suitable for disease research and drug development research of skin ulcer.

The root-associated Fusarium isolated based on fungal community analysis improves phytoremediation efficiency of Ricinus communis L. in multi metal-contaminated soils.

Phytoremediation is a widely accepted bioremediation method of treating heavy metal contaminated soils. Nevertheless, the remediation efficiency in multi-metal contaminated soils is still unsatisfactory attributable to susceptibility to different metals. To isolate root-associated fungi for improving phytoremediation efficiency in multi-metal contaminated soils, the fungal flora in root endosphere, rhizoplane, rhizosphere of Ricinus communis L. in heavy metal contaminated soils and non-heavy metal contaminated soils were compared by ITS amplicon sequencing, and then the critical fungal strains were isolated and inoculated into host plants to improve phytoremediation efficiency in Cd, Pb, and Zn-contaminated soils. The fungal ITS amplicon sequencing analysis indicated that the fungal community in root endosphere was more susceptible to heavy metals than those in rhizoplane and rhizosphere soils and Fusarium dominated the endophytic fungal community of R. communis L. roots under heavy metal stress. Three endophytic strains (Fusarium sp. F2, Fusarium sp. F8, and Fusarium sp. F14) isolated from Ricinus communis L. roots showed high resistances to multi-metals and possessed growth-promoting characteristics. Biomass and metal extraction amount of R. communis L. with Fusarium sp. F2, Fusarium sp. F8, and Fusarium sp. F14 inoculation in Cd-, Pb- and Zn-contaminated soils were significantly higher than those without the inoculation. The results suggested that fungal community analysis-guided isolation could be employed to obtain desired root-associated fungi for enhancing phytoremediation of multi-metal contaminated soils.

Combined systematic pharmacology and urine metabonomics to study the therapeutic mechanism of type 2 diabetic treated with the herbal pair of Salvia miltiorrhiza Bunge and Pueraria montana var. lobata (Willd.) Sanjappa & Pradeep.

BACKGROUND: The herbal pair of Salvia miltiorrhiza Bunge and Pueraria montana var. lobata (Willd.) Sanjappa & Pradeep (DG) is commonly used in the treatment of type 2 diabetes (T2DM) in traditional Chinese medicine (TCM). The drug pair DG was designed by Dr. Zhu chenyu to improve the treatment of T2DM. AIM: This study combined with systematic pharmacology and urine metabonomics to explore the mechanism of DG in the treatment of T2DM. METHODS: The therapeutic effect of DG on T2DM was evaluated by fasting blood glucose (FBG) and biochemical indexes. Systematic pharmacology was used to screen the active components and targets that may be related to DG. Metabonomics was established to find urinary metabolites and pathways that may be induced by DG. Finally, integrate the results of these two parts for mutual verification. RESULTS: FBG and biochemical indexes showed that DG could reduce FBG and adjust the related biochemical indexes. Metabolomics analysis indicated that 39 metabolites were related to DG for T2DM treatment. In addition, systematic pharmacology showed compounds and potential targets which were associated with DG. Finally, 12 promising targets were selected as targets for T2DM therapy by integrating the results. CONCLUSION: The combination of metabonomics and systematic pharmacology based on LC-MS is feasible and effective, which provides strong support for exploring the effective components and pharmacological mechanism of TCM.

Isolation of actinobacterial endophytes from wheat sprouts as biocontrol agents to control seed pathogenic fungi.

Seed-borne Streptomyces can transmit vertically from generation to generation and be a mutualism between the endosymbionts and hosts. The aim of this study was to isolate and characterize endophytic Streptomyces strains from wheat sprouts, and to investigate their protection against wheat seed pathogenic fungi Penicillium. Endophytic Streptomyces sp. F6 and Streptomyces sp. F39 were isolated from wheat sprouts germinated under sterile conditions. Both Streptomyces strains could produce siderophores, and showed antagonistic activities against the seed pathogenic fungi Penicillium sp. Z17. The inoculation of Streptomyces sp. F39 and F6 could protect wheat seed germination and promote seedling growth under Penicillium sp. Z17 infection. However, the protection efficiency was impacted by the Streptomyces spore concentrations, the concentration ratios of Streptomyces spores to pathogen spores, and inoculation methods. The results suggested that wheat sprouts harbored diverse endophytic Streptomyces species which derived from wheat seeds, these strains should be more likely transmitted to the next generation, and confer competitive ability to pathogens on the offspring. Owing to the more intimate correlation between sprout endophytic flora with host plants, these strains are more suitable for mature plant interiors compared with those from rhizosphere soils and root interiors.

