A phased small interfering RNA-derived pathway mediates lead stress tolerance in maize.

Phased small interfering RNAs (phasiRNAs) are a distinct class of endogenous small interfering RNAs, which regulate plant growth, development and environmental stress response. To determine the effect of phasiRNAs on maize (Zea mays L.) tolerance to lead (Pb) stress, the roots of 305 maize lines under Pb treatment were subjected to generation of individual databases of small RNAs. We identified 55 high-confidence phasiRNAs derived from 13 PHAS genes (genes producing phasiRNAs) in this maize panel, of which 41 derived from nine PHAS loci were negatively correlated with Pb content in the roots. The potential targets of the 41 phasiRNAs were enriched in ion transport and import. Only the expression of PHAS_1 (ZmTAS3j, Trans-Acting Short Interference RNA3) was regulated by its cis-expression quantitative trait locus and thus affected the Pb content in the roots. Using the Nicotiana benthamiana (N. benthamiana) transient expression system, 5'-rapid amplification of cDNA ends, and Arabidopsis heterologously expressed, we verified that ZmTAS3j was cleaved by zma-miR390 and thus generated tasiRNA targeting ARF genes (tasiARFs), and that the 5' and 3' zma-miR390 target sites of ZmTAS3j were both necessary for efficient biosynthesis and functional integrity of tasiARFs. We validated the involvement of the zma-miR390-ZmTAS3j-tasiARF-ZmARF3-ZmHMA3 pathway in Pb accumulation in maize seedlings using genetic, molecular, and cytological methods. Moreover, the increased Pb tolerance in ZmTAS3j-overexpressed lines was likely attributed to the zma-miR390-ZmTAS3j-tasiARF-ZmARF3-SAURs pathway, which elevated indole acetic acid levels and thus reactive oxygen species scavenging capacity in maize roots. Our study reveals the importance of the TAS3-derived tasiRNA pathway in plant adaptation to Pb stress.

Genome-wide identification and characterization of NAC transcription factor family members in Trifolium pratense and expression analysis under lead stress.

BACKGROUND: The NAC TF family is widely involved in plant responses to various types of stress. Red clover (Trifolium pratense) is a high-quality legume, and the study of NAC genes in red clover has not been comprehensive. The aim of this study was to analyze the NAC gene family of red clover at the whole-genome level and explore its potential role in the Pb stress response. RESULTS: In this study, 72 TpNAC genes were identified from red clover; collinearity analysis showed that there were 5 pairs of large fragment replicators of TpNAC genes, and red clover was found to be closely related to Medicago truncatula. Interestingly, the TpNAC genes have more homologs in Arabidopsis thaliana than in soybean (Glycine max). There are many elements in the TpNAC genes promoters that respond to stress. Gene expression analysis showed that all the TpNAC genes responded to Pb stress. qRT-PCR showed that the expression levels of TpNAC29 and TpNAC42 were significantly decreased after Pb stress. Protein interaction network analysis showed that 21 TpNACs and 23 other genes participated in the interaction. In addition, the TpNAC proteins had three possible 3D structures, and the secondary structure of these proteins were mainly of other types. These results indicated that most TpNAC members were involved in the regulation of Pb stress in red clover. CONCLUSION: These results suggest that most TpNAC members are involved in the regulation of Pb stress in red clover. TpNAC members play an important role in the response of red clover to Pb stress.

Flavonoid metabolism plays an important role in response to lead stress in maize at seedling stage.

