B. subtilis CNBG-PGPR-1 induces methionine to regulate ethylene pathway and ROS scavenging for improving salt tolerance of tomato.

Soil salinity severely threatens plant growth and crop yields. The utilization of PGPR is an effective strategy for enhancing plant salt tolerance, but the mechanisms involved in this process have rarely been reported. In this study, we investigated the effects of Bacillus subtilis CNBG-PGPR-1 on improving plant salt tolerance and elucidated the molecular pathways involved. The results showed that CNBG-PGPR-1 significantly improved the cellular homeostasis and photosynthetic efficiency of leaves and reduced ion toxicity and osmotic stress caused by salt in tomato. Transcriptome analysis uncovered that CNBG-PGPR-1 enhanced plant salt tolerance through the activation of complex molecular pathways, with plant hormone signal transduction playing an important role. Comparative analysis and pharmacological experiments confirmed that the ethylene pathway was closely related to the beneficial effect of CNBG-PGPR-1 on improving plant salt tolerance. Furthermore, we found that methionine, a precursor of ethylene synthesis, significantly accumulated in response to CNBG-PGPR-1 in tomato. Exogenous L-methionine largely mimicked the beneficial effects of CNBG-PGPR-1 and activated the expression of ethylene pathway-related genes, indicating CNBG-PGPR-1 induces methionine accumulation to regulate the ethylene pathway in tomato. Finally, CNBG-PGPR-1 reduced salt-induced ROS by activating ROS scavenger-encoding genes, mainly involved in GSH metabolism and POD-related genes, which were also closely linked to methionine metabolism. Overall, our studies demonstrate that CNBG-PGPR-1-induced methionine is a key regulator in enhancing plant salt tolerance through the ethylene pathway and ROS scavenging, providing a novel understanding of the mechanism by which beneficial microbes improve plant salt tolerance.

A plant growth-promoting bacteria Priestia megaterium JR48 induces plant resistance to the crucifer black rot via a salicylic acid-dependent signaling pathway.

Xanthomonas campestris pv. campestris (Xcc)-induced black rot is one of the most serious diseases in cruciferous plants. Using beneficial microbes to control this disease is promising. In our preliminary work, we isolated a bacterial strain (JR48) from a vegetable field. Here, we confirmed the plant-growth-promoting (PGP) effects of JR48 in planta, and identified JR48 as a Priestia megaterium strain. We found that JR48 was able to induce plant resistance to Xcc and prime plant defense responses including hydrogen peroxide (H2O2) accumulation and callose deposition with elevated expression of defense-related genes. Further, JR48 promoted lignin biosynthesis and raised accumulation of frees salicylic acid (SA) as well as expression of pathogenesis-related (PR) genes. Finally, we confirmed that JR48-induced plant resistance and defense responses requires SA signaling pathway. Together, our results revealed that JR48 promotes plant growth and induces plant resistance to the crucifer black rot probably through reinforcing SA accumulation and response, highlighting its potential as a novel biocontrol agent in the future.

Antifungal Activity and Possible Mechanism of Bacillus amyloliquefaciens FX2 Against the Postharvest Apple Ring Rot Pathogen.

Botryosphaeria dothidea-induced apple ring rot is one of the most serious postharvest diseases in apple production. In our preliminary work, we isolated a bacterial strain (FX2) from an infested apple orchard. Here, we confirmed the strong antifungal activity of FX2 on B. dothidea. Through phylogenetic analysis and morphological observations, we identified FX2 as a Bacillus amyloliquefaciens strain. We also found that 10% cell-free supernatant (CFS) of FX2 significantly affected mycelial growth and morphology and almost completely inhibited spore germination and germ tube elongation in B. dothidea. Furthermore, 10% CFS damaged the cell ultrastructure, resulting in a remarkable increase in cellular leakage in B. dothidea mycelia. Thus, CFS has the potential to effectively reduce in vivo B. dothidea infection, reduced lesion diameters to 64.7% compared with the control group, and reduced disease incidence by 15%. Finally, ultrafiltration, desalting chromatography, and anion exchange chromatography showed that the antifungal constituents in CFS are composed mainly of antifungal proteins. We further characterized these potential antifungal proteins via liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. Herein, we provide novel insights into the antifungal mechanisms of B. amyloliquefaciens FX2, and we highlight its potential as a novel biocontrol agent for controlling postharvest apple ring rot.

Effects of a soil collembolan on the growth and metal uptake of a hyperaccumulator: Modification of root morphology and the expression of plant defense genes.