Biochemical Characterization of a Novel Bacterial Laccase and Improvement of Its Efficiency by Directed Evolution on Dye Degradation.

Laccase is a copper-containing polyphenol oxidase with a wide range of substrates, possessing a good application prospect in wastewater treatment and dye degradation. The purpose of this research is to study the degradation of various industrial dyes by recombinant laccase rlac1338 and the mutant enzyme lac2-9 with the highest enzyme activity after modification by error-prone PCR. Four enzyme activities improved mutant enzymes were obtained through preliminary screening and rescreening, of which lac2-9 has the highest enzyme activity. There are four mutation sites, including V281A, V281A, P309L, S318G, and D232V. The results showed that the expression of the optimized mutant enzyme also increased by 22 ± 2% compared to the unoptimized enzyme and the optimal reaction temperature of the mutant enzyme lac2-9 was 5°C higher than that of the rlac1338, and the optimal pH increased by 0.5 units. The thermal stability and pH stability of mutant enzyme lac2-9 were also improved. With ABTS as the substrate, the kcat/Km of rlac1338 and mutant strain lac2-9 are the largest than other substrates, 0.1638 and 0.618 s-1M-1, respectively, indicating that ABTS is the most suitable substrate for the recombinant enzyme and mutant enzyme. In addition, the Km of the mutant strain lac2-9 (76 µM) was significantly lower, but the kcat/Km (0.618 s-1M-1) was significantly higher, and the specific enzyme activity (79.8 U/mg) increased by 3.5 times compared with the recombinant laccase (22.8 U/mg). The dye degradation results showed that the use of rlac1338 and lac2-9 alone had no degradation effect on the industrial dyes [indigo, amaranth, bromophenol blue, acid violet 7, Congo red, coomassie brilliant blue (G250)], however, adding small molecular mediators Ca2+ and ABTS at the same time can significantly improve the degradation ability. Compared to the rlac1338, the degradation rates with the simultaneous addition of Ca2+ and ABTS of mutant enzyme lac2-9 for acid violet 7, bromophenol blue and coomassie brilliant blue significantly improved by 8.3; 3.4 and 3.4 times. Therefore, the results indicated that the error-prone PCR was a feasible method to improve the degradation activity of laccase for environmental pollutants, which provided a basis for the application of laccase on dye degradation and other environmental pollutants.

Screening, cloning, enzymatic properties of a novel thermostable cellulase enzyme, and its potential application on water hyacinth utilization.

Cellulose is the cheapest, natural, renewable organic substance that is used as a carbon source in various fields. Water hyacinth, an aquatic plant rich in cellulose, is often used as a raw material in fuel production. However, natural cellulase can be hardly used in industrial production on account of its low thermal stability and activity. In this study, a metagenomic library was constructed. Then, a new cellulase gene, cel1029, was screened by Congo red staining and expressed in the prokaryotic system. Enzymatic properties of Cel1029 were explored, including optimum temperature and pH, thermal and pH stability, and tolerance against organic solvents, metal ions, and salt solutions. Finally, its ability of degrading water hyacinth was identified and evaluated. Cel1029 displayed high homology with endoglucanase in the glycoside hydrolase family 5 (GH5) and had high stability across a broad temperature range. More than 86% of its enzymatic activities were retained between 4 and 60 °C after 24 h of incubation. Single-factor analysis and orthogonal design were further conducted to determine the optimal conditions for the highest reducing sugar yield of water hyacinth. Interestingly, Cel1029 efficiently transformed water hyacinth with a reducing sugar yield of 430.39 mg/g in 22 h. These findings may open the door for significant industrial applications of a novel GH5 cellulase (NCBI Reference Sequence: MK051001, Cel1029) and help identify more efficient methods to degrade cellulose-rich plants.

[Diversity, antibacterial activites and growth promoting characteristics of endophytic fungi from sandal (Santalum album)].