BACKGROUND: Pb stress, a toxic abiotic stress, critically affects maize production and food security. Although some progress has been made in understanding the damage caused by Pb stress and plant response strategies, the regulatory mechanisms and resistance genes involved in the response to lead stress in crops are largely unknown. RESULTS: In this study, to uncover the response mechanism of maize to Pb stress phenotype, physiological and biochemical indexes, the transcriptome, and the metabolome under different concentrations of Pb stress were combined for comprehensive analysis. As a result, the development of seedlings and antioxidant system were significantly inhibited under Pb stress, especially under relatively high Pb concentrations. Transcriptome analysis revealed 3559 co-differentially expressed genes(co-DEG) under the four Pb concentration treatments (500 mg/L, 1000 mg/L, 2000 mg/L, and 3000 mg/L Pb(NO3)2), which were enriched mainly in the GO terms related to DNA-binding transcription factor activity, response to stress, response to reactive oxygen species, cell death, the plasma membrane and root epidermal cell differentiation. Metabolome analysis revealed 72 and 107 differentially expressed metabolites (DEMs) under T500 and T2000, respectively, and 36 co-DEMs. KEGG analysis of the DEMs and DEGs revealed a common metabolic pathway, namely, flavonoid biosynthesis. An association study between the flavonoid biosynthesis-related DEMs and DEGs revealed 20 genes associated with flavonoid-related metabolites, including 3 for genistin and 17 for calycosin. CONCLUSION: In summary, the study reveals that flavonoid metabolism plays an important role in response to Pb stress in maize, which not only provides genetic resources for the genetic improvement of maize Pb tolerance in the future but also enriches the theoretical basis of the maize Pb stress response.

Tetragonal crystalline MnO nanoparticles alleviate Pb stress in wheat by modulating antioxidant enzymes in leaves.

Agriculture ecosystems are seriously threatened by lead (Pb) contamination, which impacts plant growth and productivity. In this study, green synthesized manganese oxide nanoparticles (MnO NPs) using citrus peel were used for priming of wheat seeds. For the synthesis of MnO nanoparticles, peel extract of Citrus paradisi and 1 mM solution of manganese acetate were stirred and calcinated at 500 °C. Successful synthesis of MnO NPs was determined using advanced techniques. In Fourier-transform infrared spectroscopy (FTIR), the presence of amines, alkanes, aldehydes, and alcohol molecules, on the surface of MnO NPs, confirmed their stability. X-ray diffraction analysis described their average size (22 nm), while scanning electron microscopy showed tetragonal crystalline shape and nano-flowers structure of MnO NPs. Sharp peaks of energy dispersive x-ray analysis described the presence of oxygen (28.81%) and manganese (71.19%) on MnO NPs. Priming of wheat seeds with synthesized MnO NPs significantly improved the growth attributes of wheat seedlings including the size of leaf, root length, size of shoots, chlorophyll and carotenoid contents, relative water content, decreased relative electrolyte leakage, high proline accumulation and decreased concentration of malondialdehyde. Application of MnO NPs also helped plants to accumulate antioxidant enzymes in their leaves. These results proved that the priming of MnO NPs can greatly reduce lead-induced stress in wheat seedlings and these NPs can also be used for the priming of other crops.

Toxic effects of lead on plants: integrating multi-omics with bioinformatics to develop Pb-tolerant crops.

MAIN CONCLUSION: Lead disrupts plant metabolic homeostasis and key structural elements. Utilizing modern biotechnology tools, it's feasible to develop Pb-tolerant varieties by discovering biological players regulating plant metabolic pathways under stress. Lead (Pb) has been used for a variety of purposes since antiquity despite its toxic nature. After arsenic, lead is the most hazardous heavy metal without any known beneficial role in the biological system. It is a crucial inorganic pollutant that affects plant biochemical and morpho-physiological attributes. Lead toxicity harms plants throughout their life cycle and the extent of damage depends on the concentration and duration of exposure. Higher levels of lead exposure disrupt numerous key metabolic activities of plants including oxygen-evolving complex, organelles integrity, photosystem II connectivity, and electron transport chain. This review summarizes the detrimental effects of lead toxicity on seed germination, crop growth, and yield, oxidative and ultra-structural alterations, as well as nutrient absorption, transport, and assimilation. Further, it discusses the Pb-induced toxic modulation of stomatal conductance, photosynthesis, respiration, metabolic-enzymatic activity, osmolytes accumulation, and antioxidant activity. It is a comprehensive review that reports on omics-based studies along with morpho-physiological and biochemical modifications caused by lead stress. With advances in DNA sequencing technologies, genomics and transcriptomics are gradually becoming popular for studying Pb stress effects in plants. Proteomics and metabolomics are still underrated and there is a scarcity of published data, and this review highlights both their technical and research gaps. Besides, there is also a discussion on how the integration of omics with bioinformatics and the use of the latest biotechnological tools can aid in developing Pb-tolerant crops. The review concludes with core challenges and research directions that need to be addressed soon.