Soil collembolans live in close proximity to plant roots and may have a role in the phytoextraction of potentially toxic metals from contaminated soils but the underlying mechanisms remain poorly investigated. We hypothesize that soil collembolans may change the root morphology of hyperaccumulators by regulating plant physiological characteristics. Here, a pot experiment was conducted in which a cadmium (Cd) and zinc (Zn) hyperaccumulator (Sedum plumbizincicola) was grown with or without a collembolan (Folsomia candida), and plant transcriptome and hormones as well as the root characteristics of S. plumbizincicola were analyzed. F. candida promoted the growth and Cd/Zn uptake of S. plumbizincicola, the root and shoot biomass increasing by 53.3 and 34.4%, and the uptake of Cd and Zn in roots increased by 83.2 and 65.4%, respectively. Plant root morphology, total root length, root tip number and lateral root number increased significantly by 40.7, 37.2 and 33.8%, respectively, with the addition of F. candida. Transcriptome analysis reveals that the expression levels of defense-related genes in S. plumbizincicola were significantly up-regulated. In addition, the defensive plant hormones, i.e. salicylic acid in the roots, increased significantly by 338%. These results suggest that the plant in defense of the action of F. candida regulated the expression of the corresponding genes and increased the defensive plant hormones, thus modifying root morphology and plant performance. Overall, this study highlights the importance of the regulation by collembolans of plant growth and metal uptake by interaction with hyperaccumulator roots.

Strategy for Mitigating Antibiotic Resistance by Biochar and Hyperaccumulators in Cadmium and Oxytetracycline Co-contaminated Soil.

The global prevalence of antibiotic resistance genes (ARGs) is of increasing concern as a serious threat to ecological security and human health. Irrigation with sewage and farmland application of manure or biosolids in agricultural practices introduce substantial selective agents such as antibiotics and toxic metals, aggravating the transfer of ARGs from the soil environment to humans via the food chain. To address this issue, a hyperaccumulator (Sedum plumbizincicola) combined with biochar amendment was first used to investigate the mitigation of the prevalence of ARGs in cadmium and oxytetracycline co-contaminated soil by conducting a pot experiment. The addition of biochar affected the distribution of ARGs in soil and plants differently by enhancing their prevalence in the soil but restraining transmission from the soil to S. plumbizincicola. The planting of S. plumbizincicola resulted in an increase in ARGs in the soil environment. A structural equation model illustrated that mobile genetic elements played a dominant role in shaping the profile of ARGs. Taken together, these findings provide a practical understanding for mitigating the prevalence of ARGs in this soil system with complex contamination and can have profound significance for agricultural management in regard to ARG dissemination control.

Effects of biochar on the migration and transformation of metal species in a highly acid soil contaminated with multiple metals and leached with solutions of different pH.

A number of recent studies have been conducted on soil metal immobilization by biochars but there is little information on the migration and transformation of metal species in soils contaminated with multiple metals as affected by biochar and acid rain. Here, a column study investigated the effects of biochar derived from maize straw pyrolyzed at 600 °C on metal (Cu, Pb, Zn and Cd) mobility in a highly acid soil during leaching with simulated acid rain. All four metals examined were released at early stages of the leaching process and the percentages of the metals leached followed the sequence Zn > Cd > Cu > Pb. Acid rain with high acidity resulted in larger amounts of metals leached, particularly at the later stages of leaching. This enhancement of leaching by highly acidic leaching solutions was eliminated by amendment with biochar. However, the effects of biochar on metal mobility depended on metal species, with significant immobilization of soil Cu, Zn and Pb (>90%, 26% and 72%, respectively) but with no effect on soil Cd. Overall, simulated acid rain enhanced soil metal mobility and biochar reduced soil metal mobility and also alleviated the effects of acid rain. More emphasis is needed on metal speciation in the use of biochars for soil metal immobilization in areas with acid rain. The use of biochars in phytoremediation may decrease the toxicity of soil metals to the hyperaccumulator plant.

Ecotoxicity of arsenic contamination toward the soil enchytraeid Enchytraeus crypticus at different biological levels: Laboratory studies.

