Biofertilizer based on halotolerant microorganisms promotes the growth of rice plants and alleviates the effects of saline stress.

Long-term soil salinization easily contributes to soil hardness, soil nutrient imbalance, and soil microbial diversity reduction, resulting in low rice yields in the salinized fields, and microbial remediation is one of the important measures to improve salinized soil. To verify the effect of biofertilizer based on halotolerant microorganisms on promoting rice growth and alleviating saline stress, this study discussed the effects of biofertilizer on soil microbial diversity and community structure and analyzed the correlation between the formation of microbial community structure and soil nutrient factors in the salinized field. The result, in comparison with applying inorganic fertilizer (referred to as CK), showed that notably increased soil available nitrogen, available phosphorus, available potassium, and rice paddy yield (p < 0.05) and significantly decreased soil electrical conductivity (p < 0.05) were achieved via biofertilizer (referred to as G2). Additionally, the application of biofertilizer contributes to the increase in soil microbial diversity and reorganization of microbial community structure, and through the analysis of linear discriminant analysis effect size, a notable difference in relative abundance was found in 13 genera, 6 families, and 3 orders between the control group and experimental groups (p < 0.05), and by linear discriminant analysis, Desulfomonas was further identified as the differentiated indicator. The redundancy analysis showed that available phosphorus and cation exchange capacity were the key environmental factors that affected microbial community structure and composition. Through bacterial functional prediction, increased rhizosphere soil bacterial metabolism, enzyme activity, membrane transport, and other potential functions were achieved by applying biofertilizer. Therefore, the application of biofertilizer could significantly alleviate rice growth stress and increase nutrient supply capacity in saline soil. These findings provide theoretical support for soil microbial improvement technology in the salinized field.

Comparative genomic analysis of Citrobacter sp. XT1-2-2 reveals insights into the molecular mechanism of microbial immobilization of heavy metals.

BACKGROUND: In our previous study, Citrobacter sp. XT1-2-2 was isolated from high cadmium-contaminated soils, and demonstrated an excellent ability to decrease the bioavailability of cadmium in the soil and inhibit cadmium uptake in rice. In addition, the strain XT1-2-2 could significantly promote rice growth and increase rice biomass. Therefore, the strain XT1-2-2 shows great potential for remediation of cadmium -contaminated soils. However, the genome sequence of this organism has not been reported so far.  RESULTS: Here the basic characteristics and genetic diversity of the strain XT1-2-2 were described, together with the draft genome and comparative genomic results. The strain XT1-2-2 is 5040459 bp long with an average G + C content of 52.09%, and contains a total of 4801 genes. Putative genomic islands were predicted in the genome of Citrobacter sp. XT1-2-2. All genes of a complete set of sulfate reduction pathway and various putative heavy metal resistance genes in the genome were identified and analyzed. CONCLUSIONS: These analytical results provide insights into the genomic basis of microbial immobilization of heavy metals.

Combined amendment reduces soil Cd availability and rice Cd accumulation in three consecutive rice planting seasons.

The scientific application of stabilized materials has been considered an effective method for the in situ remediation of Cd-contaminated soil. This study aimed to investigate the persistence of the effect of a combined amendment of limestone and sepiolite (LS) on soil Cd availability and accumulation in rice grown in a mildly Cd-contaminated paddy field (0.45 mg/kg of Cd) over three consecutive rice seasons. 1125-4500 kg/ha of LS was applied to the soil before the first rice planting season and 562.5-2250 kg/ha of LS was supplemented before the third rice planting season. The application of LS (1125-4500 kg/ha) increased the soil pH by 0.44-1.09, 0.18-0.53, and 0.42-0.68 in the first, second, and third season, respectively, and decreased the soil acid-extractable Cd content by 18.2-36.4%, 17.7-33.5%, and 9.6-17.6%. LS application significantly decreased the Cd contents in the rice tissues. The application of 4500 kg/ha of LS decreased the Cd content in brown rice to below the National Food Limit Standard of 0.2 mg/kg (GB 2762-2017) in the three consecutive rice seasons. However, the effect of LS on the soil-rice system was significantly weakened in the third season. The supplementary application of 562.5-2250 kg/ha of LS further decreased the Cd content in brown rice by 26.1-56.5% and decreased the health risk index by 23.7-43.8%. Therefore, it was recommended to apply 4500 kg/ha of LS in the first season and to supplement 2250 kg/ha of LS in the third season to effectively guarantee the clean production of rice in three consecutive rice seasons.

Increased biomass and reduced tissue cadmium accumulation in rice via indigenous Citrobacter sp. XT1-2-2 and its mechanisms.