The aim of this study was to investigated the biological diversity, antibacterial activites and the plant growth-promoting traits of endophytic fungi of sandal (Santalum album), and to assess their potential in the development of antibacterial substances and rapid cultivation of sandal. The results of isolation and taxa analysis of endophytic fungi from sandal showed that 325 strains of endophytic fungi belonging to 16 genera of endophytic fungi were isolated from sandal (of which 86 from roots, 105 from stems and 134 from leaves). The isolation rate and colonization rate of endophytic fungi in different sandal parts showed the same pattern of change: leave>stems>roots. The diversity index of endophytic fungi in sandal roots was significantly higher than that of stems and leaves. The dominant endophytic fungi of sandal roots, stems and leaves showed significant differences. The dominant endophytic fungi of roots were Fusarium (50.00%) and Alternaria (10.47%), Alternaria (58.11%) and Acremonium (20.00%) for stems, and Pantoea (74.63%) for leaves. The antibacterial activity of 40 representative strains of sandal endophytic fungi were analyzed and the results showed that 90% of endophytic fungi exhibited inhibitory activity against at least one of the tested bacteria strains, and the strains with inhibitory activity to Escherichia coli, Enterobacter aerogenes, Shigella dysenteriae, Salmonella typhimurium, Staphylococcus aureus, and Bacillus subtilis accounted for 45.0%, 30%, 47.5%, 55%, 72.5%, and 62.5%, respectively. The sandal fungal endophytes with plant growth-promoting characteristics were screened, and 5 strains of endophytic fungi with phosphorus-solubilizing activity, 8 strains of endophytic fungi producing IAA, and 4 strains of endophytic fungi producing siderophores were found. Among them, endophytic fungus Monilia sp TXRF45 clould produced IAA and siderophores, and also show phosphate-solubilizing activity. The results indicated that the endophytic fungi of Sandal were rich in species diversity and their distribution had a certain tissue specificity. Some strains showed good antibacterial activity and growth-promoting properties, which could potentially applicable for the development of antibacterial substances and rapid cultivation of sandal.

Evaluation of Probiotic Diversity from Soybean (Glycine max) Seeds and Sprouts Using Illumina-Based Sequencing Method.

There is increasing interest in the use of plant probiotics as environmental-friendly and healthy biofertilizers. The study aimed at selecting for novel probiotic candidates of soybean (Glycine max). The bacteriome and mycobiome of soybean sprouts and seeds were analyzed by Illumina-based sequencing. Seeds contained more diverse bacteria than those in sprouts. The seeds contained similar fungal diversity with sprouts. Total 15 bacterial OTUs and 4 fungal OTUs were detected in seeds and sprouts simultaneously, suggesting that the sprouts contained bacterial and fungal taxa transmitted from seeds. The Halothiobacillus was the most dominant bacterial genus observed and coexisted in seeds and sprouts. The OTUs belonged to Ascomycota were the most dominant fungal taxa observed in seeds and sprouts. Halothiobacillus was firstly identified as endophytic probiotics of soybean. The results suggested that sprouts might contain diverse plant probiotics of mature plants and Illumina-based sequencing can be used to screen for probiotic candidates.

Evaluation of the Diversity of Probiotic Bacillus, Clostridium, and Bifidobacterium Using the Illumina-Based Sequencing Method.

Bacterial species of Bacillus, Lactobacillus, and Bifidobacterium in the intestinal tract have been used as probiotics. Selections for probiotic candidates by the culture-based approaches are time-consuming and labor-consuming. The aim of this study was to develop a new method based on sequencing strategies to select the probiotic Bacillus, Lactobacillus, and Bifidobacterium. The Illumina-based sequencing strategies with different specific primers for Bacillus, Clostridium, and Bifidobacterium were applied to analyze diversity of the genera in goat feces. The average number of different Bacillus, Clostridium, and Bifidobacterium OTUs (operational taxonomic units) at the 97% similarity level ranged from 1922 to 63172. The coverage index values of Bacillus, Clostridium, and Bifidobacterium calculated from the bacterial OTUs were 0.89, 0.99, and 1.00, respectively. The most genera of Bacillus (37.9%), Clostridium (53%), and Bifidobacterium (99%) were detected in goat feces by the Illumina-based sequencing with the specific primers of the genera, respectively. Higher phylogenetic resolutions of the genera in goat feces were successfully established. The results suggest that the selection for probiotic Bacillus, Clostridium, and Bifidobacterium based on the Illumina sequencing with their specific primers is reliable and feasible, and the core Bacillus, Clostridium, and Bifidobacterium species of healthy goats possess the potentials as probiotic microbial consortia.