Metabolomics analysis reveal the molecular responses of high CO2 concentration improve resistance to Pb stress of Oryza sativa L. seedlings.

Rice seedlings were exposed to two CO2 concentrations (400 ± 20 and 800 ± 20 µmol mol-1) and three PbNO3 concentrations (0, 50 and 100 µmol L-1) for 10 days to explore the regulatory mechanisms of elevated CO2 for Pb stress resistance. Electrical conductivity, MDA content, SOD, POD, CAT activities and metabolomics changes were studied. Results showed that: Pb stress damaged cell membrane system, electrical conductivity and MDA content increased 49.34 % and 73.27 %, respectively, and some antioxidant enzymes activities increased. Sugar, polyol, amino acid metabolism and fatty acid ß-oxidation were all enhanced to improve osmotic adjustments, maintain cell membrane stability, supply energy, nitrogen assimilates and antioxidant capacity; Under composite treatments, cell membrane damage was reduced, activities of protective enzymes increased compared with only Pb stress, POD activity increased the most (49.14 %) under severe Pb composite treatment. High CO2 caused the enhance of cells antioxidant capacity, TCA cycle intermediate products contents and fatty acid desaturation under mild Pb stress. Many sugars, polyols and amino acids contents were increased as osmotic regulatory substances by high CO2 under severe Pb stress; Secondary metabolites played an important role under Pb stress and composite treatments. The object of this study is to provide a possible molecular mechanism of rice response to Pb stress under high CO2 in the future.

Transcriptome modulation by endophyte drives rice seedlings response to Pb stress.

This study investigated the growth, SPAD value, chlorophyll fluorescence and transcriptome response of endophyte uninoculated and inoculated rice seedlings under Pb stress after treatment of 1 d and 5 d. Inoculation of endophytes significantly improved the plant height, SPAD value, Fv/F0, Fv/Fm and PIABS by 1.29, 1.73, 0.16, 1.25 and 1.90 times on the 1 d, by 1.07, 2.45, 0.11, 1.59 and 7.90 times on the 5 d, respectively, however, decreased the root length by 1.11 and 1.65 times on the 1 d and 5 d, respectively under Pb stress. Analysis of rice seedlings leaves by RNA-seq, there were 574 down-regulated and 918 up-regulated genes after treatment of 1 d, 205 down-regulated and 127 up-regulated genes after treatment of 5 d, of which 20 genes (11 up-regulated and 9 down-regulated) exhibited the same changing pattern after treatment of 1 d and 5 d. Using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) to annotate these DEGs, and it was found that many of DEGs involved in photosynthesis, oxidative detoxification, hormone synthesis and signal transduction, protein phosphorylation/kinase and transcription factors. These findings provide new insights into the molecular mechanism of interaction between endophyte and plants under heavy metal stress, and contribute to agricultural production in limited environments.

Transcriptome-Wide Identification and Response Pattern Analysis of the Salix integra NAC Transcription Factor in Response to Pb Stress.

The NAC (NAM-ATAF1/2-CUC) transcription factor family is one of the largest plant-specific transcription factor families, playing an important role in plant growth and development and abiotic stress response. As a short-rotation woody plant, Salix integra (S. integra) has high lead (Pb) phytoremediation potential. To understand the role of NAC in S. integra Pb tolerance, 53 SiNAC transcripts were identified using third-generation and next-generation transcriptomic data from S. integra exposed to Pb stress, and a phylogenetic analysis revealed 11 subfamilies. A sequence alignment showed that multiple subfamilies represented by TIP and ATAF had a gene that produced more than one transcript under Pb stress, and different transcripts had different responses to Pb. By analyzing the expression profiles of SiNACs at 9 Pb stress time points, 41 of 53 SiNACs were found to be significantly responsive to Pb. Short time-series expression miner (STEM) analysis revealed that 41 SiNACs had two significant Pb positive response patterns (early and late), both containing 10 SiNACs. The SiNACs with the most significant Pb response were mainly from the ATAF and NAP subfamilies. Therefore, 4 and 3 SiNACs from the ATAF and NAP subfamilies, respectively, were selected as candidate Pb-responsive SiNACs for further structural and functional analysis. The RT-qPCR results of 7 transcripts also confirmed the different Pb response patterns of the ATAF and NAP subfamilies. SiNAC004 and SiNAC120, which were randomly selected from two subfamilies, were confirmed to be nuclear localization proteins by subcellular localization experiments. Functional prediction analysis of the associated transcripts of seven candidate SiNACs showed that the target pathways of ATAF subfamily SiNACs were "sulfur metabolism" and "glutathione metabolism", and the target pathways of NAP subfamily SiNACs were "ribosome" and "phenylpropanoid biosynthesis". This study not only identified two NAC subfamilies with different Pb response patterns but also identified Pb-responsive SiNACs that could provide a basis for subsequent gene function verification.