Microbial remediation is a promising technique to remediate heavy metals contaminated soils. In this study, the cadmium (Cd)- resistant Citrobacter sp. XT1-2-2, isolated from heavy metals contaminated paddy soils, was investigated to evaluate the effect of this strain on soil Cd speciation, cellular Cd distribution, tissue Cd accumulation and rice biomass. The percentage of Cd2+ removal by Citrobacter sp. XT1-2-2 was up to 82.3 ± 2.1% within 240 min in the solution. The average content of soil soluble plus exchangeable and carbonate-bound fractions of Cd decreased, whereas Fe/Mn oxide-bound, organic matter-bound and residual fractions increased with bacteria inoculation. For the paddy soil inoculated with the XT1-2-2 strain, Cd concentrations of roots, culms, leaves and grains were significantly reduced by 24.1%, 46.9%, 41.5% and 66.7%, respectively. In addition, inoculation bacteria significantly increased the biomass of the roots, above-ground tissues and the rice grains. All results indicated that the XT1-2-2 strain had the ability to immobilize soil Cd and decrease Cd accumulation in rice grains. Therefore, the XT1-2-2 strain has potential for application to remediate Cd-contaminated paddy soils. It is possible to exploit a new bacterial-assisted technique for the remediation in Cd-contaminated paddy soils.

Simultaneous mitigation of tissue cadmium and lead accumulation in rice via sulfate-reducing bacterium.

The objectives of this study were to investigate the mechanism responsible for Cd and Pb immobilization by sulfate reduction to sulfide and effectiveness of decreasing Cd2+ and Pb2+ bioavailability in culture solution and paddy soils via sulfate-reducing bacterium (SRB1-1). The SRB1-1 strain, exhibiting high resistances to Cd2+ and Pb2+, was isolated from bulk soils in the metal(loid)-contaminated paddy field. During the culture of the SRB1-1 strain, the removal percentages of Cd2+ and Pb2+ from culture solution reached 99.5% and 76.0% in 72 h, respectively. The surface morphology and composition of metal precipitates formed by SRB1-1 strain were analyzed by transmission electron microscopy (TEM) and further confirmed to be CdS and PbS by X-ray diffraction (XRD) and X-ray photoelectron spectroscopy (XPS). When living SRB1-1 strain was applied in Cd and Pb-contaminated soils, the SRB1-1 strain could stably colonize using its resistance to rifampicin, and showed significantly impact on the bacterial community composition. Cd and Pb contents in rice grains were decreased by 29.5% and 26.2%, respectively, while Cd and Pb contents in the roots, culms, leaves, and husk were also decreased ranging from 19.1% to 43%, respectively. Due to growth in highly Cd and Pb contaminated soils, Cd content of the rice grains did not meet the standard for limit of Cd and Pb, but safe production of rice plants may be obtained in slightly or moderately metal(loid)-contaminated soils in the presence of the living SRB1-1 strain. These results indicated that the SRB1-1 strain could effectively reduce the Cd and Pb bioavailability in soils and uptake in rice plants. Our results highlighted the possibility to develop a new bacterial-assisted technique for reduced metal accumulation in rice grains, and also showed potential for effective synergistic bioremediation of SRB1-1 strain and rice plants in metal(loid)-contaminated soils.

Immobilization of cadmium by immobilized Alishewanella sp. WH16-1 with alginate-lotus seed pods in pot experiments of Cd-contaminated paddy soil.

This study prepared immobilized Alishewanella sp. WH16-1 using alginate and lotus seed pods as a matrix and investigated the effects of its immobilization on Cd2+ in a culture solution and in soil. Compared with the free WH16-1 strain, the immobilized WH16-1 strain possessed greater stability for long-term use and storage and higher removal ability for Cd2+ in the culture solution. A model of Cd2+ removal by the immobilized WH16-1 strain was proposed. The immobilized WH16-1 strain was incubated in the pot experiments of Cd-contaminated paddy soil for 120 days, and the pot experiments of Cd-contaminated paddy soil without the immobilized WH16-1 strain were used as a control. Compared with the control, the exchangeable and carbonate-bound Cd in the paddy soil incubated with the immobilized WH16-1 strain significantly decreased by 33.6% (P < 0.05) and 17.36%, respectively, and the Cd concentrations in the rice significantly decreased by 78.31% (P < 0.05). The results indicate that alginate-lotus seed pods can be used as excellent cost-effective cell carriers for the immobilization of Alishewanella sp. WH16-1 and that the immobilized WH16-1 strain may be applicable for the biological stabilization of Cd in Cd-contaminated paddy soil.

Impacts of a Compound Amendment on Cd Immobilization, Enzyme Activities and Crop Uptake in Acidic Cd-Contaminated Paddy Soils.