Endophytic bacterial and fungal communities transmitted from cotyledons and germs in peanut (Arachis hypogaea L.) sprouts.

Seed-borne endophytes could be transmitted into sprouts. Whether this happened in peanuts and the difference between microbial taxa in peanut germs and cotyledons remain unknown. In this research, Illumina-based sequencing was employed to investigate the microbial taxa in peanut germs, cotyledons, and sprouts. Sulfur-oxidizing bacteria was isolated and inoculated into peanut sprouts, and then, the growth of peanut seedlings was measured. The results illustrated that diverse bacteria and fungi were detected in peanut germs, cotyledons, and sprouts. The number of bacterial OTUs declined with the germination from germs and cotyledons to sprouts. However, the number of fungal OTUs increased during the seedling procedure. Seed-borne dominant bacterial genera Halothiobacillus and Synechococcus and fungal genera Humicola, Emericella, and Penicillium were detected in sprouts. Based on the endophytic community information, the Halothiobacillus strains were isolated from sprouts. Pot experiments that illustrated the growth of peanut seedlings inoculated with the strain were promoted. These results provide new understanding into plant-microbe interactions in peanut and suggest that the selection for biocontrol agents based on mycobiome and bacteriome analysis is reliable and feasible compared with the present greenhouse selection.

Fungal endophytes and their interactions with plants in phytoremediation: A review.

Endophytic microorganisms (including bacteria and fungi) are likely to interact closely with their hosts and are more protected from adverse changes in the environment. The microbiota contribute to plant growth, productivity, carbon sequestration, and phytoremediation. Elevated levels of contaminants (i.e. metals) are toxic to most plants, the plant's metabolism and growth were impaired and their potential for metal phytoextraction is highly restricted. Exploiting endophytic microorganisms to reduce metal toxicity to plants have been investigated to improve phytoremediation efficiencies. Fungi play an important role in organic and inorganic transformation, element cycling, rock and mineral transformations, bioweathering, mycogenic mineral formation, fungal-clay interactions, and metal-fungal interactions. Endophytic fungi also showed potentials to enhance phytoremediation. Compared to bacteria, most fungi exhibit a filamentous growth habit, which provides the ability to adopt both explorative or exploitative growth strategies and form linear organs of aggregated hyphae to protect fungal translocation. However, the information regarding the role of endophytic fungi in phytoremediation are incomplete, this review highlights the taxa, physiological properties, and interaction of endophytic fungi with plants in phytoremediation.

Effects of Cd- and Pb-resistant endophytic fungi on growth and phytoextraction of Brassica napus in metal-contaminated soils.

Metal-resistant endophytic fungi from roots improved phytoremediation efficacy of host plants; however, the effects of endophytic fungi from plant aerial parts on host plants are unknown. The aim of this study was to develop a feasible method to screen fungal endophytes from stems and roots of Brassica napus and to investigate effects of the endophytic fungi on growth and phytoremediation efficiency of the plant. Endophytic Fusarium sp. CBRF44, Penicillium sp. CBRF65, and Alternaria sp. CBSF68 with different traits were isolated from roots and stems of rapes grown in a metal-contaminated soil. Fusarium sp. CBRF44 (resistant to 5 mM Cd and 15 mM Pb, isolated from roots) and Alternaria sp. CBSF68 (resistant to 1 mM Cd and 10 mM Pb, isolated from stems) could produce indole-3-acetic acid (IAA) and siderophore; Penicillium sp. CBRF65 (tolerate 2 mM Cd and 20 mM Pb, isolated from roots) could not produce IAA and siderophore but showed the highest phosphate-solubilizing activities. Fusarium sp. CBRF44 and Penicillium sp. CBRF65 significantly increased the rape biomass and promoted the extraction efficacy of Pb and Cd, while Alternaria sp. CBSF68 did not show similar results. Penicillium sp. CBRF65 and Fusarium sp. CBRF44 could be frequently recovered from inoculated rape roots, while Alternaria sp. CBSF68 was scarcely recovered. The results indicate that the colonizing capacity of endophytic fungi in roots is important to improve phytoremediation efficacy of host plants.