De Novo Transcriptome Assembly, Gene Annotations, and Characterization of Functional Profiling Reveal Key Genes for Lead Alleviation in the Pb Hyperaccumulator Greek Mustard (Hirschfeldia incana L.).

Lead (Pb) contamination is a widespread environmental problem due to its toxicity to living organisms. Hirschfeldia incana L., a member of the Brassicaceae family, commonly found in the Mediterranean regions, is characterized by its ability to tolerate and accumulate Pb in soils and hydroponic cultures. This plant has been reported as an excellent model to assess the response of plants to Pb. However, the lack of genomic data for H. incana hinders research at the molecular level. In the present study, we carried out RNA deep transcriptome sequencing (RNA-seq) of H. incana under two conditions, control without Pb(NO3)2 and treatment with 100 µM of Pb(NO3)2 for 15 days. A total of 797.83 million reads were generated using Illumina sequencing technology. We assembled 77,491 transcript sequences with an average length of 959 bp and N50 of 1330 bp. Sequence similarity analyses and annotation of these transcripts were performed against the Arabidopsis thaliana nr protein database, Gene Ontology (GO), and KEGG databases. As a result, 13,046 GO terms and 138 KEGG maps were created. Under Pb stress, 577 and 270 genes were differentially expressed in roots and aboveground parts, respectively. Detailed elucidation of regulation of metal transporters, transcription factors (TFs), and plant hormone genes described the role of actors that allow the plant to fine-tune Pb stress responses. Our study revealed that several genes related to jasmonic acid biosynthesis and alpha-linoleic acid were upregulated, suggesting these components' implication in Hirschfeldia incana L responses to Pb stress. This study provides data for further genomic analyses of the biological and molecular mechanisms leading to Pb tolerance and accumulation in Hirschfeldia incana L.

Exogenously Applied Sodium Nitroprusside Mitigates Lead Toxicity in Rice by Regulating Antioxidants and Metal Stress-Related Transcripts.

Sustainable agriculture is increasingly being put in danger by environmental contamination with dangerous heavy metals (HMs), especially lead (Pb). Plants have developed a sophisticated mechanism for nitric oxide (NO) production and signaling to regulate hazardous effects of abiotic factors, including HMs. In the current study, we investigated the role of exogenously applied sodium nitroprusside (SNP, a nitric oxide (NO) donor) in ameliorating the toxic effects of lead (Pb) on rice. For this purpose, plants were subjected to 1.2 mM Pb alone and in combination with 100 µM SNP. We found that under 1.2 mM Pb stress conditions, the accumulation of oxidative stress markers, including hydrogen peroxide (H2O2) (37%), superoxide anion (O2-) (28%), malondialdehyde (MDA) (33%), and electrolyte leakage (EL) (34%), was significantly reduced via the application of 100 µM SNP. On the other hand, under the said stress of Pb, the activity of the reactive oxygen species (ROS) scavengers such as polyphenol oxidase (PPO) (60%), peroxidase (POD) (28%), catalase (CAT) (26%), superoxide dismutase (SOD) (42%), and ascorbate peroxidase (APX) (58%) was significantly increased via the application of 100 µM SNP. In addition, the application of 100 µM SNP rescued agronomic traits such as plant height (24%), number of tillers per plant (40%), and visible green pigments (44%) when the plants were exposed to 1.2 mM Pb stress. Furthermore, after exposure to 1.2 mM Pb stress, the expression of the heavy-metal stress-related genes OsPCS1 (44%), OsPCS2 (74%), OsMTP1 (83%), OsMTP5 (53%), OsMT-I-1a (31%), and OsMT-I-1b (24%) was significantly enhanced via the application of 100 µM SNP. Overall, our research evaluates that exogenously applied 100 mM SNP protects rice plants from the oxidative damage brought on by 1.2 mM Pb stress by lowering oxidative stress markers, enhancing the antioxidant system and the transcript accumulation of HMs stress-related genes.