The ecotoxicity of arsenic (As) contamination toward small soil fauna living in soil pore water such as soil enchytraeids has rarely been studied but is important in the assessment of soil pollution. Here, the endpoints of As ecotoxicity to Enchytraeus crypticus were studied at three biological levels, i.e., individual (morphology and body tissue As concentrations), population (survival, reproduction and growth) and cell biochemistry (antioxidant enzymes CAT, POD and SOD and peroxidation malondialdehyde MDA). Contact filter paper tests without soil and single species tests with OECD artificial and field soils were conducted. Arsenic contamination resulted in severe morphological pathologies in E. crypticus and the symptoms and degree of damage increased gradually with increasing As concentration and exposure time up to 48 h. The abnormal morphological effects occurred before the impairment of fecundity. The population endpoints responded to the As concentration and the EC50 values increased in the following sequence: reproduction, juvenile body weight, adult body weight, juvenile length and adult length. Changes in biochemistry parameters were induced rapidly and changed with increasing As concentration and exposure time. The activity peak values of enzymes were 3-5 times higher and the activity maximum values of MDA were 1-3 time higher than their controls. The sensitivity of enzyme activities was generally much higher than that of MDA and CAT generally showed the highest enzyme activity. The results indicate that As contamination can be very harmful to soil enchytraeids and the endpoints of the ecotoxicity tests of soil enchytraeids can be used to complement existing soil As assessment systems or may be used alone for the assessment of soil As pollution.

Acid buffering capacity of four contrasting metal-contaminated calcareous soil types: Changes in soil metals and relevance to phytoextraction.

Four different metal-contaminated calcareous soil types, Carbonati-Perudic Cambosols (CPC), Fe-accumuli-Stagnic Anthrosols (FSA), Ochri-Aquic Cambosols (OAC) and Calci-Orthic Aridosols (COA), were investigated. The acid buffering capacity and metal-releasing behaviors of the soils were explored using an acid extraction method. Soil incubation and pot experiments were conducted to investigate changes in soil metal speciation and the enhancement of phytoextraction by soil acidification. There were several to tens of times differences in acid buffering capacities between soils. Soil calcium content may represent the major buffering system as indicated by significant linear correlations between the amount of Ca2+ released and H+ addition, and metal release into solution with H+ addition showed three stages, i.e. little release, slow release and rapid release stages. Soil carbonate-bound and Fe/Mn oxide-bound Cd and Zn decreased with the addition of H+ to all four soils, but organic matter-bound and residual metals remained unchanged. Based on the intensity of acidification, the efficiency Cd and Zn phytoextraction increased substantially with the addition of H+ in the case of the CPC but not the FSA which had a higher acid buffering capacity than the CPC. Hence, it may be concluded that the acid buffering capacity and changes in soil metal fractions with acidification of contaminated calcareous soil types should be determined before phytoextraction of these soils is attempted.

Use of a hyperaccumulator and biochar to remediate an acid soil highly contaminated with trace metals and/or oxytetracycline.

Biochars and hyperaccumulators have been widely used for the remediation of trace metal contaminated soils through immobilization and phytoextraction. These two options have rarely been used simultaneously despite their potential to achieve a greater decline in trace metal availability and higher removal efficiency in polluted soils. This study investigated the combined effects of biochar and the cadmium/zinc (Cd/Zn) hyperaccumulator Sedum plumbizincicola in a pot experiment and examined the effect of an antibiotic (oxytetracycline, OTC) in an acid soil spiked with Cd/Zn alone and with OTC. Biochar amendment alone significantly decreased soil CaCl2-extractable Cd and Zn by 22.7 and 43.1%, respectively. Growing S. plumbizincicola alone resulted in 11.3% Cd and 3.88% Zn removal after ten weeks of phytoextraction. Growing S. plumbizincicola with biochar resulted in higher decreases in extractable Cd and Zn by 60.0% and 53.2%, respectively, and more than three times Cd and Zn removal efficiencies compared to growing S. plumbizincicola without biochar. The results indicate that biochar addition promoted plant growth and increased shoot trace metal concentrations, consequently increasing the removal efficiency and that soil trace metal removal by the hyperaccumulator further reduced the extractable trace metals in addition to immobilization by biochar. Biochar amendment decreased plant OTC concentrations. However, OTC showed no effect on trace metal phytoextraction. Results indicate that the simultaneous use of biochar and the hyperaccumulator can give high Cd/Zn phytoextraction efficiency in terms of both soil total and available trace metal concentrations in acid soils highly contaminated with trace metals or trace metals and OTC.

Sorption of sulfamethazine to biochars as affected by dissolved organic matters of different origin.