This study assessed the effectiveness of limestone-montmorillonite-rapeseed residue-Si fertilizer compound amendment on the bioavailability and crop uptake of cadmium (Cd) and enzyme activities in acidic paddy soils. Applying the compound amendment at ratios of 1%-3% increased soil pH by 0.1-1.9 units, decreased leaching ratios of soil Cd 4.0%-22%, and decreased exchangeable and carbonated Cd 42%-55% and 27%-49%, respectively. Organic matter-bound Cd increased 47%-62% (p < 0.05). Cadmium concentrations decreased in the roots, culms, leaves, and grains of rice grown in the Cd-contaminated soils by 37%-81%, 18%-73%, 29%-64% and 27%-63%, respectively, (p < 0.05). Catalase and urease activities increased 2.5%-63% and 3.9%-36%, (p < 0.05), respectively. Applying this compound amendment may significantly mitigate soil acidification and decrease the bioavailability and crop uptake of Cd in acidic Cd-contaminated paddy soils.

Draft Genome Sequence of Bacillus thuringiensis subsp. aizawai HD133.

We report here the 6,512,057-bp draft genome sequence of Bacillus thuringiensis subsp. aizawai HD133. This strain contains at least 6 cry genes and 13 candidate biosynthetic gene clusters.

Decontamination of multiple heavy metals-containing effluents through microbial biotechnology.

To decontaminate heavy metal-containing waste water, a microbial biotechnology was developed by using the synergy between Sulfate reducing bacteria (SRB), Bacillus cereus (B. cereus) and Camellia oleifera cake (COC). In this process the COC degradation assisted by B.cereus, created an anoxic environment and provided energy and nutrition for SRB. Both of B. cereus and SRB played significant roles through biosorption, bioaccumulation and biosurfactant production. Meanwhile, a flotation technology commonly used in many effluent treatments has been led into this system for increasing the efficiency as well. After desorption and regeneration with acid and deionized water, the biosorbents could be reused to adsorb metal ions. 97% of heavy metals removal was achieved by the proposed technology. For multiple heavy metals-containing solutions, the capacities are in the order of Cd2+>Zn2+>Cu2+.

Heterologous expression and genetic engineering of the tubulysin biosynthetic gene cluster using Red/ET recombineering and inactivation mutagenesis.

Although the tubulysin (tub) biosynthetic gene cluster has been located in two myxobacterial strains, it appears in both cases to be incomplete as obvious candidates for acyl transfer and oxidation functions are lacking. Here, we report the engineering of a heterologous expression system for the tub biosynthetic pathway from strain Cystobacter sp. SBCb004. The entire tub core cluster was reconstituted from two cosmids using Red/ET recombineering and heterologous expression achieved in strains Pseudomonas putida and Myxococcus xanthus. Availability of the heterologous expression system and the natural producer strain SBCb004 provided a platform for the functional investigation of various biosynthetic genes by targeted inactivation. In addition, BLAST analysis of SBCb004 genome data was used to identify multiple candidate monooxygenases, whose involvement in tubulysin assembly was evaluated using a combination of knockout mutagenesis and heterologous expression.

A Cry1Ac toxin variant generated by directed evolution has enhanced toxicity against Lepidopteran insects.

Cry1Ac insecticidal crystal proteins produced by Bacillus thuringiensis (Bt) have become an important natural biological agent for the control of lepidopteran insects. In this study, a cry1Ac toxin gene from Bacillus thuringiensis 4.0718 was modified by using error-prone PCR, staggered extension process (StEP) shuffling combined with Red/ET homologous recombination to investigate the insecticidal activity of delta-endotoxin Cry1Ac. A Cry1Ac toxin variant (designated as T524N) screened by insect bioassay showed increased insecticidal activity against Spodoptera exigua larvae while its original insecticidal activity against Helicoverpa armigera larvae was still retained. The mutant toxin T524N had one amino acid substitution at position 524 relative to the original Cry1Ac toxin, and it can accumulate within the acrystalliferous strain Cry-B and form more but a little smaller bipyramidal crystals than the original Cry1Ac toxin. Analysis of theoretical molecular models of mutant and original Cry1Ac proteins indicated that the mutation T524N located in the loop linking ß16-ß17 of domain III in Cry1Ac toxin happens in the fourth conserved block which is an arginine-rich region to form a highly hydrophobic surface involving interaction with receptor molecules. This study showed for the first time that single mutation T524N played an essential role in the insecticidal activity. This finding provides the biological evidence of the structural function of domain III in insecticidal activity of the Cry1Ac toxin, which probably leads to a deep understanding between the interaction of toxic proteins and receptor macromolecules.