Transcriptome profilings of two tall fescue (Festuca arundinacea) cultivars in response to lead (Pb) stress.

BACKGROUND: Lead (Pb) is one of the most toxic heavy metal environmental pollutants. Tall fescue is an important cold season turf grass which can tolerate and accumulate substantial amount of Pb. To estimate genes related to Pb response and the molecular mechanism associated with Pb tolerance and accumulation, we analyzed the transcriptome of tall fescue in response to Pb treatment. RESULTS: RNA-sequencing was performed in two tall fescue cultivars, Pb tolerant Silverado and Pb sensitive AST7001. A total of 810,146 assembled unique transcripts representing 25,415 unigenes were obtained from the tall fescue leaves. Among the panel, 3,696 differentially expressed genes (DEGs) were detected between the Pb treated (1000 mg/L) and untreated samples. Gene ontology (GO) and pathway enrichment analysis demonstrated that the DEGs were mainly implicated in energy metabolism, metabolism of terpenoids and polyketides, and carbohydrate metabolism related pathways. The expression patterns of 16 randomly selected genes were in consistent with that from the Solexa analysis using quantitative reverse-transcription PCR. In addition, compared to the common transcriptional response to Pb stress in both cultivars, the regulation of numerous genes including those involved in zeatin biosynthesis, limonene and pinene degradation, phagosome was exclusive to one cultivar. CONCLUSIONS: The tall fescue assembled transcriptome provided substantial molecular resources for further genomics analysis of turfgrass in response to heavy metal stress. The significant expression difference of specific unigenes may account for Pb tolerance or accumulation in two different tall fescue cultivars. This study provided new insights for the investigation of the molecular basis of Pb tolerance and accumulation in tall fescue as well as other related turf grass species.

Novel quantitative metabolomic approach for the study of stress responses of plant root metabolism.

Quantitative metabolomics (qMetabolomics) is a powerful tool for understanding the intricate metabolic processes involved in plant abiotic stress responses. qMetabolomics is hindered by the limited coverage and high cost of isotopically labeled standards. In this study, we first selected 271 metabolites which might play important roles in abiotic stress responses as the targets and established a comprehensive LC-MS/MS based qMetabolomic method. We then developed a novel metabolic labeling method using E. coli-Saccharomyces cerevisiae two-step cultivation for the production of uniformly (13)C-labeled metabolites as internal standards. Finally, we applied the developed qMetabolomic method to investigate the influence of Pb stress on maize root metabolism. The absolute concentration of 226 metabolites in maize roots was accurately quantified in a single run within 30 min. Our study also revealed that glycolysis, purine, pyrimidine, and phospholipids were the main metabolic pathways in maize roots involved in Pb stress response. To our knowledge, this is the most comprehensive qMetabolomic method for plant metabolomics thus far. We developed a simple and inexpensive metabolic labeling method which dramatically expanded the availability of uniformly (13)C labeled metabolites. Our findings also provided new insights of maize metabolic responses to Pb stress.

Stress-mitigating behavior of glycine betaine to enhance growth performance by suppressing the oxidative stress in Pb-stressed barley genotypes.