Sorption characteristic of sulfamethazine (SMT) to straw biochars pyrolyzed at 300°C (BC300) and 600°C (BC600), and the effect of ubiquitous DOM were investigated. Results showed that physisorption (partition) and weak chemical binding (π-π EDA interaction) dominated the sorption of SMT to BC300 and BC600, respectively. Graphene sheets in biochar played important roles in the sorption of SMT, leading to higher sorption capacity (Kf) on BC600 (1.77mg1-nLng-1) than BC300 (0.11mg1-nLng-1). Sorption amount of SMT to BC300 was not affected by polysaccharide and malic acid, while it was slightly promoted by citric acid, but dramatically increased 1.25 times by methacrylic acid through decreasing solution pH and providing new sorption sites. Humic acid and bovine serum albumin restrained the sorption of SMT to BC600, but enhanced SMT- adsorption to BC300. The chemical nature of DOM, biochar properties and antibiotic species co-determined the impact of DOM on antibiotics adsorption.

Effect of cetyltrimethyl ammonium bromide on uptake of polycyclic aromatic hydrocarbons by carrots.

This is the first study investigating the effect of cationic surfactants on the mobility of polycyclic aromatic hydrocarbons (PAHs) in aged contaminated soils and on PAH bioaccumulation in tuberous vegetables. In an aerobic soil incubation experiment, 150 mg/kg cetyltrimethyl ammonium bromide (CTMAB) decreased the bioavailability of PAHs primarily via immobilization (by 13%). In a carrot pot experiment, the effectiveness of CTMAB to reduce PAH uptake by carrots decreased with time. Accordingly, the bioavailability of PAHs in the soil decreased in the first 90 days and then increased and remained stable until harvest. In the leaching test, the leaching loss of CTMAB (15%) was lower in soils treated with small amounts of CTMAB in several applications than it was in soils (24%) treated once with CTMAB. Therefore, CTMAB, when applied in appropriate doses via addition methods, can effectively reduce the environmental risk of PAH entering humans and livestock through the food chain.

Effect of extracellular polymeric substance components on the sorption behavior of 2,2',4,4'-tetrabromodiphenyl ether to soils: Kinetics and isotherms.

Microbial extracellular polymeric substances (EPS) and persistent organic pollutants (POPs) commonly exist in the soil environment. Currently, there is a knowledge gap regarding the effect of EPS on the fate of POPs in soil. In the present study, 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) was used as a model compound to investigate the effects of bovine serum albumin (BSA) and sodium alginate (SA) - mimicking the main components of EPS - on sorption of POPs to soils, through batch experiments. Irrespective of the concentration of BSA: the addition of BSA did enhance the sorption capacity of BDE-47 to soils, due to generation of more sorption sites. For SA, it increased the sorption capacity of BDE-47 at low BDE-47 concentrations, while the presence of SA negatively affected sorption of BDE-47 at high BDE-47 concentrations. The partition effect dominates the sorption of BDE-47 to soils, but after adding either BSA or SA, the sorption of BDE-47 to soils is dominated by surface sorption. Film diffusion and intra-particle diffusion were also involved in the sorption process with and without BSA or SA, with the latter being the rate-limiting step. The heterogeneous surface and nonlinear sorption behavior of BDE-47 to soils increased in the presence of either BSA or SA. The FTIR spectra indicated that the aromatic CC, H-bonds and OH groups may be involved in the sorption process. Therefore, BSA enhanced the retention of BDE-47 to soil, while SA's influence on BDE-47 sorption to soil depended on the concentration of BDE-47.

The effect of Cu(2+) chelation on the direct photolysis of oxytetracycline: A study assisted by spectroscopy analysis and DFT calculation.

The extensive usage of OTC and Cu(2+) in livestock and poultry industry caused high residues in natural environment. Co-contamination of OTC and Cu(2+) was a considerable environmental problem in surface waters. In this study, Cu(2+) mediated direct photolysis of OTC was studied. Cu(2+) chelating with OTC was found to greatly inhibit OTC photodegradation. To reveal the chelation mechanism of OTC-Cu complexes, multiple methods including UV-Vis absorption spectra, Infrared (IR) spectra, mass spectroscopy, and density functional theoretical (DFT) modeling were performed. Four OTC-Cu complexes were proposed. Cu(2+) preferably bond to O11O12 site with the binding constants logK = 8.19 and 7.86 for CuHL+ and CuL±, respectively. The second chelating site was suggested to be O2O3 with the binding constants of logK = 4.41 and 4.62 for Cu2HL3+ and Cu2L2+, respectively. The suppressed quantum yield of OTC by Cu2+ chelation was accused for their intra-/inter-molecular electron transfer, by which the energy in activated states was distributed. The occurrence of electron transfer between BCD ring and A ring also from BCD ring to Cu was evidenced by the TD-DFT result only for the OTC-Cu complexes. Besides, the cyclic voltammetry measurement also suggested one OTC-Cu(II)/OTC-Cu(I) redox couple. These results suggested that the persistence of OTC in environmental surface waters will probably be underestimated for neglecting the chelating effect of Cu2+. The photolysis quantum yield of OTC-Cu complexes, as well as the specific molar absorption constants, the equilibrium binding constants of Cu2+ with OTC could contribute to more accurate kinetic models of OTC.