Engineering the robustness of Saccharomyces cerevisiae by introducing bifunctional glutathione synthase gene.

Robust, high-yielding Saccharomyces cerevisiae is highly desirable for cost-effective cellulosic ethanol production. In this study, the bifunctional glutathione (GSH) synthetase genes GCSGS at high copy number was integrated into ribosomal DNA of S. cerevisiae by Cre-LoxP system. Threefold higher GSH contents (54.9 µmol/g dry weight) accumulated in the engineered strain BY-G compared to the reference strain. Tolerance of BY-G to H2O2 (3 mM), temperature (40 °C), furfural (10 mM), hydroxymethylfurfural (HMF, 10 mM) and 0.5 mM Cd(2+) increased compared to reference strain. Twofold higher ethanol concentration was obtained by BY-G in simultaneous saccharification and fermentation of corn stover compared to the reference strain. The results showed that intracellular GSH content of S. cerevisiae has an influence on robustness. The strategy is used to engineer S. cerevisiae strains adaptive to a combination of tolerance to inhibitors and raised temperature that may occur in high solid simultaneous saccharification and fermentation of lignocellulosic feedstocks.

Characterization of cattle fecal Streptomyces strains converting cellulose and hemicelluloses into reducing sugars.

To characterize Streptomyces isolated from cattle feces for converting lignocellulose into reducing sugars, five Streptomyces strains were screened. All the strains could convert lignocellulose into reducing sugars. The strain A16 accumulate 3.3-folds more reducing sugars on cottonseed shells treated with ethanol than without the treatment (P < 0.05). The five strains did not accumulate more reducing sugars on rice straws and wheat brans than those on cottonseed shells. Compared with A10 alone, the microbial combination of F1 + A10 accumulated 19, 61, and 25 % less reducing sugars on cottonseed shell, rice straw, and wheat bran than those by A10 solely, respectively (P < 0.05). Further studies indicated that the activities of avicelase and xylanase were not correlated with the reducing sugar amount accumulated by the test strains. Strain A7 could produce more cellular lipids with xylose and glucose as the sole carbon sources. This study shows the potential for Streptomyces strains from herbivore feces to convert lignocelluloses into lipids and reducing sugars for fuel production.

Characterization of Cd-, Pb-, Zn-resistant endophytic Lasiodiplodia sp. MXSF31 from metal accumulating Portulaca oleracea and its potential in promoting the growth of rape in metal-contaminated soils.

The aim of this study was to characterize the features of a Cd-, Pb-, and Zn-resistant endophytic fungus Lasiodiplodia sp. MXSF31 and to investigate the potential of MXSF31 to remove metals from contaminated water and soils. The endophytic fungus was isolated from the stem of Portulaca oleracea growing in metal-contaminated soils. The maximum biosorption capacities of MXSF31 were 3.0 × 10(3), 1.1 × 10(4), and 1.3 × 10(4) mg kg(-1) for Cd, Pb, and Zn, respectively. The biosorption processes of Cd, Pb, and Zn by MXSF31 were well characterized with the pseudo-second-order kinetic model. The biosorption isotherm processes of Pb and Zn by the fungus were fitted better with the Langmuir model, while the biosorption processes of Cd was better fitted with the Freundlich model. The biosorption process of MXSF31 was attributed to the functional groups of hydroxyl, amino, carbonyl, and benzene ring on the cell wall. The active biomass of the strain removed more Cd, Pb, and Zn (4.6 × 10(4), 5.6 × 10(5), and 7.0 × 10(4) mg kg(-1), respectively) than the dead biomass. The inoculation of MXSF31 increased the biomass of rape (Brassica napus L.), the translocation factor of Cd, and the extraction amount of Cd by rape in the Cd+Pb-contaminated soils. The results indicated that the endophytic fungus strain had the potential to remove heavy metals from water and soils contaminated by multiple heavy metals, and plants accumulating multiple metals might harbor diverse fungi suitable for bioremediation of contaminated media.