The toxicity of lead (Pb) in agricultural soil is constantly increasing as a result of anthropogenic activities. Pb is one of the most phytotoxic metals in soil that accumulates in plant tissue, resulting in yield loss. It is currently becoming more popular to supplement glycine betaine (GB) for Pb-induced stress tolerance in crop plants. Currently, no report describes the use of GB as a stress mitigator for growth attributes and stress-specific biomarkers in barley plants under Pb stress conditions. Hence, the present research was designed to examine the stress-mitigating behavior of GB on various growth attributes including germination percentage, seed vigor index (SVI), radicle length, plant biomass (fresh and dry), shoot and root length, physiological attributes such as relative water content (RWC), and stress-specific biomarkers like electrolyte leakage (EL), and H2O2 content of two barley varieties viz. BH959 and BH946 at three Pb stress treatments (15 mM, 25 mM, and 35 mM), with and without GB (2 mM) supplementation in natural conditions. The present investigation showed that at the highest Pb stress (35 mM), the germination rate was reduced to zero, and the growth attributes and RWC of both barley varieties were also reduced as compared to the non-stressed plants (control) with an increase in Pb treatment. However, EL up to 70% and H2O2 content up to 30% increased with an increase in Pb stress concentration indicated by ROS accumulation, resulting in more oxidative stress. Additionally, GB application alleviated the toxic effect of Pb stress by improving the rate of germination by 33.3% and growth performance by reducing the ROS accumulation in terms of reducing stress biomarkers H2O2 by 25%, and EL by 12%. It has been revealed that the application of GB can minimize or reduce the toxic effects caused by Pb toxicity in both varieties, positively modulating plant growth performances and lowering oxidative stress. This research may provide a scientific basis for assessing Pb tolerance in barley plants and developing alternative approaches to protecting them from the severe effects of Pb toxicity.

Exogenous methyl jasmonate enhanced kenaf (Hibiscus cannabinus) tolerance against lead (Pb) toxicity by improving antioxidant capacity and osmoregulators.

In this study, the effects of exogenous methyl jasmonate (MeJA) on metal uptake and its ability to attenuate metal toxicity in kenaf plants under Pb stress were investigated. The experiment was conducted with five different MeJA concentrations (0, 40, 80, 160, and 320 µM) as a foilar application to kenaf plants exposed to 200 µM Pb stress. The results revealed that pretreatmen of MeJA significantly increased plant dry weight, plant height, and root architecture at all concentrations tested, with the most significant increase at 320 µM. Foliar application of MeJA at 160 µM and 320 µM increased the Pb concentrations in leaves and stems as well as the translocation factor (TF) from root to leaf. However, the bioaccumulation factor in the shoot initially decreased and then increased with increasing MeJA concentration. By increasing enzymatic (SOD, POD, and CAT) and non-enzymatic (AsA and non-protein thiols) antioxidants, MeJA pretreatment decreased lipid peroxidation, O2- and H2O2 accumulation and recovered photosynthetic pigment content under Pb stress. Increased osmolytes (proline, sugar, and starch) and protein content after MeJA pretreatment under Pb stress restore cellular homeostasis and improved kenaf tolerance. Our results suggest that MeJA pretreatment modifies the antioxidant machinery of kenaf and inhibits stress-related processes that cause lipid peroxidation, hence enhancing plant tolerance to Pb stress.

Lead Tolerance and Remediation Potential of Four Indocalamus Species in Lead-Contaminated Soil.

Indocalamus plants are low-growing shrubby bamboos with growth advantages, such as high biomass and strong resistance, and they are rich in germplasm resources in southern China. This study conducted soil lead (Pb) stress experiments on Indocalamus latifolius (Keng) McClure (LA), Indocalamus hunanensis B.M. Yang (HU), Indocalamus chishuiensis Y.L. Yang and Hsueh (CH) and Indocalamus lacunosus Wen (LC). Five Pb treatments (0, 500, 1000, 1500 mg·kg-1 Pb, and 1000 mg·kg-1 Pb + 1000 mg·kg-1 ethylenediamine tetraacetic acid (EDTA)) were established. EDTA was applied to explore the tolerance mechanism of different Indocalamus species after absorbing large amounts of heavy metals. The results were as follows: (1) under Pb treatment, the total relative biomass of LA, HU and LC was <100%, whereas the total relative biomass of CH was >100%; (2) after applying EDTA, the bioconcentration coefficient, translocation factor, and free proline content of the four Indocalamus species increased; and (3) the Pb mobility and distribution rates of the underground parts of the four Indocalamus species were consistently greater than those of the aboveground parts. The Pb mobility and distribution rates in the stems increased after applying EDTA, while those in the leaves decreased, as the plants tended to transfer Pb to their stems, which have lower physiological activity than their leaves.

Isolation and screening of phosphorus solubilizing bacteria from saline alkali soil and their potential for Pb pollution remediation.