Changes in metal availability, desorption kinetics and speciation in contaminated soils during repeated phytoextraction with the Zn/Cd hyperaccumulator Sedum plumbizincicola.

Phytoextraction is one of the most promising technologies for the remediation of metal contaminated soils. Changes in soil metal availability during phytoremediation have direct effects on removal efficiency and can also illustrate the interactive mechanisms between hyperaccumulators and metal contaminated soils. In the present study the changes in metal availability, desorption kinetics and speciation in four metal-contaminated soils during repeated phytoextraction by the zinc/cadmium hyperaccumulator Sedum plumbizincicola (S. plumbizincicola) over three years were investigated by chemical extraction and the DGT-induced fluxes in soils (DIFS) model. The available metal fractions (i.e. metal in the soil solution extracted by CaCl2 and by EDTA) decreased greatly by >84% after phytoextraction in acid soils and the deceases were dramatic at the initial stages of phytoextraction. However, the decreases in metal extractable by CaCl2 and EDTA in calcareous soils were not significant or quite low. Large decreases in metal desorption rate constants evaluated by DIFS were found in calcareous soils. Sequential extraction indicated that the acid-soluble metal fraction was easily removed by S. plumbizincicola from acid soils but not from calcareous soils. Reducible and oxidisable metal fractions showed discernible decreases in acid and calcareous soils, indicating that S. plumbizincicola can mobilize non-labile metal for uptake but the residual metal cannot be removed. The results indicate that phytoextraction significantly decreases metal availability by reducing metal pool sizes and/or desorption rates and that S. plumbizincicola plays an important role in the mobilization of less active metal fractions during repeated phytoextraction.

Effects of pH and metal ions on oxytetracycline sorption to maize-straw-derived biochar.

Biochars produced from biomass residues have been recognized as effective sorbents to hydrophobic compounds, but knowledge on sorption of antibiotics to biochar and its mechanisms are still inadequate. Sorption of oxytetracycline (OTC) in aqueous solution to maize-straw-derived biochar, and the effect of pH and metal ions, was investigated in batch experiments, and the main sorption mechanisms were elucidated using FTIR and zeta potential measurements. The results showed that sorption of OTC on biochar was highly pH-dependant. The amount of sorbed OTC first increased and then decreased with increasing pH, and maximum sorption was achieved at pH 5.5. Cu(2+) enhanced the sorption of OTC, while Pb(2+) slightly reduced the sorption under acidic conditions. Other metal ions had no significant effect on the sorption of OTC to biochar. Surface complexation, through π-π interaction and metal bridging, was the most important sorption mechanism although cation exchange might have played a role.

Bioavailability assessment of hexachlorobenzene in soil as affected by wheat straw biochar.

Biochar incorporation with soil could increase sorption of organic contaminants, thereby reducing their bioavailability. In this study, the effects of wheat straw biochar on the sorption, dissipation and bioavailability of hexachlorobenzene (HCB), a typical persistent organic pollutant (POP), were investigated in laboratory experiments. We observed that HCB sorption by biochar was 42 times higher than that by soil and the sorption isotherm was linear for the concentration range studied. Biochar amendments reduced HCB dissipation, volatilization and earthworm (Eisenia foetida) uptake of HCB from soil. Hydroxypropyl-ß-cyclodextrin extraction correlated better with the earthworm bioassay than butanol extraction of HCB in biochar-amended soil. The results of both chemical extraction and earthworm bioassay indicate that biochar amendment of soil resulted in a rapid reduction in the bioavailability of HCB, even for the 0.1% biochar application rate. This suggested that wheat straw biochar could potentially be used in immobilizing POPs in contaminated sites.