Effects of Cd, Pb, Zn, Cu-resistant endophytic Enterobacter sr CBSB1 and Rhodotorula sp. CBSB79 on the growth and phytoextraction of Brassica plants in multimetal contaminated soils.

To survey the effects of endophytic Enterobacter sp. CBSB1 and Rhodotorula sp. CBSB79 resistant to Cd2+, Pb2+, Zn2+, and Cu2+ on the growth and phytoextraction of Brassica, the endophytes were isolated by surface- sterilized methods and characterized. The CBSB1 significantly increased 44.2% of the dry weight of Brassica napus in the multimetal contaminated soil (P < 0.05) and showed no effect or declined the dry weight of B. alboglabra, B. campestris ssp. chinensis var. cummunis, B. campestris ssp. chinensis var. utilis cv. Youqing12, B. campestris ssp. chinensis var. utilis cv. Lvbao701 plants. The dry weights of B. napus, B. campestris ssp. chinensis var. utilis, and B. alboglabra showed a significant increase when the CBSB79 was inoculated (P < 0.05). In general, inoculation with bacteria and yeast did not greatly alter the metal concentration in plant tissues. Compared to Enterobacter sp. CBSB1, the yeast Rhodotorula sp CBSB79 showed higher potentials to improve extraction efficacy of Cd, Pb, Zn, and Cu by Brassica seedlings in the field.

Enhancement of phytoremediation of Cd- and Pb-contaminated soils by self-fusion of protoplasts from endophytic fungus Mucor sp. CBRF59.

The aim of this study was to isolate protoplasts from endophytic fungi and to carry out self-fusion of protoplasts for their enhancement of metal resistance. Self-fusant CBRF59T3 with resistance to 25 mM Cd(II) was constructed by self-fusion of inactivated protoplasts from Mucor sp. CBRF59. The inoculation of CBRF59 and CBRF59T3 improved significantly the availability of Cd(II) and Pb(II) in the soil. Compared with CBRF59, CBRF59T3 inoculation increased the content of water-soluble Cd(II) by 24%. The dry weight of rape inoculated with CBRF59 and CBRF59T3 was both higher than that of the uninoculation rape. Inoculation of CBRF59T3 further increased the dry weight of rape by 62% than CBRF59 in the higher Cd(II)-+Pb(II)-contaminated soil. Compared with CBRF59, CBRF59T3 inoculation increased the concentration of Cd(II) in rape shoots by 35-189% in Cd(II)- and Cd(II)-+Pb(II)-contaminated soils. The inoculation of CBRF59T3 also enhanced the translocation of Cd(II) from roots to shoots and increased the amount of extracted Cd(II) by rape. The results indicated that the mutants constructed by protoplast fusion is a feasible and efficient method to improve stress tolerance of uncharacterized fungi for phytoremediation of soils contaminated by heavy metals.

Characterization of Cd- and Pb-resistant fungal endophyte Mucor sp. CBRF59 isolated from rapes (Brassica chinensis) in a metal-contaminated soil.

To better understand the characteristics of fungal endophytes in the development of effective phytoremediation of heavy metals, the objectives of this study were to isolate a fungal endophyte tolerant Cd and Pb from rape roots grown in a heavy metal-contaminated soil, to characterize the metal-resistant fungal endophyte, and to assess its potential applications in removal of Cd and Pb from contaminated solutions and experimental soil. The isolate CBRF59 was identified as Mucor sp. based on morphological characteristics and phylogenetic analysis. From a Cd solution of 2.0mM, the maximum biosorption capacity of Cd by dead biomass of Mucor sp. CBRF59 was 108 mg g(-1). Under the same conditions, the bioaccumulation capacity of Cd by active biomass of the strain was 173 mg g(-1). The bioaccumulation capacity of Pb by active biomass of the strain was significantly lower than that by dead biomass in the initial Pb concentrations from 1.0 to 2.0mM. The ratio of Pb to Cd and initial pH values in the mixed Cd+Pb solutions affected the bioaccumulation and biosorption capacities of the metals by CBRF59. The addition of the active mycelia of CBRF59 significantly increased the availability of soil Pb and Cd by 77% and 11.5-fold, respectively. The results showed that the endophytic fungus was potentially applicable for the decontamination of metal-polluted media.