The high pH and salinity of saline alkali soil not only seriously restrict the growth of crops, but also aggravate the pollution of heavy metals. The fixation of heavy metals and the regulation of pH by phosphorus solubilizing microorganisms may become a new way to repair heavy mental and improve saline alkali soil. In this study, a saline-alkali resistant bacteria (CZ-B1, CGMCC No: 1.19458) was screened from saline-alkali soil, and its tolerance to salt/alkali/lead stress was investigated by shaking flask experiment. The strain was identified as Bacillus amyloliquefaciens by morphology and 16S rRNA gene sequence analysis. The optimum growth temperature of CZ-B1 is about 35°C-40℃. The maximum salt stress and pH that it can tolerance are 100 g/L and 9 respectively, and its tolerance to Pb2+ can reach 2000 mg/L. The phosphorus release amount of CZ-B1 to Ca3(PO4)2 within 72 h is 91.00-102.73 mg/L. The phosphate solubilizing index in PVK agar medium and NBRIP agar medium are more than 2, which can be defined as phosphate solubilizing bacteria. Moreover, the dissolution of CZ-B1 to phosphorus is mainly attributed to tartaric acid, citric acid and succinic acid in inorganic medium. In addition, the removal rate of Pb2+ by CZ-B1 can reach 90.38% for 500 mg/L. This study found that CZ-B1 can immobilize Pb through three biological mechanisms (organic acid, extracellular polymers and mineralization reaction). The release of succinic acid (10.97 g/L) and citric acid (5.26 g/L) may be the main mechanism to promote the mineralization reaction of CZ-B1 (phosphate and oxalate) and resistance to Pb stress. In addition, the high enrichment of Pb2+ by EPS can increase the rate of extracellular electron transfer and accelerate the mineralization of CZ-B1. The screening and domestication of saline-tolerant phosphorus-solubilizing bacteria not only help to remediate Pb contamination in saline soils, but also can provide P element for plant growth in saline soil.

Application of exogenous glycinebetaine alleviates lead toxicity in pakchoi (Brassica chinensis L.) by promoting antioxidant enzymes and suppressing Pb accumulation.

Lead (Pb) poses an adverse effect on plant growth and development. Glycinebetaine (GB) plays an important role in plants response to stress environment. The study was performed to examine the potential of exogenous GB (0.5, 1, 2, and 5 mM) in alleviating Pb toxicity, the physiological and biochemical responses in pakchoi under 100 µM Pb stress by hydroponic experiment. Pb stress significantly decreased the growth, contents of pigment and mineral nutrient, and activities of antioxidative enzymes (CAT, SOD, and APX) in roots and shoots of pakchoi, while it caused a significant increase in Pb and ROS accumulation both in roots and shoots of pakchoi in comparison to the control. Exogenous application of GB improved leaf and root length, fresh and dry weight, mineral nutrient, and pigment contents of pakchoi under Pb stress. GB also effectively enhanced antioxidative enzyme activities and the accumulation of proline, soluble sugar, and GB and reduced the oxidative stress and Pb contents in shoots and roots of pakchoi. Principle component analysis (PCA) provided useful information on the classification of Pb tolerance according to the response to GB. Overall, the 1 mM GB was more effective to ameliorate the detrimental impacts of Pb stress. These findings suggested that GB application might be considered an effective strategy for alleviating Pb toxicity and enhancing the tolerance of pakchoi plants under Pb stress.

Lead Responses and Tolerance Mechanisms of Koelreuteria paniculata: A Newly Potential Plant for Sustainable Phytoremediation of Pb-Contaminated Soil.

Phytoremediation could be an alternative strategy for lead (Pb) contamination. K. paniculata has been reported as a newly potential plant for sustainable phytoremediation of Pb-contaminated soil. Physiological indexes, enrichment accumulation characteristics, Pb subcellular distribution and microstructure of K. paniculata were carefully studied at different levels of Pb stress (0-1200 mg/L). The results showed that plant growth increased up to 123.8% and 112.7%, relative to the control group when Pb stress was 200 mg/L and 400 mg/L, respectively. However, the average height and biomass of K. paniculata decrease when the Pb stress continues to increase. In all treatment groups, the accumulation of Pb in plant organs showed a trend of root > stem > leaf, and Pb accumulation reached 81.31%~86.69% in the root. Chlorophyll content and chlorophyll a/b showed a rising trend and then fell with increasing Pb stress. Catalase (CAT) and peroxidase (POD) activity showed a positive trend followed by a negative decline, while superoxide dismutase (SOD) activity significantly increased with increasing levels of Pb exposure stress. Transmission electron microscopy (TEM) showed that Pb accumulates in the inactive metabolic regions (cell walls and vesicles) in roots and stems, which may be the main mechanism for plants to reduce Pb biotoxicity. Fourier transform infrared spectroscopy (FTIR) showed that Pb stress increased the content of intracellular -OH and -COOH functional groups. Through organic acids, polysaccharides, proteins and other compounds bound to Pb, the adaptation and tolerance of K. paniculata to Pb were enhanced. K. paniculata showed good phytoremediation potential and has broad application prospects for heavy metal-contaminated soil.

NTA-assisted mineral element and lead transportation in Eremochloa ophiuroides (Munro) Hack.

Lead (Pb) is one of the most toxic and harmful pollutants to the environment and human health. Centipedegrass (Eremochloa ophiuroides (Munro) Hack.), an excellent ground cover plant for urban plant communities, exhibits the outstanding lead tolerance and accumulation. Nitrilotriacetic acid (NTA) is an environmentally friendly chelating agent that strengthens phytoremediation. This study explored the effects of different NTA concentrations on the absorption and transportation of mineral elements and Pb in centipedegrass. Following exposure to Pb (500 µM) for 7 days in hydroponic nutrient solution, NTA increased root Mg, K, and Ca concentrations and shoot Fe, Cu, and Mg concentrations and significantly enhanced the translocation factors of mineral elements to the shoot. Although NTA notably decreased root Pb absorption and accumulation, it significantly enhanced Pb translocation factors, and the Pb TF value was the highest in the 2.0 mM NTA treatment. Furthermore, the shoot translocation of Pb and mineral elements was synergistic. NTA can support mineral element homeostasis and improve Pb translocation efficiency in centipedegrass. Regarding root radial transport, NTA (2.0 mM) significantly promoted Pb transport by the symplastic pathway under the treatments with low-temperature and metabolic inhibitors. Meanwhile, NTA increased apoplastic Pb transport at medium and high Pb concentrations (200-800 µM). NTA also enhanced the Pb radial transport efficiency in roots and thus assisted Pb translocation. The results of this study elucidate the effects of NTA on the absorption and transportation of mineral elements and Pb in plants and provide a theoretical basis for the practical application of the biodegradable chelating agent NTA in soil Pb remediation.

Transcriptomics-based analysis of genes related to lead stress and their expression in the roots of Pogonatherum crinitum.

Revealing plants' tolerance and transport genes to heavy metal stress play an important role in exploring the potential of phytoremediation. Taking the heavy metal lead (Pb) hyperaccumulator plant Pogonatherum crinitum (Thunb.) Kunth as the research object, a hydroponic simulation stress experiment was set up to determine the physiological indicators such as antioxidant enzymes and non-enzymatic antioxidants in the roots of P. crinitum under different Pb concentrations (0, 300, 500, 1000, 2000 mg·L-1). RNA-Seq was performed, the Unigenes obtained by transcriptome sequencing were enriched and annotated by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases, and the differential expression genes (DEGs) of root were screened and verified by quantitative real-time polymerase chain reaction (qRT-PCR). The results are as follows: with the increase of Pb concentration, superoxide dismutase (SOD), catalase (CAT), and ascorbic acid (AsA) content increased. Peroxidase (POD), malondialdehyde (MDA), and ascorbic acid-glutathione (AsA-GSH) cycles showed low promotion with high inhibition. A total of 38.21 Gb of bases were obtained by transcriptome sequencing, and the base quality of each sample reached Q20 and Q30, accounting for 90%, making the sequencing results reliable. Combined with transcriptome sequencing, functional annotation, and qRT-PCR validation results, 17 root Pb-tolerant genes of P. crinitum were screened out, which were related to antioxidation, transportation, and transcription functions. Moreover, qRT-PCR verification results under different Pb stress concentrations were consistent with the transcriptome sequencing results and changes in physiological indicators. In brief, the root of P. crinitum can adapt to the Pb stress environment by up-regulating the expression of related genes to regulate the physiological characteristics.